.· INTERNATIONAL BIDDETERIORATION

BuLLETIN

CATOMANCE LIMITED

manufacturers of

mustox* for the preservation of

timber, textiles paper, cordage

plastics and specialised applications

* myswx is the registered trade mark of

Catomance Limited 94 BRIDGE ROAD EAST, WELWYN GARDEN CITY,

HERTS., ENGLAND. Telephone: Welwyn Garden 24373/8

Volume 18 Number 1 Spring 1982 ISSN 0020-6164

I NTERNA 1/0NAL BIODETERIORA 1/0N

BuLLETIN

CONTENTS

Biodeterioration Society Newsletter

Page No.

Abstracts of Papers presented at a Symposium on Modern Methods for Detecting Microbial Spoilage iii-vii

Airborne Algae around Singapore 1-5 Y.C. Wee Algues dans I' air auteur de Singapour Algeo in der Luft von Singapur Algas del aire en los alrededores de Singapur

The Effects of Metals upon the Inhibitory Activities of Cutting Fluid Preservatives 7-12 E. 0. Benne//. i.E. Gannon and !. U. Onyekwelu Les effets des metaux sur !'action inhibitrice des produits de preservation des liquides de coupe Der einfluss von Metallen auf die hemmenden Eigenschaften von Schutzmitteln fiir Kiihl- und Schmierstoffe El efecto de metales sobre Ia actividad inhibitoria de liquidos lubricantes antisepticos

Subterranean Termite Control: Results of Long-term Tests. 13-18 Raymond H. Beal and Ralph W. Howard Contrille des termites souterrains: resultat d'essais a long terme Bekiimpfung von Erdtermiten: Ergebnisse von Langzeitversuchen Control subterraneo de termitas: resultados de pruebas a largo plazo

A Means of Dispersal of Botryodip/odia theobromae 19-21 S.K. Ogundana Un moyen de dispersion de Botryodiplodia theobromae Zur Miiglichkeit der Verbreitung von Botryodip/odia theobromae Un medic de dispersion de Botryodip/odia theobromae

Shorter Communications 22

Degradation of Tree Bark by Thermophilic and Thermotolerant Fungi from Nigerian Municipal Compost 23·24 C./.C. Ogbonna. G.J.F. Pugh and H.O.W. Eggins Degradaci6n de Corteza de arbol por bongos termofilos y termotole~antes en Compost en Nigeria. Der Abbau von Baumrinde durch thermophile und thermotolerante Pilze in stiidtischem Miill in Nigeria Degradaci6n de Corteza de arbol por hongos termofilos y termotolerantcs en Compost en Nigeria.

The Biodeterioration of Ethanediol Antifreezes by Penicillium simplicissimum 25·2(; R.Dale. T. Rubidge and G. Williams • La deterioration d'antigels ethanediols par Penicillium simplicissimum Der biologische Abbau von Athandioi-Frostschutzmitteln durch Penicillium simplicissimum Biodeterioracion de anticongelante de etandiol por Penicillium simplicissimum

INTERNATIONAL BIODETERIORATION

BULLETIN

BIODETERIORATION CENTRE UNIVERSITY OF ASTON ST. PETER'S COLLEGE, SALTLEY, BIRMINGHAM B8 3TE.

Editor-in-Chief of Biodeterioration Centre Journals Dr H 0 W Egg ins Editors Professor T A Oxley and Christine Allsopp Business Manager Dr D Allsopp

The Editors are able to call upon the assistance of an Editorial Board whose members are in Britain, various countries of Europe, and the U.S.A.

NOTES FOR CONTRIBUTORS The International Biodeterioration Bulletin is published four times per year (Spring, Summer, Autumn and Winter). Typescript contributions should be sent to the Editors, at the above address. The Bulletin acts as a vehicle for the publication of original works, incuding reviews. on all aspects of biodeterioration, i.e., deterioration of materials, artefacts or facilities, of economic importance by living organisms, which include microorganisms, insects, rodents, birds, higher plants, etc. Articles on biodegradation, that is conversion of materials to less objectionable, more easily disposable, or higher value products by living organisms, are also published. Contributions are published only in. English. Each article must be accompanied by a summary in 50- 150words which will be translated into French, German and Spanish. Native speakers of these languages are invited to submit their summaries in their own language; in certain circumstances complete articles may be submitted in French, German or Spanish and will be translated into English for publication. Illustrations must be very clearly drawn, normally larger than the size finally desired. The suggested final size should be clearly indicated but the the Editors reserve the right to vary this in the interest of economy and

As far as possible diagrams will be reduced to single column width (80 mm) or to half page (170 mm). In any event, neither these nor half tone photographs can exceed full page (260 by 170 mm). Authors should bear in mind that it is generally more convenient for readers if legends which accompany diagrams or photographs appear with them on the same page and should proportion their illustrations accordingly. Lettering on diagrams will normally be inserted by the printer; authors are therefore asked to insert lettering or symbols in pencil on the originals or preferably, in ink on a photocopy. All articles are submitted by the Editors to one or more independent referees for advice on their clarity, originality, and general suitability for publication, but the final decision whether or not to publish an article rests with the editors. If articles are rejected the substance of the referee's report will usually be communicated to the author and in suitable cases the Editors will be pleased to help suthors to improve their papers with a view to possible publication. Bibliographic references are indicated in the text by author names (no initials} and year only, viz: Reese and Levinson (1952); or: Darby et al .• (1968) and in the bibliography in strict alphabetical order of first authors names, thus:

or:

Reese E T and Levinson H G ( 1952) Comparative study of the breakdown of cellulose by microorganisms Physiologica Plantarum 5: 354-366

Darby Fi.T. S1mmons ·E G and Wiley B J (1968) A survey of fungi in a military aircraft fuel supply system. International Biodeterioration Bulletin~( 1 I ~9-:J1

References to books, conference proceedings, etc. should quote first the author(s)or editor(s), then the year of publication and title followed by the city in which it is published and the name of the publisher. As far as possible titles of journals should be given in full except for such abbreviations as 'Journ'., 'Proc'., 'Trans.' etc. 20 reprints will be sent free of charge to the first named author unless otherwise instructed. Any number (normally not more than 50) of additional reprints may be purchased it ordered sufficiently in advance. An order form and price will be sent giving about one month's notice.

clarity. MAJOR SUBSCRIBERS

ALBRIGHT & WILSON (MFGI LTD., Oldbury Division, P.O. Box 3, Oldbury, Warley, Wares .. England.

BRITISH INSULATED CALLENDERS­CABLES LIMITED, 38 Wood Lane, London, Wl2, England.

BP CHEMICALS INTERNATIONAL LIMITED.

CATOMANCE LIMITED. Welwyn Garden City, Hertfordshire, England; manufacturers of speciality chemicals for the textile, paper, timber,leather industries, etc., including fungiCides, bactericides and insecticides.

FARBENFABRtKEN BAYER A.G •• Leverkusen, Germany; manufacturers of dyestuffs, industrial chemiculs. synthetic fibres, pharmaceutical and agricultural chemicals and preserva­tives for wood, foodstuffs and technical products.

STERLING INDUSTRIAL LIMITED. Chapel town, Sheffield, England,

HALDANE CONSULTANTS /LIMITED. 27 Dawkins Road, Poole, Dorset, BH15 4JB; consultants in industrial microbiology and microbiological deterioration.

ii

IMPERIAL CHEMICAL INDUSTRIES LIMITED, Agricultural Division, Billingham, Co. Durham, England.

LUCAS AEROSPACE LTD., Shaftmoor Lane, Birmingham, England. B2B asw.

NATIONAL COAL BOARD, Coal House Lyon Road, Harrow, Middlesex, England.

RENTOKIL LIMITED, East Grinstead, West Sussex, England.

lnt£"rnat[onal Biod£'taioration Bul/£"tin ISSN 0020..6164 18(1) Spring 1982

BIODETERIORATION SOCIETY NEWSLETTER

Summer Meeting and A.G.M.

The annual summer meeting will be held at Portsmouth Polytechnic on July 8th and 9th 1982. The subject of the main symposium will be:

Timber Deterioration and its Prevention

The Annual General Meeting of the Society will be held in connection with this meeting at 9 am on Friday 9th July.

Another important event will be the first session of presentations for the award of the Bunker Memorial Prize. This is timed for 4 pm on Thursday 8th July.

Wood Preservation Group to meet in Australia

The 'I.R.G.' (International Research Group in Wood Preservation) is to hold its 14th Annual Meeting in May 1983 in Australia. The meeting will be held during the week 9 - 13th May and will be followed by a technical-cum-sightseeing tour down the east coast from Surfer's paradise to Melbourne. Chairman of the organising committee is Dr. Harry Greaves of the C.S.I.R.O. Division of Building Research, Graham Road, Highet!, Vic. 3190. The Secretary General of I.R.G. is Mr. Ron Cockcroft whose office is at:

Drottinig Kristinas viig 47 C S - 114 28 Stockholm, Sweden

Abstracts of papers presented at a

Symposium on Modern Methods for Detecting Microbial Spoilage

held at Long Ashton Research Station, University of Bristol on

15- 16 April 1982

Title: A New Look at Older Methods

Author: R.R. Davenport

Address: Long Ashton Research Station University of Bristol Long Ashton, BRISTOL BSI8 9AF

Abstract

The advance of technology has influenced some trends in microbiological methods. For example, automated methods are becoming more commonplace. This is desirable and indeed essential for industries where large 'volumes' of materials and/or products have to be checked for microbial contamination. However, modern methods have limitations (e.g. expensive apparatus and trained operators/interpreters), whereas some reliable conventional procedures can be very economic and give results which can indicate more than one feature. Too often conventional microbiology has been applied directly from one set of organisms to another without due regard to microbial and environmental characteristics.

This paper re-examines some topics (e.g. swabs, enrichment cultures, microscopy and incubation temperatures) which can benefit those investigators working with low budgets, particularly in industries and developing countries.

iii

Title: Bryn C. Hughes

Address: Department of Science Bristol Polytechnic Coldharbour Lane Frenchay, BRISTOL

Abstract

Adenosine triphosphate (ATP) can be assayed quantitatively and rapidly using the light production from the luciferin-luciferase enzyme complex. Although this reaction has optimum conditions for maximal light output (egionic concentrations, pH and temperature), it has been successfully employed in the estimation of the toxic effects of specific heavy metal treatments upon unicellular algal suspensions. In addition, this technique has shown good correlation with other screening methods, when antifouling leachate solutions have been used as the toxic sample. In both studies, a short exposure period of 24 hours was employed; a significant reduction in screening time when compared to existing toxicity testing methods.

ATP estimates have been widely used to determine biomass in various environments. Levels of microbial contamination in DIESO fuel samples have been difficult to quantify, however , preliminary studies have shown that ATP estimates from such fuel samples compare favourably with corresponding plate count determinations.

Biod('t~riOWihlll s~l(,'ictr Newsletter, Spring 1982.

Title: Experience with Dairy Products with Particular Reference to Impedance Methods

Author: D.W. Matthews

Address: The Nestle Co. Ltd.

Ab.\'/I'(IC/

Quality Assurance Deparrment St. George's House Croydon Surrey CR91NR

Bacteria in pasteurised cream and cream desserts, and yeasts in yoghurt, were estimated impedimetrically with a Bactometer 32 instrument. A direct comparison with standard plate counts of impedance detection time was made and a good correlation was found except for counts of less than 100 per gram where sampling error and the type of microbial flora present influence the results. The detection time for various creams and desserts was compared with their keeping quality in storage at a market simulation temperature of II °C. Products with a shorter detection time (indicating that there are more bacteria present) showed a greater degree of deterioration. The detection time determined at 30°C in the Bactometer correlated quite well with spoilage of the product after storage at li°C. Impedance estimation gives us a chance to tailor growth media more to the characteristics of the products under examination and thereby estimate the ability of the microbial flora present to spoil the food.

Title: Some Particular Problems of Testing Emulsions

Author: E.C. Hill '

Address: University College, Department of Microbiology, P.O. Box 97, Cardiff CFI IXP

Ab11rau

A recent innovation to speed up conventional plate counting has been the introduction of the "Spiral Plater". Experience has shown that if metal working emulsions containing preservatives are tested there may be sufficient carry over of the antimicrobial, particularly in the inner whirls of the spiral, to inhibit colony formation. The method then underestimates the total count. One major benefit of "Spiral Plater" counting has been , that to isolate the dominant organisms, one need only locate the outer end of the spiral and work inwards, picking off colonies in turn.

A foreseeable advance in controlling microbial spoilage of metal-working emulsions would be the

iv

development of an in-line system for detecting microbial contamination and growth. Emulsions present particular difficulties; several methods have been investigated and. with certain reservations. the impedance method selected for development.

Title: Bactec 460 Automated Radiometric Microbiology System

Author: Jayne M. Lewis

Address: Laboratory lmpex Ltd. Lion Road Twickenham, Middlesex.

Abstract

The Bactec 460 is an automated radiometric microbiology system. It utilizes a culture vial containing 30 ml of a culture medium in which many of the substrates arc labelled with a radioactive isotope, carbon-14. These vials are inoculated with a sample and incubated. If microorganisms are present in the sample, they will utilize the substrates and carbon-14 is released. This diffuses into the gaseous layer at the top of the bottle. The Bactec instrument automatically removes this head space gas and examines it for radioactivity, at a rate of one bottle per minute. Applications of the system are being developed to include sterility testing, limit testing, and microbial enumeration.

Title: The Direct Epifluorescence Filter Technique

Author: G.L. Pettipher

Address: National Institute for Research in Dairying Shinfield. Reading, Berks.

Abs/racl

The Direct Epifluorcscence Technique (DEFT) which was originally developed for milk, is a rapid, sensitive, direct method for enumerating microorganisms using membrane filtration and epifluorescence microscopy. It takes less than 30 minutes to complete, costs less than £0.40 per sample in disposables, and is suitable for milks containing between 0.5 x 104 and 108 bacteria per mi. For use on foods a preliminary pre-filtration through a 5].tm pore size nylon filter of a 'stomached' suspension is necessary to avoid difficulty caused by debris. With this pre-treatment DEFT can be used for meat, fish, vegetables, cakes, spices and powders with a maximum sensitivity of 0.6 x 105 cells per gram and correlates well with the unfiltered plate count, especially for fresh meat and fish.

Title: Modern Methods for Detecting Microbial Growth on Surfaces.

Author: A.M. Paton

Address: School of Agriculture, University of Aberdeen, 581 King Street, Aberdeen, AB9 IUD

Abstract

The increasing availability of the epilluorescence microscope offers excellent opportunities fort he rapid and detailed examination of surfaces associated with biodegradation. Using a mixed lluorescent stain containing acridine orange and a cationic optical brightener directly on suitable surfaces or on adhesive tape impressions, fluorescent emissions at either short or long UV wavelengths clearly visualise the fungal or bacterial microllora.

lmrnmtimwl Bimlctl'rirmllirm Bullt•tiniSSN 0020-6164 18(1) Spring 19!U

v

Title: The Detection and Quantilication of Fungal Spoilage on Material

Author: Susan Ward

Address: C.L.E.A.R.

Abstract

Huntingdon Road Laboratories, Cambridge. CB3 ODJ

Degraded samples of cotton duck and painted/varnished surfaces show fluorescence under UV light corresponding to viable fungal hyphae after being stained with fluorescein diacetate (FDA). This nuorcscence can be quantified using a densitometer to scan the sample. By staining liquid cultures of fungi with FDA (repli£ate llasks being used for dry weight determinations) the lluorescent counts obtained for the degraded samples can be quantified in terms ofmg dry weight of fungus.

Degraded plastic and rubber samples show little, if any, viable hyphae, however the pitting and discolouration produced on the surfaces as a result of fungal attack can be measured by a densitometer and used to produce a semi-quantitative estimation of growth which is more accurate and reproducible than current British Standard test ratings for measurement of biodeterioration.

BIODETERIORATION CENTRE SHORT COURSE PROGRAMME 1982

VC 825- 5-14 July 1982

Biodeterioration

A comprehensive residential course of lectures, demonstrations, seminars, visits and practical work giving a basic background in many aspects of materials biology. The course, which is based on a previous UNESCO­UNEP sponsored course will be of benefit to postgraduate biologists and industrialists in biological industries, such as biocide manufacturers and consulting scientists.

Cost: including full board residential accommodation - £250.00

NOTE: Some sponsored places may be available to studellls from Developing Countries/

VC 826- 27 (& 28) September 1982

Moisture in Materials

Problems caused by moisture in materials and subsequent biological effects will be covered, together with techniques for monitoring and measurement. A second day on identification of spoilage fungi is planned as an optional extra.

Cost: including lunch and refreshments - £35.00

Identification course including Ol'ernight accommodation - £50.00 extra

VC 827 - 12 October I 982

Chemical Preservation of Materials

Lectures on various aspects of chemical preservatives for both solid and liquid systems will be given and there will be a commercial exhibition hy manufacturers. This course will be held either at the Biodeterioration Centre or in a hotel in the Midlands.

Cost: including lunch and refreshments In he announced but probably in the region of £50.00

VC 828 - 26 October I 982

Toxins in Food and Feedstuffs

The course will concern itself with toxic agents of biological origin in both human and animal foodstuffs. Special emphasis will be given to Mycotoxins.

Cost: including lunch and refreshments - £35.00

VC 829- 9 November 1982

Weed Control

Various aspects of weed control (physical. chemical and biological) will be considered both in agricultural and amenity situations. Of interest to agriculturalists, local authorities and building conservators.

Cost: including lunch and refreshments - £35.00

All courses, unless otherwise stated, are held at the Biodeterioration Centre. Further details of courses and booking forms may be obtained from:

Dr D Allsopp, Course Organiser, Biodeterioration Centre, University of Aston in Birmingham, St. Peter's College, College Road, Saltley, Birmingham BS 3TE.

Telephone: 021-328-5950 Telex: 336997 (Mark .for allen/ion Biodeterioration Centre)

The Centre reserves the right to amend this programme and to cancel any courses should there be insuf{icielll support to run them economically.

vi

lntemaliollal Hiodl'tl'rioration Bulletin ISSN 0020-6164 18(1) Spring 1982

THE BIODETERIORATION SOCIETY

APPLICATION FOR MEMBERSHIP

I wish to apply for membership of the Biodeterioration Society I enclose a Cheque/Postal Order/Money Order* for £4.50/£3.00* as my subscription for the calendar year 19 ....

(Please type or write very clearly)

Surname (with usual title) .............................................................. .

Forenames .................................................................. , ........ .

Address ............................................................................. .

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Place of work ........................................................................ .

Name of one person, a member of the Society, who is known to you, whom we may approach as your sponsor

Cheques should be made payable to the Biodeterioration Society. Please send this completed form, with your remittance, to the Secretary of the Society:

Mrs. Joan Maw, Secretary, The Biodeterioration Society C. P. Snow Building, The Hatfield Polytechnic, P.O. Box 109 HATFIELD Herts. ALIO 9AB

• Please delete whichever does not apply.

£4.50 for members in Great Britain and Ireland (includes meetings fee) £3.00 for members in other parts of the world (who may be asked by local meetings secretaries in their

own countries to pay an additional meetings fee for meetings organised in their area).

Reduced Rate Subscriptions to Journals Published by the

Biodeterioration Centre

As a member of the Biodeterioration Society you will be entitled to a reduced rate personal subscription to the International Biodeterioration Bulletin and/or the bibliographic journal Biodeterioration Research Titles (B.R.T.) Currently this reduction amounts to about 44%. The rates are given inside the back cover of the International Biodeterioration Bulletin.

If you are interested in this possibility, please tick the box and return this part of the form with your application to the secretary.

I may be interested in a personal subscription to either or both of LB. B. and B.R.T.

Do not send money for publications with this form. You will be invoiced directly by the Biodeterioration Centre. You will be asked to undertake that your subscription is for your own personal use and not for the Institution with which you are associated.

vii

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t

I

lntemational Biodetaiorarion Bulletin ISSN 0020-6164 18(1) Spring 1982

AIRBORNE ALGAE AROUND SINGAPORE

Y C Wee'

Summary

21 samplings of algae were collected from 71 air samplings obtained from various locations in Singapore and two from Malaysia. Of these, 18 species from II genera were blue-greens. The most prevalent species was Schizothrix ca/cico/a, a filamentous form, occurring in 80% of the samples. The unicellular Anacystis (A. therm(J/1,\·, A. montana and A. dimidiata) made up the next three most common species. There were, in addition, two species from two genera of green algae and one species of diatom. Weekly air samples showed a peak number of colonies per sample towards the last week of April 1981, with a second, lower, peak in August 1980. No definite correlation was seen between rainfall and the amount of airborne algae but heavy rain before or during the sampling reduced the amount of algal pollution.

Algues dans l'air,autour de Singapour

On a collectC 21 especes d'algues iipurtir de 71 prCii:vements d'airen divers sites ii Singapour et 2en Malaisie. Parmi elles, I 8 espi::ces dans II genres Ctaient des algues vert-blue. L'espece dominante fut Schizothrix ca/cico/a, de forme filamenteuse, dans 80% des Cchantillons. L'unicellulaire Anacystis (A. thermo/is, A. montana, et A. dimidiata) forma les 3 autrcs espCccs lcs plus courantcs. II y a, en plus, deux espi::ces de deux genres d'algues vertes et une espi::ce de diatomee. Les prCii::vements d'air hebdomadaires ant montrC un pic des colonies vers Ia derniC:re semained'Avril 1981 avec un second pic plus faible en Aolit 1980. On n'a pas vu de correlation dCfinie entre les chutes de pluie et le montant des algues dans !'air mais, de fortes pluies avant ou pendani le prl:li:vement, a rl:duit le taux de Ia pollution par les algues.

Introduction

Microscopic algae are widespread. Their presence in Singapore has been demonstrated in various habitats: in different types of soils (Johnson, 1962; 1973a); in bodies of water (Johnson, 1973b; 1978); on bark of trees along the wayside (Johnson and Awan, 1972); and on walls of buildings in built-up areas (Lee and Wee, 1981). Their presence in most cases can be beneficial. Heteroceous blue-green algae fix free nitrogen, thus improving the fertility of the soils in which they exist. On trees, they help modify the habitat, paving the way for the subsequent establishment of epiphytic ferns and orchids, whose presence enhances the aesthetic value of these wayside trees (Wee, 1978). In ponds and lakes these algae are the primary producers, the starting points of many food chains and food webs. However, on walls of buildings they disfigure the facade, making early redecoration necessary (Wee and Lee, 1980).

The abundance of such habitats supporting prolific algal growth must result in cells and bits of filaments of these organisms polluting the air. As far back as 1937

Algcn in der Luft mn Singapur

Einundzwanzig Algenarten wurden aus 71 Luflproben aus vcrschiedenen Gebieten in Singupur und und zwei aus Malaysia bcstimmt. Achtzehn Arten, die zu II Gattungen gehOrten, waren blaugri:ine Algen. Die hllufigste Art war Sclrizuthrix ca/cicolu, cine Fadenalge, die in 80% der Proben auftrat, gefolgt von 3 einzelligen Anacysti.f-Arten (A. thermo/is. A. montana und A. dimidiata). Auaerdem gab es zwei Arten aus zwei Gattungen von Grilnalgen und cine Diatomeen-Art. WOchentlich der luft entnommene Proben wiesesn die groate Anzahl von Kolonien pro Probe ungeHi.hr in der lctzten Aprilwochc 1981 auf. Ein zweites weniger ausgeprfigtes Maximum trat im August 1980 auf. Es gab keine eindeutige Korrelation zwischen den Niederschlagen und der Anzahl der Algen in der Luft, aber heftige Niederschlage vor oder wah rend ver Probenahme verminderten die Menge der Algen in der Luft.

Algas del Aire en los Alrededores de Singapur

Apartir de 71 muestra de airc procedentcs de varias localidades de Singapur y 2 de Mal asia. Se han aislado 21 especie de algas. De estas 18 especies de II generos fueron verde azuladas, Ia especie mas extendida fue Schizotlrrix ca/cicola, filamentosa que apareci6 en el 80% de las muestras. Las especies Anacysris thermo/is, A. montana y A. dimitliata fueron las tres especies siguientes. Hubo ademas 2 especies de 2 generos de algas verdes y una especie de diatomea. Muestras de aire semanales tubieron un nlimero maximo de colonias en Ia ultima semana de Abril de 1981, siguiendole en cantidad, Agosto de algas en el aire. Pero una pricipitaciOn elevada antes o durante Ia toma de muestras rcdujo Ia poluci6 par algas.

airborne algae were shown to exist when Overeem pioneered their collection from an airplane at various altitudes over the Netherlands. Sub'sequent work by others confirmed the presence of viable protozoa and fungal spores, in addition to algae, in the air (Brown. Larson and Bold, 1964; Gregory, Hamilton and Sreeramulti, 1955;Hamilton, 1959; Schlichting.l961). The presence of airborne algae if in sufficient quantity, can pose problems, as they can cause inhalent allergy. It was with this in mind that a series of surveys were initiated in February 1980 with the objective of finding out the extent of algal pollution in the air around Singapore. Also, such information would contribute to our understanding of algal infestation of building walls, especially the initial colonisation and the subsequent spread.

Materials and Methods

The air was sampled for algae by exposing a petri dish half-filled with Bold's enrichment medium (Nicholas and Bold, 1965) solidified with I% agar. The dish was

Department of Botany, National University of Singapore. Kent Ridge. Singapore 1025, Republic of Singapore. (Received, October 1981).

Airborne: Algae around Singapore Y C Wcc 1

exposed from a moving car travelling at approximately 50 km/hour for a specific time. The exposed dish was incubated in an inverted position in the laboratory under 12 hours daily nuorescent lighting for two weeks. The inverted position was necessary to prevent water condensing on the agar surface. This would result in more colonies developing, as the film of water would disperse the algae all over the agar surface. The colonies formed after two weeks were then examined under the microscope and the algae identified. The number of colonies of each species was also noted.

Initially, various areas around Singapore were sampled to study the species and quantities of algae occurring in the air around the island. In such samples the dishes were exposed for five minutes and samplings were made at various times of the day. As far as possible, rainless days were chosen. A total of 19 localities were sampled to represent the various areas around Singapore. Two samples from Kuala Lumpur, Malaysia, were included for comparison. All samplings were done during February-April 1980.

Subsequently, weekly samples from a single area were made. A stretch of road from the former Bukit Timah Campus of the University to the author's home was chosen as this route was travelled daily. Prior to choosing the time of sampling, six samples were taken on each of two days in May 1980 between 0800 and 2200 hours. Based on the results from this, evening sampling was decided upon. Weekly sampling on every Monday from May 1980 to May 1981 was carried out between 1700 and 1800 hours. Where it was not possible to sample on a particular Monday, the following day was substituted. In this series of samplings the petri dishes were exposed for about 10 minutes along the route of approximately 10 kilometres. The weather condition at the time of sampling was noted.

Results and Discussion

Results of the samplings made from the various localities are given in Tables I, 2 and 3. A total of 21 species of algae from 71 locations in Singapore and two in Malysia were identified. There were only 14 genera, of which 112 were Cyanophyta (blue-green). .r::;chizothrix calcico/a, a common blue-green in most marine. freshwater and sub-aerial habitats, and of wide distribution around the world ( Drouet, 1908), was the commonest species. The other species present in significant numbers were species of Anac:l'.Wis, another blue-green.

In other parts of the world, by far the greatest quantity and di,·ersity of airborne algae have been collected by Brown. Larson and Bold ( 1964) from around Austin, Texas, who foiund 38 genera of Chlorophyta, 17 of Cyanophyta and 7 of Chrysophyta. The commonest fenera collected by Brown el at were those of Chlorophyta: Ch/orc/la. Ch/orosarcinopsis, Namwch/ori\', Protocoa·us and Trebouxia. Similarly.

1

Overbeen (1937) found Ch/orococcum common, and Schlichting ( 1961) reported the prevalence of Ch/orococcum and Ch/orella.

In the present work the greatest number of species found on a single petri dish coming from a single sampling, was six. and the greatest number of algal colonies developing was 37. There were eight samples, one of which came from Malaysia, which were free from algae. There was no indication of correlation between species of algae and the sampling locality, which included built-up urban, industrial, and rural areas. Nor was there any definite correlation between monthly rainfall and fluctuations in airborne algae in Singapore (Figure 1). However, it was noted that the numbers of airborne algae were low if substantial rain fell earlier during the day of sampling, or if it was raining heavily during the time of sampling. Slight drizzle during sampling, however, did not lower the algal content of the air. On the other hand, a low algal count did not necessarily indicate that rain had fallen earlier.

Airborne algae are relevant to the study of allergens. Atmospheric pollen has long been known to cause allergic response in susceptible individuals (Hill, Smart and Knox, 1979; Knox; 1979; Sharwood, 1935; Specht, Brouwer and Derrick, 1975). Similarly fungal spores have been reported to cause allergy (Clarke and Mason, 1964; Frey, Cross and Durie, 1963; Gale, 1969). The role of airborne algae as causative agents in inhalent allergies has been recognised but its study has not been intensive (Bernstein and Safferman, 1970; McElhenney, Bold, Brown and McGovern, 1962; McGovern, Hayward and McElhenney, 1966). The present investigation of algae present in the air around Singapore may, therefore, prove useful in future allergic studies.

The total absence of Trcntepohlia adoral a from all the samples is rather surprising. This green alga grows profusely on the walls of many high-rise buildings in Singapore (Wee and Lee, 1980). It is wind dispersed as the sporangia are detachable and thus blown away to infest other buildings. The absence of this alga may be due to its relatively slow growth rate. It has been observed in the laboratory that any blue-green species would literally overgrow it within a few days. On the other hand, T. odorala may have seasonal sporangia I production and the timing of the present samplings may not have coincided with these.

On walls, T. odora/a paves the way for the appearance of other algae, mainly blue-greens. Although this species was not detected in the air samples, the blue­greens that appear subsequently on walls were common. Anacystis thermalis and 4. montana. two unicellular species common on walls. were the most common algae after S. calcico/a. Of the other blue­green algae collected from the air, many were also found on walls. It is thus obvious that airborne algae are responsible for the spread of these organisms which proliferate on walls of buildings.

_::. ~ ..c ·;;

-~ a ·;; " ~c " :::: ~ ~ " ~

Table I. a "' ::;: t::l ::: :::: '" " ~ N

" 0 -~ Qd :;:: > :::: - :S! r::_o::: :::: ·c: v

"' .;: E ~ ~ § Qd

"' s:: . t::l ·-· s: t::l .::: 0 -~ t(J 0 'G ~

"' 0 ~ :§>.~ ~ ~ -~~t: ~ 0 v - 1:::0 :: ~ § c. " Number of colonies and species of algae collected from 19Jocations in "'iio 2" :::.--::: 1: .-~ ~ ..:::: :._ ~ ::::: ~ ci. u ~ c "' ~"""": ~0 " ~ '- '-·~

Singapore and two in Malaysia. -~ ::c § - :.:;: {':! ·- :::o .... I::: .... ·- ci. 0 0 c. -...c :::- t::l ..... Cl - :::: -~ ~- ~ - ..... v '"' ~

"' .... , -~ ffl -~ ~ E ~ -;:: ~ ~ ~:- ~ :g -~ ~ ~ g ~ ~ e. 0 0 .,. "' ] - "' ;::; c ::::; ..c "' :::~ ~. ~c-..o~a -E::~csE ~~ " " 00 Locality Time (hrs) Type of Area Weather " 01) t·i t·:j ·~< " ~eJ'Jr_.,'l-.-t:lO ~;:E~eJ'J (Sz.~O ~0. g -;; -;;

""~ g- ~-::; t· ~ ~ 0 g ~ ~ ._ ~ ~ ~ ._ "'E 0 ~ en ~

~ ~ ;:.; Conditions <5:l " ~ ~ "' "' <. 0o '<'<<0 vOov

"' 0 N

0900 g Adam Road Urban Hot, cloudless - - - - - - - - - 0 0 z Dalvcy Road 0800 Urban Hot, cloudless - - - - - - - - 0 0 "' - -~ Holland Road 0800 Urban Hot, cloudless 3 3

~ - - - - - - - - I

Clementi Road 0800 Urban Hot, cloudless - - - 2 - - - - - - - 2 I <S Braddell Road 0900 Urban Hot, cloudless 5 - 4 - - 7 - - - - - 16 3 c -~ Ayer Rajah Road 1000 Urban Hot, cloudless 3 2 2 - - - - - 7 3 ~ .e Holland Road 1000 Urban Hot, cloudless I I - - 2 - - - I - - - 5 4 ~ ~ Bukit Timah Road 1300 Urban Hot, cloudless I - - - - - - - - - - - I 1 -E:

Upper Serangoon Road 1300 Urban Hot cloudless 2 2 1 "' - - - - - - - - - - -c

Quccnsway 1330 Urban Hot, cloudless 2 - - 1 - 2 - 5 3 c - - - -0 -~ Telok Blangah ~

~ Road 1300 Urban Rained in "' morning 2 I 2 I - - - - 1 7 5 Macpherson Road 1600 Urban Hot, cloudless 4 I I - - 3 - - - - - 9 4 East Coast Road 1700 Rural Rained in

morning 2 1 - 3 I - - - - - - 7 4 Upper Changi Road 1530 Rural Light drizzle - I - - - - - - 1 I Upper Bukit Timah Road0800 Rural Drizzling I - 2 - - I 3 - - I - - - 8 5 Yio Chu Kang Road 0900 Rural Hot, cloudless 1 - 1 - - - - - - - - - 2 2 Punggol Road 1300 Rural Hot, cloudless 3 I - - I - - - - 5 3 Pan Island Expressway 1700 Forest Rained in

morning 3 2 - - 1 - - - - - - - 6 3 Off Jardine Steps 1000 Ocean Light drizzle 1 - - - - _I - 2 2 Highway into Kuala

Lumpur, Malaysia 1200 Plantations Hot. cloudless - - - - - - - - 0 0 Kuala Lumpur city

area,Malaysia 1230 Urban Hot, cloudless 2 3 I - - - - 1 7 4

TOTAL: 33 14 II 7 6 6 8 3 2 1 1 1 1 1 95 No. of occurrences (out of 21): 14 10 6 4 4 3 2 1 1 % occurrences: 66.7 47.6 28.6 19.0 19.0 14.3 9.5 4.8 4.8 4.8 4.8 4.8 4.8 4.8

Airborne Algae around Singapore Y.C.Wee ...

Table 2.

Number of colonies and species of algae collected at six different times on two days along the same stretch

of Bukit Timah Road.

Time No. of colonies No. of species (hours)

Day! Day II Day! Day II

0800 3 2

1100 3 3 2 3

1400 5 4 3 2

1700 8 6 4 4

2000 2 2 2

2200 5 4 3 2

Figure I

Number of algal colonies per weekly samples (shaded) and monthly rainfall (unshaded) for May 1980- May 1981 (ar =after rain; d =drizzle; r =heavy rain).

40 400

30 300

"' !'! z g2o 0

:200 ...

u ... 0

0 z

10 100

0 0

MJJASON OJFMAM 1980 19SI

3 •

Table 3.

Frequency occurrences of algae and total number of colonies developing from the 52 samples taken along Bukit Timah

Road.

Species Occurrences No. (%) colonies

Schizothrix calcicola (Agardh) Gamont 81 !52 Anacystis thermo/is (Menegh.) Dr. & 69 92 Daily Anacystis montan (Lightf.) Dr. & Daily 38 41 Anacystis dimidiata Dr. & Daily 33 41 Tetracystis sp. II 18 Coccoch/oris stagnina Sprengel 10 Westiellopsis pro/ijica Janet 8 Anabaena oscillarioides Bory 6 Calothrix parietina (Nageli) Thuret 6 Chlorococcum sp. 6 Scytonema hofmanni Agardh 4 Mastigoc/adus laminosus Cohn 4 Schizolhrix mexicana Gamont 4 Nostoc commune Yauch. 4 Fischerel/a ambigua (Nageli) Gamont 4 Schizothrix friesii (Agardh) Gamont 2 Schizothrix rubella Gamont 2 Anacystis marina Dr. & Daily 2 Oscillatoria /utea Agardh 2 Navicula sp. 2 Nostoc spumigena (Mert.) Dr. 2

Acknowledgements

I wish to thank the director, Meterological Service, Singapore, for providing the rainfall data.

4

REFENCES

Bernstein, I. L. and Safferman, R. S. (1970). Viable algae in house dust.

Nature, London, 227: 851-852.

Brown, R. M. Jr., Larson, D. A. and Bold, H. C . (1964) ..

Airborne algae: their abundance and heterogeneity.

Science, 143: 583-585.

Clarke, P. S. and Mason, P. E. (1964). Allergenic mould spores in Tasmania.

Medical Journal of Australia, 1: 192-194.

Drouet, F. (1968). Revision of classification of Oscillatoriaceae.

Fulton Press, Lancaster, Pennsylvania.

5 4 5 4 3 4 3 2 2 2 I I I I I I

Frey, D., Cross, D. 0. and Durie, E. B. (1963). Investigation of a series of samples of house dust for the presence of fungi: correlation with previous investigations of airborne fungi and sensitivity tests on patients.

Mycopathologia, 19: 83-86.

Gale, A. E. (1969). Review of air spora analysis in relation to allergic respiratory disease.

South Australian Clinics, 4: 56-61.

Gregory, P. H., Hamilton, E. D., and Sreeramulti, T. ( 1955).

Occurrence of the alga G/oeocap.l'a in the air. Nature, London, 176: 1270.

Hamilton, E. D. (1959). Studies on air spora.

Acta allergologica, 13: 143-175.

Hill, D. J .. Smart, I. J., and Knox, R. B. (1979). Childhood asthma and grass aerobiology in Mel borne.

Medical Journal of Australia, I: 426-429.

Johnson, A. (1962). Precursory studies on the epiterranean soil algae of Singapore and Malaya.

Gardens' Bulletin, Singapore. 19: 379-384.

Johnson, A. ( 1973a). A survey of the occurancc of epitcrranean soil algae in Singapore Island.

Gardens' Bulletin, Singapore. 26: 289-302.

Johnson, A. ( 1973b). The microllora of the University of Singapore pond.

Singapore National Academy of Science Journal, 3: 241-252.

Johns0n, A. ( 1978). The distribution of freshwater algae in Singapore.

I nstitutc of Natural Sciences. Nanyang University, Occasional Paper No. 35. 23pp.

Johnson, A. amd Awan, B. (1972). The distribution of epiphytes on Fagraca .fhtgrmu and .\'11'ietenia macrophylla.

Malayan Forester. 35: 5-12.

Knox, R. B. ( 1979). Pollen and allergy.

Edward Arnold, London.

Lee, K. B. and Wee, Y. C. (1980). A guide to the algae growing on walls around Singapore. (In press).

hll£'rnatirmal Hiotlt•terirmuion Bulletin ISSN 0020-6164 I X( I) Spring l9X2

5

Maynard, Nancy G. (1968). Significance of airborne algae.

Zeitschrift fur allgemeine Mikrobiologie, 8: 225-226.

McElhenney, T. R., Bold, H. C., Brown, R. M. Jr. and McGovern, J.P. (1962).

Algae, a cause of inhalent allergy in children. Annals of Allergy, 20: 739-743.

McGovern, J.P., Hayward, T. J. and MeEihenney. T. R. (1966).

Airborne algae and their allcrgenicity. II. Clinical and laboratory multiple correlation studies with four genera.

Annals of allergy, 24: 145-149.

Nicholas, H. W. and Bold, H. C. (1965). Trichosarcina polymorpha gen. et sp. nov.

Journal of Phycology, 1: 34-38.

Overeem. M.A. van. (1937). On green organisms occuring in the lower troposphere.

Recueil des travaux botaniques neerlandais, 34: 389-439.

Schlichting. H. E. (1961). Viable species of algae and protozoa in the atmosphere.

Lloydia. 24: 81-88.

Shamood. M. M. (1935). Pollen content of Melbourne air during the hay fever season of August 1933-March 1934.

Medical Journal of Australia. I: 326-332. ·

Specht. R. L.. Brouwer. Y. M. and Derrick. E. H. ( 1975).

Seasonal waves of asthma: a possible botanical cause.

International Journal of Bioclimatolog~ and Biometeorology. 19: 28-36.

Wee. Y. C. (1978). Vascular epiphytes of Singapore's wayside trees.

Gardens' Bulletin, Singapore. 31: 114-126.

Wee, Y. C. and Lee. K. B. (1980). Proliferation of algae on surfaces of buildings in Singapore.

International Biodetcrioration Bulletin. lb (4): 113-117.

SPECIALISED BIBL

Prepared and published by the Biodeterioration Centre

University of ASTON

Specialised bibliographies are produced from the Biodeterioration Information Centre's document collection from 1965 as listed in the bibliographic journals Biodeterioration Research Titles (B.R.T.) and Waste Materials Biodeterioration (W.M.B.).

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Glass, Ceramics, and Optical Equipment - 34 references

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Microbial Corrosion of Metals - 291 references

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6

lntemalimwl Biodelaioratitm Bulletin ISSN 0020-6164 18( I) Spring 1982

THE EFFECTS OF METALS UPON THE INHIBITORY ACTIVITIES OF CUTTING FLUID PRESERVATIVES

E. 0. Bennett', .1. E. Gannon' and I. U. Onyekwelu 1

Summary

The effects of 15 different metals upon the inhibitory propcrtic~ of 12 cutting Ouid preservatives v.rcre studietl. Thirteen metal'> increased the inhibitory properties of Grotan liD II while four improved the action or Ucarcidc 250. The other metal~ had no deleterious effect upon the two inhibitors.

Kathan 886 and Proxel CRL produced mixed rc<,ults with a few metals improving theicr inhibitory properties or having no significant effect, while the bulk of the metals reduced the activities of the compounds.

Lcs errccls des rnCtaux sur !'action inhibilrice des produits de preservation des liquides de coupe

On a CtudiC les effets de 15 rnCtaux di!Ti:rents sur lcs propriCtCs inhibitrices de 12 produits de prCservation des liquids de <:oupes. Treize mi:taux ant augmentC les proprii:tCs inhibitrices du Grotan HD II alors que quatre ont amCiiori: !'action de I'Ucarcide 250. Les autres mCtaux n'ont pas eu d'cffct dCICthe sur les deux inhibiteurs.

le Kathan 886 et le Proxel CRL ant donnC des rCsultats mixtes avec quelques mi:taux, amCliorant leurs propriCtCs inhibitrices ou n'ayant pas d'effet significatif alors que Je gros des mCtaux a rCduit l'activitC des composes.

Introduction

In view of the fact that metals are the major contaminants of working cutting fluids, it is surprising that there are few studies in the literature pertaining to the effects of these materials on rancidity control. Iron chips have been added to test systems employed to study the efficacy of various antimicrobial agents (Pivnick and Fabian, 1953; Wheeler and Bennett, 1956; Kitzke and McGray, 1959; Flemming and Baker, 1959; Himmelfarb and Scott, 1968; Rossmoore and Williams, 1971; Rogers, Kaplan and Beaumont, 1975). These studies did not employ the preservative both in the presence and absence of the metal in order to detect the effect of the metal itself upon preservative action.

There is only one report pertaining to the effect of metals upon the efficacy of cutting fluid preservatives (Bennett, 1973). This study involved only 5 metals and demonstrated that iron, aluminium, zinc, magnesium and copper could influence preservative action.

The objective of this present investigation was to study the effects of most metals involved in machining operations, both in the pure form as well as in alloy, upon preservative action in a representative number of cutting fluids.

Ucr Einnun um .\lctallcn auf die hcmmcndcn Eigcn!-ichartcn mn ~chutzmittcln fUr Kiihl- und Schmicr~tofrc

Die Wirkung \on 15 ~·erschicdencn Mctallen auf die hemmenden Ligcn<,chaften \On Schutzmitteln fUr SchneidOiissigkeiten wurde untcr~ucht. Dreizehn Mctalle erhOhten die hemmenden Eigcnschaften von Grotan HD I I, wiihrcnd vier die Wirkungsweisc von Ucarcide 250 verst1irkten. Die anderen Metalle hatten keincn schadlichcn EinfluB auf die Z\vci Schutzmittel.

Kathan 886 und Proxel CRLergaben unterschicdlichc Ergebnisse. wobei einige Metallc die hemmenden Eigenschaften verbcsserten oder keine ausgeprligtc Wirkung zeigten, wfihrend der grOilte Teil der Metalle die Wirksamkeit dieser Verbindungen verringerte.

El Efccto de Mctales Sobrc Ia Acth·idad lnhibitoria de Liquidos Lubricantcs Antiscpticos

Se han estudiado el efccto de 15 metales diferentcs sabre las propiedades inhibitorias de 12 Jiquidos lubricanrcs. Trece metales incrementan las propiedades inhibitorias del Grotan HD II mientras 4 mejoran Ia acciOn del Ucarcidc 250. Los otros metales no tienen ningun efecto sabre 2 inhibidores el Kathan 886 y Proxel CRL produjeron resultados mezclados con unos pocos metales mejorando sus propiedades inhibitorias o no teniendo efecto significante mientras que Ia mayoria de los metales rcdujeron Ia actividad de los compuestos.

Experimental procedure

The test units consisted of wide mouthed glass containers of sufficient size to hold approximately I litre of liquid. Each unit was aerated at all times by employing a capillary pipette. The amount of aeration was controlled by a valve so that uniform "rolling" of the coolant was obtained in the units.

Five hundred ml of tap water (approximately 120 ppm hardness), the desired concentration of preservative (wt/vol or vol/vol) and 1.0 g of a metal was added to the appropriate test unit. The metals employed in the study were purchased from chemical supply houses and were of purified quality. Each was in powder form except titanium which was in the form of a sponge.

The coolant concentrate was added ( 15.0 ml) and thoroughly mixed until a uniform emulsion was formed. The coolant was diluted to 600.0 ml by adding additional tap water and the liquid level was carefully marked upon the container. Once each week, distilled water was added to bring the liquid level back to this mark. Distilled water was used in order to avoid a build-up of organic salts over a period of time.

The units were inoculated with 1.0 ml each of a bacterial inoculum and a mould inoculum. and

Department of Biology, University of l-louston, Houston, Texas. 77004. USA.

(Received, September 1981)

7

The Effects of Metals upon the Inhibitory Activities of Culting Fluid Pre.st:rvativcs E. 0. Bennett . J. E. Gannon and I. U. Onyekwelu

reinoculated once a each week with a 1.0 ml of a 50/50 mixture of the two inocula. The organisms in the inocula originally came from spoiled samples of industrial cutting fluids. The bacterial inoculum, while subject to some variation, consisted predominantly of different species of pseudomonads with lesser numbers of Paracolobacterium, Proteus, and Klebsiella species. The mould culture consisted mostly of Fusarium and Cephalosporium species with minor numbers of Candida and Monilia. The bacterial inoculum was maintained in a petroleum based cutting fluid known to be highly susceptible to bacterial attack and was aerated at all times.

The mould inoculum was maintained in a synthetic coolant known to be highly susceptible to slime formation under constant aeration. Each week the containers were shaken vigorously and approximately three-fourths of the coolant removed and fresh fluid added in order to maintain vigorous growth. Neither of the two inocula have been grown on any substrate other than cutting fluids for several years. The bacterial inoculum contained between 25 and I 00 million cells per ml while the mould culture contained between 100,000 and 250,000 units/mi.

Once each week. each unit was examined for its microbial content using standard microbiological procedures designed to enumerate bacteria and moulds. Subculture media were incubated at 35°C and read after 48 hours. Test were continued until two consecutive counts in excess of 100,000 organisms per ml were obtained or until visible slime formed in the units. All units which contained less than 100,000 organisms/ml or did not develop slime were studied for 105 days.

Every effort was made to provide maximum challenge to the chemicals employed to treat the coolants. All units were open to aerial contamination, no effort was made to prevent introduction of organisms via the compressed air and the water used to dilute the coolants was not sterilized. These conditions simulated, as far as possible, those encountered under industrial usage.

All counts, inoculations, subculturing, make-up, and any other practice was done at the same time each week in order to minimize any variation in results from this source. All experiments were done in duplicate and where the data are of significant importance, experiments were repeated a number of times.

Since this investigation constitutes a continuation of previous investigations pertaining to antimicrobial agents in cutting fluids, additional controls were included. These. consisted of four units containing a common cutting fluid preservative (Milidin TI-10) in a petroleum base product (Shell Dromus B). These controls arc kept under test at all times; they fail in 21 to 28 days due to mould growth and have done so for several years. These controls functioned normally during the test period.

8

Preservatives employed

The preservatives employed in this investigation included the following products. Each was used at a concentration of 1000 ppm except Kathan 886 which was used at a level of 100 ppm. Throughout, the concentrations stated are those of the products as received; they arc not the concentration of the active ingredients.

Dowicide A A 97% active powder of sodium a-phenyl-phenate produced by the Dow Chemical Company, Midland, Mich., USA.

Tris Nitro A 50% solution of tris (hydroxymethyl)-nitromethanc produced by the IMC Chemical Group, Des Plaines, lll., USA.

Grotan A 78.5% solution of hexahydro-1, 3, 5-tris (2-hydroxyethyl)-s-triazine produced by the Lehn and Fink Industrial Products Division of Sterling Drug, Inc., Montvale, N. J., USA.

Milidin Tl-10 A solution (concentration not given) of hexahydro-1,3,5 -tris (2-hydroxyethyl)-s-triazine-iodine complex produced by the DeMille Chemical Corp., Jersey City, N.J., USA.

Vancide TH A 95% solution ofhexahydro-1, 3, 5-triethyl-s-triazine produced by the R. T. Vanderbilt Co. Inc., New York, N.Y., USA.

Doll'icil 75 A 67.5% active powder of 1-(3-chloroallyl)-3, 5, 7-triaza-1-azoniaadamantane chloride produced by the Dow Chemical Company, Midland, Mich., USA.

Bioban P-1487 A solution containing 70% 4-(2-nitrobutyl) morpholine and 20% 4, 4-(2-ethyl-2-nitromethylene) dimorpholine produced by IMC Chemical•Group, Des Plaines, Ill., USA.

Sodium Omadine A 40% solution of sodium pyridine thiol-1-oxide produced by the Olin Corp., Stamford, Conn., USA.

Proxel CRI. A 30% solution of I, 2-benziosothiazolin-3-one produced by ICI United States Inc., Wilmington, Del., USA.

Kathan 886 A solution containing 8.6% of 5-chloro-2-methyl-4-isothiazolin-3-one and 2.6% 2-methyl-4-isothiazolin-3-one and 2.6% 2-methyl-4-isothiazolin-3-one produced by Rohm and Haas Company, Philadelphia, Pa., USA.

Grotan HD 1/ A powder containing 2-chloro-N-(hydroxymethyl)­acetamide (39%), sodium tetraborate (41%), potassium iodide (0.39%), produced by the Lehn and Fink Industrial Products Division of Sterling Drugs, Inc., Montvale, N.J., USA.

Ucarcide 250 A 50% solution of gluteraldehyde produced by Union Carbide Corp., New York, N. Y., USA.

Cutting fluids employed

lrmco 310 (emulsion) International Relining and Manufacturing Co., Evanston, Ill., USA.

Shamrock (synthetic) F. E. Anderson Oil and Chemical Co., Portland, Conn., USA.

Dascool (emulsion) D. A. Stuart Oil Co., Scarborough, Ont., Canada.

Sanson 7080NB (emulsion) Henry E. Sanson & Sons, Inc., Bristol, Pa., USA.

Quaker 18A (synthetic) Quaker Chemical Corp., Conshohocken, Penn., USA.

Results

The results of this investigation are presented in Tables I, 2 and 3. Only two preservatives, Grotan HD II and Ucarcide 250, were not adversely affected by metals. More importantly, the anti-microbial properties of Grotan HD II were significantly increased in the presence of all but two of the metals. It may be noted that those preservatives which are generally known as formaldehyde releasers were almost uniformly adversely affected by metals.

The patterns produced by the individual metals are of interest. For example, iron produced no significant effect upon the inhibitory properties ofGrotan HD II, Urcacide 250, or Kathon 886, but adversely affected the action of all the other preservatives. Molybdenum improved inhibition in Grotan HD II, Urcacide and Kathon 886; it had no significant effect upon Proxel CRL, Tris Nitro, or Bioban P-1487 and produced a deleterious effect upon other preservatives.

Statistical studies were conducted to determine if valance(s), molecular weights, or other factors influenced the specific effects. No correlation with any other factor could be established. Even closely related preservatives such as Grotan and Milidin TI-10 exhibited quite different results with the same metal.

Discussion

It has been known for many years that the presence of iron increases the oxidation of hydrocarbons (Zuidema, 1946) and reduces the working lives of cutting fluids (Flemming & Baker, 1959). It also has

lnternationa/Biodeterioration Bulletin ISSN 0020·6164 18(1) Spring 1982

9

been noted that the presence of iron, magnesium or aluminium ions decreases the useful lives of rolling oils (Tripathi, 1975). In regards to iron, the sulphur content of the fines may be the most important factor in deterioration. Fines containing no more than 0.1% sulphur caused practically no spoilage while lines containing higher levels of sulphur caused emulsions to spoil in a short time.

Metallic ions influence deterioration in several ways. First, metallic ions can stimulate microbial growth (Zuidema, 1946; Englander and Corden, 1971) particularly in an alkaline environment. Second, there are numerous references in the literature concerning the well established fact that metals can influence the action of antimicrobial agents (Weinberg, 1957).

The results of this investigation show that metals generally interfere with the action of most cutting fluid preservatives. In certain instances, they can improve the inhibitory properties of a few preservatives.

The results show that Grotan HD II and Ucarcide 250 would be the preservatives most likely to control rancidity particularly in situations where the coolant becomes heavily loaded with lines. It must be remembered that none of the preservatives currently available are effective in all products. The tables can be

Table I

Effects of Metals on the Inhibitory Properties of Cutting Fluid Preservatives

% increase in activity in presence of I.Og of metal in 500ml

METAL GROTAN HD UCARCIDE II 250

Copper 74.6 76.1 Molybdenum 80.0 58.1 Magnesium 56.8 20.5 Zinc 30.9 41.0 Cobalt 47.7 • Aluminium 38.3 • Manganese 38.3 • Cadmium 33.6 • Titanium 33.6 • Nickel 30.9 • Tin 30.9 • Lead 30.9 • Tungsten 24.8 • Iron • • Chromium • •

• No significant difference from control containing no metal.

Results represent means of duplicate determination with 5 different cutting fluids diluted 1-40.

The Effects of Metals upon the Inhibitory Activities of Cutting Fluid Preservatives E. 0. Bennett , J. E. Gannon and I. U. Onyekwelu

Table 2

Effects of Metals on the Inhibitory Properties of Cutting Fluid Preservatives

% increase(+) or decrease(-) in activity in presence of I.Og of metal in 500ml

0-PHENYL PROXEL SODIUM METAL KATHON886 PHENATE CRL OMADINE

Copper +30.1 +22.0 • -82.1 Magnesium -50.0 -46.9 +30.7 -53.4 Zinc -42.2 -57.6 • -89.3 Cadmium +23.5 -53.2 +46.8 -39.0 Iron • -56.3 -22.4 -89.3 Nickel -55.2 -56.3 -42.4 -74.9 Cob~lt • -56.3 -40.1 -80.6 Tin -30.8 -46.9 -31.3 -49.8 Lead +33 • • +55.8 Aluminium -45.7 -53.2 • -53.4 Titanium -26.4 -46.9 -33.5 -60.6 Chromium -18.5 -53.2 -33.5 -74.9 Manganese • -65.7 -18.0 -56.0 Molybdenum +20.2 -84.4 • -85.7 Tungsten -17.2 -77.2 -24.6 -46.2

• No significant difference from control containing no metal.

Results represent means of duplicate determinations with 5 different cutting fluids diluted 1-40.

Table 3

Effects of Metals on the Inhibitory Properties of Cutting Fluid Preservatives

% decrease in activity in presence of I.Og of metal in 500ml

METAL GROT AN TRIS MILIDIN VANCIDE DOWICIL BJOBAN TH75 P-1487 NITRO Tl-10

Copper 69.2 33.9 60.5 27.8 68.9 24.8 Magnesium 40.6 44.3 54.8 29.7 36.3 58.3 Zinc 73.9 68.7 37.8 46.3 57.8 70.8 Cadmium 59.6 16.5 • 31.5 40.0 26.2 Iron 71.5 86.1 58.6 29.7 24.5 62.4 Nickel 54.9 33.9 20.9 22.3 44.5 24.8 Cobalt 23.7 59.3 60.5 30.8 51.2 55.5 Tin 47.8 68.7 • 24.7 51.2 38.8 Lead • 61.7 • • 22.3 66.6 Aluminium 66.8 30.4 37.8 48.2 77.8 • Titanium 71.1 23.4 18.0 24.1 42.3 45.7 Chromium 45.2 23.4 • 29.7 40.0 44.3 Manganese 76.3 64.7 57.6 37.1 46.7 66.6 Molybdenum 59.6 • 26.5 53.8 37.8 • Tungsten 28.7 40.8 71.8 27.8 31.2 55.5

10

used to select the preservative which is least affected by the metal being worked. For example, if an individual required a preservative for a coolant used to machine magnesium and it was found that Grotan HD II or Ucarcide 250 were ineffective in the product, then the tables show that Proxel CRL would possibly be the preservative to consider for use. In some instances, the individual might have to select a preservative which is least adversely affected by the metal being worked.

It has been reported that magnesium ions increased the inhibitory properties of glutaraldehyde (Ucarcide 250) (Gorman and Scott, 1979) and this was noted in this investigation. The effects of metals upon the action of phenolics has been reviewed (Bennett, 1959), and it was noted that copper increases the action of these preservatives, and this too has been observed in the study. Cobalt has been reported to increase the inhibitory action of phenolics; however, this was not noted in this investigation.

An additional factor of interest is that it has been found that Grotan HD II exhibits about a four fold increase in activity when employed with EDTA. If metals increase the inhibitory properties of this preservative, then their removal "via the use of a chelating agent should neutralise the effect.

Experiments employing Grotan HD II with EDT A and Grotan HD II with metals were repeated a number of times and both potentiate the inhibitory properties of the preservative. It is likely that the mode of action of Grotan HD II and metals is different than that of the preservative and EDTA upon the cells.

The results open for further consideration, the practice of using iron fines in laboratory tests to establish the efficacy of a chemical as a cutting fluid preservative. Since iron had generally a negative effect upon preservative action, the presence of this metal in routine screening tests could eliminate chemicals which might be favourably affected by other metals.

In the development of a new cutting fluid preservative, it appears advisable to study the effects of metals upon the compound but these test should follow those which indicate general activity in these products. In order to reduce developmental costs, it would be fortunate if a standard mixture of metallic powders could be developed for such tests in order to avoid studying each metal individually.

It should be noted that it may be possible to improve the antimicrobial properties of certain cutting fluid preservatives by adding small concentrations of metallic salts to the products. For example, a copper salt might increase the antimicrobial properties of Grotan HD II, Ucarcide 250 and Kathan 886.

lntema!ional BioclcH·rioration Bulll'tin ISSN 0020-6164 18(1) Spring 1982

II

References

Bennett, E. 0. (1959) Factors affecting the antimicrobial activity of phenol.

Advan. Appl. Microbial. 1: 123-7.

Bennett E. 0. (1973) Microbiological aspects of metalworking fluids.

S.M. E. Technical Paper No. MR73-826; 1-127.

Englander, C. M. and Carden, M. E. (1971) Stimulation of mycelial growth of Endothia parastica by heavy metals.

Appl. Microbial., 22: 1012-1016.

Fleming, C. D. and Baker, R. J. (1959) Controlling the spoilage of water soluble cutting fluids.

Paper presented at the 14th ASLE annual meeting, Buffalo, N. Y., Preprint No. 59AM 3A-2.

Gorman, S. P. and Scott, E. M. (1979) Effect of inorganic cations on the biocidal and cellular activity of gluteraldehyde.

J. Appl. 'Bact., 47: 463-468.

Himmelfarb, P. and Scott, A. (1968) Simple circulating tank test for evaulation of germicides in cutting fluid emulsions.

Appl. Microbial., 16: 1437-1438.

Kitzke, E. D. and McGray, R. J. (1959) Coolant microbiology: the role of industrial research.

Paper presented to the 14th ASLE national meeting, Buffalo, N.Y., Paper No. 59AM 3A-/

Onyekwelu, I. U., Bennett, E. 0. and Gannon, J. E. (1981)

The effective life of preservatives in cutting fluid concentrates

Tribiology International, i4: 1-9.

Pivnick, H. and Fabian, F. W. (1953) Methods for testing the germicidal value of chemical compounds for disinfecting soluble oil emulsions.

Appl. Microbial., 1: 204-207.

Rogers. M. R., Kaplan, A. M. and Beaumont, E. (1975)

A laboratory in-plant analysis of a test procedure for biocides in metal-working fluids.

Lub. Eng., 31: 301-310.

The Effects of Metals upon the Inhibitory Activities of Cutting Fluitl Prescr\'ati\'cs E. 0. Benncll 1 • .1. E. Gannnn 1 and I. U. Onyekwclu 1

Rossmoore, H. W. and Williams, B. W. (1971) An evaluation of a laboratory and plant procedure for preservation of cutting fluids.

International Biodeterioration Bulletin 7: 55-60.

Rossmoore, H. W. and Rossmoore. L.A. (1979) The identification of a defined microbial inoculum for the evaluation ofbiocides in water­based metal-working fluids.

ASLE preprint N.79-AM-1A-2: 1-5.

Tripathi, K. C. ( 1975) A new method to improve the life and performance of aluminium hot rolling mills.

Aluminium, 51: 574-579.

Weinberg, E. D. (1975) The mutual effects of antimicrobial compounds and metallic cations.

Bact. Reviews. 21: 46-68.

Wheeler, H. 0. and Bennett, E. 0. (1956) Bacterial inhibitors for cutting oil.

Appl. Microbial., 4: 122-126.

Zuidema, H. H. (1946) Oxidation of lubricating oils.

Chern. Reviews. 38: 197-226.

12

ACKNOWLEDGEMENTS

The following L·ompanies ;JrL' acknowledged for their co-opcmtiun amltinancial support which h:\\'1: made this p<.lpcr possible.

Ab Karlshams Oljcf;1hrikcr. Sweden: Andcr:-;on Oil omd ChcmiL·al Comp;my. Inc .. Porti<.Hlll. Connecticut: Hem\ Kemi Ah .. Swctkn: BcilUL'L'll & Co .. SPA .. Torino. Italy: Blaser & Cu .. /\G .. SwitZL·rland: Buckcvc Lubricants. Hcdford. Ohio; Chesson Oil Compan~. Inc .. l.;1ke Chmlcs. Louisiana: D. A. Stuart Oil Compan~. Chicago. Illinois: DcMilleChcmicoJI Corporation, Jersey Cit). Nc\\ Jersey: Do her Chemical Corpomtion. Midlothi;m. Illinois: Henry E. S;mson & Sons. Inc .. Bristol. Pcnnsylvunia: Gunther Schmidt. Hamburg. Germany; IMC Chemical Group. Des Pl<tincs. Illinois: International Chemical Company. Philadelphia. Pennsyh·<mia; International Refining & Manufacturing Co .. Evanston. Illinois: J. R. Schneider Company Inc., Corle Madera. California; Lehn and Fink Products Company, Montvale, New Jersey; Merck & Company, Inc., Rahway, New Jersey; Metalworking Chemicals & Equipment, Inc., Chester, New Jersey; Monroe Chemical Company. Inc., Hilton, New York: Monsanto Industrial Chemical Company. St Louis. Missouri; Olin Corporation, New Haven, Connecticut; Oil Kraft. Inc., Cincinnati, Ohio; Pillsbury Chemical & Oil. Inc .• Detroit. Michigan: Polar Chip, Inc., Irvine, California; Process Research Products, Pennington, New Jersey; Rohm and Haas Company. Philadeplphia, Pennsylvania; S. C. Johnson & Son. Inc .. Racine. Wisconsin; S. H. Mack and Company,lnc., StCharles. Illinois: Sino!, Snc .• Torino, Italy; TRW., Inc., Jamestown. New York: T.upmatic Corporation. Irvine, California; Tower Oil & Technology Company. Chicago. Illinois; Ry-Chemie Vertriebs-GmbH. Germany: Union Oil Company of California. Brea. California: Van Straaten Chemical Company, Chicngo. Illinois.

J

htf('I'IWtfrmal Biodeterioration Bulletin ISSN 0020-6164 18(1) Spring 1982

SUBERRANEAN TERMITE CONTROL: RESULTS OF LONG TERM TESTS

Raymond H. Bcal 1 and Ralph W. Howard 1

Summary

For more than 32 years, aldrin, chlordane, dieldrin, and heptachlor have been field tested in southern Mississippi soils to cv:1luatc resistance to subterranean termite (Reticulitames spp.) :mack. At the U.S. Department of Agriculture recOmmended dosage level (5. I liters/m 2 f I pint/ft2

] of0.5 to 1.0 percent active ingredient), all four insecticides are still 100 percent effective. Chemical analysis of treated soils arter 16 years ofwCatheringshows similar relationships between parts per million (ppm) of remaining insecticide and initial application rate for all rour termiticides.

ContrOie des termites soutcrrains: rfsultat d' cssais :i long terme

Depuis plus de 32 ans, I' aldrine, le chlordane, Ia dieldrine et [' heptachlor ant CtC essayCs en cssais de champs dans lc Missisipi de sud pour estimer Ia resistance a I' attaque du termite souterrain (Re~iculitermes spp.). Au niveau recommande par le Department US de I' agriculture (5.1 l/m2 fl pint (US)/pied2

,] de 0.5 a 1.0% de matiere active), taus les quatre insecticides sont encore it 100% efficaces. l' analyse chimique de sols traites,apri:s 16 ans aux agents atmosphCriques, montre des rapports semblables entre les parties par million (ppm) d' insecticide restant et le taux initial d' application pour les quatre termiticides.

Introduction

Recommendations for insecticides to control subterranean termites (USDA, 1980) are based on long-term studies being conducted by the U.S. Department of Agriculture, Forest Service, Forestry Sciences Laboratory in southern Mississippi. This paper reports the latest results of studies installed in 1958, as well as earlier studies (1948) in which several concentrations of aldrin, chlordane, dieldrin, and heptachlor are still! 00 percent effective after 28 and 32 years. In addition, chemical analysis of these four insecticides and their metabolites after 16 years of field exposure are presented.

Methods and Materials

Test plots and treatmellls. Aldrin, chlordane, dieldrin, and heptachlor were installed in 1958 at various concentrations and rates of application in a randomized block design with treatments replicated 10 times. Tests were limited to five concentrations: 0.031, 0.125, 0.250, 0.500, and 1.0 percent active ingredient (A. I.) applied at a rate of 5.1 liters/m2 (I pt/ft'). Each replicate consisted of a 43 x 43 em ( 17 x 17 in) surface area of treated soil with a 15 x 15 em (6 x 6 in) untreated pine board in the center. The soil beneath the board and in a band 7.6 em (3 in) wide around the

The authors are entomologist and chemist, respectively, at the Porestry Sciences Laboratory, P.O. Box 2008 GMF, MS 39503 (Received, November 1981)

13

BckUmprung ''on Erd termilcn: Ergebnisse mn Langzeit nrsuchL•n

.Seit mehr als 32 Jahrcn werden Aldrin. Chlordan, Dieldrin und Heptachlor in verschicdenen Erdcn in Frcilandvcrsuchen in SOd­Mississippi aufihre Wirksamkcit gegcn Erdtermiten (Reticulilermes spp.) geprOft. Bei den vom U.S.S Ministcrium ror Land-wirtschaft empfohlenen Einbringmengen (5.1 11m2 mit 0.5 his I % Wirksubstanz) waren aile vier lnscktizidc noch 100% ig wirksam. Eine chcmische Behandlung der behandelten ErdbOden nach 16 Jahren Witterungseinnun zeigte ein fihnlichcs Verhiiltnis 7.wischcn den ppm-Werten von wiedergefundencm Insektizid und urspriinglicher Anwendungsmengc aller ·vier Tcrmiten­Schutzmittel.

Control subterraneo de termitas: resultados de prucbas a largo plazo

Durante mas de 32 aiios, aldrin. clordano, dieldrin y heptacloro. han sido estudiados en suelos del sur de Mississippi pard evaluar su resistencia al ataque de termitas subterraneas. (Reliculitermes spp.). A Ia concentraci6n recomcndada par el Departamento de Agriculture de E.E.U.U. (5.1 L/m2 de 0.5-1 %de ingrediente activo). los cuatro insecticidas son aun crcctivos. Los analisis quimicos de suelos tratados tras 16 alios de permanencer al intemperie muestran similar rclaci6n de insccticida permanente y el inicial para los cuatre insecticidas.

board were not disturbed when soil samples were taken-for residue analysis. A total of 200 soil samples were taken from treated plots. Ten samples were also taken from untreated controls.

The test plots are located in sandy loam soil on the Harrison Experimental Forest 30 km (20 miles) north of Gulfport, Mississippi. This area receives an average annual rainfall of 175 em (69 inches) and an average annual maximum temperature of 36°C (97°F). Major vegetation consists of a pine overstory mixed with hardwoods. Major termite species present are Reticulitermes jlavipies and R. virginicus.

The effectiveness of the treatments was determined by annual examination of the pine boards on the treated soil. One hundred percent effectiveness denotes that none of the boards on the treated soil on any of the 10 replicates of a given treatment had been attacked or damaged by termites.

Sampling technique. Core samples, 2.5 em (I in) in diameter and 5 em (2 in) deep, were taken 15 em (6 in) out from the center of the bait board at all four corners and combined to make one sample. As the samples were taken, they were placed either in glass containers

Suberranean Termite Control: Results of Long Term Tests Raymond H. Bcotl omd Rill ph W. !Inward

Table I

Soil treatments that are still 100% effective against Reticu/itermes sp. in ground-board tests established in

1948-52 in southern Mississippi

Years in test as of 1980

32

Chemical

Chlordane (tech.) in No.2 fuel oil

Chlordane (tech.) in water emulsion

31 Aldrin (actual) in water emulsion

Dieldrin (actual) in water emulsion

Concn

(%)

1.0 2.0 2.0

2.0 2.0

0.5 1.0

0.25 0.5 1.0

28 Heptachlor (actual) in water emulsion 1.0

2.0

5.1 = I pint (US)/ft2; 10.2 = 2 pint (US)/ft2

Rate of applica­

tion1 (liters 1m2

)

10.2 5.1

10.2

5.1 10.2

10.2 10.2

10.2 10.2 10.2

5.1 5.1

14

Table 3

Soil residue levels of dieldrin remaining after 16 years of field exposure

Initial application rate 5.1 Jiters/m2 surli1ce area = I pt( US)/ft 2

Treatment Parts per million dieldrin in soil

Concen- (%) After

trations At time 0 16 yrs (± SD)

1.0 500.0 402.8 (134.3) 0.5 250.0 I 10.6 (18.6) 0.25 125.0 27.5 (8.7) 0.125 62.5 0.9 (0.3) 0.031 15.6 1.0 (0.4)

Table 4

Soil residue levels of aldrin and its metablite after 16 years of field exposure

Initial application rate 5.1 Jiters/m2 surface area = I pt(US)/ft2

Treatment

Concen­tration (%)

1.0 0.5 0.25 0.!25 0.031

At Parts per million aldrin and time dieldrin in soil

Aldrin after Dieldrin after 0 16 yrs (± SD) 16 yrs (± SD)

500.0 250.0 125.0 62.5 15.6

56.5 (9.22) 7.1 (1.!2) 1.3 (0.18) 0.3 (0.05) 0.02 (0.003)

181.1 ( 10.89) 33.9 (1.58) 9.8 (0.63) 4.0 (0.46) 0.4 (0.053)

International Biodeteriormion Built• lin ISSN 0020-6164 18( I) Spring 19~2

Table 2

Evaluation of insecticides applied as water emulsions in ground-hoard to:sts in 195K in southern Mississippi

Formulation Rate %ground bo01rds undamaged by t~:rmitcs alicr exposure li.lr indle<llo:d years (approx..% by wt) ' application

(liters/m~) 2 3 4 6 8 10 12 14 \6 JK 211 22

Aldrin 0.002 5.1 RO 30 \0.2 90 70 60 40 20.4 \\)(1 KO 70 70 50

0.04 5.1 \00 60 40 10.2 100 100 90 60 (10 411 20.4 \00 100 90 811 70 20

0.008 5.1 90 KO 711 50 10.2 J\)(1 )(}() 611 511 20.4 \00 90 911 811 811 50

0.016 5.1 \00 100 100 711 711 411 10.2 \00 100 100 \00 )(){} 811 411 20.4 \00 100 100 \110 911 K11 611 511

0.031 5.1 \00 \00 90 90 90 811 50 10.2 100 100 100 \00 \00 \\)(1 100 811 811 511 20.4 \00 ·100 100 100 \00 )()() )()() \IMI )Of) 70 411

0.062 5.1 100 100 100 \00 \DO (()() HKI I 00 911 70 611 511 10.2 \00 \00 \00 \00 \00 100 100 \IMI \\)(\ 90 811 511 20.4 100 100 100 100 100 100 100 IIMI IIXl lCKI 100 KO 811

0.\25 5.1 100 \00 1110 100 100 100 \00 \00 HXl 90 811 70 711 10.2 \00 \00 \00 \00 100 100 \110 IIMI JIM I II XI IlK I 100 IDO 20.4 \00 100 100 100 \00 \00 \110 JIM\ 100 ]()() \110 1\Kl )()()

Chlordane 0,002 5.1 \00 80 70 50 10.2 \00 80 50 20.4 100 100 80 60 30

0.008 5.1 \00 50 10.2 \00 60 60 50 20.4 \00 90 80 811 611 40

0.031 5.1 100 80 80 80 70 50 \0.2 \00 100 \00 \00 \IMI 911 811 611 511 20.4 \00 100 100 100 \00 \00 90 90 60 40

0.125 5.1 100 100 100 100 \00 \00 90 80 60 40 \0.2 100 100 100 100 100 100 100 100 90 711 60 50 20.4 100 100 100 100 \110 100 1110 \110 100 )()() l!IO 90 90

0.250 5.1 100 100 100 100 100 100 100 100 100 100 90 KO 811 10.2 \00 100 100 \00 100 100 100 I<Xl )()() )()() \\)(1 100 IIXl 20.4 100 100 100 1110 100 JIM\" 100 100 1110 100 100 \00 100

Dieldrin 0.002 5.1 "

100 80 80 50 \0.2 100 80 70 50 20.4 100 100 90 70 60 50

0.004 5.\ 90 70 60 40 10.2 80 60 50 20.4 100 100 90 80 80 60 60 50

0.008 5.1 \00 90 80 60 50 10.2 100 100 100 90 60 40 20.4 \110 100 100 100 90 90 KO 80 611 411

0.016 5.1 \IMI 90 90 80 60 411 10.2 \110 100 100 100 90 90 90 90 60 511 20.4 \(10 100 100 100 90 90 90 90 80 80 60 40

0.031 5.1 100 100 90 90 90 80 711 70 40 10.2 100 100 \00 100 100 100 100 100 \00 90 70 50 20.4 100 100 100 \00 100 100 JIM\ 100 \00 811 80 80 HO

0.063 5.1 \00 100 \110 100 100 100 100 \110 JIM\ \110 \00 KO 811 10.1 100 100 100 100 \00 100 100 IIMI IIMI 1110 IIMI 20.4 100 100 \110 100 90 90 90 90 90 90 90 90 90

0.125 5.1 100 100 100 100 \110 100 IIMI 100 100 90 90 90 90 \0.2 100 100 100 1110 100 100 J(IO 100 \110 \()() \00 100 100 20.4 100 100 100 100 100 100 100 100 100 100 100 100 IIMI

Heptachlor 0.002 5.1 100 90 70 50 0.004 5.1 100 90 50 0.008 5.1 \00 100 100 60 40 0.016 5.1 \00 100 100 90 90 70 50 0.031 5.1 100 100 100 100 100 90 KO 40

10.2 100 100 100 100 100 100 100 100 90 KO 40 20.4 \00 100 \00 100 100 100 \110 100 100 90 KO HO 811

0.063 5.1 \00 100 100 100 100 100 90 90 KO 60 50 10.1 100 100 100 100 100 100 100 UXI IIMI \00 90 911 911 20.4 \00 1011 100 1011 100 100 100 100 100 100 \110 100 )()()

0.125 5.1 \011 100 100 1011 100 100 JIM\ 1011 90 811 KO 50 \0.2 \00 100 100 1011 100 100 100 100 100 1110 100 l!IO 100 20.4 100 100 100 \011 \00 100 100 \00 \110 ]()() \110 100 ){)()

15

Subc:rranl'an Tc:rmill' (\Hurul: R~,.·:-;ult~ of long T~.·nu T~:>l:-\ Raymond H. Bc:al and Ralph W. llnw:ml

Table 5

Soil residue lc\'cls of heptachlor and its metabolites after 16 years of field exposure

Total ppm of Total ppm of heptachlor and its metaholites after 16 yrs (± SD) Treatm~:nt heptachlor and

"'r its metabolites Heptachlor Heptachlor- y-chlordanc a-chlordane at T = 0 epoxide

1.0 500.0 148.1 (32.32) 17.6 (3.38) 74.9 (X.98) 12.9 ( 1.80)

0.5 250.0 20.4 (5.16) 17.1 (4.0) 39.0 (9.2) 8.3 ( 1.87)

0.25 125.0 2.3 (0.371 7.8 (1.26) 14.8 ( 1.28) 3.0 (0.26)

0.125 62.5 0.5 (0.13) 2.4 (0.43) 3.0 (0.37) 1.6 (0.80)

0.031 15.6 0.02 (0.008\ 0.4 (0.06) - 0.2 (0.05) 0.03 (0.008)

Table 6

Soil residue levels of chlordane and its metabolites after 16 years of field exposure

Total ppm of Total ppm of chlordane and its metabolites after 16 yrs (± SD) Treatment chlordane and

I"() its metabolites Heptachlor- y-chlordane a-chlordane Nonaehlor at T = 0 -epoxide

1.0 500.0 0.0 (0.00) 26.3 (3.26) 27.6 (3.14) 9.5 ( 1.08)

0.5 250.0 0.0 (0.00) 6.0 (0.68) 6.8 (0.75) 2.4 (0.27)

0.25 125.0 0.3 (0.07) 2.9 (0.32) 3.5 (0.35) 1.3 (0.12)

0.125 62.5 0.2 (0.02) 0.8 (0.10) 1.0 (ll.ll) 0.5 (0.05)

0.031 15.6 0.1 (0.01) 0.1 (0.03) 0.1 (0.04) 0.1 (0.02)

16

or in aluminum-lined bags. Samples were taken in sequence from the lowest concentration of insecticide to the highest concentration to minimize cross­contamination.

Preparation of samples for analysis. When all 210 samples had been collected, each was sifted through a No. 10 mesh screen to remove roots, rocks, and other debris. The soil was then thoroughly mixed and I 00 g extracted as described below. The extracts were stored at -20~C until analyzed by gas chromatography.

Extraction and analysis of samples. Insecticidal residues were extracted from the soil as described by Carter and Stringer (1970). Spiked samples gave recovery efficiencies of at least 90 percent. Gas chromatographic analyses of extracts were made on either a Micro-Tek Model GC-2000MF equipped with a 130 millicurie tritium foil electron capture detector operated in a pulse mode, or a Tracor Model MT-220 chromatograph equipped with a Ni-63 electron capture detector operated in a nonpulse mode. All glass 6.4-mm outside diameter (o.d.) x 180-cm columns contained either 3 percent SE-30 on 60/70 mesh Chromport XXX or 3 percent DC-200 on !00/!20 mesh Gas Chrom 0. Argon-methane (95:5) was used as the carrier gas on the Micro-Tek instrument; nitrogen was used for the Tracor instrument. All results are reported as parts per million (ppm) of insecticidal residues in the soil.

Results and Discussion

As shown in Table !, treatment concentrations that ranged from half as much to twice as much as currently recommended and registered for subterranean termite control (0.5% aldrin, !.0% chlordane, 0.5% dieldrin, and 0.5% heptachlor) (USDA, 1980) are still 100 percent effective after 28 to 32 years. Quantitative soil analyses for movement of chemical residues were made from soil taken in and near the areas treated in !948-52. These analyses were completed when the tests were !7- to 2!-years-old and indicated that the insecticide had moved only a few inches (Smith, !969) during exposure to the elements. There is no reason to expect any significant additional movement of these materials although no further evaluation has been done. Since insecticides are placed under buildings where there is a minimum of weathering, erosion, or other disturbance, the treatment presents a minimal · hazard to man.

The degree of effectiveness of lesser concentrations is given in Table 2 for an interval of I to 22 years in ground-board tests. All treatments at concentrations and rates of application higher than those show in this table are still giving I 00 percent control; those with less insecticide failed within a few years. Though lower concentrations have prevented termites from penetrating soil for a number of years, it is recommended that consumers use the previously mentioned concentration because variations in climate, soil type, and termite species may alter effectiveness. These materials, applied at a rate of 5.!

International Biodeterioration Bulle/in ISSN 0020-6164 18(1) Spring 1982

17

liters/m2 (I pt/ft2) as recommended by the U.S.

Department of Agriculture ( !980), will provide protection from termites for at least 32 years for chlordane, 31 years for aldrin and dieldrin, and 28 years for heptachlor.·

Comparison of results by analysis of variance with different volumes of liquid carrier indicates effectiveness is not altered greatly by the amount of water used, but is related to the actual amount of insecticide placed in the soil. For example, treatments with concentrations of 0.002, 0.004, and 0.008 percent aldrin applied at rates of 20.4, !0.2, and 5.! liters/m2

(4, 2, and I pt/ft2) of soil contain the same quantity of

toxicant and all three treatments have given protection from termite attack for about the same length of time (Table 2).

These studies are still in progress and will be continued as long as any of the applications of insecticide give termite control.

· Chemical examination of selected treatments which had been in test for !6 years showed similar relationships between ppm of remaining chemical and initial application rates for all four insecticides (Tables 3-6). No metabolites of dieldrin were detected, but polar compounds would not have been recovered by our technique. Aldrin, as expected, underwent significant conversion to its oxidation product dieldrin. Heptachlor also was oxidized readily (yielding heptachlor epoxide), with more oxidation apparently occurring at the lower treatment rates than at the higher ones. Little heptachlor epoxide was found in the chlordane treatments, but significant quantities of nonachlor were detected. In none of the treatments was any unusual derivatives of the parent insecticides found. Derivatives that were found differed not in type but in level from those previously reported in agricultural studies. These levels, whiie greater, were not correspondingly higher than what might be expected in light of the higher rates applied for termite control purposes.

References

Carter, F.L. and Stringer, C.A. (!970) Residues and degradation products of technical heptachlor in various soil types. J. Economic Entomology 63(2): 625-628.

Johnston, H.R. (!960) Soil treatments for subterranean termites. U.S. Department of Agriculture Forest Service Occasional Paper 152 (revised)

Smith, V.K. (1969) Termite control and the natural environment. Proc. British Wood Preserving Association termite symposium, Cambridge, England, 1969: 101-104.

Subcrraneun Termite Control: Results of Long Term Tests Raymond H. Beal and Ralph W. Howard

U.S.Department of Agriculture, Science and Education Administration and Forest Service (1980)

Guidelines for the control of insect and mite pests of foods, fibers, feeds, ornamentals, livestock, households, forests, and forest products

U.S. Department of Agriculture Handbook No. 571. 796 p.

18

Pcsticidl• prccaution:try statement

l11is publication rcporls rcse<Jrch involving pesticides. It docs not contain recommcndmions for their us'-'. nor docs il imply that the uses discussed here have h'-'en rcgistcrL·d. All uses of pesticides must be registered by uppropriutc Stntc and/or Fcdcml agencies before they can he recommended.

CAUTION: Pcslicides can he injurious to humans. domestic :mimnls. desirable plants. and lish ~r other wildlife- if they arc not homdlcd or :1pplied properly. Usc all pl-sticidcs selectively and carefully. Follow recommended practices for the disposiJI of surplus pesticides and pesricidc conluiners.

Note: Mention in this paper of u company or trade numc is solely to identify muterial used and does not imply endorsement by the U.S. Department of Agriculture.

/menuuimwl Hiodt•taiorarion Hulh•till ISSN 0020-6164 1~-lt I) Spring 1982

A MEANS OF DISPERSAL OF BOTRYODIPLODIA THEOBROMAE

S. K. Ogundana 1

Summary

Pycnidiospores und pycnidia of Botryodiplodia theohromnr Pat. were fed to albino rats, cockroaches and grasshoppers, and thrir faeces examined for the presence and viability of the: fungus. The studies showed that the spores and pycnidia. after passage through the digestive tracts of the animals, still retained their viability. It is therefore h~·rcby suggested that animals play a role in the dispersal of B. theohmmae.

Un moyen de dispersion de Botryodiplodia thcobromac

On a alimentC des rats albinos, des blattes et des sauterelles avec des pycnidiospores et des pycnides de Bolryudiplodia theobroma'' ct examine les ICces pour voir Ia presence ct Ia viabilite du champignon. Les l:tudcs ont montrC que k'S spores et les pycnides, apri:s passage d<tnds le tube digestif des animaux ont conserve leur viabilite. On suggCre, en consequence, que les animaux jouent un rOle dans Ia dispersion de n. throhromae.

Introduction

Botryodiplodia theobromae Pat. has been shown to be an important plant pathogen in the tropics (Wardlaw, 1935; Meredith, 1961; Olofinboba, 1967) and has also been shown to be a major cause of storage rots of rootcrops in Nigeria (Okafor, 1966; Adeniji, 1970; Ogundana, Naqvi and Ekundayo, 1975). Although a considerable interest has been shown in B. rheobromae as an important plant pathogen, there is scanty literature on its life history.

Evidence has shown that even the non-sporulating Sclerotium rolfsii can be introduced into disease-free areas by cattle and sheep previously pastured on infected beet roots and tops (Leach and Mead, 1963). The present study, therefore, investigates the possible role of some animals in the dispersal of B. theobromae. Since rats, grasshoppers and cockroaches are commonly found in yam stores they may, perhaps, be a source of infection of yams by storage rots. Accordingly, the present study was undertaken to investigate the possible role of these animals in the dispersal of B. theohromae.

Materials and methods

The animals employed in this investigation were the albino rats, Musmusculus domeJticus L.: the cockroach, Peri'p/aneta americana L.; and the grasshopper, Zonecerus varigatus. L. A pair of each animal was confined in an appropriate cage previously sterilised. The rats were fed on recommended rat cubes

DepL t•l Micrubiulogy. Uni\crsity of lfe. lle-lfc. Nigeria {Rc .. :ciwd Nuvembt.•r 19!-ll)

19

Zur Miil!lichkcit dcr Vcrhrcitung \'On Botr)'diplodio theobronrae

Pyknidiosporcn und Pyknidien von Botrydiplodia tht·o!Jromue Pat. wurden an Albino-Rattcn. Schaben und Heuschreckcn vcrfOtlcrt, und dcren Exkremente wurden auf Vorhandensein und Lebcnsfiihigkcit dieses Pilzcs untersucht. Die Vcrsuche zcigtcn, d<tB die Sporen unc.J Pyknidien nach der Passage durch den Verdauungstrnkt dcr Ticrc ihre lebcnsfahigkeit behieltcn. Es wird deshalb hicr dar:mf hingcwicsen, daB die Tiere eine Rolle in der Vcrbreitung von B. tht•obromae spiclcn kiinntcn.

Un mcdio de dispcr:son de Dot.-yodiplodia lhcobromae

Las pegnidiosporas y pignidios de Botryodiplodia tltt•obromuc fueron suministr:tdas a ratas albinas, cocarachas ysaltamontes y sus heccs examinadas buscando Ia prest:ncia y viabilidad del bongo. Los cstudios han mostrado qu~ las csporas y pignidius tras pasur el tractn dig~stim de los ani males, rctienen su viabilidud Iuque sugi~re que los ;.mimales jucgan un papel en Ia dispersiOn de H. th('nhro~mu·.

and the cockroaches were given moistened cassava powder (gari) in sterile petri dish. The gras~hoppers were fed with sterilised cassava leaves (Manihot esculenta).

Faecal pellets were collected from the animals and incubated on malt extract agar to observe what types of fungi could be isolated. The exercise was to ascerlan that B. theobromae was not previously present in the gut of these animals.

Feeding wilh spore suspension

Sproulation of B. theobromae cultures was induced by incubating them in light chambers (Ekundayo and Haskins, 1969). A suspension of pyknidiospores was obtained by picking the pycnidia into a beaker containing sterile water. The pycnidia were then broken open to release the spores and mycelial fragments were separated by filtration through loose non-absorbent cotton wool. ·

Rat cubes were soaked for 2 minutes in the spore suspension and used for feeding the rats. About 10 ml of the spore suspension was mixed with 10 g of gari in a sterile petri dish and placed inside the cage containing the cockroaches. For the grasshoppers, surface sterilised cassava leaves soaked in the spore suspension were supplied as food in a sterile petri dish lined with sterile filler paper, also soaked in the spore suspension. The cages containing the animals were left on the laboratory bench at room temperature (26±l 0 C).

A Means of Dispersal of Bon:l·odiplodia Theobromm• S. K. Ogundana

Feeding was carried out for three consecutive days at intervals of 12 hours, and at each feeding new food materials with freshly prepared spore suspension were used. Subsequent to the three days, the animals were fed without spore suspension.

Feeding with pycnidia

To investigate the possibility of dispersal by means of pycnidia, another set of rats was used. One boiled egg was sliced lonitudinally into four strips and a total of 40 pycnidia stuck into the slices and fed to the rats. This feeding was carried out three times.

Collection and examination of the animal faeces

The droppings of the insects fed with spore suspension were collected, washed in three changes of sterile distilled water, blotted between sterile filter papers and incubated on malt extract agar at 28°C. The surface portion of the faeces voided by the rats was removed and a sample for incubation taken from near the centre.

A second method of faecal collection was adopted in the case of the rats. Each rat was held in one hand by the back of the neck and the anal region swabbed with cotton wool moistened with methylated spirit and finally cleaned up with another cotton wool moistened with sterile distilled water. Still holding the rat in the same position, a harder gripping pressure at the neck region caused fresh faeces to be voided (Needham, 1977) and these were collected directly into a sterile petri dish.

The examination of faeces of the rats fed with pycnidia was based on the methods of Akueshi and Oso ( 1976) whereby the droppings were collected and distributed into McCartney bottles (5 per bottle)continuing20ml of sterile distilled water and shaken for 30 tninutes in a wrist action flask shaker. The contents of each bottle were poured into petri dishes and any intact pycnidia

.. picked out. A pycnidiospore suspension was obtained from these as described earlier. A drop of the spore suspension was then inoculated onto malt extract agar and incubated.

The droppings were examined daily until no more growth of B. theobromae occurred.

Examination of the alimentary canal of rats

Seven days after the rats stopped producing faeces with B. theobromae, the alimentary canal was dissected out and its contents examined for presence of the pathogen spores. The various regions of the canal were also cut open and incubated on malt extract agar.

Results

The faeces from the animals prior to feeding with spore suspension or pycnidia gave rise to some bacteriaa and Absidia sp. but no B. theobromae were

20

present. The rat cubes, examined before being inoculated with the spore suspension also did not show the presence of the fungus.

Hyphae of B. theobromae grew out of the droppings of all the animals fed with spore suspension irrespective of the method of faecal collection. The B. theobromae isolated from the droppings produced pycnidia and spores similar to those of the original isolate when subcultured on m"alt extract agar. Viable spores were still encountered six days after feeding with spore suspension had ceased. In the experiment where pycnidia were used, the rats picked 97 out of the 120 pycnidia supplied. The residue from the faecal pellets contained some intact pycnidia. Both the residue from the faecal pellets, and the pycnidiospore suspension obtained from the recovered pycnidia, showed mycelial growth of the fungus when incubated.

The contents of the dissected alimentary canal, as well as the various regions of the gut of the rat showed no growth of the fungus.

Discussion

The passage of inert material through the digestive tract of animals has been studied by some investigators. Ficke and Melchers (1929) showed that some corn smut and sorghum smut spores retained their viability after passage through the digestive tract of horses. Akueshi and Oso (1976) also found some sclerotia of Sclerotium rolfsii retained their viability after passage through the difestive tracts of cockroaches and rats. That the spores of B. theobromae still retained their viability after passage through the digestive tracts of the animals employed in the present study not only agrees with the results obtained by others in similar work (Ficke and Melchers, 1929; Akueshi and Oso, 1976) but also suggests that the animals could introduce the pathogen to new areas. In this way the pathogen can be widely distributed since viable spores were still obtained from the animals six days after feeding with spore suspension had been discontinued, a period during which the animals could cover wide areas or long distances.

Faeces from animals which had been fed with either spores or pycnidia, developed the fungus. This shows that the fungus can be dispersed in both forms, indicating great efficiency in distribution. Examination of the residue from the faecal pellets of the rats fed with pycnidia showed that some pycnidia were fragmented, since B. theobromae also grew from the contents in the absence of intact pycnidia. This suggests that pycnidia, if picked up by animals, could be dispersed in two ways, either in the intact form, or disintegrated to release spores.

References

Adeniji, M.O. (1970) Fungi associated with storage decay of yams in Nigeria Phytopathology 60: 590-592

Akueshi, C.O. and Oso, B.A. (1976) Studies on Sclerotium rolftii II Observations on a means of sclerotia dispersal. Nigerian Journal of Plant Protection 2: 40-42

Arinze, A.E., Naqvi, S.H.Z. and Ekundayo, J.A. (1975)

Storage rot of sweet potato (Ipomoea batatus) and the effect of fungicides on extracellular cellulolytic and pectolytic enzymes of the causal organisms. International Biodeterioration Bulletin 11(2): 41-47

Ekundayo, J.A. and Daniels, I.M. (1973) Cassava rot and its control Trans. Britisl) Mycological Society 61: 27-32

Ekundayo, J.A. and Haskins, R.H. (1969) Pycnidia production by Botryodiplodia tlzeobromae Pat. Canadian Journ. of Botany 47: 1153-1156

Ficke, C.H. and Melchers, L.E. (1929) The effect of digestive processes of animals on the viability of corn and sorghum smut spores. Journ. of Agricultural Research, 38: 633-645

Leach, L.D. and Mead, S.W. (1936) Viability of sclerotia of Sclerotium rolfsii after passage through the digestive tract of cattle and sheep. Journ. of Agricultural Research 53: 519-526

International Biodeteriorotion Bulletin ISSN 002G-6164 18(1) Spring 1982

21

Meredith, D.S. (1961) Botryodiplodia theobromae Pat. and Nigrospora sp in the air of Jamaican banana plantation. Nature 190: 555-557

Needham, J .R. ( 1977) The collection of animal faeces for laboratory examination. Journ. Institute of Animal Technicians 28: 63-65.

Ogundana, S.K., Naqvi, S.H.Z. and Ekundayo, J.A. (1970)

Fungi associated with the soft rot of yams (Dioscorea spp) in storage in Nigeria. Trans. British Mycological Society 54: 445-451.

Okafor, N. (1966) Microbial rotting of stored yams (Dioscorea spp) in Nigeria. Experimental Agriculture 2: 179-182.

Olofinboba, M.O. (1967) Carbohydrate balance in the xylem of Antiaris africana and its relation to the incidence of blue­stain organism in the felled timber. PhD Thesis, University of Ibadan, Nigeria.

Wardlaw, C.W. (1935) Diseases of the Banana. Macmillan, London.

Shorter Communications Spring 1982

SHORTER COMMUNICATIONS

It has been decided, in response to a number of suggestions, that International Biodcterioration Bulletin should in future carry "Shorter Communications". It is suggested that none shall not normally exceed two sides of one sheet including any illustrations and all references. No summary will be included, and therefore only the titles will be translated for inclusion on the title page.

The most detailed case for including this form of communication has come from the Biodeterioration Laboratory at Woolwich Arsenal (Ministry of Defence) in the following terms: The categories of work we suggest for publication in this way would be:

I. Small pieces of work which are largely confirmatory and, without breaking new ground, extend our understanding of the scope of biodeterioration.

2. Work which gives rise to negative results but has importance in being so.

3. Observations of biodeterioration on less important or unexpected substrates which lead to no extended study.

4. Work which leads to the application in a new way of established physical, chemical or biological techniques to the study of biodeterioration.

To these categories can be added observations which are believed to be novel but which neither the author nor any of his colleagues is able to follow up. Also, since time will be saved in preparation of these pieces for the press, they can expect quicker publication than more formal papers and hence may serve to establish priority.

T.A. Oxley, Editor.

22

hrtcmaJimwl Himll'tC'rionuionllul/ctiniSSN 0020-6164 18(1) Spring \982

HIE DEGRADATION OF TREE BARK BY THERMOPHILIC AND THERMOTOLERANT FUNGI FROM NIGERIAN MUNICIPAL COMPOST

C.I.C. Ogbonna , G.J.F. Pugh and H.O.W. Eggins

DCgradation d' Ccorccs d' nrbrcs par des chnmpignons thcrmophilcs et thermoto!Crants de compma municipal NigCrian.

Introduction

Bark presents currently one of the most intractable problems facing the wood conversion industries in Nigeria. The amount of bark produced, almost 6% of the wood, is so high that continuous removal from all mill sites is required. Different outlets for utilisation of the bark have been proposed such as tannin extraction, production of fire briquettes, and burning of bark as fuel. With increasing interest in recycling of waste products, the trend for use of bark ns a substrate for horticultural purposes has become important.

If wood bark is left lying on the soil there is a tendency for the heavy rain which falls in Nigeria to leach out the mineral nutrients it contains into the subsoil. Such mineral nutrients thus are rendered inaccessible to land plants. But if the bark were to be composted, many of the mineral nutrients would be incorporated into the fungal mycelia and only gradually released to the soil, thereby becoming available to plants.

Bark is resistant to decay and therefore to com posting. The present experiment was therefore carried out to investigate the degradation of Nigerian wood bark by thermophilic and thcrmotolerant fungi. It was considered necessary to employ fungi of strains which had been isolated from a com posting system, therefore municipal compost was used as their source. (Ogbonna 1980).

Materials and methods

Bark from the African timber tree, Conarium schweinfurthii. was obtained from the African Timber and Plywood factory, Sapele, Nigeria. 50 g dry weight of bark was weighed into each of three 500 ml sterile conical flasks. Another 50 g portion of bark which had been pulverised was weighed into each of three additional sterile 500 ml flasks. The flasks were closed with cotton wool bungs. The six nasks were then heated to 80°C and then cooled to -I8°C. This procedure was repeated seven times at daily intervals; preliminary experiments had confirmed that such treatment killed the initial fungal nora of the bark.

Each flask was inoculated with I ml of equal dilutions of spores of the following fungi all of which has been isolated from composted municipal waste and had been shown to be strongly cellulolytic.

Dcr Abbau von Baumrindc durch thermophile und thcrmotolcr:Jntl.! Pilzc in stlldtischcm MUll in Nigeria.

DegradaciOn de Cortcza de :1rbol por hongos tcrmofilos y tcrmotolcrantcs en Compost en Nigcri;~.

Chaetomium !hcrmophi/c• var. coprophilc (Cooney and Emerson) Mycdiophthora tilcrmopilila (Apinis) v Oorschot Aspergillus fumigatlls (Fresnius} Sporotrichum pulverulellllllll (Novobranova)

The bark in the llasks was moistened with sterile distilled water to bring it to 65o/r moisture content, and the llasks were then incubated at 40°C. The flasks were left for a period of six months, after which there was no further production of offensive odours. Reisolation experiments were then carried out to ensure that all four of the test fungi were still present and that there had not been any other fungal contamination. Loss in weight was considered to be due solely to growth of the test fungi. pH was determined at the beginning and end of the experiment.

Results

The fungal reisolation experiments confirmed that the test fungi were still present at the end of the experiment and that there were no fungal contaminants. There was considerable weight reduction in the two sets of composted bark, the greater loss being in the pulverised bark. The results are given in the table, each figure being the mean of three llask results.

Discussion

These results show that wood bark can be com posted if suitable microorganisms are used. The pH of the pulverised bark was tending towards the alkaline side at the end of the experiment which may indicate that the bark composting was nearing completion. Jann. Howard and Salle (1959) reported that organic material, if fully com posted, will maintain an alkaline pH for twenty-four hours. They also reported that such a compost will not undergo anaerobic fermentation on storage.

Bark is a difficult material to compost, probably because of its high carbon/nitrogen ratio, which makes it unattractive to sr~prophytic microorganisms. and also partly because two of its main constituents, cork and lignin, arc resistant to decay. The greater weight loss observed for the pulverised bark may have been due to the greater surface area created for colonisation by the microfungi.

Dcpartmcm of Biological Sciences, Univcr:;ity of Aston. Birmingham BX 7ET. England Dr. Oghonna's prcscnt address: Depart men! or Botany. University of .los, .los. Nigeria.

(Received Augusl 19!-il)

23

The Dcgralbtlull ofTil'L" Bark by ThL"rll1ophilic :.md Thcrmotolcrant Fungi from Nigcri:m Municipal Compost C.I.C.Ogbonna ,G.J.F. Pugh and H.O.W. bggm~

Table Weight losses of Nigerian wood

bark during composting (means of three determinations)

Initial Final dry weight dry weight

Pulverised bark 50.0 g 10.50 g

Non-pulverised bark 50.0 g 17.30 g

While composted bark might be suitable for mushroom cultivation, probably the best outlet is as a compost for usc in horticulture. It makes a long­lasting mulch for horticultural crops giving good weed suppression and soil moisture retention. If it is first pulverised it has a tidier appearance and is probably Jess resistant to penetration by min. It is often claimed that if granulated bark is dug into the soil it can improve texture and fertility by increasing aeration and by assisting the retention of moisture. However, untreated bark dug into the soil may reduce the amount of available N in the soil because of its high C:N ratio.

24

Weight loss Initial Final percent pH pH

79.0 6.2 7.2

65.4 6.2 6.8

References

Jann, G.J., Howard, D.H. and Salle, A.J. (1959) Method for the determination of completion of com posting Applied Microbiology 7: 271-275.

Ogbonna, C.I.C. (1980) The effects of Nigerian fungi on Nigerian wood PhD Thesis, University of Aston.

/ntNnatimwl Himh•f,·l'iorcuirm Bulletin ISSN 0020-6164 18{1) Spring 191':2

THE BIODETERIORATION OF ETHANEDIOL ANTIFREEZES BY PENICILLIUM SJMPL/CJSSIMUM

R. Dale 1 2 T. Rubidge 1 and G.R. Williams 1

La dCtCrioration d' anligcls clhancdiols par Penicillium .~imp/iciuimum.

Ocr biologischc Abbau von A:thandioi·Frostschutzmitlcln durch Penicillium simplicluimum

Introduction

Ethanediol is the major constituent of most antifreeze formulations (ca 90% v/v). At antifreeze concentrations of 10% (v/v) the formulation depresses the freezing point of water to about -4°C and at 20% (v/v) to -10°C. As ethanediol is an, alcohol readily oxidised by microbes (Lamont & Stadtman, 1963) cooling systems- might become colonized. This phenomenon has been observed in a tank containing ethanedioJ: Problems associated with colonisation of cooljngsystems might include lowered concentration of 3ctive ingredient, corrosion caused by secreted metabolic waste products and system blockage by fungal mats. Microbial growth is most probable with infrequently used systems. Systems used frequently, such as motor car radiators, will tend to be sterilized through the fungicidal effect of the heat and pressure developed. This paper describes studies of the deterioration of two antifreeze formulations by Penicillium simplicissimum.

Experimental

Antifreeze formulations

Ethanediol at 90% (v/v) was the active ingredient. Antifreeze I also contained triethanolammonium orthophosphate, sodium mercaptobenzthiazole and sodium fluorescein. Antifreeze 2 contained sodium sebacate and benzotriazole, as well as ethanedio!.

• Respirometry

Absorbtion of metabolically produced carbon dioxide by sodium hydroxide was measured using a Warburg apparatus. Cell suspensions (0.5 cm3

) were mixed with distilled water (2.0 cm3

), phosphate buffer (100 Jl mol, pH 7.0) and appropriate concentrations of the compounds studied. Respiration rates were measured in a SEA model OM-K metabolic oxygen meter.

Diodctcrioracion de anticongclantc de ctanodiol por P£'nicilllum .tinrp/ic·i.uinlll/1!

Analytical methods

Ethanediol: NAD oxidoreductase and glycolaldehyde: NADH oxidoreductase activities were determined using the method of Racker ( 1949), acetaldehyde: NAD oxidoreductase activity was determined using the method of Black ( 1951 ). Protein was determined using coomassie blue reagent (Bradford, '1976), and the products of ethanediol degradation by the methods of Jones and Turner (1973) and Gokay

. (1978).

Results and Discussion

Penicillium simplicissimum. P.pinophilum and Acremonium sp. were isolated from a tank containing degraded solution of Antifreeze 2 -used at 10% v/v ethanediol. The ability of these organisms to degrade the individual components of the formulations was investigated. P. simplicissimum was selected for further study because it showed greatest growth on ethanediol as sole source of carbon and energy and was capable of growth on sodium fluorescein. It was unaffected by triethanolammonium orthophosphate and inhibited by sodium mercaptobenzthiazole at concentrations greater than 0.3% (w/v). Warburg manometry was used to study the mechanism of degradation 'of ethanedio!. The organism was able to oxidise ethanediol, glycolaldehyde and acetaldehyde as shown by carbon dioxide evolution at rates above endogenous. Cell suspensions possessed the following enzyme activities (~ moles substrate metabolised. min -•. mg _, protein) NAD oxidoreductase { 15.9). glycolaldehyde NADH oxidoreductase (5.9) and ethanediol: NAD oxidoreductase (0.6). Only one volatile degradation product was detected: It was identified as acetaldehyde by thin layer chromatography. These data are consistent with catabolism of ethanediol via glycolaldeh~·de and acetaldehyde to acetate with subsequent metabolism via the tricarboxylic acid cycle. ·

Biodeteriomtion Labormory. Materials Quality Assurance Directorate. Royal Arsenal East. Woolwich. London SE 18.

Present address: School of Biological Sciences, Thames Polytechnic. Woolwich, London SEI8

25

The Biodeterioration of Ethanediol Antifreezes by Penicillium Simplidnimum R. Dale f. Rubidge and G.R. Williams

I 0 . -1 -1 n mo es 2. nun .mg dry wt cells

1'0

o·s

0'6

0'2

0 I 2 3 4 5 6 7 8 9 10 11 12 13 14 IS

FIGURE I. Respiration rate of P. simplicissimum incubated concentration % v/v with Antifreeze I( 0 ), Antifreeze 2( o) & ethanediol (•).

When the respiration rate of P. simplicissimum was studied using the oxygen meter the organism was shown to be equally able to oxidise both Antifreeze 2 and ethanediol at concentrations up to about 10% (v/v) with optima at 5-7% (v/v) and 2 to 5% (v/v) respectively (Figure I). Respiration rates in excess of endogenous were detected with Antifreeze I only to about 5% (v/v) with an optimum at 3% (v/v). Thus in the absence of sodium mercaptobenzthiazole or other inhibitory components, ethanediol antifreezes are susceptible to degradation by P. simplicissimum up to a concentration of 10% (v/v). Where sodium mercaptobenzthiazole is included in the formulation (BS 3150) a final inhibitory concentration of at least 0.015% (w/v) equivalent to an antifreeze concentration of 5% (v/v) is required to prevent biodeterioration.

References

Black, S. (1951) Yeast aldehyde dehydrogenase. Archives of Biochemistry and Biophysics, 34, 886-897.

26

BS 3150: 1959 Am. I· Ethanediol Antifreeze, Type I. British Standards Institution.

Bradford, M. (1976) A rapid and sensitiVe method for the quantnauon of microgram quantities of protein utilizing the principle of protein · dye binding. Analytical Biochemistry, 72, 248-254.

Gokay, S.F. (1978) The microbiological degradation of ethers. PhD Thesis, University College, Cardiff, Wales.

Jones, A. and Turner, J.M. (1973) Microbiological metabolism of amino alcohols. Biochemical Journal, 134, 167·182.

Lamont, G.W. and Stadtman, E.R. (1963) Fermentation of ethylene glycol by Clostridium glyco/icum. Journal of Bacteriology, 85, 35~362.

Racker, E. (1949) Crystalline alcohol dehydrogenase from bakers' yeast. Journal of Biological Chemistry, 177, 18J.I92.

·.

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ST. PETER'S COLLEGE COLLEGE ROAD, SAL TLEY

BIRMINGHAM 88 3TE

Telephone: 021-328 5950 Telex: 336997 UN lAST G

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