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CULTURE-NEGATIVE

ENDOCARDITIS

5/3/2012



Definition

Blood culture-negative IE is defined as endocarditis without

etiology following inoculation of at least three independent

blood samples in a standard blood culture system with

negative cultures after seven days of incubation and

subculturing



Incidence

Accounts for about 30% of all cases of endocarditis

worldwide

In developed countries-

Netherlands (1%), the USA (5%), Sweden (12%), the UK (15%), France (18%)

In developing countries-

Khan et al (JPMA 60:24; 2010) - 46.7% IE causes were culture

negative ( Pakistan)Ravi et al (Am heart J 2011)- 59% CNE (India)



Incidence

Variations in incidence accounted for by

(i) differences in the diagnostic criteria used

(ii) specific epidemiological factors, as for fastidious zoonotic

agents

(iii) variations in the early use of antibiotics prior to blood

sampling

(iv) differences in sampling



Factors Contributing to Sterility of Blood

Cultures

Antibiotic administration preceding blood cultures

More in developing countries due to unrestricted usage and availability of

antibiotics without prescription This situation arises in patients who received antibiotics for

unexplained fever before any blood cultures were performed

and in whom the diagnosis of IE was not considered

Causative organisms are most often oral streptococci , S.aureus or CNS.



Factors Contributing to Sterility of

Blood Cultures

Impact of Taking Antibiotics Before Blood Cultures Are Obtained

It¶s the leading cause of BC-negative IE.

Reduces the recovery rate of bacteria by 35-40%.

The length of time the cultures will remain negative is determined by

the susceptibility of the organism and the duration of antibiotic use

before blood cultures are drawn.

Cultures of pts who receive longer course of high dose, bactericidalantibiotics may remain negative for weeks.

If the initial cultures were negative after only a few days of antibiotics,

infective endocarditis pts may have a positive bacterial culture after

only several days without antibiotics.




Cultures

Fastidious slow-growing bacteria

Nutritionally variant streptococci, fastidious Gram-negative bacilli

of the HACEK group (Haemophilus parainfluenzae,

Aggregatibacter(previously Hemophilus) aphrophilus,

Aggregatibacter (previouslyActinobacillus )

actinomycetemcomitans,

Cardiobacterium hominis, Eikenella corrodens, Kingella kingae)and Brucella




Cultures

Intracellular bacteria

As Coxiella burnetii, Bartonella, Chlamydia, Tropheryma whipplei,

the agent of Whipple¶s disease

Non-bacterial organisms i.e. fungi

Patient factors

Prosthetic valves, renal failure, Pacemakers, Indwelling

intravenous catheters, immunocompromised states



Non-Infectious Causes of CNE

2.5% of CNE in recent series

Antiphospholipid syndrome: Primary or Secondary (e.g. SLE or

malignancies)

Neoplasia Associated:Atrial Myxoma, Marantic endocarditis,Carcinoid

AI associated: Rheumatic carditis, SLE, PA N, Behcet¶s disease

Postvalvular surgery: Thrombus, stitch, or other postsurgical

changes Miscellaneous: eosinophilic heart disease, ruptured mitral chordae,

myxomatous degeneration

Fournier et al. CID. 2010



Factors Contributing to Sterility of

Blood Cultures



Modified Duke criteria



Modified Duke criteria

By applying serology in Dukes criteria

8.9% patients upgraded from possible IE to definite IE

7.3% patients upgraded from rejected IE to possible IE

J Clin Microbiol 2005:43



Possible microbiological causes of CNE

HACEK organisms- traditionally thought to be the common

agents of culture-negative endocarditis but can be easily

isolated with current blood culture systems when incubated for

at least five days

The most common agents of CNE are fastidious organisms

(eg, zoonotic agents and fungi) and Streptococcal spp. in

patients who have received previous antibiotic treatment.

C. burnetii - a relatively common cause of IE in Europeancountries

Bartonella

Fungi



Possible microbiological causes of CNE

Coxiella burnetii 3-48%

Bartonella species 10-28%

Staphylococcus species 2-11%

Streptococcus species 1-6%

HACEK 0.5-3%

Fungi 1-6%

Candida,Aspergillus, Cryptococcus, endemic fungi, others

Tropheryma whipplei 0.3-3%

Others: Legionella, Chlamydia, Brucella

Fournier. CID.

2010;96.



When to suspect ..



In India

C. burnetii and Bartonella spp. were responsible for

8% of all infectious endocarditis cases and 14% of

blood culture±negative cases (Chennai)Emerg Infect Dis. 2008 July; 14(7): 1168±1169

There are also reports of brucella, trophyrema

(whipple¶s) endocarditis in India.



Value of positive cultures

Positive cultures have been a cornerstone in the diagnosis of

IE

Selection of appropriate antibiotic therapy based on thesensitivity pattern

Requirement to cover all likely organisms (in CNE) ±

increased chances of drug toxicities

Reported increased mortality and complications of CNE insome series



To increase yield of cultures

Avoid prior antibiotic usage

Atleast 3 sets

Blood subcultures on chocolate agar

Prolonged incubation may be needed

If the patient is clinically stable , antibiotics can be withheld

awaiting fresh cultures



Diagnosis

Special culturing techniques (eg, shell vial and lysis

centrifugation)

Molecular techniques (eg, polymerase chain reaction) Serologic assays

Histopathology evaluation of valvular tissue when surgical

excision is performed



Molecular techniques

Polymerase chain reaction ± most data for diagnosis of CNE

come from its use in the setting of explanted valve tissue but

can also be used in serum..

There are broad-range PCR techniques for amplifying16SrDNA (for bacteria) or 18SrDNA (for fungi), which can

then be sequenced for pathogen identification

The most frequently identified bacteria have included

streptococci, staphylococci, Bartonella spp., and T. whipplei .

There are real-time PCR techniques for diagnosis of specific

pathogens including Bartonella spp., C. burnetii (Q fever), and

Whipple's disease



Molecular techniques

Sensitivity of PCR for pathogen identification in CNE ranges

from 40 to 60 percent, with specificity nearing 100 percent

Limitations

False negative results - preoperative antibiotics, which probably decreasethe diagnostic yield of valve tissue PCR

False positive results - persistent nonviable bacteria months or years after

apparent cure , if valve tissue is contaminated in the process of surgical

removal or transport

³ A positive PCR test from valve tissue must be correlated with theknown propensity of the detected microbe to cause IE´



Serologic assays

The etiologic agents in culture-negative IE best identified by

serology include Coxiella burnetii, Bartonella spp.,

Chlamydophila (formerly Chlamydia) spp., Legionella spp.,

and Brucella spp

Coxiella burnetii phase I antibody titers >800 are diagnostic

for Q fever endocarditis

Bartonella IgG levels 800 are diagnostic for Bartonella spp.

IE . However, patients with Q fever may have low-level cross-

reacting antibodies to Bartonella spp. and/or Chlamydophila



Special culture techniques

Lysis centrifugation - improve yield of fastidious organisms

(eg, Brucella spp. and fungi) . This culture system contains

components that lyse leukocytes and erythrocytes, as well as

inactivate plasma complement and certain antibiotics, allows

release of intracellular microorganisms, and the centrifugation

step concentrates the organisms which can then be plated

directly onto supportive medium.

Shell vial cell culture - Of blood or excised heart valves

allows the isolation of Coxiella burnetii, Bartonella spp., T.whipplei, and Brucella spp. and has been useful in identifying

the etiologic agent in CNE.



Histopathology

Can be useful for pathogen identification of infectious

endocarditis as well as noninfectious mimickers includingmarantic endocarditis, rheumatic endocarditis, myxoma and

others.



Multimodal strategy

Comprehensive Diagnostic Strategy for Blood Culture±Negative

Endocarditis:A Prospective Study of 819 New Cases. Fournier et al CID

2010

Multimodal strategy incorporating serological, molecular, and

histopathological assays to investigate specimens from 819 patientssuspected of having BCNE

A causative microorganism was identified in 62.7%, and a noninfective

etiology in 2.5%

Blood was the most useful specimen, providing a diagnosis for 47.7% of

patients by serological analysis (mainly Q fever and Bartonella infections)

PCR of blood and Bartonella±specific Western blot methods diagnosed 7

additional cases. PCR of valvular biopsies identified 109 more etiologies,

mostly streptococci, Tropheryma whipplei, Bartonella species, and fungi

Histological analysis or searching for antinuclear antibodies yielded non

infectious cause in 2.5% of the patients



Therapy

Suspected NVE with confounding antibiotic therapy

Ampicillin ± Sulbactam plus Gentamycin

Or

Vancomycin plus Gentamycin and Ciprofloxacin

Suspected PVE

Vancomycin plus Gentamycin, Cefepime and Rifampicin



Therapy



Therapy

If no response to empirical antibiotic therapy,

surgical intervention both as therapy and to obtain

specimens is recommended.



Thank you

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