Aus dem Institut für Tierzucht und Tierhaltung

der Agrar- und Ernährungswissenschaftlichen Fakultät

der Christian-Albrechts-Universität zu Kiel

CAMPYLOBACTER SPP., YERSINIA SPP. AND

SALMONELLA SPP. AS ZOONOTIC PATHOGENS IN

PIG PRODUCTION

Dissertation

zur Erlangung des Doktorgrades der Agrar- und Ernährungswissenschaftlichen Fakultät

der Christian-Albrechts-Universität zu Kiel

vorgelegt von

Master of Science

TANJA WEHEBRINK

aus Rahden, Nordrhein-Westfalen

Dekan: Prof. Dr. Joachim Krieter Erster Berichterstatter: Prof. Dr. Joachim Krieter

Zweiter Berichterstatter: Prof. Dr. Edgar Schallenberger

Tag der mündlichen Prüfung: 3. Mai 2007

Die Dissertation wurde mit dankenswerter finanzieller Unterstützung der H. Wilhelm Schaumann Stiftung, dem Ministerium für Soziales, Gesundheit, Familie,

Jugend und Senioren des Landes Schleswig-Holstein und der Arbeitsgruppe Lebensmittelqualität und -sicherheit (QUASI) der Agrar- und Ernährungswissenschaftlichen

Fakultät der Christian-Albrechts-Universität zu Kiel angefertigt

TABLE OF CONTENTS

GENERAL INTRODUCTION

………………………………………………………………………………………………… 1

CHAPTER ONE

Campylobacter spp. und Yersinia spp. beim Schwein: ein Überblick

………………………………………………………………………………………………… 3

CHAPTER TWO

Prevalence of Campylobacter spp. and Yersinia spp. in the pig production

………………………………………………………………………………………………. 19

CHAPTER THREE

Campylobacter spp.: Risk factor analysis in fattening farms

………………………………………………………………………………………………. 37

CHAPTER FOUR

Simulation study on the epidemiology of Salmonella spp. in the pork supply chain

………………………………………………………………………………………………. 53

GENERAL DISCUSSION

………………………………………………………………………………………………. 73

GENERAL SUMMARY

………………………………………………………………………………………………. 85

ZUSAMMENFASSUNG

………………………………………………………………………………………………. 89

1

GENERAL INTRODUCTION

Any disease and/or infection which is naturally "transmissible from vertebrate animals to

man" is classified as a zooanthroponosis according to EU-directive 92/117 (1992). To date,

over 200 zooanthroponoses have been described, involving all types of agents bacteria,

parasites and viruses (Krauss et al., 2004). The main part of zoonotic agents is represented by

bacterial pathogens. Every year millions of people become sick because of food-borne

zoonoses such as salmonellosis, campylobacteriosis or yersiniosis causing fever, diarrhoea,

abdominal pain, malaise and nausea. Other bacterial zoonoses are: anthrax, brucellosis, E.

coli-infections, leptospirosis, plague, shigellosis and tularaemia. The second group of

zooanthroponoses causing pathogens are parasites. In Latin America for example, 100 out of

100,000 inhabitants are estimated to suffer from cysticercosis (World Health Organisation,

2007). Other parasitical zoonoses are echinococcosis/hydatidosis, toxoplasmosis and

trematodosis (Heeschen, 2005). The third class of zoonotic pathogens are viruses. Rabies is a

disease of carnivores and bats mainly transmitted to humans by bites. Almost all persons

severely exposed to rabid animals will die if left untreated. An estimated number of 55,000

persons, mainly children, die of this disease in the world every year (World Health

Organisation, 2007). Other viral zoonoses are avian influenza, Crimean-Congo haemorrhagic

fever, ebola and Rift Valley fever (Krauss et al., 2004).

As bacterial pathogens are mainly responsible for zoonoses the following thesis concentrates

on this important group, especially gram-negative enterobacteriaceae. Zooanthroponoses even

though the estimated number of unreported cases is much higher than of the reported ones.

These zooanthroponoses affect hundred thousands of people especially in developing

countries, although most of them can be prevented.

The aim of the present thesis was to contribute to a better understanding of the bacterial

zoonotic pathogens Campylobacter spp., Yersinia spp. and Salmonella spp. causing disease in

humans and animals and to use this information to assess and manage the risk to animals and

humans.

CHAPTER ONE summarises several studies emphasising the importance of Campylobacter

spp. and Yersinia spp. as widespread pathogens in the pig production chain. First, the

taxonomy and the pathogen character are described, and second, prevalence in the pig

production is reported.

2

The objective of CHAPTER TWO was to gather further information about the prevalence of

Campylobacter spp. and Yersinia spp. at different stages of the pig production chain via

cultural isolation. Samples were taken from sows, suckling piglets, growing and finishing

pigs, carcasses, raw meat, forage and their environment (separating plate, feeding trough).

A further purpose in CHAPTER THREE was to increase the knowledge about the sources of

infection from Campylobacter spp. and their qualitative and quantitative importance in pig

production. Analysis of the data from questionnaires from the corresponding pig farms

provided first indications of factors which may influence the prevalence of Campylobacter

spp. in herds.

CHAPTER FOUR includes an exploration of possible measures that can be implemented in

farrowing and fattening units to control the introduction and reduce the prevalence of

Salmonella in finishing pigs. A stochastic state-transition simulation model was established to

gather further information about the influence of the risk factors in the different pig

production stages on the Salmonella spp. prevalence in fattening pigs. Furthermore, the

influence of preventive arrangements of the immunisation of sows, and additionally, of

pathogen-free purchased gilts on the Salmonella spp. prevalence in the farrowing and

fattening unit were determined.

References

EU-directive 92/117 EWG des Rates vom 17. Dezember 1992 über Maßnahmen zum Schutz

gegen bestimmte Zoonosen bzw. ihre Erreger bei Tieren und Erzeugnissen tierischen

Ursprungs zur Verhütung lebensmittelbedingter Infektionen und Vergiftungen.

Amtsblatt L62 vom 15.3.1993.

Heeschen, W.H., 2005. Zoonosen und lebensmittelbedingte Erkrankungen. Systematische

Übersicht der wichtigsten Bakterien, Viren und Parasiten. Behr’s Verlag, Hamburg.

Krauss, H., Weber, A., Appel, M., Enders, B., Graevenitz, v.A., Isenberg, H.D., Schiefer,

H.G., Slenczka, W., Zahner, H., 2004. Zoonosen. Von Tier zu Mensch übertragbare

Infektionskrankheiten. 3. Auflage, Deutscher Ärzte-Verlag GmbH, Köln.

WHO, 2007. World Health Organisation, http://www.who.int/en.

3

Chapter One

Campylobacter spp. und Yersinia spp. beim Schwein:

ein Überblick

TANJA WEHEBRINK1, NICOLE KEMPER

1, ELISABETH GROSSE BEILAGE2

and JOACHIM KRIETER1

1Institute of Animal Breeding and Husbandry

Christian-Albrechts-University

D-24118 Kiel, Germany 2University of Veterinary Medicine Hannover

Fieldstation for Epidemiology

D-49456 Bakum, Germany

Accepted for publication in Züchtungskunde

4

1. Einleitung

Campylobacter spp.- und Yersinia enterocolitica- Infektionen zählen neben den Salmonella

spp.- Infektionen zu den häufigsten gemeldeten Infektionskrankheiten des Menschen, die

durch Lebensmittel übertragbar sind und Darminfektionen hervorrufen können. Im Jahr 2006

führten laut Robert Koch-Institut (2007) Infektionen durch Campylobacter spp. zu 51.764

Erkrankungsfällen in Deutschland. Bei Yersinia spp. lag die Höhe der Erkrankungsfälle im

gleichem Jahr bei 5.135. Besondere Bedeutung kommt hier vor allem den thermophilen

Spezies Campylobacter (C.) jejuni und C. coli zu, welche am häufigsten von an Enteritis

erkrankten Personen isoliert wurden. Yersinia (Y.) enterocolitica ist neben Y. pestis und Y.

pseudotuberculosis eine der drei humanpathogenen Yersinia-Spezies. Hier ist das Bioserovar

4/O:3 von besonderer Bedeutung, da dieses die Hauptursache humaner Yersiniosen im

europäischen Raum ist.

Beide Erkrankungen sind vor allem Kleinkindererkrankungen, bei der Campylobacteriose ist

eine zweite Erkrankungshäufung im frühen Erwachsenenalter zu erkennen. Hauptsächlich

äußern sich die Erkrankungen mit Durchfällen, aber auch schwere oder klinisch inapparente

Verläufe sind zu beobachten. Ebenso sind Spätfolgen, wie beispielsweise das Erythema

nodosum (Neubauer et al., 2001a), möglich. Bei Campylobacter spp. ist die geringe

Infektionsdosis von 500-800 Keimen noch hervorzuheben (Black et al., 1988). Für beide

Keime besteht seit Inkrafttreten des Infektionsschutzgesetzes im Jahr 2001 Meldepflicht. Da

beide Infektionen beim Schwein in der Regel symptomlos verlaufen (Bätza, 1996) und somit

weder im Bestand noch auf dem Schlachthof bei der Schlachttier- und Fleischuntersuchung

erkannt werden, ist es möglich, dass diese Zooanthroponosenerreger in die Lebensmittelkette

gelangen.

Der vorliegende Artikel liefert eine Literaturzusammenfassung über diese zwei wichtigen

Zooanthroponosenerreger, verdeutlicht die Erregereigenschaften und liefert einen Überblick

über die Prävalenzen in der Schweineproduktion.

2. Geschichte und Taxonomie

2.2 Campylobacter-Spezies

Der Kinderarzt Theodor Escherich beschrieb 1886 spiralig gewundene Bakterien, welche er

aus dem Darminhalt von Säuglingen mit Diarrhoe isoliert hatte. Zwei Jahre später gelang ihm

die Isolierung von ebenfalls spiralförmigen Darmbakterien von an Durchfall erkrankten

Katzen, welche er Vibrio felinus nannte (Escherich, 1886). Im Jahre 1919 wurden diese

5

Bakterien auch bei abortierten Rinderfeten nachgewiesen und als Vibrio fetus bezeichnet

(Smith und Taylor, 1919). Jones et al. (1931) fanden Vibrionen bei an Winterdysenterie

erkrankten Kälbern und nannten sie aufgrund ihrer Ähnlichkeit zu Vibrio fetus, aber dem

Vorhandensein von andere Antigeneigenschaften, Vibrio jejuni. Weitere mikroaerophile

Mikroorganismen wurden im Colon von dysenterischen Schweinen gefunden und wegen ihrer

Vibrionenähnlichkeit mit der Bezeichnung Vibrio coli versehen (Doyle, 1944). King fand

1957 zwei Gruppen von Vibrionen in Blutkulturen von Patienten, die an einer

hämorrhagischen Darmentzündung erkrankt waren. Die eine Gruppe war Vibrio fetus sehr

ähnlich, die andere Gruppe beschrieb er als „related Vibrios“. Sebald und Veron (1963)

stellten fest, dass sich die DNA dieser beiden Gruppen von der DNA der Gattung Vibrio im

Guanin- und Cytosingehalt unterschied. Aufgrund dieser Erkenntnis wurde dieser neuen

Spezies der Genusname Campylobacter gegeben, der aus dem Griechischen stammt und

„gebogener Stab“ bedeutet. Die ersten Isolierungen aus Stuhlproben von an Durchfall

erkrankten Patienten gelangen Anfang der siebziger Jahre (Butzler et al., 1973). Die

Entwicklung verbesserter Nachweisverfahren für Campylobacter spp. führte zunehmend zu

einer weltweiten Wahrnehmung insbesondere von C. coli und C. jejuni als bakterielle

Enteritiserreger beim Menschen (Kist, 2002).

2.3 Yersinia-Spezies

Im Jahre 1934 wurde die erste anerkannte Beschreibung von Yersinia enterocolitica in den

USA durch MCiver und Pike (1934) verfasst. Sie berichteten unter den Namen

Flavobacterium pseudomallei über einen kleinen gramnegativen Kokkobazillus, welcher aus

zwei Gesichtsabzessen einer Farmbewohnerin isoliert worden war. Sie hielten es aber für

wahrscheinlicher, es mit einer atypischen Form eines bereits bekannten Erregers zu tun zu

haben als mit einer neuen Spezies. Fünf Jahre später schenkten Schleifstein und Coleman

(1939) der Beschreibung von MCiver und Pike Beachtung, als sie einen Keim untersuchten,

der Ähnlichkeit mit Actinobacillus lignieresii und Pasteurella pseudotuberculosis hatte. Der

Keim wurde aus dem Darminhalt isoliert, deshalb schlugen sie den Name Bacterium

enterocoliticum vor. Der Gattungsname Yersinia wurde im Jahre 1944 durch Van Loghem zu

Ehren von Alexandre Yersin, welcher 1894 in Hongkong während einer Pestepidemie den

Erreger der Pest (ehemals Pasteurella pestis, heute Yersinia pestis) entdeckte, begründet. Im

Jahre 1964 wurde das Bacterium enterocoliticum in Y. enterocolitica umbenannt und in die

Familie der Enterobacteriaceae eingegliedert (Fredriksen, 1964). Im Jahr 1980 wurden vier

Yersinia-Spezies etabliert: Y. enterocolitica, Y. intermedia, Y. frederiksenii und Y. kristensinii

6

(Brenner et al., 1980). Um eine Einteilung hinsichtlich Pathogenität und Epidemiologie der Y.

enterocolitica-Isolate zu erhalten, wurde eine Zuordnung zu Biovaren geschaffen. Wauters et

al. (1987) nahmen, aufgrund unterschiedlicher Substratverwertung, die Einteilung in sechs

Biovare vor: 1A, 1B, sowie Biovar 2 bis 5. Biovar 1A fasst den überwiegenden Teil der bis

dahin als apathogenen eingeschätzten Isolate zusammen. Die Biovare 2, 3, 4 und 5 enthalten

die pathogenen europäischen, die Biovare 1B die pathogenen in Amerika isolierten Stämme.

Die tierpathogenen Stämme gehören stets zu den Biotypen 3 oder 5 (Aleksic und Bockemühl,

1990). Zusätzlich zur Einteilung in Biovare wird in der Routinediagnostik eine Einteilung in

Serovare vorgenommen. Die Serotypisierung basiert überwiegend auf O-Antigenen

(Oberflächenantigen), seltener auf den H- (Geißel-) oder F- (Fimbrien) Antigen. Für Yersinia-

Spezies wurden bis heute 60 O- Gruppen gefunden, wovon 28 auf Y. enterocolitica entfallen

(Aleksic und Bockemühl, 1990). Während bestimmte O- Antigene bei verschiedenen Spezies

vorkommen, sind die H- Antigene Spezies-spezifisch und können daher auch zur direkten

Identifizierung der Yersinia - Arten herangezogen werden. Bislang wurden 18 H- Faktoren

bei Y. enterocolitica definiert. Hier sind bestimmte Kombinationen von H- Antigenfaktoren

signifikant für pathogene Serotypen und können bei der Unterscheidung pathogener und

apathogener Stämme hilfreich sein. Nach Befunden von Aleksic und Bockemühl (1990) sind

die pathogenen Serotypen O:3, O:9 und O:5,27 von Y. enterocolitica stets mit den H-

Antigenen a,b; a,b,c; a,b,c,v; a,c; c oder b,c kombiniert. Der H- Antigenkomplex H: b,e,f,i

kommt hingegen bei den fast ausschließlich in den USA auftretenden pathogenen Serovaren

O:8; O:4,32; O:18; O:20 und O:21 von Y. enterocolitica vor (Aleksic und Bockemühl, 1990).

3. Spezifische Eigenschaften

3.1 Campylobacter-Spezies

Nach Garrity et al. (2002) untergliedert sich die Familie der Campylobacteriaceae in Gattung

I Campylobacter, Gattung II Arcobacter und Gattung III Sulfurospirillum. Zur Zeit sind 16

Spezies und 6 Subspezies von Campylobacter spp. anerkannt (On et al., 2001). Die

humanpathogenen Campylobacter können in zwei Hauptgruppen eingeteilt werden: in die

Durchfallerreger wie C. jejuni, C. coli, C. lari, C. upsaliensis und in die Erreger

extraintestinaler Infektionen wie C. fetus (Hu und Kopecko, 2003).

Bakterien der Gattung Campylobacter sind gramnegative, schlanke, kommaförmige,

sporenlose Stäbchenbakterien, die ca. 0,2-0,5 µm breit und 0,5-5 µm lang sind (Rolle und

Mayr, 2002). Sie können eine oder mehrere helikale Windungen besitzen und maximal bis zu

7

acht µm Länge erreichen. Die Bildung kurzer Ketten ist ebenfalls möglich. Sie erscheinen

auch s-förmig und in älteren Kolonien können kokkoide Zellen auftreten. Charakteristisch ist

die korkenzieherartige Bewegung, die durch die polare monotriche Begeißelung entsteht.

Campylobacter spp. haben einen respiratorischen Stoffwechsel, verwerten Kohlenhydrate

weder fermentativ noch oxidativ (d.h. sie sind „asaccharolytisch“) und ernähren sich von

Zwischenprodukten aus dem Tricarbonsäurezyklus und von Aminosäuren (Anonymus, 1994).

Eisen wird ebenfalls von Campylobacter spp. als essentieller Nährstoff benötigt (PARK, 2002).

Die Vermehrung findet in Temperaturbereichen von 32°C bis 46°C bei mikroaerophilem

Klima mit ca. 5% O2, 10% CO2 und 85% N2 statt (Hunt et al., 2001). Vermutlich ist die

Mikroaerophilie auch ein Resultat der Adaption von Campylobacter spp. an die

atmosphärische Zustände im Darm von warmblütigen Tieren und Vögeln (Park, 2002). Die

minimale Wachstumstemperatur liegt bei thermophilen Campylobacter spp. bei 31°C bis

32°C. Unter 30°C sind die Keime nicht mehr wachstumsfähig. Somit ist eine Multiplikation

während der Handhabung oder Lagerung von Lebensmitteln bei Zimmertemperatur

ausgeschlossen (Jacobs-Reitsma, 2000). Die Ursache für die fehlende Vermehrung außerhalb

des Tierkörpers und unterhalb von 30°C kann möglicherweise an der fehlenden Produktion

von Kälteschockproteinen liegen (Parkhill et al., 2000). In kontaminierten Substraten haben

thermophile Campylobacter spp. bei niedrigen Temperaturen eine höhere Überlebensfähigkeit

als bei höheren Temperaturen. Während sie bei 4°C mehrere Wochen lebensfähig sind,

sterben sie bei Temperaturen von 55°C ab (Wundt und Kasper, 1982).

Die Keime sind sehr empfindlich gegenüber Trockenheit. Sie überleben nur kurze Zeit in

trockener Atmosphäre. Bei aW-Werten kleiner als 0,97 sterben die Keime schnell ab.

Lebensfähige Keime können nur von feuchten Oberflächen isoliert werden. Die Kombination

aus Temperatur und Luftfeuchtigkeit scheint eine essentielle Rolle für das Überleben der

Keime zu spielen (Doyle und Roman, 1982).

Der pH-Wert des umgebenden Milieus beeinflusst das Überleben von Campylobacter spp. in

Abhängigkeit von der Zeit und der Temperatur. Das pH-Optimum liegt bei Werten zwischen

6,5 und 7,5, das Maximum bei pH 9. Werte von über pH 9 und unter pH 4 führen zum

raschen Absterben, besonders bei höheren Temperaturen (Gill und Harris, 1983).

Campylobacter spp. lassen sich leicht durch ultraviolette Strahlen und Röntgenstrahlen

abtöten. Gegen UV-Strahlen ist C. jejuni empfindlicher als Escherichia coli und Y.

enterocolitica (Butler et al., 1987).

Thermophile Campylobacter spp. können unter schwierigen Umgebungsbedingungen einen

besonderen Zustand einnehmen, in dem sie lebensfähig aber nicht kultivierbar sind. Dieses

8

viable-but-nonculturable-Stadium (VBNC) ist auch bei anderen humanpathogenen Erregern

wie z.B. Escherichia coli, Salmonella enteritidis, Vibrio cholerae und Legionella

pneumophila bekannt (Tholozan et al., 1999). Einige Autoren beschreiben diese Form als eine

Art Schutzzustand, bei dem sich die Keime in einem Ruhestadium befinden und später, unter

besseren Bedingungen, wieder wachsen können. Andere Verfasser bezeichnen dieses Stadium

als eine degenerative Form des beginnenden Zelltodes. Die Bakterien bleiben in diesem

Zustand aber infektionsfähig.

Bei der Identifizierung der unterschiedlichen Campylobacter-Spezies wird zwischen

genotypischen und phänotypischen Methoden unterschieden (Nachamkin et al., 2000). Bei

den phänotypischen Methoden handelt es sich um relativ einfache, oft angewandte Tests, die

auf dem Nachweis von biochemischen Reaktionen, verschiedenen Wachstumsparametern,

Resistenzprofilen gegenüber Antibiotika und serologischen Verfahren beruhen. Jedoch

verhalten sich Campylobacter spp. biochemisch inert, was die Differenzierung und

Unterscheidung der Spezies erschwert. Die einzige Reaktion zur biochemischen Reaktion der

beiden Spezies C. jejuni und C. coli ist die Hippurathydrolyse. Genotypische Methoden

basieren auf dem Nachweis stabiler chromosonaler Unterschiede, die reproduzierbar und stark

diskriminierend sind. Vor allem für die Typisierung von Stämmen und für die

epidemiologische Fragestellung eignen sich diese Methoden gut.

3.2 Yersinia-Spezies

Yersinia ist eine Gattung innerhalb der Familie der Enterobacteriaceae und umfasst derzeit

elf verschiedene Spezies. Yersinia pestis, der Erreger des „schwarzen Todes", einer Infektion,

die im Mittelalter epidemisch auftrat, ist heute aus unseren Breitengraden verschwunden

(Kayser et al., 1993), während Y. pseudotuberculosis und vor allem Y. enterocolitica als

Erreger der menschlichen Yersiniose in den letzten Jahren zunehmend an Bedeutung

gewonnen hat (Bottone, 1999). Yersinia enterocolitica ist jedoch nicht ausschließlich als

humanpathogener Erreger einzustufen. Neben pathogenen Vertretern dieser Spezies existieren

noch eine Reihe von apathogenen Umweltisolaten, die diagnostisch abgegrenzt werden

müssen (Neubauer et al., 2001b).

Yersinia enterocolitica ist ein gramnegatives, fakultativ anaerobes, pleomorphes, peritrich

begeißeltes Stäbchenbakterium, das eine Länge von 1,0-5,0 µm erreicht. Yersinien sind

oxidasenegativ, katalasepositiv und reduzieren Nitrat und Nitrit (Aleksic und Bockemühl,

1990). Sie kommen ubiquitär vor und bilden keine Kapseln oder Sporen (Knapp, 1988).

9

Bei unter 28°C sind sie beweglich, darüber jedoch nicht, da die Geißeln in der Regel nur bei

Temperaturen unter 30°C gebildet werden (Rolle und Mayr, 2002). Yersinia enterocolitica ist

psychrotrop, das bedeutet, dass eine Vermehrung bei Kühlungstemperaturen bis 0°C möglich

ist. Die optimale Wachstumstemperatur beträgt +30°C, wobei die Obergrenze der

Vermehrungsfähigkeit bei +43°C liegt.

Zur Bestimmung des Serotys sind zwischenzeitlich kommerzielle Test erhältlich, die auf einer

Agglutinationsreaktion beruhen. In der Diagnostik des weltweit am häufigsten beim

Menschen isolierten Serovars Y. enterocolitica Serotyp O:3 ist eine biochemische

Charakterisierung zusätzlich zur Serotypisierung unumgänglich, um eine sichere Aussage

über die klinische Relevanz eines Isolates (insbesondere bei klinischem Material und

Umweltproben) treffen zu können, da dieses Serovar auch bei anderen verwandten Yersinien-

Spezies oder Stämme des Biotyps 1A anzutreffen ist (Hoofar und Holmvig, 1999). Die

bakteriologische Diagnostik pathogener Y. enterocolitica-Isolate ist bis heute mit hohem

zeitlichen Aufwand verbunden, und mögliche Diagnosen können aufgrund mangelnder

Spezifität und Sensitivität der zur Zeit verfügbaren Testsysteme immer nur unter Vorbehalt

gestellt werden oder bedürfen in ihrer Interpretation eines hohen Maßes an Expertise. Bis

heute ist trotz der hohen zoonotischen Bedeutung des Erregers keine einheitliche Methode

zum bakteriologischen Nachweis pathogener Y. enterocolitica-Isolate beschrieben.

Erschwerend kommt hinzu, dass allein die Vielzahl der bis heute beschriebenen

Untersuchungen zu widersprüchlichen Ergebnissen führt (Arnold, 2002).

4. Prävalenzen und epidemiologische Aspekte in der Schweineproduktion

4.1 Campylobacter spp. und Yersinia spp. beim Schwein

Thermophile Campylobacter spp. scheinen keine Bedeutung für Erkrankungen bei Schweinen

zu haben (Altekruse und Swerdlow, 2002). Der Keim ist wahrscheinlich der normalen

Darmflora zuzurechnen (Görgen et al., 1983). Beim Schwein ist C. coli die verbreitetste

Spezies mit Nachweisraten von bis zu 100%. Einzelne Untersuchungen zeigen aber auch, dass

C. jejuni in bestimmten Beständen sehr häufig isoliert werden kann (Young et al., 2000).

Viele Untersuchungen belegen, dass sich Campylobacter spp. häufig aus dem Kot gesunder

Schweine isolieren lassen. Zu diesem Ergebnis kam auch Gaull (2002), der bei der Beprobung

von Mast- und Schlachtschweinen Nachweisraten zwischen 70% und 93% ermittelten.

Ähnliche Prävalenzen stellte auch Weijtens (1996) in einer Studie fest, in der er

Mastschweine im Verlauf einer Mastperiode auf Campylobacter spp. untersuchte. Dabei

10

wurde bei 98% der elf Wochen alten Schweine Campylobacter spp. aus dem Kot isoliert,

wobei ein Rückgang auf 85% zum Zeitpunkt der Schlachtung zu verzeichnen war. Die

Ursache für den Rückgang liegt möglicherweise in der stabileren Darmflora älterer Tiere, die

das Wachstum von Campylobacter spp. behindert. Aber auch ein Futterwechsel während der

Mast könnte als Erklärung hierfür angeführt werden. Es konnte beobachtet werden, dass

einzelne Tiere an aufeinander folgenden Beprobungsterminen unterschiedliche

Untersuchungsergebnisse aufwiesen. Dass Schweine während der Mast eine Campylobacter-

Freiheit erlangen und sich anschließend reinfizieren, scheint jedoch unwahrscheinlich.

Vielmehr ist von einer intermittierenden Ausscheidung auszugehen, die auf einer heterogenen

Verteilung des Erregers infolge chemischer Anziehungskräfte beruht. Da sowohl

Untersuchungen von Weijtens (1996) und Gaull (2002) gezeigt haben, dass Schweine schon

zu Beginn der Mastperiode Campylobacter spp. im Kot aufweisen, ist der primäre

Infektionszeitpunkt bereits im Ferkelalter zu suchen. Sauen in Ferkelerzeugerbetrieben weisen

häufig hohe Infektionsraten von bis zu 100% auf und können durch erregerhaltige

Ausscheidung einen massiven Infektionsdruck in ihrer Umwelt aufbauen (Weijtens, 1996).

Während die Ferkel zum Zeitpunkt der Geburt noch Campylobacter-frei sind, steigen die

Prävalenzen schon in den ersten Lebenswochen erheblich an. Wenn auch die Aufstallung der

Ferkel zunächst einen Einfluss auf die Höhe der Prävalenz in den ersten Lebenswochen zu

haben scheint (Ferkel in Ställen mit Fußbodenheizung weisen geringere Belastungen auf),

relativieren sich die Unterschiede am Ende der Aufzuchtphase und erreichen Nachweisraten

von 90% und mehr (Gaull, 2002).

Das Schwein ist seit langem als Reservoir von humanpathogenen Yersinia enterocolitica der

Serovare O.3, O:9 und O:5,27 bekannt (Johannessen et al., 2000). Beim Schwein selbst tritt

die Yersiniose überwiegend bei Jungtieren klinisch apparent auf. Ältere Tiere gelten als

asymptomatische Träger des Keims (Neubauer et al., 2001b). Serologische Untersuchungen

in Norwegen zeigen, dass 86% der untersuchten reinen Mastbestände positiv waren,

wohingegen die Herdenprävalenz bei geschlossenem System mit 53,1% erheblich niedriger

lag (Skjerve et al., 1998). Die Yersinia spp.-Infektion wird durch Zukauf und anschließender

fäkal-orale Kontamination sowie durch infiziertes Sperma oder Abortmaterial nach

intrauteriner Infektion verbreitet. Nach Ansicht der Autoren ist das Transportfahrzeug eine

wichtige Kontaminationsquelle. Auch der Einsatz von Stroheinstreu birgt nach ihrer

Auffassung ein erhöhtes Risiko. Dagegen konnte nach ihrer Auswertung die Herdenprävalenz

durch den Einsatz einer Unterdruckventilation sowie einer manuellen Fütterung gesenkt

werden. Skjerve et al. (1998) kamen zu dem Schluss, dass die Risikominimierung für eine Y.

11

enterocolitica-Infektion durch die strikte Trennung von infizierten und nicht-infizierten

Beständen zu erreichen ist. Bottone (1997) isolierte von klinisch gesunden Schweinen

pathogene Y. enterocolitica. Dabei lag die Isolationsrate aus dem Rachen weit höher als jene

aus dem Kot. Im Jahr 2000 wurden in Süddeutschland an einem Schlachthof 50

Schlachtschweine untersucht, hierbei konnten in 60% der Tonsillen und 10% der Kotproben

Y. enterocolitica 4/O:3 nachgewiesen werden (Fredriksson-Ahomaa et al., 2000). Weitere

Daten, die aus Deutschland stammten, wurden 2004 erfasst. Dabei waren 45,5% der

untersuchten Schweinemastbestände in Bayern serologisch positiv (Hensel et al., 2004).

Verlaufsuntersuchungen vom Ferkel bis zum adulten Tier liegen bislang noch nicht vor, aber

es ist bekannt, dass bei Sauen viel seltener Y. enterocolitica aus den Tonsillen zu isolieren ist.

So untersuchten Korte et al. (2004) Tonsillen von Mastschweinen und Sauen von sieben

verschiedenen Schlachthöfen. Während bei den Mastschweinen 56% positiv waren, konnte

bei den Sauenproben nur bei 14% der Erreger nachgewiesen werden.

4.2 Campylobacter spp. und Yersinia spp. im Schweinefleisch

In Lebensmittelproben wurden in Deutschland im Jahr 2001 nach Mitteilung von elf

Bundesländern in einer von insgesamt 159 untersuchten Schweinefleischproben

Campylobacter spp. nachgewiesen. Von 16 Anlassproben wies keine ein positives Ergebnis

auf (Hartung, 2002). In den USA untersuchten Zhao et al. (2001) Fleischprodukte aus 59

Fleischtheken verschiedener Supermarktketten auf das Vorkommen von Campylobacter spp..

Dabei war in 1,7% der Proben vom Schwein der Erreger nachweisbar. Oosterom et al. (1985)

gehen davon aus, dass positive Campylobacter-Nachweise am Schlachtkörper weniger durch

den ursprünglichen Keimgehalt im Darm der Tiere hervorgerufen werden, sondern vielmehr

Kreuzkontaminationen durch Oberflächen und Arbeitsgeräte in der Schlachthalle darstellen.

Deutlich stärker als das Fleisch dieser Tierart sind ihre Lebern belastet. So konnten Kramer et

al. (2000) in 71,1% der Schweinelebern thermophile Campylobacter nachweisen. Als Ursache

für diese hohe Prävalenz vermuteten sie eine Kreuzkontamination, da die Lebern zu mehreren

Kilogramm in jeweils einem Paket unter Luftabschluss verpackt wurden.

Die Nachweisrate von Y. enterocolitica in rohem Schweinefleisch ist mit Ausnahme von

Schweinezungen und –innereien gering (Beer, 1995), die Prävalenz im Hackfleisch, für

welches in manchen Regionen Kopffleisch und Tonsillen verwendet werden, ist jedoch hoch

(Tauxe et al. 1987). Über das Vorkommen von Y. enterocolitica in hitzebehandelten

Schweinefleischprodukten liegen nur wenige Studien vor (Hank, 2003). Bisher wurden keine

pathogenen Stämme aus hitzebehandelten Produkten isoliert. Dennoch wurden apathogene Y.

12

enterocolitica Stämme nachgewiesen. Dies zeigt, dass bei mangelhafter Hygiene eine

Kreuzkontamination von rohen zu hitzebehandelten Produkten möglich ist.

5. Schlussfolgerung

Wie die gezeigten Studien verdeutlichen, sind Campylobacter spp. und Yersinia spp. Keime,

die in Schweinebeständen in Europa weit verbreitet sind und daher ein Risiko für die

Gesundheit des Menschen darstellen. Daher sind weitere infektionsepidemiologische Studien

notwendig, um das vom Schwein ausgehende Gefahrenpotential für die menschliche

Campylobacter spp.- und Yersinia spp. -Infektion abschätzen zu können. Hierbei sind

Langzeitstudien erforderlich, um offene Fragen bezüglich der Epidemiologie und der

Eintragsquellen beider Erreger zu klären. Auch fehlen Informationen über die

Erregerprävalenz in der gesamten Produktionskette beim Schwein. Diese sind notwendig, um

festzustellen, auf welcher Produktionsstufe eine Erregerbekämpfung sinnvoll ist, um den

Eintrag zu minimieren. Es sollte geklärt werden ob der Einsatz einer Impfung zur

Erregerreduktion auf Bestandsebene praktikabel ist, oder eine Änderung der Schlachttechnik

einen positiven Einfluss hat. Da Campylobacter-Keime in der Lage sind, den VBNC-Status

einzunehmen, kann die Frage des Überlebens des Erregers auf der

Schlachttierkörperoberfläche nicht völlig geklärt werden. Über die Mechanismen und

Bedeutung des VBNC-Status bei Campylobacter spp. sollten weitere Untersuchungen

vorgenommen werden. Zur Zeit gibt es weder für Campylobacter spp. noch für Yersinia spp.

einen „Gold Standard“ in der Analysetechnik. Dies erschwert die Vergleichbarkeit der

verschiedenen Studien untereinander. Somit ist zusätzlich die Entwicklung für eine sichere,

schnelle, einfach durchführbare und kostengünstige Erregerdiagnostik unabdingbar.

Zusammenfassung

Die vorliegende Arbeit diente als Literaturübersicht über Campylobacter spp. und Yersinia

spp. in der Schweineproduktionskette. Es wurde zum einen die Systematik und die

Erregereigenschaften dieser zwei weltweit bedeutenden Zooanthroponoserreger dargestellt.

Zum anderen wurde über die herrschende Prävalenzen in der Produktionskette beim Schwein

berichtet. Es wird deutlich, dass Schweine häufig Träger humanpathogener Campylobacter

spp. und Yersinia spp. sind und eine Kontamination ihres Fleisches während des

Schlachtprozesses möglich ist. Allerdings sind humanpathogene Campylobacter spp. und

Yersinien spp. relativ selten im Fleisch nachweisbar. Eine größere Gefahr stellen Innereien

13

dar. Um Schweinefleisch noch sicherer zu machen, sollte in Zukunft versucht werden, die

Epidemiologie des Erregers genauer aufzuklären, um somit die Ursache der

Erregerausbreitung zu erkennen und geeignete Gegenmaßnahmen ergreifen zu können.

Schlüsselwörter: Campylobacter spp., Yersinia spp., Schwein, Literaturübersicht

Abstract

This review summarises several studies emphasising the importance of Campylobacter spp.

and Yersinia spp. in the pig production chain as widespread pathogens. First, taxonomy and

pathogen character of these world-wide important pathogens were described, and second,

prevalence in the pig production was reported. Obviously, pigs are often carriers of

Campylobacter spp. and Yersinia spp. causing infections in humans. Contamination during

the slaughtering process is possible. However, pathogenic Campylobacter spp. and Yersinia

spp. are comparatively infrequently isolated from meat. A bigger health risk is represented by

entrails. Concluding, to increase pork safety, further epidemiological studies are urgently

needed to determine the origin of pathogens and to take counteractive measures.

keywords: Campylobacter spp., Yersinia spp., pigs, review article

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19

CHAPTER TWO

Prevalence of Campylobacter spp. and Yersinia spp. in the

pig production

TANJA WEHEBRINK1, NICOLE KEMPER

1, ELISABETH GROSSE BEILAGE2

JOACHIM KRIETER1

1Institute of Animal Breeding and Husbandry

Christian-Albrechts-University

D-24118 Kiel, Germany 2University of Veterinary Medicine Hannover

Fieldstation for Epidemiology

D-49456 Bakum, Germany

Submitted for publication in Berliner und Münchner Tierärztliche Wochenschrift

20

Abstract

The aim of this study was to determine the prevalence of Campylobacter spp. and Yersinia

spp. in a total of 1,040 faecal samples taken from animals at different ages from four

farrowing and twelve fattening herds. In the farrowing unit, faeces were collected from 68

sows (faecal samples) and 256 suckling piglets (rectal swab samples). Further samples were

collected from 362 growing and 354 finishing pigs (rectal swab samples). Additionally, 56

feed and environmental samples were collected.

During the slaughtering process, 122 pigs and their carcasses respectively, were sampled three

times. First, rectal samples were taken with swabs during the lairage. Second, the samples

were taken from the carcass before entering the chilling room. The same method was repeated

in the chilling room twelve hours after starting the chilling.

Finally, 86 raw meat samples were taken from 34 retail stores.

Campylobacter spp. were isolated in sows (33.8%), piglets (80.9%), growing (89.2%) and

finishing (64.7%) pigs. Yersinia spp. were detected in growing (15.2%) and finishing (13.3%)

pigs only. During lairage, Campylobacter spp. were identified from pig faeces from all farms

whereas Yersinia spp. were detected in pigs from just two herds. After twelve hours of

chilling neither Campylobacter spp. nor Yersinia spp. were detected. In raw meat samples,

Campylobacter spp. were isolated from one liver sample and Yersinia enterocolitica from two

meat samples (mince and cutlet). Common slaughter techniques and hygiene procedures may

be effective tools to reduce the risk of contamination and recontamination of meat products

since Campylobacter spp. and Yersinia spp. were found only sporadically in raw meat

samples.

keywords: Campylobacter spp., Yersinia spp., cultural isolation, pig production chain,

zoonotic pathogens

21

1. Introduction

Campylobacter (C.) spp. are among the most common bacterial causes of enteric diseases

worldwide. Members of the genus Campylobacter colonize the gastrointestinal tract of a

broad range of animals as commensals. In contrast, they are associated with disease in

humans. In Germany, the Robert Koch-Institute registered 61,823 cases of people suffering

from campylobacteriosis in 2005 (Robert Koch-Institut, 2006). Out of the 16 species within

the genus Campylobacter, the thermophilic species C. jejuni and C. coli are of special

importance as zoonotic agents with regard to human health. Infections with C. spp. in humans

are mainly related to consumption of contaminated food, especially chicken products.

Another important source of food-borne infections is raw or insufficiently cooked pork.

Furthermore, surface water used for drinking purposes can serve as a source of infection.

Besides Salmonella spp. and Campylobacter spp., Yersinia (Y.) spp. is another important

zoonotic pathogen from the list of human diseases (Aleksic and Bockemühl, 1990) with 5,600

registered infections in Germany in 2005 (Robert Koch-Institut, 2006). Together with Y.

pestis and Y. pseudotuberculosis, Y. enterocolitica represents pathogenic Yersinia species

with a certain risk to human health. Most cases of yersiniosis in Europe are related to

bioserovar 4/O:3 Y. enterocolitica is thought to be a significant food-borne pathogen,

although pathogenic isolates have been isolated from food infrequently, except from edible

pig offal (De Boer, 1995). In case-control studies, a correlation has been demonstrated

between the consumption of raw or undercooked pork and yersiniosis (Satterthwaite et al.,

1999). The main infection source for Y. enterocolitica bioserovar 4/O:3 is raw pig meat, for

pigs serving as natural carriers of this bioserovar (Fredriksson-Ahomaa et al., 2001).

Both infections are infant diseases with a clear infection peak in children up to two years and

a second incidence peak for campylobacteriosis in early adulthood. Diarrhoea is symptomatic

for both campylobacteriosis and yersiniosis, but severe or clinically unapparent courses of

disease are possible as well. In contrast to its importance as a human pathogen, the

understanding of the pathomechanisms of Campylobacter spp.-associated diseases is still

relatively poor (Vlient and Ketley, 2001). In the same way, this applies for the epidemiology

of Y. enterocolitica infections, as it is complex and poorly understood (Fredriksson-Ahomaa

and Korkeala, 2003).

The objective of this study was to gather further information about the prevalence of

Campylobacter spp. and Yersinia spp. at different stages of the pig production chain via

cultural isolation. Samples were taken from sows, suckling piglets, growing and finishing

pigs, carcasses, raw meat, forage and their environment (separating plate, feeding trough).

22

2. Materials and Methods

2.1 Materials

Table 1 shows the number of herds at every stage of the production chain and the number of

samples taken.

Table 1

Study design

production stage number

of samples farrowing units

4 herds

fattening units

12 herds

slaughterhouse

4 herds

retail

34 retail stores

sows 68 - - -

piglets 256 - - -

pigs - 716 366 -

forage 8 26 - -

environment 10 12 6 -

raw meat samples - - - 86

2.1.1 Farrowing and fattening units

During the period from November 2004 till June 2005 data were collected from four

farrowing and twelve fattening herds. The ZNVG (Vermarktungsgemeinschaft für Zucht- und

Nutzvieh, Neumünster) supplied a list of several farms. The herds for the present study were

selected based on the herd size and the relationships between farrowing and fattening. Due to

practical limitations, the study design was arranged in the following way: The number of

sows in the farrowing herds was between 150 to 650 sows and the fattening herds had

fattening places for 350 to 2000 animals. In three cases, a supply relationship between

farrowing and fattening unit existed. In all herds, pigs were kept under conventional

conditions.

The sampling size for each herd was calculated on the herd size and expected prevalence

according to the formula from Noordhuizen et al. (1997). The expected prevalence of

Campylobacter spp. and Yersinia spp., taken from literature, was appointed by the sows and

23

fattening pigs with 80% and 60% respectively and in the retails with 0.2% or rather 2%. The

absolute accuracy was 14% and the probability of error was 5%.

In the farrowing unit, faeces were collected in the farrowing house from 68 sows (faecal

samples) and 256 suckling piglets (rectal swab samples). In three herds, 17 sows and

additionally 85 or 86 suckling piglets (five or six piglets per litter) per herd were sampled and

in one herd only 17 sows. The selection of piglets was random and the time of sampling the

piglets was before weaning. Every pig was given a numbered ear tag enabling individual

identification at all times.

In the fattening unit, samples (rectal swab samples) were collected from 362 growing and 354

finishing pigs. In twelve herds, between 29 and 31 animals were sampled per herd. Eight pigs

died during the fattening period. The observation of 91 pigs from the farrowing unit could be

continued over the whole fattening period.

Additionally, 56 environmental and feed samples were collected in both production stages.

The environmental samples were taken from the separating plate and the feeding trough. Feed

samples consisted of forage for piglets, for sows in early and late pregnancy, and for pigs at

the beginning or end of fattening, respectively.

2.1.2 Slaughterhouse

All investigations concerning slaughter pigs and carcasses were carried out at a commercial

abattoir. The slaughterhouse was visited four times for samplings in the period from April

until June 2005. Four herds, sampled at different times, were the origin of the pigs

investigated at the slaughterhouse. Altogether, 122 pigs were sampled three times during the

slaughtering process. First, rectal samples were taken with swabs during the lairage. Second,

for the carcass surface, swabs moistened with a 0.9% NaCl dilution were used to sample an at

least 100cm2 large sampling field on the belly by rubbing with the necessary compression.

The samples were taken from the carcass before entering the chilling room. Third, the same

method was repeated in the chilling room twelve hours after beginning the chilling process.

During the slaughter process environmental samples were taken from diverse equipment

(knives, saws etc.).

Twenty-nine out of the 122 pigs at slaughterhouse level had also been sampled as piglets and

as fatting pigs representing a complete sampling passage at every step of the production

chain.

24

2.1.3 Retail

From June till July 2005, 86 raw meat samples were taken from 34 retail stores in two

different towns. In 13 butcher’s shops, twelve mince 13 escalope and two liver samples were

bought. In 21 discount shops, the sample material composed on the one hand of 16 mince, 19

escalope, one liver and one kidney portion from the self-service counter, and on the other

hand of eleven mince and eleven escalope samples from the sales counter.

2.2 Methods

After collection, samples were stored at 4°C and taken to the laboratory (Zentrale Einrichtung

Medizinaluntersuchungsamt und Krankenhaushygiene, Hygiene-Institut, Kiel) within four

hours and processed directly after arrival. Cultural methods were used to test all samples for

Campylobacter spp. and Yersinia spp., including differentiation of subspecies.

2.2.1 Detection of Campylobacter spp.

To isolate Campylobacter species, 1g of faeces or the swab sample was inoculated in 9ml

Preston broth (Oxoid). After incubation for 24 hours in a microaerophilic atmosphere (5%

oxygen, 10% carbon dioxide, 3% hydrogen and 82% nitrogen) at 37°C, a loop of the enriched

suspension was plated on Preston agar (Oxoid) and incubated for 48 hours under the above-

mentioned microaerobic conditions at 37°C. Campylobacter-like colonies were analysed by

Gram staining and catalase and oxidase tests (Hippurathydrolysis: ISO 10272, 1995,

modified), and biochemical reactions were assessed (ApiCampy; bioMerieux).

2.2.2 Detection of Yersinia spp.

Cultural isolation of Yersinia spp. was performed by adding 1g of faeces or the swab sample

to 9ml of Gram-negative broth (Becton & Dickinson) and incubating for 48 hours at 21°C.

One loop of broth was then plated on Yersinia-selective agar (Difco, CIN-Agar; CIN =

Cefsulodin-Irgasan-Novobiocin) and incubated for another 48 hours at 21°C. Colonies with

the typical bull’s eye appearance were subcultured on blood agar and Gram-stained and

biochemical tests were subsequently carried out by using API 20E (bioMerieux). Serum

agglutination was performed with isolates identified as Y. enterocolitica to detect serovars

O:3 and O:9 (ISO 10273, 1994, modified).

25

2.2.3 Statistical evaluation

Calculation of the animal prevalence and the 95% confidence intervals within the production

stage was performed with the PROC SURVEYMEANS procedure from the software package

SAS® (2002).

3. Results

3.1 Farrowing unit

In three of the four herds at the farrowing level, Campylobacter spp. was isolated from the

sow samples. Overall, Campylobacter spp. (total) were isolated in 33.8% (n = 23) of the sows

and in 80.9% (n = 207) of the piglets (Table 2). In six cases (2.3%), both pathogens, C. coli

and C. jejuni, were simultaneously isolated from the piglet samples and one sow (1.5%) was

infected with both subspecies too. No Yersinia spp. were detected in any of these samples in

the farrowing unit.

Table 2

Prevalence of Campylobacter spp. and Yersinia spp. in sows and suckling piglets

sows (n = 68) suckling piglets (n = 256)

% 95% C.I.1 % 95% C.I.

Campylobacter coli 30.9 19.6-42.1 71.1 65.5-76.7

Campylobacter jejuni 4.4 0-9.4 12.1 8.1-16.1

Campylobacter total2 33.8 22.3-45.4 80.9 76.0-85.7

Yersinia spp. 0 - 0 -

195% Confidence Interval 2Campylobacter total = C. coli and/or C. jejuni

Regarding the risks of vertical infection, Figure 1 points out that an infected sow does not

necessarily lead to infected piglets or that an uninfected sow automatically means a pathogen-

free piglet. For example, in herd ‘3’ sows (n = 17) were free from Campylobacter spp. and

Yersinia spp., but in the piglets (n = 85) C. coli was isolated in 21.2% (n = 18) and C. jejuni in

36.5% (n = 31) of cases.

26

0

20

40

60

80

100

sow piglet sow piglet sow piglet sow

herd 1 herd 2 herd 3 herd 4

farrowing unit

pre

va

len

ce

(%

)C. coli C. jejuni C. total¹

1C. total = C. coli and/or C. jejuni

Figure 1

Prevalence of the different pathogens in farrowing unit section (total sampled: 17 sows and 85

piglets per farm)

Additionally, Figure 2 shows the relationship between infected or non-infected sows and their

piglets in detail on the basis of the litters. Out of the 68 regarded litters, in 1.5% of the cases

neither sows nor piglets were infected. In 23.5%, Campylobacter spp. was detected in sows

and the whole tested piglets per litter. Notable is the fact that non- infected sows have

nevertheless infected piglets so piglets from non-infected sows were positive for

Campylobacter spp..

0

5

10

15

20

25

30

35

0 1 2 3 4 5 6

number of infected piglets

rel.

fre

qu

en

cy

(%

)

infected sow not infected sow

Figure 2

Relationship between Campylobacter total (C. coli and/or C. jejuni) infected sows and

infected piglets (n = 68 litters)

27

Campylobacter spp. and Yersinia spp. were not isolated neither in feed nor in environmental

samples.

3.2 Fattening unit

Campylobacter spp. were detected in all herds in growing and finishing pigs. Yersinia

enterocolitica O:3 were detected in the faeces of growing pigs in three of the twelve herds

only. Yersinia spp. were isolated in finishing pigs in six of the twelve herds. The prevalence

of Campylobacter spp. (total) and Yersinia spp. (total) in growing pigs were 89.2% (n = 323)

and 15.2% (n = 55), respectively (Table 3). While the prevalence of Campylobacter spp. was

slightly lower (64.7%; n = 229) in finishing pigs, that of Yersinia spp. was nearly the same

(13.3%; n = 47) as in growing pigs. Table 3 shows the decrease in prevalence of C. coli and

C. jejuni and Yersinia spp. during the fattening period. Furthermore, it illustrates the minor

role of Yersinia spp. in fattening herds.

Table 3

Prevalence of Campylobacter spp. and Yersinia spp. in growing and fattening pigs

growing pigs (n = 362) finishing pigs (n = 354)

% 95% C.I.1 % 95% C.I.

Campylobacter coli 71.3 66.6-76.0 28.0 23.3-32.7

Campylobacter jejuni 25.7 21.2-30.2 42.1 36.9-47.3

Campylobacter total2 89.2 86.0-92.4 64.7 59.4-69.4

Yersinia enterocolitica O:3 11.1 7.8-14.3 12.2 8.7-15.6

Yersinia enterocolitica O:9 0.3 0-0.8 0 -

Yersinia paratuberculosis 0 - 1.1 0-2.2

Yersinia enterocolitica 3.9 1.9-5.9 0 -

Yersinia total3 15.2 11.5-18.9 13.3 9.7-16.8

195% Confidence Interval 2Campylobacter total = C. coli and/or C. jejuni

3Yersinia total = Y. enterocolitica O:3 and/or Y. enterocolitica O:9 and/or

Y. paratuberculosis and/or Y. enterocolitica

In 28 cases (7.7%), both pathogens, C. coli and C. jejuni, were simultaneously isolated from

the piglet samples at the beginning of the fattening period. Twenty finishing pigs (5.6%) were

infected with both subspecies, too.

28

Campylobacter spp. total (C. coli and/or C. jejuni) was detected at both sampling times from

206 (28.8%) pigs and Yersinia spp. total (Y. enterocolitica O:3 and/or Y. enterocolitica O:9

and/or Y. paratuberculosis and/or Y. enterocolitica) from two pigs (0.3%) during the whole

fattening period.

Campylobacter spp. and Yersinia spp. were not isolated neither in feed nor in environmental

samples.

3.3 Slaughterhouse

During lairage, Campylobacter spp. were isolated from faeces of pigs (n = 68) from all farms

but Yersinia spp. were detected in pigs (n = 7) from two herds only. Before chilling

Campylobacter spp. were isolated from swabs taken from the carcass surface of pigs (n = 24)

from three farms. Yersinia spp. were detected in pigs (n = 1) from only one herd. After twelve

hours of chilling, neither Campylobacter spp. nor Yersinia spp. were isolated from swabs.

The prevalence of Campylobacter spp. (total) decreased during the three sampling phases

from 55.7% (lairage) to 19.7% (before chilling) to 0% (after 12 h chilling), and those of

Yersinia (total) fell from 5.7% to 0.8% to 0% (Table 4).

Table 4

Prevalence of Campylobacter spp. and Yersinia spp. in the slaughterhouse

lairage before chilling after chilling n = 122

% 95% C.I.1 % 95% C.I. % 95% C.I.

Campylobacter coli 27.9 19.8-35.9 10.7 5.1-16.2 0 -

Campylobacter jejuni 36.9 28.2-45.6 9.8 4.5-15.2 0 -

Campylobacter total2 55.7 46.8-64.7 19.7 12.5-26.8 0 -

Yersinia enterocolitica O:3 5.7 1.6-9.9 0.8 0-2.4 0 -

Yersinia total3 5.7 1.6-9.9 0.8 0-2.4 0 -

195% Confidence Interval 2Campylobacter total = C. coli and/or C. jejuni

3Yersinia total = Y. enterocolitica O:3 and/or Y. enterocolitica O:9 and/or

Y. paratuberculosis and/or Y. enterocolitica

In lairage, eleven pigs (9.0%) were carriers of C. coli and C. jejuni and before chilling one

animal (0.8%). During both sampling times (lairage and before chilling), Campylobacter total

29

was detected in 13 pigs (5.3%). Campylobacter spp. and Yersinia spp. were not isolated in

equipment samples.

3.4 The production chain from the piglet to the carcass after chilling

From 91 pigs, it was possible to obtain information from the farrowing to the fattening unit.

Out of these, data from 29 animals were acquired at the different stages of the pig production

chain. Figure 3 illustrates the declining tendency of Campylobacter spp. in the whole

production chain and the low prevalence of Yersinia spp. in fattening herds.

0

20

40

60

80

100

piglets growing

pigs

finishing

pigs

lairage³ before

chilling³

after

chilling³

production chain

pre

va

len

ce

(%

)

C. total¹ Y. total²

1C. total = C. coli and/or C. jejuni

2Y. total = Y. enterocolitica O:3 and/or Y. enterocolitica O:9 and/or

Y. paratuberculosis and/or Y. enterocolitica 3n = 29

Figure 3

Prevalence of the different pathogens in the whole pig production chain (n = 91)

In the farrowing unit, C. coli was detected in 69.2% (n = 63) and C. jejuni in 12.1% (n = 11)

of cases.

During the fattening period, the prevalence of C. coli decreased from 54 growing pigs

(59.3%) to eleven finishing pigs (12.4%). The prevalence of C. jejuni rose from 15 growing

pigs (16.5%) to 43 finishing pigs (48.3%). In the same period, three growing pigs (3.3%) were

carriers of Y. enterocolitica and four finishing pigs (4.5%) were carriers of Y. enterocolitica

O:3.

30

In lairage, eleven pigs (37.9%) were infected with C. coli and five pigs (17.2%) with C.

jejuni. Additionally C. coli was detected on two carcasses before chilling (6.9%).

Only in four piglets (4.4%), four growing (4.4%) and five finishing pigs (5.5%) were both

pathogens C. coli and C. jejuni detected simultaneously.

From the 91 pigs, Campylobacter spp. total could be identified in 33 (36.3%) animals during

the farrowing and fattening unit. Seven pigs (24.1%) were carriers of Campylobacter spp.

total as piglet, growing and finishing pigs and as living pigs in lairage. From only one pig

could Campylobacter spp. be isolated in all steps of the production chain from piglet to

carcass before chilling.

3.5 Retail

Campylobacter coli was isolated from only one liver sample, and Y. enterocolitica, from two

meat samples (mince and cutlet). The pathogens could not be detected in the other 83

samples.

4. Discussion

The aim of this study was to gather further information about the prevalence of

Campylobacter spp. and Yersinia spp. at the different stages of the pig production chain by

using culture isolation methods.

The results from the farrowing unit point out that compared with sows (33.8%) the prevalence

of their piglets is very high (80.9%). Alter et al. (2005) did not find such a high detection rate.

Whereas no Campylobacter spp. was detectable in the faeces of piglets on the day of birth,

Campylobacter spp. incidence rose within seven days to 32.8%. After transfer to the nursery

unit, the prevalence increased to 56.6%. Jensen et al. (2006) detected high prevalence of

Campylobacter spp. in organic outdoor pigs. All pigs (n = 47) shed Campylobacter (103-107

CFUg-1 faeces) from the age of 8-13 weeks. C. jejuni was found in 29% of pigs in three

consecutive trails and always in minority to C. coli (0.3%-46%). On the basis of the results

from the present project, it becomes obvious that there is no relationship between infected

sows and the infection of their piglets with Campylobacter spp.. This fact clarifies that

sampling of sows alone is useless without taking the piglets into account. Yersinia spp. seems

to play a negligible role in farrowing herds. This is in accordance with another study detecting

Yersinia spp. only during the fattening period but not in sows and piglets (Kasimir, 2005).

The fact that Y. enterocolitica was not isolated in the farrowing unit but first at the beginning

31

of the fattening period is evidence that the cause of infection has to be looked for in the

fattening unit.

On the basis of the results in the fattening unit, it becomes obvious that a stable gut flora from

older pigs can cause a decrease in prevalence. Other studies e.g. from Weijtens et al. (1993;

1999) approve this effect. In this study, the amount of Campylobacter was at 104 cfu/g

excrement at the beginning of the fattening period and about 102 cfu/g excrement at the end of

the fattening period. Also Young et al. (2000) detected a higher prevalence in 14-day-old

piglets compared to gilts. In the present project, the detection rates from Y. enterocolitica in

growing and finishing pigs are moderate (15.2% vs. 13.3%). The low Yersinia-prevalence in

this production stage can be attributed to the persistence of Yersinia spp. in the palatine tonsil

and intermittent shedding. A robust gut flora in older pigs might be the reason for lower

pathogen prevalence due to competition. Pilon et al. (2000) sampled faeces from 20 different

farms. The prevalence of Y. enterocolitica were between 0% and 46.9%. Bush et al. (2003)

detected 12.8% Y. enterocolitica in 2664 faecal samples and Kasimir (2005) described

isolation rates between 0% and 65.4%. However, factors influencing the shedding of

pathogens can rarely be determined definitely, but pigs carrying certain pathogens are

consequently an infection source.

Neither in the environmental nor in the feed samples were Campylobacter spp. and Yersinia

spp. isolated. One reason therefore can be found in the method of detection. Especially for

environmental and animal feed samples, the cultivation method seems to be inferior compared

to Polymerase-Chain-Reaction (PCR), because the low numbers of pathogenic strains in these

samples can often be suppressed by a distinct satellite flora (Fredriksson-Ahomaa and

Korkeala, 2003).

Despite the high prevalence in lairage (Campylobacter total: 55.7%) at the slaughterhouse,

none of the examined pathogens was detected after chilling. Apparently, the chilling of

carcasses and the associated dehydration of the surface area reduce the number of

Campylobacter spp. In this way, an effective minimisation of the infection risk via the food

store chains is possible. But a residual risk attributed to the VBNC status (viable but non

culturable) status could not be denied, enabling certain strains to be still viable without being

identifiable through cultivation. Malakauskas et al. (2006) showed that 28 (63.6%) of the 44

samples collected at the slaughterhouse were contaminated by Campylobacter spp. 23.4% (28

of 120) isolates were identified as C. jejuni (19 from carcasses and nine from slaughter line

surface) and 76.6% (92 of 120) isolates as C. coli (28 from faeces, 47 from carcasses and 17

from slaughter line surfaces). The results suggest that cross-contamination originated in the

32

gastro-intestinal tract of the slaughtered pigs and that the cross-contamination happened

during the slaughter process (Malakauskas et al., 2006). The prevalence of Yersinia spp. in the

slaughterhouse were low (lairage: 5.7% vs. before chilling: 0.8%). One reason for this effect

is that Yersinia spp. persists in the tonsils and will be shed with the faeces discontinuously.

For consumer protection purposes it is noteworthy that in the present project C. coli was

isolated from one liver sample only. The prevalence of Yersinia spp. in raw meat samples

were very low, too. Further studies confirm this result. For example, Arnold et al. (2004)

detected the pathogen in 0.5% of mince samples and Fredriksson-Ahomaa et al. (2001) in

12%. A higher rate was detected only in samples from offal, tongues and palatine tonsils

(Fredriksson-Ahomaa et al., 2001). The low detection rate of Yersinia spp. in raw meat can

also be due to methodological difficulties. In food samples, analysed by cultivation methods

and PCR, the PCR technology recorded a higher prevalence (Fredriksson-Ahomaa and

Kokkeala 2003). Other detection methods are for example DNA hybridisation,

immunofluorescence tests and serotyping. In conclusion, it has to be stated that none of the

methodologies published hitherto is sufficient regarding the reliable detection of pathogenic

Yersinia spp. strains. Therefore, only conditionally fast and safe enrichment and cultivation

methods are available at the moment to detect yersiniosis. With regard to hygiene, one major

point of concern is the ability of Yersinia spp. to survive in raw meat for a long time because

they are viable at temperatures of 4°C. Lack of reasonable care in kitchen hygiene, especially

in private households, can easily lead to cross-contaminations.

Besides C. coli, C. jejuni was laboratory-confirmed in this examination. The isolation of C.

jejuni from pig samples was described by other studies as well. For example, Stich-Groh

(1982) and Young et al. (2000) identified 23.4% and 76.3% respectively, Campylobacter spp.

as C. jejuni. In these assays, hippurathydrolysis served as a confirmation method. This

technique is based on the ability of C. jejuni to hydrolyse hippurat, a biochemical reaction C.

coli is not capable of. One major problem of this method is the possible loss of this ability

during the life span of C. jejuni, causing false positive results with regard to C. coli. But it can

be possible that in some farms or in geographical regions C. jejuni is described as common in

pigs (Kasimir, 2005).

5. Conclusion

The aim of this study was to analyse the prevalence of Campylobacter spp. and Yersinia spp.

at the different stages of the pig production chain via cultural examination. Samples were

33

taken from sows, suckling piglets, growing and finishing pigs, carcasses, raw meat, forage

and their environment (separating plate, feeding trough).

High prevalence of Campylobacter spp. were found in suckling, growing and finishing pigs.

The observed prevalence from the farrowing unit confirm the conclusion that a pathogen-free

sow does not necessarily mean pathogen-free piglets. Yersinia spp. infections in farrowing

units can be neglected. Additionally it can be pointed out that the prevalence of both

pathogens decrease with the increasing age of animals in the fattening unit. The fact that both

examined pathogens were found only sporadically in food indicates that common slaughter

techniques and hygiene procedures are effective tools to reduce the risks for contamination or

recontamination of meat products.

The most important risk factors responsible for the spread of Campylobacter spp. and

Yersinia spp. in the farrowing and fattening unit should be identified in further studies.

Acknowledgements

This research was financially supported by the H. Wilhelm Schaumann Stiftung, the

Ministerium für Soziales, Gesundheit, Familie, Jugend und Senioren des Landes Schleswig-

Holstein and the Arbeitsgruppe Lebensmittelqualität und -sicherheit (QUASI) from the

Faculty of Agricultural and Nutritional Science, Christian-Albrechts-University, Kiel.

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specific one-step PCR-assay fort he rapid discrimination of enteropathogenic Yersinia

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Yersinia enterocolitica und Campylobacter spp. in Schweinebeständen von der Geburt

bis zur Schlachtung sowie Genotypisierung ausgewählter Isolate. (Diss. med. vet.).

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Pilon, J., Higgins, R., Quessy, S., 2000. Epidemiological study of Yersinia enterocolitica in

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Weijtens, M.J.B., Bijker P.G., van der Plas, J., Urlings, H.A., Biesheuvel, M.H., 1993.

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36

37

CHAPTER THREE

Campylobacter spp.: Risk factor analysis in fattening pig

farms

TANJA WEHEBRINK1, NICOLE KEMPER

1, ELISABETH GROSSE BEILAGE2

and JOACHIM KRIETER1

1Institute of Animal Breeding and Husbandry

Christian-Albrechts-University

D-24118 Kiel, Germany 2University of Veterinary Medicine Hannover

Fieldstation for Epidemiology

D-49456 Bakum, Germany

Accepted for publication in Archives of Animal Breeding

38

Abstract

There is a lack of information about the prevalence and origins of the important zoonotic

pathogen Campylobacter spp. in the different stages of the pig production chain. The aim of

this study was to gather further information about the sources of infection with

Campylobacter spp. and their qualitative and quantitative importance in pig production. For

statistical analysis, 1,040 results from the bacteriological examination for Campylobacter spp.

were evaluated with questionnaires from four farrowing and twelve fattening units. The

prevalence was determined via faeces and swab samples with regard to certain farm

production parameters. Thereby 30.8% of the sows and 80.9% of their piglets were carriers of

Campylobacter spp.. In the fattening unit, the prevalence at the beginning of the fattening

period was 89.2% and at the end 64.7%. As a result of the small sample size in the farrowing

unit it was not possible to perform a risk analysis which yielded significant conclusions. In the

fattening stage, the following risk factors had a significant effect (p≤0.05) on Campylobacter

spp. prevalence: sampling time, number of fattening places per herd, mixed farming, floor

space design, feed origin, antibacterial and anthelmintic treatment. These results show that

housing and management have a possible influence on the Campylobacter spp. prevalence

and should be investigated further.

keywords: Campylobacter coli / jejuni, pig, fattening units, risk analysis, odds ratio

Zusammenfassung

Titel der Arbeit: Campylobacter spp.: Risikoanalyse in Schweinemastbetrieben

Über die Prävalenzen und Eintragsquellen des Zoonosenerregers Campylobacter spp. in den

verschiedenen Produktionsstufen der Schweineerzeugung existieren bisher nur wenige

Informationen. Die vorliegende Studie soll zur Aufdeckung produktionsspezifischer

Risikofaktoren und ihrer Analyse hinsichtlich der qualitativen und quantitativen Bedeutung

beitragen. Für die statistische Analyse wurden 1.040 Ergebnisse der bakteriologischen

Untersuchung auf Campylobacter spp. im Zusammenhang mit den Informationen aus einem

Fragebogen aus vier Ferkelerzeuger- und zwölf Mastbetrieben ausgewertet. Die Prävalenzen

des Erregers wurden mit Hilfe von Kot- und Abstrichtupferproben vor dem Hintergrund

verschiedener Betriebsbedingungen ermittelt. Dabei wurden bei 33,8% der Sauen und bei

80,9% der Ferkel Campylobacter spp. nachgewiesen. In der Produktionsstufe Mast betrug die

Prävalenz am Mastanfang 89,2% und am Mastende 64,7%. Aufgrund des geringen

Datenmaterials konnte auf der Produktionsstufe Ferkelerzeugung keine Risikoanalyse

durchgeführt werden. Folgende Faktoren hatten auf den Mastbetrieben einen signifikanten

39

Einfluss (p≤0,05) auf die Campylobacter Prävalenz: Zeitpunkt der Probeentnahme, Anzahl

Mastplätze, Mischbetrieb, Bodengestaltung, Futterherkunft, Einstallbehandlung und

anthelminthische Behandlung. Die Ergebnisse veranschaulichen, dass eine Reduzierung der

Campylobacter spp. Prävalenz durch betriebliche Haltungs- und Managementfaktoren

möglich ist. Dieses Phänomen sollte weiter untersucht werden.

Schlüsselwörter: Campylobacter coli / jejuni, Schwein, Mastbetriebe, Risikoanalyse, Odds

Ratio

1. Introduction

Infections caused by Campylobacter spp. (C.) are prevalent worldwide. Campylobacter jejuni

and C. coli are by far the most common Campylobacter species infecting humans. Both

species are associated with clinically indistinguishable diarrhoea in humans (Nachamkin,

2003). In Germany, the Robert Koch-Institute registered 61,823 cases of humans suffering

from such an infection in 2005. However, C. jejuni is implicated in about 85% of the cases of

human campylobacteriosis, with the remaining cases being primarily caused by C. coli

(Friedman et al., 2000).

Campylobacter spp. are part of the normal gut microflora in many food-producing animal

species, including chickens, turkeys, swine, cattle and sheep (Blaser, 1997). For instance, C.

jejuni is more commonly isolated from chickens and cattle, while C. coli is more common

among swine (Young et al., 2000). Transmission to humans appears to occur primarily

through the consumption of contaminated poultry products, unpasteurised milk products and

meat products (Effler et al., 2001; Friedman et al., 2004). In addition to the consumption of

undercooked meat, cross-contamination to other food products may play a significant role in

the number of illnesses observed. The infective dose (number of organisms sufficient to cause

infection) in humans can be very low. Only 800 colony-forming units of specific strains can

lead to Campylobacter infection (Black, 1988).

According to the regulations of the “White Paper on Food Safety” (Europäisches Weissbuch

zur Lebensmittelsicherheit, 2000), the farmer and the participating manufacturing industry in

the food production should have the main responsibility for food safety. Now and in future,

this adds up to the demand for preventive measures in primary production following the

principle “from the producer to the consumer”. This leads to a consolidated need for the

detection of relations between pathogen prevalence in the herds and the herd management and

husbandry. Determination of various important entry routes and spreading factors provides

useful decision guidance for all production units in the meat production chain to minimise the

40

transmission of zoonotic pathogens. For these reasons, this study was conducted with the aim

to determine the prevalence of Campylobacter spp. in farrowing and fattening units by the

collection of faeces and rectal swabs. Further risk factors for the occurrence of

Campylobacter spp. in farrowing and fattening units should be observed via environmental

and feed samples from the checked herds and questionnaires in the corresponding pig farms.

2. Material and Methods

Four farrowing and twelve fattening farms provided the basis for the present study. The

sampling size on every farm was calculated according to the formula from Noordhuizen et al.

(1997). In total, 1.040 faecal or swab samples respectively from pigs of all ages from

farrowing and fattening units were analysed. Additionally, 56 environmental and feed

samples were collected.

Cultural methods were used to test all samples for Campylobacter spp., including the

differentiation of subspecies. The bacterial detection of Campylobacter spp. proceeds from

ISO 10272 (1995) with following biochemical differentiation of C. coli and C. jejuni.

Calculation of the intraherd and animal prevalence and the 95%-confidence intervals within

the production stage was performed with the PROC SURVEYMEANS procedure from SAS®

(2002).

On every farrowing and fattening farm, data collection was carried out with the aid of a

questionnaire. Besides the general farm information, detailed data about the housing system,

management, state of health and aspects of disease surveillance were acquired. In

consideration of the bacteriological results, these data contributed to a hazard analysis to

detect the origin and spread of Campylobacter spp. infections.

The statistical analysis was performed with a generalised linear model. At first the

management-specific parameters were tested respectively with the χ2-test regarding the

influence on the pathogen prevalence. Every parameter having a value p<0.3 in the χ2-test and

an adequate distribution was included in the generalised linear model. The GENMOD

procedure from the software package SAS® (2002) was reviewed for significance (p≤0.05).

For the estimation, a binomial distribution and a logistic link function (i.e. logistic regression)

were assumed. As a result of the small sample size in the farrowing unit, it was not possible to

perform a risk analysis which yielded significant conclusions. From the fattening unit, the

following fixed effects were considered in the model: sampling time (growing pigs, finishing

pigs), herd organisation (number of fattening places, mixed farming), housing system and

41

forage (floor space design, feed origin) and health (antibacterial and anthelmintic treatment).

The estimates (ê) from the risk factors were transformed into odds ratios (OR = exp (ê)) and

the 95%-confidence intervals were calculated. A low absolute frequency in the least sub

classes from some factors did not allow a statistical analysis with logistic regression. For the

factors having a p-value ≤0.05 in the χ2-test, the odds ratios and 95%-confidence intervals

were calculated separately.

3. Results

3.1 Prevalence

3.1.1 Sows and suckling pigs

Campylobacter (C.) spp. were isolated in 33.8% of the sows and in 80.9% of the piglets

(Figure 1). Neither pathogen was isolated from the environmental and feed samples.

30.9

4.4

33.8

71.1

12.1

80.9

0

20

40

60

80

100

C. coli C. jejuni C. total¹

pathogen

pre

va

len

ce

(%

)

sows (n = 68)

suckling pigs (n = 256)

1C. total = C. coli and/or C. jejuni

Figure 1

Prevalence of Campylobacter spp. in sows and suckling pigs (Prävalenz von Campylobacter

spp. bei Sauen und Saugferkeln)

Table 1 shows the prevalence of Campylobacter spp. in pigs of the farrowing unit at herd

level. Notable is the fact that in herd 4 no sows are carriers of the pathogen but some of their

piglets are. In herd 3, no piglets were sampled, therefore no results for this production stage

appear in Table 1.

42

Table 1

Prevalence of Campylobacter spp. in pigs of the farrowing unit at herd level (Prävalenz von

Campylobacter spp. in der Ferkelerzeugung auf Betriebsebene)

sows1 suckling pigs2

prevalence (%) 95% C.I.3 prevalence (%) 95% C.I.

herd 1 C. coli 23.5 1.1-46.0 96.5 92.5-100.0

C. jejuni - - - -

C. total4 23.5 1.1-46.0 96.5 92.5-100.0

herd 2 C. coli 94.1 81.6-100.0 95.3 90.8-99.9

C. jejuni - - - -

C. total 81.6 81.6-100.0 95.3 90.8-99.9

herd 3 C. coli 5.9 0-18.4 - -

C. jejuni 17.6 0-37.9 - -

C. total 17.6 0-37.9 - -

herd 4 C. coli not sampled not sampled 21.2 12.3-30.0

C. jejuni not sampled not sampled 36.5 26.0-46.9

C. total not sampled not sampled 50.6 39.7-61.4

1n = 17 per herd 2n = 85 or 86 per herd 395% Confidence Interval 4

C. total = C. coli and/or C. jejuni

3.1.2 Fattening pigs

The prevalence of Campylobacter spp. in growing pigs was 89.2% and in finishing pigs

slightly lower with 64.7% (Figure 2). Neither pathogen was isolated from the environmental

and feed samples.

43

71.3

25.7

89.2

28.0

42.1

64.7

0

20

40

60

80

100

C. coli C. jejuni C. total¹

pathogen

pre

va

len

ce

(%

)growing pigs (n = 362)

finishing pigs (n = 354)

1C. total = C. coli and/or C. jejuni

Figure 2

Prevalence of Campylobacter spp. in growing and finishing pigs (Prävalenz von

Campylobacter spp. am Mastanfang bzw. Mastende)

Campylobacter spp. were detected on all farms in growing and finishing pigs (Figure 3). Herd

10 was the farm with the lowest Campylobacter spp. prevalence (54.8% in growing pigs and

19.4% in finishing pigs). In herd 9, no growing pig was pathogen-free (n = 29). There was

still a high prevalence at the second sampling time in comparison to the other herds with

81.5%. Nearly the same results were achieved by herd 12 with 100% (n = 31) carriers of

Campylobacter spp. at the beginning of fattening period and 80.6% at the end of growing

time. In every herd the prevalence decreased from the first sampling time to the second. Only

in herd 3 did the prevalence increase from 75.9% to 86.2%.

44

0

20

40

60

80

100

1 2 3 4 5 6 7 8 9 10 11 12

herd2

pre

vale

nce (

%)

C. to

tal1

growing pigs finishing pigs

1C. total = C. coli and/or C. jejuni

2herd = 29 to 31 sampled pigs per herd

Figure 3

Prevalence of Campylobacter spp. in the fattening pigs at herd level (Prävalenz von

Campylobacter spp. bei Mastschweinen auf Betriebsebene)

3.2 Risk factors

For the statistical risk factor analysis in the fattening unit, 716 results from the bacteriological

examination were evaluated in context with the questionnaire data from the twelve fattening

herds. Twenty factors were tested regarding their influence on the prevalence of

Campylobacter. Significant effects were shown for the following factors: sampling time,

number of fattening places, mixed farming, floor space design, feed origin, antibacterial and

anthelmintic treatments (Table 2).

45

Table 2

Significant risk factor and further risk factors: fattening unit (Signifikante Risikofaktoren und

weitere Einflussfaktoren bei Mastschweinen)

risk factor p-value prevalence (%) OR1 95% C.I.2

date sampling time <.0001 growing pigs 89.2 4.64 3.11-6.93 finishing pigs 64.7 1 -

herd organisation number of fattening places 0.052 < 1000 places 80.0 1.44 1.00-2.08 > 1000 places 74.3 1 - mixed farming 0.015 stall separated 74.6 0.61 0.41-0.92 stall not separated 82.0 1 -

housing system and forage floor space design 0.001 fully slatted floor 74.4 0.35 0.20-0.95 <50% slatted floor 74.8 0.56 0.32-0.97 plan floor without bedding 84.7 1 - feed origin 0.001 own forage 70.3 0.41 0.24-0.68 purchase forage 79.4 1 -

health antibacterial treatment 0.028 yes 74.6 0.66 0.45-0.96 no 79.7 1 - anthelmintic treatment 0.003 yes 83.9 1.99 1.25-3.18 no 74.8 1 -

source3

own piglets 73.3 0.26 0.09-0.75 steadier farrowing herds 76.1 0.32 0.13-0.76 purchase breeding herds 90.3 1 -

feed consistency3

meal 70.3 0.63 0.42-0.96 granule 81.0 1.23 0.60-2.54 pellets 78.0 1 -

blank dwell time3

>10 days 90.5 3.53 1.82-6.86 <10 days 74.5 1 -

1odds ratio 295% Confidence Interval 3further risk factor in the fattening unit

46

Over the fattening period the Campylobacter spp. prevalence decreased. At the beginning the

odds ratio increased by a factor of 4.46 (Table 2).

The risk factor fattening places per herd was differentiated between farms size under 1000

pigs and alternatively over 1000 pigs. The bacteriological results show that pigs from farms

with less than 1000 fattening places had a prevalence of 80.0% and those from larger farms a

prevalence of 74.3%. The chance to isolate Campylobacter spp. from pigs from smaller herds

increased by a factor of 1.44.

Housing in separated stalls is another preventive influence. When the animals on mixed farms

were kept in separated stalls the chance of a positive bacteriological result decreased

(OR = 0.61).

Pigs which were kept on a plan floor without bedding had the highest prevalence in

comparison to the other flooring systems. In this housing system, the chance of obtaining a

positive result was highest.

An antibacterial treatment at the beginning of the fattening period was implemented on seven

herds. The following antibiotics were used for this treatment: Amoxicillin, Tetracycline and

Sulfonamide. The chance of a positive finding decreased when the animals were treated with

antibacterial substances during this time period (OR = 0.66).

On four herds, anthelmintics were used at the beginning of fattening period. The appliance of

Ivermectin, Flubendazol and Levamisolhydrochlorid was adopted for deworming. The chance

of obtaining a positive result rose by a factor of 1.99 when anthelmintics were administered.

Further risk factors ‘source of piglets’, ‘feed consistency’ and ‘blank dwell time’ had an

influence on the prevalence of Campylobacter spp., too. The chance of obtaining a positive

result from the bacteriological investigation was smaller from fattening pigs in a closed herd

system (OR = 0.26). Furthermore, the following cases were preventive: feeding meal

(OR = 0.63) instead of granule or pellets and blank dwell time under 10 days.

4. Discussion

The results from the present study prove that Campylobacter spp. are of increasing

importance in farrowing and fattening units: high prevalence of Campylobacter spp. were

found in suckling, growing and finishing pigs (Wehebrink, 2006). Other studies also confirm

these results (Kasimir, 2005; Gaull, 2002).

The occurrence of Campylobacter spp. in subsequent samples of pigs and sows was often

variable in this analysis. As known from further studies the Campylobacter spp. prevalence

may vary because the physiological status of the animal and external factors can influence the

47

intestinal flora. The ability of Campylobacter spp. to colonise the intestinal tract of pigs is

probably subject to the various factors influencing the colonisation resistance of the gut

(Ruckebusch et al., 1991). Furthermore, the virulence of the Campylobacter spp. strains

(re)infecting the pigs may also alter the bacteriological results (Weijtens et al., 1999).

The prevalence estimates on basis of bacterial findings must be questioned critically. Because

of the intermittent shedding at animal level the bacterial detection in faecal samples can create

a false image of the prevalence at herd level. Additionally, during sampling and laboratory

processing, the pathogen’s sensibility to environmental influences can decrease the detection

rate.

The bacteriological analysis showed that in some herds as far as 100% of the pigs had contact

with Campylobacter spp.. In contrast to Young et al. (2000), a successful abatement strategy

can be doubted due to high general prevalence and the infection of piglets during the first

weeks of life.

Based on the zoonotic directive (Nr. 2160/2003), a monitoring for Campylobacter spp. is

mandatory. It should take place at an adequate stage of the food chain. Control has to be

directed primarily at the prevention of colonisation of farm animals by means of the

implementation of Good Hygienic Practice (GHP), biosecurity measures and husbandry

practices incorporating Hazard Analysis Critical Control Point (HACCP) based on risk

management systems (Whyte et al., 2002). Because of this, the objective of this study was to

obtain more information about the risk factors influencing the prevalence of this pathogen. As

a result of the small sample size in the farrowing unit, it was not possible to perform a risk

analysis which yielded significant conclusions. In the fattening unit the attention was focused

additionally on risk factors which do not reach the significant limitation of the 5% probability

error because of the small sample size. Effects which exceeded the housing and management

factors were not acquired in the questionnaire and could not consequently be regarded in the

evaluation. Because of this the results should only be regarded as tendencies.

One important influencing factor could be the sampling time. Because of the steady state of

immunity the chance of a positive Campylobacter spp. result is higher in growing pigs than to

finishing pigs. Additionally, transport stress, changing the forage and status conflicts can raise

the faecal shedding of this pathogen in growing pigs.

In contrast to recent studies, risk factor analysis in the fattening unit demonstrated a

significant influence on the Campylobacter spp. detection rate for the ‘number of fattening

places’. The chance of obtaining a positive Campylobacter spp. result is higher when animals

are held in smaller herds (<1000 places). This result did not conform to Gaull (2002). He

48

detected that the factor ‘number of animals’ hardly has any influence on Campylobacter spp.-

positive animals.

Separating the herds in ‘mixed farming’ is a useful method to decrease pathogen transmission.

In contrast to our study, Boes et al. (2005) could not assert this effect: investigation of the

occurrence and diversity of C. jejuni infections in finisher pigs in herds with combined cattle

or poultry production and herds only producing pigs showed no evidence of transmission of

C. jejuni from cattle or poultry to pigs in mixed production herds. Herd prevalence of C.

jejuni was 8.3%, whereas C. jejuni and C. coli were isolated from 0.8% and 92.0% of pigs,

respectively. In mixed production herds, C. jejuni predominated in cattle (42.7%) and poultry

(31.6%), whereas C. jejuni was only isolated from 1.3% to 2.5% of pigs in these herds.

A lower Campylobacter spp. detection rate is not promoted by a plan floor without bedding

and purchase forage. One reason for the higher prevalence in housing systems with plan floor

is the intensive contact of the pigs with their faeces for a longer time. With regard to

purchased forage, the origin is often uncertain: whether the forage comes directly from the

forage producer or whether several forage chandlers are interposed, increasing the risk of

contamination, remains often unknown.

A further result from the questionnaire analysis was that an arranged antibacterial treatment

but no anthelmintic treatment was preventive against Campylobacter spp. infections. These

results must be questioned critically because it is not known first which health status in detail

can be found in the different herds and, second, what the antimicrobial resistance of

Campylobacter spp. is. Further studies will be needed to explain these two risk factors.

Despite the fact that forage in granule form is heated during the manufacturing process, the

chance of obtaining a positive Campylobacter spp. result rose by a factor of 1.23 in this form

of forage feeding.

The fact that a blank dwell time under ten days is better for the pathogen prevalence than a

blank dwell time over ten days can be related to recontamination after disinfection and

cleaning.

Other studies found risks factors which could not be proven in this study. For example, Gaull

(2002) discovered that a factor such as different ‘husbandry’ hardly has any influence on

Campylobacter spp.-positive animals. ‘Feed’ and ‘number of pig delivering farms’ are not

risk factors either (Weijtens et al., 1993). Schuppers et al. (2005) detected that important risk

factors contributing to the prevalence of resistance strains were shortened tails, lameness, skin

lesions, feed without whey, and ad libitum feeding. Multiple antimicrobial resistance was

more likely in farms which only partially used an all-in all-out system, or a continuous-flow

49

system compared to a strict all-in all-out animal-flow. Presence of lameness, ill-thrift, and

scratches at the shoulder in the herd also increased the odds for multiple resistance. Thus, the

results from Schuppers et al. (2005) showed that on finishing farms which maintained a good

herd health status and optimal farm management the prevalence of antimicrobial resistance

was also more favourable.

In the present study, only a few factors could be identified as potential risk factors. For further

clarification of risk factors comprehensive assessment and transmission devolution studies are

required.

Acknowledgement

This research was financially supported by the H. Wilhelm Schaumann Stiftung, the

Ministerium für Soziales, Gesundheit, Familie, Jugend und Senioren des Landes Schleswig-

Holstein and the Arbeitsgruppe Lebensmittelqualität und -sicherheit (QUASI) from the

Faculty of Agricultural and Nutritional Science, Christian-Albrechts-University, Kiel.

References

Black, R.E.; Levine, M.M., Clements, M.L., Hughes, T.P., Blaser, M.J., 1988. Experimental

Campylobacter jejuni infection in humans. J. Infect. Dis. 157 (3), 472-479.

Blaser, M.J., 1997. Epidemiologic and clinical features of Campylobacter jejuni infections. J.

Infect. Dis. 176 (Suppl. 2), 103-105.

Boes, J., Nersting, L., Nielsen, E.M., Kranker, S., Enøe, C., Wachmann, H.C., Baggesen,

D.L., 2005, Prevalence and Diversity of Campylobacter jejuni in Pig Herds on Farms

with and without Cattle or Poultry. J. Food Prot. 68, 722-727.

Effler, P., Ieong, M.C., Kimura, A., Nakata, M., Burr, R., Cremer, E., 2001. Sporadic

Campylobacter jejuni infections in Hawaii: associations with prior antibiotic use and

commercially prepared chicken. J. Infect. Dis.183 (7), 1152-1155.

Friedman, C.R., Neimann, J., Wegener, H.C., Tauxe, R.V., 2000. Epidemiology of

Campylobacter jejuni infections in the United States and other industrialized nations.

In: Nachamkin, I.; Blaser, M.J.: Campylobacter. 2nd ed. Washington, D.C., American

Society for Microbiology Press., 121-138.

50

Friedman, C.R., Hoekstra, R.M., Samuel, M., Marcus, R., Bender, J., Shiferaw, B., 2004. Risk

factors for sporadic Campylobacter infections in the United States: a case-control study

in FoodNet sites. Clin. Infect. Dis. 38 (Suppl. 3), 285-296.

Gaull, F., 2002. Vorkommen thermophiler Campylobacter spp. bei Schweinen im Betrieb und

auf dem Schlachthof, auf Putenschlachttierkörpern und in Lebensmitteln tierischen

Ursprungs – Typisierung der Isolate mit molekularbiologischen Fingerprintingmethoden

und Vergleich der Isolate untereinander und mit humanen Isolaten. (Diss. med. vet.).

Univ. Leipzig.

International Organization for Standardization, 1995. International Standard 10272.

Kasimir, S., 2005. Verlaufsuntersuchungen zum Vorkommen potentiell humanpathogener

Yersinia enterocolitica und Campylobacter spp. in Schweinebeständen von der Geburt

bis zur Schlachtung sowie Genotypisierung ausgewählter Isolate. (Diss. med. vet.).

Univ. Leipzig.

Nachamkin, I., 2003. Campylobacter and Arcobacter. In: Murray, P.R., Baron, E.J., Pfaller,

M.A., Jorgensen, J.H., Yolken, R.H.: Manual of clinical microbiology, ASM Press,

Washington, DC, 902-914.

Noordhuizen, M., Frankena, K., Graat, E., 1997. Animal health care and public health issues.

In: World Congress on Food Hygiene, The Hague/Netherlands, Proc., 59.

Robert Koch-Institut, 2006. Epidemiologisches Bulletin Nr. 3.

Ruckebusch, Y., Phaneuf, L.P., Dunlop, R., 1991. Microflora and immunology of the

digestive tract. In Physiology of Small and Large Animals ed. Ruckebusch, Y.; Phaneuf,

L.P., Dunlop, R., Philadelphia: Becker, 198-208.

SAS Institute Inc., 2002, User’s Guide (release 8.1.), Cary, NC, USA.

Schuppers, M.E., Stephan, R., Ledergerber, U., Danuser, J., Bissing-Choisat, B., Stärk,

K.D.C., Regula, G., 2005. Clinical herd health, farm management and antimicrobial

resistance in Campylobacter coli on finishing pig farms in Switzerland. Prev. vet. Med.

69, 189-202.

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November 2003 zur Bekämpfung von Salmonellen und bestimmten anderen durch

Lebensmittel übertragbaren Zoonoseerregern.

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Wehebrink, T., Kemper, N., grosse Beilage, E., Krieter, J. (2006). Prevalence of

Campylobacter spp. and Yersinia spp. in the pig production. Prev. Vet. Med.

(submitted).

Weijtens, M.J.B., Bijker, P.G.H., van der Plas, J., Urlings, H.A.P., Biesheuvel, M.H., 1993.

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Quart. 15, 138-143.

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52

53

CHAPTER FOUR

Simulation study on the epidemiology of Salmonella spp. in

the pork supply chain

TANJA WEHEBRINK, NICOLE KEMPER

and JOACHIM KRIETER

Institute of Animal Breeding and Husbandry

Christian-Albrechts-University

D-24118 Kiel, Germany

54

Abstract

Pork can be regarded as an important source of food-borne salmonellosis. The objective of

this research was to gain insight into the epidemiological effects of different strategies in the

farrowing and finishing units to improve the food safety of pork with respect to the

prevalence of Salmonella spp. in finishing pigs. Therefore a stochastic transition model was

designed depending on prevalence in the population (sows = 0.5% to 65%; rearing pigs = 2%

to 95%), infection risks (farmer = 0% to 10%; rodents = 0% to 5%; feed = 0% to 10%; and

dust = 0% to 5%), the immunisation schedule of sows (yes/no) and the purchase of pathogen-

free gilts (yes/no). The simulation model generated an integrated pig production chain with

linkages between the stages farrowing, rearing and fattening. Within each herd, dynamic

patterns of Salmonella infections were simulated. The simulation covered a time interval of

24 months.

The results in the present study showed that preventive measures must first be introduced in

the fattening unit because at this production stage preventive measures regarding the different

risk factors had the highest influence on the prevalence of Salmonella spp.. The risk factor

‘farmer’ represented an exception as the influence of this factor was higher in the rearing unit

(22.8% vs. 17.1%). The distribution over management interventions in the finishing stages

was in the following order: farmer (p-value: 0.0004-0.0443), feed (p-value: 0.03-0.46), dust

(p-value: 0.33-0.66) and rodents (p-value: 0.71-0.92). Immunisation against Salmonella spp.

in sows represents a good strategy to decrease prevalence of Salmonella spp. in the fattening

unit.

keywords: farrowing and fattening unit, Monte Carlo Simulation, risk factors, Salmonella spp.

55

1. Introduction

In industrialised countries, Salmonella enterica is a frequent cause of food-borne infections

(Gebreyes et al., 2006). About 15-25% of all human salmonellosis cases worldwide can be

attributed to the consumption of contaminated pork and pork products (van Pelt and

Valkenburgh, 2001). Up to now, more than 2,500 serovars of Salmonella enterica have been

recognized (Farmer, 1999). Two non-host-adapted serovars common in animals and humans

are S. (Salmonella) Enteritidis and S. Typhimurium, which have been reported as two of the

top food-borne infections in developed countries (Poppe et al., 2002). In Germany, 52,245

salmonellosis cases were registered in 2005. Compared with cases in 2004, a decrease of

8.3% was assessed. Salmonella Enteritidis was analysed in 68% and S. Typhimurium in 25%

of the infections (Bätza, 2006).

Contamination of pork products is related to asymptomatic intestinal carriage of Salmonella

spp. by living pigs arriving at the slaughterhouse. To reduce the risk of pork contamination,

some countries have established monitoring programs to identify pig farms with a high

proportion of market hogs carrying Salmonella spp., followed by steps towards the reduction

of the on-farm prevalence (Christensen et al., 2002). These actions will be mandatory in the

future and especially in Europe, where several laws have recently been announced. In this

context, directive 92/117/EEC was abolished and replaced by directive 2003/99/EC on the

monitoring of zoonoses and zoonotic agents (Anonymous, 2003). Furthermore, a regulation

for the control of Salmonella spp. in pig production was established (Anonymous, 2007).

Control of Salmonella spp. in pork can be accomplished at all levels of production including

pre-harvest (farm level). In order to limit and control Salmonella spp. occurrence in a swine

herd, it is initially necessary to conduct epidemiological studies, first to determine the

prevalence of Salmonella spp. and identify possible risk factors, and consequently to

implement and monitor control programs (Mousing et al., 1997). Since it is not possible to

test all individual interventions in practice, computer simulation is an attractive way to

explore the effect of prevalence variations (Dijkhuizen and Morris, 1997).

The present research included an exploration of possible measures that can be implemented in

a farrowing and fattening unit to control the introduction and reduce the prevalence of

Salmonella in finishing pigs. A stochastic state-transition simulation model was established to

gather further information about the influence of the risk factors in the different pig

production stages on the Salmonella spp. prevalence in fattening pigs. Furthermore, the

influence of preventive arrangements of the immunisation of sows, and additionally, of

56

pathogen-free purchased gilts on the Salmonella spp. prevalence in the farrowing and

fattening unit were determined.

2. Materials and Methods

2.1 General conception

In accordance with Krieter (2004), the simulation model includes an integrated pig production

chain with vertical linkages between the three stages farrowing, rearing and fattening of pigs.

Herd size at the farrowing stage was set to 210 productive sows, at the fattening level 1,500

places per farm were assumed implying 2.7 production cycles per year. In the farrowing

stage, feeder pigs were produced, which are passed on to the fattening stage at a live weight of

28 kg. Animals were slaughtered at a live weight of 115 kg. The model starts with the

generation of the sows’ performance. The production cycle was 150 days, based on the

gestation of 115 days, lactation length of 28 days and seven days from weaning to breeding.

Because of these facts, the number of litters per sow and year was 2.3. Litter size and piglet

mortality was simulated over ten litters with non-linear patterns for litter size born alive and

piglet mortality (Brandt, 1984). An average of 10.2 piglets were born alive and piglet

mortality varied between 13.3% and 18.1%. The culling percentage of sows was defined by a

40% replacement rate. The purchased gilts were integrated into the herd with 180 days.

Postweaning mortality was 1%, during fattening the mortality rate rose to 3%. The simulation

model includes possible crowding effects (e.g. stress, higher infection risk) between the

different stages of the production chain. After weaning, three litters were housed in one large

group within the farrowing farm at one time. In the fattening unit, the piglets were split into

two batches after transport. As shown by van der Wolf (2000), the in-herd Salmonella spp.

prevalence fluctuates within a given period. To make allowances for this variation, the

duration of the simulation was extended to 24 months.

2.2 Entry and spread of Salmonella spp. in the farrowing and fattening unit

The model considered several alternatives for Salmonella spp. entry in the production chain

(Table 1). The assumptions were based on literature about Salmonella spp. in the pork chain

(van der Wolf, 2000; Stege et al., 2001; Meyer et al., 2005). The probability of an infection

due to the risk factors varied between stages and depended on the prevalence in the

population. An important source of Salmonella spp. entry of all stages was the acquisition of

infected animals from the preliminary stage. Latently infected animals entering the herd

57

unnoticed due to the lack of clinical symptoms can intermittently shed Salmonella spp. via

their faeces. The farmer himself is the highest risk factor. For example, he can introduce

Salmonella spp. in the barn via boots, overalls and other implements and he is additionally

responsible for the spread of the pathogen in the herd. The feasibility of an infection due to

the farmer was simulated from 0% to 15%. Another living vector are rodents. In the model, it

was assumed that rodents mainly initialise and maintain the contamination cycle at the farm.

The probability of an infection due to rodents ranged from 0% to 5 %.

The occurrence of Salmonella spp. in feed is mostly a consequence of recontamination during

production, transport or storage. The probability of an infection due to feedstuff ranged from

0% to 10%.

The concluding risk factor in the simulation program is dust. The pathogen Salmonella spp. is

able to survive in dust at room temperature for four years (Selbitz, 2002). Thus, dust is

responsible for a re-infection of a cleaned and disinfected barn. The chance of a positive result

from the pigs ranged from 0% to 5% (Table 1). These four assumed risk factors were higher

in breeding farms compared to finishing farms (due to e.g. all-in all-out, cleaning and

disinfection).

Table 1

Description of the model inputs

Description of variables mean min max

Production farrowing unit

sows 210 piglet number born alive per litter1, piglet 10.2 8.6 11.5 mortality (piglets), % 15.3 13.3 18.1 lactation period, days 28 weaning-to-oestrus interval, days 7 litters per sow and year, n 2.3 replacement rate, % 40 rearing unit (7 to 28kg live weight) mortality, % 1 production cycle 6.1 finishing unit (>28 to 115kg live weight) mortality, % 3 production cycle 2.7

58

Table 1

Description of the model inputs (cont.)

Salmonella spp. introduction farrowing unit

sows prevalence replacement gilts, % -2 0 90 probability of an infection due to, % biotic vector: farmer -2 0 15 rodents -2 0 5 abiotic vector: feed -2 0 10 dust -2 0 5

rearing unit (weaning piglet) probability of an infection due to, % biotic vectors: farmer -2 0 10 rodents -2 0 1 abiotic vectors: feed -2 0 2 dust -2 0 2

fattening unit probability of an infection due to, % biotic vectors: farmer -2 0 10 rodents -2 0 2.5 abiotic vectors: feed -2 0 10 dust -2 0 5

Salmonella spp. transmission farrowing unit sows, piglets – suckling period probability – excretion of S. spp. via faeces by infected sows -2 10 95 probability – infection of piglets by sows infected -2 20 90 rearing unit crowding, no. litters per pen 3 crowding factor, c3 0.50 probability for the infection from pen to pen, % 30 fattening unit crowding, no. pen per pen (weaning) 2 crowding factor, c3 0.80 probability for the infection from pen to pen, % 60

1depending on parity, weighted with frequency of parity distribution 2depending on the prevalence 3explanation, see text

In the stochastic state-transition model, groups of pigs move through the pork supply chain

and may become infected with Salmonella spp.. A pig can have two states with respect to

Salmonella spp. over the expected 24 months with four transitions respectively: free stays

free, free changes to infected, infected changes to free or infected stays infected. The

transmission of Salmonella spp. in the vertical production chain depends on the Salmonella

spp. status of the sow at the farrowing unit. Suckling piglets can be infected by perinatal

59

contamination and the faeces of the sow. The probability that infected sows excreted

Salmonella spp. with the faeces ranged between 10% and 95%, depending on the general

Salmonella status of the farm. The risk of a piglet becoming infected by the contaminated

faeces of the sow varied from 20% to 90%. Each alteration of the stage caused an increase in

prevalence due to crowding. Crowding was considered from lactation to weaning and from

weaning to finishing. Two patterns of the spread of infection within units were taken into

account. First, if a pen had a known number of animal infected (nj) after arrival, the

proportion of infected animals (pui) was updated with:

pui = ni/N+c[ni/N(1-exp-(1-ni/N))]

N is the total number of animals per pen and c the weighting factor depending on the stage

considered. Parameter c diminishes the probability of spreading the infection at the weaning

stage (0.50) and increases the risk at the fattening level (0.80). Secondly, infections spread

from adjacent pen to adjacent pen within a barn due to faeces and other vectors (e.g. boots,

overalls and other implements). The probability of pen-to-pen transmission was set at 30% at

weaning and 60% at finishing. The Salmonella transmission from barn to barn was neglected

in the model.

2.3 Simulation scenarios

The data in Table 2 represent the different scenarios of Salmonella spp. introduction, e.g. by

sows, weaning piglets and fattening pigs.

The basic scenario has approximately the same values as the ‘low scenario’. Additionally,

‘middle scenarios’ and ‘high scenarios' of a probability of infection due to biotic and abiotic

vectors were simulated.

60

Table 2

Parameters of simulation scenarios

parameter value low middle high

Salmonella spp. introduction probability of an infection due to, % farrowing unit sow biotic vectors: farmer 0.50 7.00 15.00 rodents 0.25 1.50 4.00 abiotic vectors: feed 0 5.00 10.00 dust 0.30 2.50 4.90 immunisation schedule (sows) 10.00 purchase of gilts 10.00 rearing unit weaning piglet biotic vectors: farmer 0.50 5.00 10.00 rodents 0.25 0.75 1.00 abiotic vectors: feed 0.05 1.00 2.00 dust 0.07 1.75 1.90 fattening unit pig biotic vectors: farmer 0.80 6.50 9.50 rodents 0.15 1.75 2.50 abiotic vectors: feed 0.50 5.50 10.00 dust 0.13 3.50 5.0

2.4 Statistical analysis

The significance of systematic environmental effects on the prevalence of Salmonella spp.

was investigated in a generalized linear model. The following risk factors were considered in

the model: farmer, rodents, feed and dust with four classes respectively and two fixed effects

were additionally assumed: immunisation of sows (yes/no) and purchase of pathogen-free

gilts (yes/no) (see Table 2). The analysis was performed with the SAS procedure GENMOD

(SAS® 2002). For the estimation, a gamma distribution and a logistic link function (i.e.

logistic regression) were assumed. In total, 100 farms were simulated with 100 replicates.

61

*

3. Results

3.1 Basic situation

Figure 1 presents the prevalence of the animals from the farrowing to the fattening unit in

consideration of the baseline parameters. In the farrowing unit, the prevalence from sows

(17.1%) and suckling piglets (14.4%) was higher than in the following production stages.

14.4

17.1

12.3 11.9

0

5

10

15

20

suckling

piglets

sows weaning pigs fattening

pigs

pig production

pre

va

len

ce

(%

)

Salmonella spp.

*standard error

Figure 1

Prevalence (LS-Means) of Salmonella spp. in the basic situation

3.2 Simulation scenarios

Farmer

The risk factor ‘farmer’ is a synonym for a very complex introduction- and spread-risk.

For example, a farmer can introduce Salmonella spp. in the barn via boots, overalls and other

implements. Additionally, he can be responsible for the spread of the pathogen in the herd

because he is a biotic vector for cross-contaminations. Figure 2 shows the farmer’s influence

with different probability of infection (0.5%-15.0%) in the farrowing / rearing / fattening unit

on the prevalence of Salmonella spp. in fattening pigs.

62

*

11.713.712.7

11.2

17.1

22.8

15.017.1

11.8

0

5

10

15

20

25

30

low middle high

probability of an infection (%)

pre

va

len

ce

fa

tte

nin

g

pig

s (

%)

farrowing unit rearing unit fattening unit

*standard error

Figure 2

Salmonella spp. prevalence (LS-Means) in fattening pigs depending on the risk factor

‘farmer’ at the different production stages

In total, the increasing of Salmonella spp. from the fattening pigs during the different

scenarios was moderate (11.7% vs. 12.7% vs. 13.7%) when the infection source was in the

farrowing unit. A huge influence on the pathogen prevalence was when the introduction

happened in the rearing unit. In this case, the prevalence rose from 11.2% over 17.1% to

22.8%.

In every production stage, the farmer was a significant risk factor (p-value = 0.0055 farrowing

unit; p-value = 0.0004 rearing unit; p-value = 0.0443 fattening unit) on the Salmonella spp.

prevalence in fattening pigs.

Rodents

Another biotic vector are rodents. In the model, it was assumed that rodents mainly initialise

and maintain the contamination cycle at the farm. Rodents had no significant influence on the

prevalence in fattening pigs (p-value: 0.71-0.92). Figure 3 shows that the Salmonella spp.

prevalence of fattening pigs only varied slightly during the different scenarios (between

11.1% to 12.8%). Apparently, there is no impact as to where the introduction takes place and

with which tendency (0.25%-4.0% probability of an infection).

63

* 11.7 11.8 12.411.1 11.6 11.911.3

12.3 12.8

0

2

4

6

8

10

12

14

low middle high

probability of an infection (%)

pre

va

len

ce

fa

tte

nin

g

pig

s (

%)

farrowing unit rearing unit fattening unit

*standard error

Figure 3

Salmonella spp. prevalence (LS-Means) in fattening pigs depending on the risk factor

‘rodents’ at the different production stages

Feed

The occurrence of Salmonella spp. in feed is mostly a consequence of recontamination during

production, transport or storage. Feed only had a significant influence (p-value = 0.03) in the

fattening unit. The highest prevalence was reached in the scenario when the fattening pigs

were contaminated with feed (Figure 4). Although the sows had nearly the same infection

probabilities in the middle (5.0% vs. 5.5%) and high scenarios (10.0% vs. 10.0%), the

prevalence in fattening pigs was at 12.1% (middle infection probability) and 13.0% (high

infection probability) not as high as when the infection happened in the fattening unit. The

Salmonella spp. prevalence reached in this case 15.6% and 17.1%.

64

*

12.113.0

10.511.9

13.211.5

17.115.6

11.5

0

5

10

15

20

low middle high

probability of an infection (%)

pre

va

len

ce

fa

tte

nin

g p

igs

(%)

farrowing unit rearing unit fattening unit

*standard error

Figure 4

Salmonella spp. prevalence (LS-Means) in fattening pigs depending on the risk factor ‘feed’

at the different production stages

Dust

The pathogen Salmonella spp. is able to survive in dust at room temperature for four years.

Thus, dust is responsible for a re-infection of a cleaned and disinfected barn. Dust had no

significant influence on the prevalence in fattening pigs (p-value: 0.33-0.66). Figure 5 points

out that the highest prevalence (14.3%) was reached when the barn was not strictly cleaned in

the fattening unit.

65

11.5 12.1 12.312.9

10.7

12.714.3

11.5

13.3

02

46

810

1214

16

low middle high

probability of an infection (%)

pre

va

len

ce

fa

tte

nin

g

pig

s (

%)

farrowing unit rearing unit fattening unit

*standard error

Figure 5

Salmonella spp. prevalence (LS-Means) in fattening pigs depending on the risk factor ‘dust’

at the different production stages

Preventive measures

Additionally, in the farrowing unit, it was simulated which preventive measure is more

effective. Figure 6 clarifies that the sow immunisation is more crucial than the purchase of

pathogen-free gilts. The prevalence in animals in the scenario with ‘purchase of pathogen-free

gilts’ were between 2.6% and 4.3% higher compared to the scenario ‘sow immunisation’.

Neither the immunisation nor the purchase of pathogen-free gilts had a significant influence

(p-value = 0.76 vs. p-value = 0.85) on the Salmonella spp. prevalence in the pig production.

*

66

* 7.5

13.0

7.0 7.3

11.8

15.6

10.4 10.3

0

5

10

15

20

suckling piglets sows weaning pigs fattening pigs

production stage

pre

va

len

ce

(%

)immunisation pathogen free gilts

*standard error

Figure 6

The effects of immunisation and purchase of pathogen-free gilts (LS-Means)

4. Discussion

The simulation model generates an integrated production chain starting with the purchase of

sows at the farrowing unit and closing with the finishing pig in the fattening unit. At each

stage, Salmonella spp. may enter the production chain by different vectors (e.g. latently

infected animals, feed etc.), the transmission is affected be the status of the sow, crowding

effects and pen-to-pen infections. Assumptions about the entry and spread of Salmonella spp.

were derived from the literature.

Due to the lack of further information in the literature about the different parameters and their

infection probabilities, for instance the crowding effect, various scenarios were simulated

within biological limits. Based on the simulation of best- and worst-case scenarios,

parameters representing the most important effects influencing or lowering the prevalence in

fattening pigs were determined. Finally, these parameters were adjusted to the prevalence at

the different production stages as known from the literature.

The estimated prevalence in the basic scenario is confirmed by numerous prevalence studies.

For instance, in a Lower-Saxonian study by Quante (2000), in 79 out of 88 examined farms,

less than 20% of the sows were serologically tested positive, in seven 20%-40% and on two

farms more than 40% were positive. Meyer et al. (2005) analysed 1,498 blood samples of

sows serologically, showing positive results for 17.1% of the samples. Regarding fattening

67

pigs, on 96 farms Stege et al. (2000) showed at least one positive serological sample in 65.6%

of the farms. The average intraherd prevalence was 2%, the highest Salmonella spp.

prevalence was reached with 32% positively tested animals. In another study, 11% of 1,760

tested blood samples from fattening pigs reacted ELISA positive (van der Wolf, 2000). Meyer

et al. (2005) reported 301 (11.4%) positive serological results out of 2,642 blood samples

from fattening pigs.

The aim of the present paper focused on further information about the influence of the risk

factors (biotic vectors = farmer, rodents; abiotic vectors = forage, dust) in the different pig

production stages on the Salmonella spp. prevalence in fattening pigs. Furthermore, the

influence of preventive arrangements of immunisation of sows, and additionally, of pathogen-

free purchased gilts on the Salmonella spp. prevalence in the farrowing and fattening unit

were determined.

The results in the present study clarify that the greatest influence on the Salmonella spp.

detection rate is in the fattening unit. This conforms with an expert survey in the Netherlands

and Denmark by van der Gaag (2002). The ranking of the management interventions in the

primary stages shows that most of the emphasis is placed on reducing or preventing the

spread of Salmonella spp. within the farm. Two stages in the chain (finishing and

slaughtering) are expected to be able to most effectively improve the food safety of pork with

respect to Salmonella spp. (van der Gaag, 2002).

The interaction between Salmonella spp., host, and environment is influenced by various

factors. Thus, especially in the rearing unit, the farmer plays a major role in Salmonella spp.

transmission in this unit and later Salmonella spp. prevalence in the fatting unit (11.2%-

22.8%). One reason for this has to be seen in the incompletely developed immune response in

the pigs at an age between 21 and 80 days. Maternal immunoglobulines transferred to the

piglets with the colostrum are supposed to have vanished at this point of time while the

individual antibody production is only slowly increasing. Especially in the fattening unit, the

crowding effect has to be regarded, implicating a higher infection risk between the different

stages of the production chain.

Entry of pathogens can be brought about via abiotic factors such as contaminated equipment

and other vectors. Additional spreading of Salmonella spp. is provoked by cross-

contamination during daily work processes (Blaha, 1993). This fact is confirmed by other and

the presented research. Barber et al. (2002) and Rajic et al. (2005) reported that Salmonella

spp. were detected in 11% and 39% of boot samples, respectively. However, as part of

68

hygienic-lock facilities combined with all-in all-out production (Lo Fo Wong et al., 2004),

clean farmers’ boots might contribute to reducing the risk of Salmonella spp. infections.

Further potential risk factors for Salmonella spp. are rodents and wild birds (Zheng et al.,

2007). Wild fauna as well as other domestic animals living on the farm or in the surrounding

environment may introduce and transmit Salmonella spp. through direct contact with pigs, or

by faecal contamination of feed or farm equipment. Rodents are known to be carriers of

Salmonella spp. (Leirs et al., 2004). Reported prevalence of Salmonella spp. positive rodents

were between 4% and 30% (Böhm, 1993). Contrary to these facts, in the present simulation

rodents had hardly any influence on the prevalence The Salmonella spp. prevalence in

fattening pigs was in every scenario at a low level (11.7%-12.8%) and there were no high

variations. The prevalence in fattening pigs rose with the respected increase in the infection

probability in the fattening unit from 11.5% over 15.6% to 17.1%. The reason for this has to

be seen in the estimated low entry risk and the impact at a relatively late stage of the

simulation.

In addition to the presented Monte Carlo simulation, many studies have shown that the type of

feed appears to be strongly associated with the presence of Salmonella spp.. For example,

Cook and Miller (2005) reported that farms feeding home-mixed rations had a lower

seroprevalence of Salmonella spp. (OR = 0.77) in a study including 1,806 farms. On the other

hand, Harris et al. (1997) found a higher prevalence of Salmonella spp.-contaminated

homemade feed than purchased feed on farm. The quality and hygiene of homemade feed

might vary in the studies, and furthermore, Harris et al. (1997) investigated only 30 farms.

Purchased feed might constitute a risk of introducing Salmonella spp. in the herd. The

importance of the risk factor feed could be confirmed in the present study.

Another risk factor is dust. It can be responsible for re-infection of cleaned and disinfected

barns because the pathogen is able to survive in dust at room temperature for four years

(Selbitz, 2002). In the present study, dust had a higher influence on the prevalence in fattening

pigs than rodents, stressing the fact that strict cleaning of barns is a basis for good health

management.

The immunity scenario showed that the immunisation of sows is an opportunity for

Salmonella spp. abatement. The Salmonella-Typhimurium-alive vaccine is well established

and effective due to the fact that 70% of Salmonella infections in pigs are related to S.

Typhimurium (Enneking, 2005). In this scenario, immunisations are the preferable method of

reduction of infections. In practice, in most cases these measures are not economically

justifiable and only appropriate in problem herds.

69

The results of the simulation show that the purchase of pathogen-free gilts is only reasonable

with the simultaneous improvement of hygiene and management conditions.

The presented Monte Carlo simulation takes into consideration the multi-factorial sources of

Salmonella spp. infection. In order to control Salmonella spp. in pigs, quantified possible risk

factors are needed to develop effective management strategies in pig herds (Zheng, et al.

2007). Additionally the simulation study indicated that most single intervention and control

measures are not effective enough to reduce or remove a Salmonella spp. infection or

contamination from a herd. It is therefore recommended that a herd-specific intervention and

control strategy be formulated, based on a combination of measures which are both practically

and economically feasible in a herd. A multi-factorial infection such as a Salmonella spp.

infection requires a multi-level approach of intervention and control, i.e. between and within

herds, as well as between and within pigs. The results from the presented study suggest that

improvements to all steps from stable to table need to be considered, and the most

economically optimal solution should be chosen. To identify this, an economic optimisation

model should be carried out, probably individually for each production stage.

5. Conclusion

A stochastic state-transition simulation model was established to gather further information

about the influence of the risk factors at the different pig production stages on the Salmonella

spp. prevalence in fattening pigs.

The results in the present study showed that preventive measures must first be introduced in

the fattening unit because at this production stage preventive measures regarding the different

risk factors had the highest influence on the prevalence of Salmonella spp.. The risk factor

‘farmer’ represented an exception as the influence of this factor was higher in the rearing unit.

The distribution over management interventions in the finishing stages was in the following

order: farmer, feed, dust and rodents. Immunisation against Salmonella spp. in sows

represents a good strategy to decrease the prevalence in the fattening unit. The results of this

simulation emphasise once more the outstanding importance of optimised hygiene

management.

70

Acknowledgements

This research was financially supported by the H. Wilhelm Schaumann Stiftung, the

Ministerium für Soziales, Gesundheit, Familie, Jugend und Senioren des Landes Schleswig-

Holstein and the Arbeitsgruppe Lebensmittelqualität und -sicherheit (QUASI) from the

Faculty of Agricultural and Nutritional Science, Christian-Albrechts-University, Kiel.

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Manninen, K., Dewey, C.E., McEwen, S.A., 2005. Longitudinal study of Salmonella

species in 90 Alberta swine finishing farms. Vet. Microbiol. 105, 47-56.

SAS Institute Inc., 2002. User’s Guide (release 8.1.), Cary, NC, USA.

Selbitz, H.J., 2002. Bakterielle Krankheiten der Tiere. In: Rolle, M., und A. Mayr (Hrsg.):

Medizinische Mikrobiologie, Infektions- und Seuchenlehre. 7 Aufl. Verlag Enke,

Stuttgard.

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Prevalence of subclinical Salmonella enterica infection in Danish finishing pig herds.

Prev. Vet. Med. 44, 175-188.

Stege, H., Jensen, T.K., Møller, K., Bækbo, P., Jorsal, S.E., 2001. Risk factors for intestinal

pathogens in Danish finishing pig herds. Prev. Vet. Med. 50, 135-146.

van der Gaag, M.A., Huirne, R.B.M., 2002. Elicitation of expert knowledge on controlling

Salmonella in pork chain. Chain and network science, 135-147.

van der Wolf, P.J., 2000. Salmonella in the pork production chain: feasibility of Salmonella-

free pig production. Department of Pig Health of the Animal Health Service, Utrecht,

The Netherlands, PhD Thesis.

van Pelt, W., Valkenburgh, SM., 2001. Zoonoses and zoonotic agents in humans, food,

animals and feed in the Netherlands. Available: www.keuringsdienstvanwaren.nl.

Zheng, D.M., Bonde, M., Sørensen, J.T., 2007. Associations between the proportion of

Salmonella seropositive slaughter pigs and the presence of herd level risk factors for

introduction and transmission of Salmonella in 34 Danish organic, outdoor (non-

organic) and indoor finishing-pig farms. Liv. Sci. 106, 189-199.

73

GENERAL DISCUSSION

Introduction

Pork can be regarded as an important source for food-borne campylobacteriosis, yersiniosis

and salmonellosis. All these agents are carried by pigs without any clinical signs, food

products represent a potential source of human infections. In order to control these pathogens

in pigs a quantification of possible risk factors and the development of effective management

strategies in pig herds is needed. The aim of the present thesis was to contribute to a better

understanding of these bacterial pathogens causing disease both in humans and animals and to

use this information to assess and manage the risk to animals and humans.

A purpose of this thesis was to increase the knowledge about the epidemiology of the

occurrence of Campylobacter spp. and Yersinia spp. in farrowing and fattening herds with

particular emphasis on bacteriological findings. Analysis of the data from questionnaires

provided first indications of factors which may influence the prevalence of Campylobacter

spp. and Yersinia spp. in herds.

Another objective of this thesis was to gain insight into the epidemiological effects of

different strategies in the farrowing and finishing units to improve the food safety of pork

with respect to the prevalence of Salmonella spp. in finishing pigs. Therefore a stochastic

transition model was designed depending on farm size, prevalence in the population, rearing,

infection risks, the immunisation schedule of sows and the purchase of pathogen-free gilts.

The simulation model generates an integrated pig production chain with linkages between the

stages farrowing, rearing and fattening with Monte Carlo methods.

The outline of the GENERAL DISCUSSION is focused in the first section on the results from the

bacteriological analysis of Campylobacter spp. and the indicated risk factors. In the second

section, the main emphasis is laid on the prevalence of Yersinia spp. and in the third section

on the different measures to prevent the spread of Salmonella spp. simulated with a stochastic

transition model.

Campylobacter spp.

The results from the thesis (CHAPTER TWO) prove that Campylobacter spp. are of increasing

importance in farrowing and fattening units. High prevalence of Campylobacter spp. were

found in suckling (80.9%), growing (89.2%) and finishing pigs (64.7%). Other studies also

confirm these results (Görgen et al., 1983; Weijtens et al., 1993; Gaull, 2002).

74

In the farrowing unit the prevalence of piglets (80.9%) was very high compared with the

prevalence of sows (33.8%). One explanation could be the stable gut flora from older animals.

The same effect was observed by Weijtens et al. (2000). They described a sow herd with a

minor Campylobacter spp. status (0-22% during 22 month). This farm started breeding with

SPF-animals (specific pathogen-free), but the hygienic regime was not strict enough to avoid

pathogen contamination. However, the prevalence remained at a low level.

The main infection route is the transmission from animal to animal. Gaull (2002) showed that

an infection from sow to piglet is possible, showing 100% prevalence in piglets within 24

hours after birth. This fact cannot be confirmed however. On the basis of the results from the

present thesis, it becomes obvious that there is no relationship between infected sows and the

infection of their piglets with Campylobacter spp.. This fact clarifies that sampling of sows

alone is useless without taking the piglets into account.

On the basis of the results in the fattening unit, it becomes obvious that a stable gut flora from

older pigs can cause a decrease in prevalence (89.2% vs. 64.7%). Other studies e.g. from

Weijtens et al. (1993; 1999) confirm this effect.

Furthermore, it could be noticed that in every herd in the fattening unit Campylobacter spp.

excretion was intermittent. This path of excretion was described in other studies, too (Gaull,

2002; Kasimir, 2005). The occasionally pathogen-free status and the following re-infection

could be one explanation for these effects. Weijtens et al. (1999) stated that the pigs are not

pathogen-free when there is no detection rate. They suspected that the pathogen further

existed in the intestinal villi.

A total pathogen eradication in pig herds seems to be utopian, however for chicks several

studies and strategies for pathogen reduction have been described (Kasimir, 2005).

Immunisation seemed to be successful (Rice et al., 1997). Another opportunity for chicks is

Competitive Exclusion. There are no studies on Competitive Exclusion in pigs, but this

method may be successful in that species, too (Weijtens, 1996). As an alternative, Weijtens et

al. (2000) demonstrated that it is possible to keep Campylobacter spp. at a low level or to

arrange a pathogen-free pig herd. The principle is based on Campylobacter spp.-free sows

(from specific pathogen-free herds) in solidly cleaned and disinfected cots, which have been,

when possible, vacant for some time. Sows are, as a result of their robust gut flora, less

susceptible for the pathogen compared to piglets. Additionally, frequently practised housing

in crates prevents coprophagy. The negative sows cannot infect their piglets.

Despite the high prevalence in the faeces, low detection rates on carcasses between 2.9% and

36.5% are described in the literature (Nesbakken et al., 2002; Pearce et al., 2003; Kasimir,

75

2005). In the present thesis, the detected Campylobacter spp. prevalence decreased from

55.7% in the lairage to over 19.7% on the carcasses before chilling to 0% after twelve hours

chilling. In most of the literature, Campylobacter spp. is mostly not detectable after chilling,

too (Chang et al., 2003; Pearce et al., 2003). Other studies have shown that after chilling

equally high prevalence are possible. For example, Gebreys et al. (2003) isolated

Campylobacter spp. from 29% of the carcasses after chilling. It is known that Campylobacter

spp. can survive the chilling process on chicken skin because the skin is sulcate, clammy and

the follicles feature the opportunity to survive it (Kasimir, 2005). A high prevalence of

Campylobacter spp. on pig carcasses is implausible because pig skin with its relatively flush

surface wipes off during the chilling process.

For consumer protection purposes it is noteworthy that in the present project C. coli was

isolated from one liver sample only.

Besides C. coli, C. jejuni were laboratory-confirmed in this examination. The isolation of C.

jejuni from pig samples has been described by other studies as well. For example, Stich-Groh

(1982) and Young et al. (2000) identified 23.4% and 76.3% respectively, Campylobacter spp.

as C. jejuni. In these assays, hippurathydrolysis served as a confirmation method. This

technique is based on the ability of C. jejuni to hydrolyse hippurat, a biochemical reaction C.

coli is not capable of. One major problem of this method is the possible loss of this ability

during the life span of C. jejuni, causing false positive results with regard to C. coli. But it can

be possible that in some farms or in some geographical regions C. jejuni is described as

common in pigs (Kasimir, 2005).

As a result of the small sample size in the farrowing unit (CHAPTER THREE) it was not

possible to perform a risk analysis which yielded significant conclusions in this production

stage. Thus, further risk factors for the occurrence of Campylobacter spp. in fattening units

should be observed via environmental and feed samples from the checked herds and

questionnaires in the corresponding pig farms. Neither in the feed nor in the environmental

samples was Campylobacter spp. detected. In the fattening stage, the following risk factors

had a significant effect (p≤0.05) on Campylobacter spp. prevalence: sampling time, number

of fattening places per herd, mixed farming, floor space design, feed origin, antibacterial and

anthelmintic treatment. These results show that housing and management have a possible

influence on Campylobacter spp. prevalence and should be investigated further. Weijtens et

al. (2000) found out that feed, water and biotic vectors including humans are permanent risk

factors for piglets. As a result of the low moisture content, feed can be excluded as a risk

factor (Altekruse and Swerdlow, 2002). The risk of introducing the pathogen via water can be

76

reduced by using chlorinated water. The combination of keeping away birds, rodents and

insects with a strict hygiene management routine should prevent or at least limit the risk

factors at the farrowing and fattening units. Furthermore, Kasimir (2005) found that the age of

the cots and the corresponding infection-pressure have no influence on pathogen incidence.

Recapitulating, it can be mentioned that many facts are known about Campylobacter spp.

epidemiology. It seems to be possible to hold the pathogen prevalence at a low herd level.

Other methods to create a pathogen-free herd with SPF-animals (specific pathogen-free) and

with a really strict hygiene regime are associated with high costs for the farmers. Because of

the probably low impact on human health, such arrangements, such as herd decontamination,

make no sense. Against this background, the discussion of general Campylobacter spp.

abatement is essential, especially with regard to effective preventive adoption at a certain

stage of the production chain. One point speaking against such an implementation is the fact

that human campylobacteriosis is caused by C. jejuni in only 90% of cases. Only 5% to 10%

of the cases are caused by C. coli (Tam et al., 2003). The main source for C. jejuni is chicken

meat, while the infection source for human C. coli-infection is unclear. This pathogen is often

isolated from pigs but also from turkey hens (Kramer, 2000). Despite the high isolation rate in

pig tonsils and faeces samples, pork (besides offal) is hardly contaminated with

Campylobacter spp.. One reason for this is the effective chilling of the carcasses in

combination with drying the skin after slaughtering. Campylobacter infections are often

sporadic single-diseases, so the search for the infection source is very difficult.

The advantage of abatement is the reduction of the potential health risk for humans, because

at the moment it cannot be estimated how often the VBNC status (viable-but-not-cultivable)

is present on the carcasses and in the meat. Further studies are urgently needed to gather

further information on the VBNC mechanism of Campylobacter spp..

With regard to Campylobacter spp., consumer education is important. There is a cross-

contamination risk from chicken and maybe from pig meat in combination with bad kitchen

hygiene because the infection dose is very low with 500 to 800 pathogens (Black et al., 1988).

Yersinia spp.

Yersinia spp. seems to play a negligible role in farrowing herds because neither in suckling

piglets nor in sows was the pathogen detected. This is in accordance with another study

detecting Yersinia spp. only during the fattening period but not in sows and piglets (Kasimir,

2005). Whereas Korte et al. (2004) in contrast reasoned from their study that sows are an

important infection source for the pig herd.

77

The fact that Y. enterocolitica was not isolated in the farrowing unit but could not be isolated

until the beginning of the fattening period is evidence that the cause of infection has to be

looked for in the fattening unit. In this production stage, the prevalence of Y. enterocolitica

were between 0% and 46.9%. Bush et al. (2003) detected 12.8% Y. enterocolitica in 2,664

faecal samples and Kasimir (2005) described isolation rates between 0% and 65.4%. The

prevalence at farm level arranged variably. Some farms had a prevalence of 0% at the

beginning of fattening period and at the end nearly 100% or vice versa. It is ambiguous as to

why the pathogen diffuses in some herds at a high level and not in other herds in turn,

although some pigs are infected there, too. It is described in the literature that the pathogen

can only be separated in a certain period after infection (Nielsen et al., 1996). Furthermore, a

re-infection of the pigs is impossible, because of the gut-generated immunity (Fukushima,

1983). This is the reason why the prevalence decreased from the first (15.2%) to the second

(13.3%) sample time in the fattening unit.

The prevalence of Yersinia spp. in the slaughterhouse was low (lairage: 5.7% vs. before

chilling: 0.8%). One reason for this effect is that Yersinia spp. persists in the tonsils and will

be shed with the faeces discontinuously. Anderson (1988) described the influence of different

eviscerate techniques in relation to carcass contamination in the slaughterhouse. By the

manual gut cut down, he found on the medial hind leg significantly more (26.3%) Yersinia

enterocolitica O:3 than by the use of the bung cutters (13.4%). The contamination was lowest

when behind the bung cutter the rectum was closed with a plastic bag. Nesbakken et al.

(1994) produced similar results. The faeces has no influence on carcass contamination.

Bornadi et al. (2003) could not isolate Yersinia spp. on 150 carcass samples while the rectum

was not closed with a plastic bag. The detection rate in the faeces was very low with 4.0%.

The tonsils contaminate the carcasses only marginally. Of higher significance is the

contamination of the pluck per tonsils (Fredriksson-Ahomaa et al., 2001). The authors found

that the pluck had a higher prevalence than the kidney.

Neither in the environmental nor in the feed samples were Yersinia spp. isolated. One reason

therefore can be found in the method of detection. Especially for environmental and animal

feed samples, the cultivation method seems to be inferior compared to Polymerase-Chain-

Reaction (PCR), because the low numbers of pathogenic strains in these samples can often be

suppressed by a distinct satellite flora (Fredriksson-Ahomaa and Korkeala, 2003).

Yersinia spp. is only sporadically found in meat samples. One reason is that it is not easy for

the pathogen to flourish against the natural meat micro-flora. But the pathogen is able to

survive in raw pig meat for a long time (Fukushima and Gomyoda, 1986). A higher risk factor

78

is offal. Bucher et al. (2001) were able to isolate Yersinia enterocolitica 4/O:3 from 75% of

tongues, 70% of hearts and 25% of livers. When these contaminated offal are further

processed at home and the knives or the workplace are not cleaned correctly, the carry-over

risk to other food exists.

As a result of the small pathogen detection rate in the farrowing and fattening unit it was not

possible to perform a risk analysis which yielded significant conclusions. But in the literature

there is research about the risk factors. For example, a Norwegian study about risk factors of

Yersinia spp. in pig production shows that herds with only fattening pigs have a higher

prevalence than farrowing-to-finishing herds (Skjerve et al., 1998). The purchase of animals

and the associated animal assortment are the highest risk factor for pathogen diffusion in the

herd. Also cats and straw litter raise the infection risk. A pathogen decrease was realised with

low-pressure aeration, hygienic methods such as a disinfected mat in the entrance area and

feeding per hand. When the pathogen is in the herd it is persistent. It is not really known how

eradication methods have an influence of Yersinia status. The elimination from carrier-

animals inside a herd is not effective (Skjerve et al., 1998). During 2001, a Swiss study dealt

with the prevalence of Yersinia spp. in pork herds with different animal husbandries. In this

case, the application of medicine feed as a prophylaxis for pigs at the beginning of the

fattening period was a high risk factor for occurrence of Yersinia spp. in the herd

(Ledergerber et al., 2003). In a Canadian study, 1,944 environmental samples were analysed

(Pilon et al., 2000). From only 17 (0.6%) could Yersinia spp. be isolated. Per farm only one

genotype was isolated. Because of this fact the authors came to the conclusion that external

causes of a pathogen risk factor are only of little importance. Pathogen isolation from

environmental samples is very difficult, because the pathogen concentration is very low and

the concentration of company flora is very high (Fredriksson-Ahomaa and Korkeala, 2003).

In conclusion, knowledge about the epidemiology of Yersinia spp. is currently very limited.

Concrete arrangements do not exist, but the purchase of pigs from different herds, the

application of herd-specific vaccination in problem herds and the forceful compliance of

hygiene methods seem to be steps into the right direction. A monitoring implementation can

help define high-contaminated herds from low-contaminated. The main focus has to be placed

on the slaughterhouse. Herds with a high prevalence should be slaughtered separately at the

end of a slaughter day and their co-products should only be brought to the market heat-

treated. Furthermore, the slaughter technique discontinuing the had completely with tongue

and tonsils, is a preventive method (Christensen and Lüthje, 1994). Likewise, the

improvement of hygiene standards at the slaughterhouse is very important for a generation of

79

safety food. Regular education is important to improve the hygienic awareness of the

assistants (Bucher, 2001).

In addition, consumers have to be educated about the contact risk of pig meat, mainly offal.

Especially the risk from cross-contamination in their own kitchens is often underestimated.

Stochastic transition model for the epidemiology of Salmonella spp. in the pork supply

chain

Besides the importance of Salmonella spp. for public health, another aspect is the cost

generated by human salmonellosis. A working document of the European Commission

estimated that costs linked to food-borne salmonellosis ranged between 560 millions and € 2.8

billion in Europe, where Salmonella spp. was estimated to be responsible for nearly 166,000

cases in 1999 (Anonymous, 2001). However, indirect incentives such as the increased interest

in food safety and the large competition on the (international) market for pork, are of

increasing importance, since 1.25 million tons of pork are exported annually (ZMP, 2007).

Therefore, it is important to obtain more insight into the trade-off between prevalence

reduction and associated costs. Currently, the most common perspective on food safety and

human salmonellosis is the stable-to-table concept, acknowledging that each link in the food

production chain has a share in the responsibility of reducing the risk of food-borne disease.

Hence, for an effective control resulting in a satisfying reduction in the end product, the entire

supply chain must be involved (Lammerding and Fazil, 2000).

The presented thesis (CHAPTER FOUR) includes an approach for possible measures that can be

implemented in the farrowing and fattening unit to control the introduction and reduce the

prevalence of Salmonella spp. in finishing pigs. A stochastic state-transition simulation model

was established to gather further information about the influence of the risk factors in the

different pig production stages on the Salmonella spp. prevalence in fattening pigs.

Furthermore, the influence of preventive arrangements of the immunisation of sows, and

additionally, of pathogen-free purchased gilts on the Salmonella spp. prevalence in the

farrowing and fattening unit were determined. The application of risk analysis methods in the

assessment of microbial contamination of foods is relatively new. It offers a potential

overview of the interrelationship between the different processes which influence the

contamination of food items. This is in contrast to more specific detailed experiments which

only provide information about selected area. However, risk analysis models depend on data

preferably from such studies, in order to provide reliable estimates. All models are reduced

80

explanations of the real world. The more sophisticated the model, the more precisely the real

world may be explained.

The results in the present thesis showed that preventive measures must be affected in the

fattening unit because at this production stage the risk factors have the highest influence on

the prevalence of Salmonella spp.. Van der Gaag et al. (1999) identified the fattening farm as

the most important stage to achieve a reduction in Salmonella spp. prevalence, too.

There are several ways to reduce the Salmonella prevalence in a herd. For example, changes

in feeding practise, installation of adequate rodent control and improvements in hygiene

(Alban and Stärk, 2005). The model simulated the following distribution over management

interventions to reduce the prevalence of Salmonella spp. in fattening pigs: farmer (p-value:

0.0004-0.0443), feed (p-value: 0.03-0.46), dust (p-value: 0.33-0.66) and rodents (0.71-0.92).

Immunisation against Salmonella spp. in sows results in a good effect on the prevalence

reduction in the pig production. The exact quantitative effects of separated interventions on

the introduction and spread of Salmonella spp. and the course of infection are very difficult to

quantify precisely. Still, it is known that a package of multiple interventions leads to a

reduction of Salmonella spp. prevalence (Bagger and Nielsen, 2001).

In conclusion, the (pre-) harvest stages of the pork supply chain cannot ensure a zero

prevalence of contaminated carcasses (van der Gaag, 2004). Thus, the next stages (processing,

storage at retail and storage and preparing the pork by the consumer) are also important. For

instance, the consumer can reduce the risk of food-borne salmonellosis by cool storage and

thorough heating of the pork and avoiding cross-contamination in the kitchen (Gorman et al.,

2002). Nevertheless, by reducing the prevalence of contaminated carcasses, the risk for the

consumer should be decreased since less contaminated pork enters the consumer’s kitchen.

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GENERAL SUMMARY

This thesis focuses on information about the prevalence and origins and preventive measures

of the important zoonotic pathogens Campylobacter spp. and Yersinia spp. in the different

stages of the pig production chain. Furthermore, the epidemiological effects of different

strategies in the farrowing and finishing units with respect to the prevalence of Salmonella

spp. in finishing pigs were evaluated by simulation.

CHAPTER ONE summarises several studies emphasising the importance of Campylobacter

spp. and Yersinia spp. as widespread pathogens in the pig production chain.

First, the taxonomy and the pathogen character of these internationally important pathogens

are described, and second, prevalence in the pig production is reported. Obviously, pigs are

often carriers of Campylobacter spp. and Yersinia spp. causing infections in humans.

Contamination during the slaughtering process is possible.

However, pathogenic Campylobacter spp. and Yersinia spp. are comparatively infrequently

isolated from meat. A greater health risk is represented by entrails. In conclusion, to increase

pork safety, further epidemiological studies are urgently needed to determine the origin of

pathogens and to take counteractive measures.

The objective of CHAPTER TWO was to determine the prevalence of Campylobacter spp. and

Yersinia spp. in a total of 1,040 faecal samples taken from animals at different ages from four

farrowing and twelve fattening herds. In the farrowing unit, faeces were collected from 68

sows (faecal samples) and 256 suckling piglets (rectal swab samples). Further samples were

collected from 362 growing and 354 finishing pigs (rectal swab samples). Additionally, 56

feed and environmental samples were collected. During the slaughtering process, 122 pigs

and their carcasses respectively were sampled three times. First, rectal samples were taken

with swabs during the lairage. Second, the samples were taken from the carcass before

entering the chilling room. The same method was repeated in the chilling room twelve hours

after starting the chilling. Finally, 86 raw meat samples were taken from 34 retail stores.

Campylobacter spp. were isolated in sows (33.8%), piglets (80.9%), growing (89.2%) and

finishing (64.7%) pigs. Yersinia spp. were detected in growing (15.2%) and finishing (13.3%)

pigs only.

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During lairage, Campylobacter spp. were identified from pig faeces from all farms whereas

Yersinia spp. were detected in pigs from just two herds. After twelve hours of chilling neither

Campylobacter spp. nor Yersinia spp. were detected.

In raw meat samples, Campylobacter spp. were isolated from one liver sample and Yersinia

enterocolitica from two meat samples (mince and cutlet).

Common slaughter techniques and hygiene procedures may be effective tools to reduce the

risk of contamination and recontamination of meat products since Campylobacter spp. and

Yersinia spp. were found only sporadically in raw meat samples.

The aim of CHAPTER THREE was to gather further information about the sources of infection

with Campylobacter spp. and their qualitative and quantitative importance in pig production.

For statistical analysis, 1,040 results from the bacteriological examination for Campylobacter

spp. were evaluated with questionnaires from four farrowing and twelve fattening units. The

prevalence was determined via faeces and swab samples with regard to certain farm

production parameters.

Thereby, 30.8% of the sows and 80.9% of their piglets were carriers of Campylobacter spp..

In the fattening unit, the prevalence at the beginning of the fattening period was 89.2% and at

the end 64.7%.

As a result of the small sample size in the farrowing unit, it was not possible to perform a risk

analysis which yielded significant conclusions.

In the fattening stage, the following risk factors had a significant effect (p≤0.05) on

Campylobacter spp. prevalence: sampling time, number of fattening places per herd, mixed

farming, floor space design, feed origin, antibacterial and anthelmintic treatment. These

results show that housing and management have a possible influence on the Campylobacter

spp. prevalence and should be investigated further.

In CHAPTER FOUR the objective was to gain insight into the epidemiological effects of

different strategies in the farrowing and finishing units to improve the food safety of pork

with respect to the prevalence of Salmonella spp. in finishing pigs.

Therefore a stochastic transition model was designed depending on prevalence in the

population (sows = 0.5% to 65%; rearing pigs = 2% to 95%), infection risks (farmer = 0% to

10%; rodents = 0% to 5%; feed = 0% to 10%; and dust = 0% to 5%), the immunisation

schedule of sows (yes/no) and the purchase of pathogen-free gilts (yes/no).

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The simulation model generated an integrated pig production chain with linkages between the

stages farrowing, rearing and fattening. Within each herd, dynamic patterns of Salmonella

infections were simulated. The simulation covered a time interval of 24 months.

The results in the present study showed that preventive measures must first be introduced in

the fattening unit because at this production stage preventive measures regarding the different

risk factors had the highest influence on the prevalence of Salmonella spp..

The risk factor ‘farmer’ represented an exception as the influence of this factor was higher in

the rearing unit (22.8% vs. 17.1%). The distribution over management interventions in the

finishing stages was in the following order: farmer (p-value: 0.0004-0.0443), feed (p-value:

0.03-0.46), dust (p-value: 0.33-0.66) and rodents (p-value: 0.71-0.92). Immunisation against

Salmonella spp. in sows represents a good strategy to decrease prevalence of Salmonella spp.

in the fattening unit.

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89

ZUSAMMENFASSUNG

Die Ziele der vorliegenden Arbeit bestanden zum Einen aus der Erfassung und Bewertung

von Prävalenzen und Eintragsquellen der Zooanthroponosenerreger Campylobacter spp. und

Yersinia spp. in den verschiedenen Produktionsstufen der Schweineerzeugung. Zum Anderen

wurde eine Simulation der Ausbreitung von Salmonella spp. und der Einfluss präventiver

Maßnahmen auf die Prävalenz bei Mastschweinen in Ferkelerzeuger- und Mastbetrieben

vorgenommen.

KAPITEL EINS umfasst eine Literaturübersicht über Campylobacter spp. und Yersinia spp. in

der Schweineproduktionskette. Zunächst wurde die Systematik und die Erregereigenschaften

dieser zwei weltweit bedeutenden Zooanthroponoserreger dargestellt. Im Anschluss wurden

die bisher festgestellten Prävalenzen in der Produktionskette beim Schwein aufgezeigt. Es

wird deutlich, dass Schweine häufig Träger humanpathogener Campylobacter spp. und

Yersinia spp. sind und somit eine Kontamination ihres Fleisches während des

Schlachtprozesses möglich ist. Allerdings sind humanpathogene Campylobacter spp. und

Yersinia spp. relativ selten im Fleisch nachweisbar. Eine größere Gefahr stellen Innereien dar.

Zur Sicherung der hygienischen Unbedenklichkeit von Schweinefleisch sollte in Zukunft

versucht werden, die Epidemiologie der Erreger genauer aufzuklären um die Ursache der

Erregerausbreitung zu erkennen und geeignete Gegenmaßnahmen ergreifen zu können.

KAPITEL ZWEI zeigt die ermittelten Prävalenzen von Campylobacter spp. und Yersinia spp.

aus insgesamt 1.040 Kotproben von Tieren unterschiedlichen Alters auf vier Ferkel- und

zwölf Mastbetrieben. In der Ferkelerzeugung wurden 68 Sauen (Kotproben) und 256 Ferkel

(rektale Abstrichtupferproben) beprobt. Weitere Proben wurden von 362 Schweinen am

Mastanfang und 354 Schweinen am Mastende (rektale Abstrichtupferproben) entnommen.

Zusätzlich wurden 56 Futter- und Umweltproben gesammelt. Während des Schlachtprozesses

wurden 122 Schweine und ihre Schlachtkörper insgesamt dreimalig beprobt. Zuerst wurden

rektale Abstrichtupferproben im Wartebereich entnommen. Die zweite Beprobung erfolgte

am Schlachttierkörper direkt vor der Kühlung und noch einmal nach 12 Stunden Kühlung. In

34 Verkaufsstätten wurden abschließend 86 rohe Fleischwarenproben erworben und beprobt.

Campylobacter spp. wurden in Sauen (33,8%), Saugferkeln (80,9%), Schweinen am

Mastanfang (89,2%) und Mastschweinen am Mastende (64,7%) nachgewiesen. Yersinia spp.

wurden nur bei Schweinen am Anfang (15,2%) bzw. am Ende der Mastperiode (13,3%)

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analysiert. Im Wartebereich des Schlachthofes wurde Campylobacter spp. in Kotproben von

allen Mastbetrieben nachgewiesen, wohingegen Yersinia spp. nur in Schweinen von zwei

Betrieben entdeckt werden konnten. Nach zwölf Stunden Kühlung wurden weder

Campylobacter spp. noch Yersinia spp. nachgewiesen. In den rohen Fleischwarenproben,

wurden Campylobacter spp. in einer Leberprobe analysiert und Yersinia spp. von zwei

Fleischproben (Hackfleisch und Schnitzel).

Anscheinend sind die gebräuchlichen Schlachttechniken und Hygieneprozeduren effektiv

genug um das Risiko einer Kontamination bzw. Rekontamination von Fleischprodukten zu

reduzieren, da die Erreger nur sporadisch in den Fleischproben nachgewiesen werden

konnten.

KAPITEL DREI sollte zur Aufdeckung produktionsspezifischer Risikofaktoren und ihrer

qualitativen und quantitativen Bedeutung beitragen. Für die statistische Analyse wurden 1.040

Ergebnisse der bakteriologischen Untersuchung auf Campylobacter spp. im Zusammenhang

mit den Informationen aus einem Fragebogen aus vier Ferkelerzeuger- und zwölf

Mastbetrieben ausgewertet. Die Prävalenzen des Erregers wurden mit Hilfe von Kot- und

Abstrichtupferproben vor dem Hintergrund verschiedener Betriebsbedingungen ermittelt.

Dabei wurden bei 33,8% der Sauen und bei 80,9% der Ferkel Campylobacter spp.

nachgewiesen. In der Produktionsstufe Mast betrug die Prävalenz am Mastanfang 89,2% und

am Mastende 64,7%. Aufgrund des geringen Datenmaterials konnte auf der Produktionsstufe

Ferkelerzeugung keine Risikoanalyse durchgeführt werden.

Folgende Faktoren hatten auf den Mastbetrieben einen signifikanten Einfluss (p≤0,05) auf die

Campylobacter spp. Prävalenz: Zeitpunkt der Probeentnahme, Anzahl Mastplätze,

Mischbetrieb, Bodengestaltung, Futterherkunft, Einstallbehandlung und anthelminthische

Behandlung. Die Ergebnisse veranschaulichen, dass eine Reduzierung der Campylobacter

spp. Prävalenz durch betriebliche Haltungs- und Managementfaktoren möglich ist. Aus diesen

Ergebnissen resultiert weiterer Forschungsbedarf.

Ziel des KAPITEL VIER war es, einen Einblick in die epidemiologischen Effekte

verschiedener Strategien zur Qualitätssicherung in Ferkel- und Mastbetrieben und deren

Einfluss auf die Salmonellenprävalenz bei Mastschweinen zu gewinnen.

Dafür wurde ein stochastisches Simulationsmodell in Abhängigkeit der Populationsprävalenz

(Sauen: 0,5% bis 65%; Läufer: 2% bis 95%), des Infektionsrisikos (Betreuungspersonal = 0%

bis 10%; Schadnager = 0% bis 5%; Futter = 0% bis 10% und Staub = 0% bis 5%), der

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Impfung der Sauen (ja/nein) und dem Zukauf pathogen-freier Jungsauen (ja/nein) konstruiert.

Das Simulationsmodell generiert ein integriertes Produktionssystem beim Schwein mit den

Stufen Ferkelerzeugung, Aufzucht und Mast über einen Zeitintervall von 24 Monaten.

Die vorliegende Studie zeigt, dass präventive Maßnahmen zuerst in der Mast erfolgen

müssen, da dort die größten Effekte auf die Salmonellenprävalenz erzielt wurden. Der

Risikofaktor Betreuungspersonal bildete dabei eine Ausnahme, da er im Flatdeckbereich eine

höhere Prävalenz bei den Mastschweinen verursachte (22.8% vs. 17.1%).

Die Aufteilung der Managementmaßnahmen in der Mastschweineproduktion wurde in

folgender Reihenfolge vorgenommen: Betreuungspersonal (p-Wert: 0.0004-0.0443), Futter

(p-Wert: 0.03-0.46), Staub (p-Wert: 0.33-0.66) und Schadnager (p-Wert: 0.71-0.92). Die

Immunisierung der Sauen gegen Salmonella spp. wirkten im Mastbereich

prävalenzreduzierend.

DANKSAGUNG

An dieser Stelle möchte ich mich bei den Menschen bedanken, die zum Gelingen dieser

Arbeit beigetragen haben.

Ich danke Herrn Prof. Dr. Joachim Krieter für die Überlassung des Themas, die Unterstützung

bei der Abfassung der Dissertation sowie für die Möglichkeit, meine Ergebnisse auf

unterschiedlichen Tagungen im In- und Ausland vorzustellen.

Herrn Prof. Dr. Edgar Schallenberger danke ich für die Übernahme des Koreferats.

Frau Priv. Doz. Dr. Elisabeth grosse Beilage von der Außenstelle für Epidemiologie der

Tierärztlichen Hochschule Hannover möchte ich für die hilfreiche Beratung und für die

Bereitschaft zum Korrekturlesen danken.

Ein besonders großes Dankeschön geht an Frau Dr. Nicole Kemper für ihre Hilfsbereitschaft

und Unterstützung, der ständigen Bereitschaft zum Korrekturlesen und ihrer wertvollen

Anregungen bei der Anfertigung dieser Arbeit.

Der Vermarktungsgemeinschaft für Zucht- und Nutzvieh (ZNVG, Neumünster) danke ich für

die Unterstützung bei der Auswahl der Betriebe.

Allen Landwirten und dem Schlachthof Jensen (Oldenburg i.H.) möchte ich herzlich für die

Teilnahme an der Untersuchung danken. Die unkomplizierte Art und Hilfsbereitschaft lassen

mich die Besuche auf den Höfen in guter Erinnerung behalten.

Das Projekt wurde ermöglicht durch die finanzielle Förderung der H. Wilhelm Schaumann

Stiftung, dem Ministerium für Soziales, Gesundheit, Familie, Jugend und Senioren des

Landes Schleswig-Holstein und der Arbeitsgruppe Lebensmittelqualität und -sicherheit

(QUASI) der Agrar- und Ernährungswissenschaftlichen Fakultät der Christian-Albrechts-

Universität zu Kiel, denen ich herzlich danke.

Für die schöne Zeit am Institut, dem guten und freundschaftlichen Arbeitsklima danke ich

allen Kollegen. Besonders möchte ich meinen „Containerkollegen“ danken, allen voran Lotti,

Imke und Diane. Danke, dass ich mit meinen Sorgen und Nöten bei euch immer auf offene

Ohren gestoßen bin und Danke für das freundschaftliche Verhältnis und die moralische

Unterstützung.

Der größte Dank gilt meiner Familie, die es mir durch ihren Rückhalt und ihrem

entgegengebrachten Verständnis ermöglicht hat, meine Promotion erfolgreich zu beenden.

LEBENSLAUF Name: Tanja Wehebrink Geburtsdatum: 01.05.1978 Geburtsort: Rahden Staatsangehörigkeit: deutsch Familienstand: ledig Eltern: Heinz Wehebrink, Inge Wehebrink (geb. Timm)

Schulbildung: 1984 – 1988 Grundschule Varl 1988 – 1994 Freiherr-vom-Stein-Realschule, Rahden 1994 – 1997 Söderblom Gymnasium, Espelkamp Abschluss: Allgemeine Hochschulreife

Berufsausbildung: 1997 – 1999 Landwirtin

1997 – 1998 Betrieb Ernst Flömer in Gestringen (Milchvieh)

1998 – 1999 Betrieb Friedhelm Lange in Hille (Ferkelerzeugung u. Mast)

Studium: 1999 – 2002 Studium Agrarwissenschaften mit der Fachrichtung Tierproduktion an der Christian-Albrechts-Universität zu Kiel Abschluss: Bachelor of Science

2002 – 2004 Studium Agrarwissenschaften mit der Fachrichtung Tierproduktion an der Christian-Albrechts-Universität zu Kiel Abschluss: Master of Science

Berufliche Tätigkeit: seit Juni 2004 Wissenschaftliche Mitarbeiterin am Institut für Tierzucht und Tierhaltung der Christian-Albrechts-Universität zu Kiel bei Herrn Prof. Dr. Joachim Krieter