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June 1986, Volume 36, Issue 6

Original Article


Zumara Sami  ( National Institute of Health, Islamabad. )
Sabiha Roohi  ( National Institute of Health, Islamabad. )
Mubashir A. Khan  ( National Institute of Health, Islamabad. )
Abdul Ghafoor  ( National Institute of Health, Islamabad. )


A total of 282 food samples of different varieties were analysed for microbiological con­tamination.
High miàrobial load and presence of patho­genic organisms renders the poor hygienic stan­dard of the foods examined. Mean viable bacterial count was found to be 1.2x108/gm food and 49% of the samples were declared unfit for human consumption.
Pathogenic bacteria recovered included E. co/i (14.9%), Staphylococcus aureus (1.8%), Clostridium perfringens (1 .1%) and Salmonella (0.7%). Bacillus cereus (0.3%) was also incriminated.
Foods from street side shops and vendors  were heavily contaminated (70%). Middle and upper class restaurants also indicated a high rate (40%)of microbial contamination (JPMA 36:141 1986).


Food can be an important vehicle for trans­mission of a broad spectrum of diseases. The transmission of human diseases through food, water and waste water is a global problem, particu­larly of developing countries. Awareness of micro­biological health hazards arising from the consum­ption of contaminated food has grown in recent years and has resulted in national and international intensification of food hygiene programmes.1
The interaction between nutrition and infection is of fundamental importance to public health workers, especially in developing countries like Pakistan, because infections take a heavy toll ‘of human life. Epidemiological studies indicate that most infections make the nutritional status worse if the individual is already consuming a deficient diet. 2
Malnutrition in combination with endemic diarrheal disease is one of the most significant health problems among children in many developing countries1. A correlation between contaminated food, water and diarrhea has also been proved.3
One thousand seven hundred and three outbreaks of foodborne origin (97,590 cases) were reported by state Health Department and other agencies during 1967 through l97l. 4During this period, staphylococcal intoxication, salmonellosis and Clostridium perfringens gas­troentritis were the diseases most frequently re­ported.5
Microorganisms contaminating food may be non pathogenic and yet may cause chemical changes that render food unit for human consump­tion or may be pathogenic and cause infections or food poisoning when consumed.6
Keeping in view the importance of the subject, and to know the microbial status of foods in this region this study on microbiological contamination of different foods was undertaken,’ at National Institute of Health, Islamabad.


1. Collection of food specimens:
Foods of public health importance were mainly analysed for microbiological contaminants. A total of two hundred and eighty two (282) samples investigated in the study were collected from different hotels, restaurants, institutional canteens, railway station and small food industries in Rawalpindi and Islamabad area.
Fifty grams of each food sample was asep­tically placed in wide mouth glass stoppered flasks and immediately transported to the laboratory for microbiological examination.
2. Primary isolation media:
a. Plating media:
Viable bacterial count was per­formed on plate count agar. SSA (Difco) and Mac Conkey agar
(Difco) were .used for the isolation of members of the enterobacteria­ceae and other gram negative or­ganisms. Thiosulphate Citrate Bile Salts sucrose agar TCBS (Difco) was used for recovery of V. cholerae and V. paraheamolyticus. However, isolation of gram positive aerobic organisms was carried out on Baird parker medium (Oxoid) and anaerobic organisms on Blood agar i.e 10% defibrinated sheep blood’ in nutrient agar (Difco). Egg yolk agar (Oxoid) was medium of choice for Bacillus cereus. Fungal species were recovered on Sabo­rauds dextrose agar (Difco).
b. Enrichment medium:
Different enrichment media were used according to the organisms recovered. Tetrathionate broth (Oxoid) was used for Salmonella and Shigella organisms while alka­lline peptone water pH 8.6 (Difco) for Vibrio cholera. Phosphate buffer saline pH 7.2 (Difco) was used for the enrichment of Yer­sinia enterocolitica.
For anaerobic organisms fluid thiogly­collate medium and cooked meat medium were employed. Glucose broth and MacConkey’s broth were also used for gram positive and gram negative ‘organisms respectively.
Twenty five gram of food sample weighed aseptically was blended with 225 ml of PBS pH 7.2. From this homogenate serial ten fold dilutions were made from lxl01 to lxl06.
Methodology used for isolation and identi­fication of organisms and viable plate counts are given in flow diagrams I, II & III.7 Viable aerobic counts were determined by the pour plate method.7 Coagulase positive stap­hylococcus were identified by using standard methods8. Members of enterobacteriaceae were identified systek system No. 1 by Eiken chemical, Japan and scheme proposed by Edward and Ewing. 9 Other organisms were isolated and identi­ fled using methods given by Topley and Wilson10


Two hundred and eighty two samples of different varieties of food including raw, cooked and processed foods were screened for the micro­biological contamination. The detail description of foods examined and microbiological contamina­tion from different types of food is given in table I.

Each food sample yielded 0-3 microbial agents. Out of 282 foods examined, 138 (49%) were found to be contaminated with different bacterial agents.

Table II also indicates the percentage of microbial contaminants found in different varieties of food. The highest percentage of contamination was found in raw foods, raw meats and dried.
The total viable count in aerobic plates ranged upto lx 10 /gm of the food when incubated at 37°C, whereas the mean viable plate count of the examined foods was 1.2x108 / gm (Table II). Of the examined food samples 51% were found fit for human consumption having counts within the permissible range or were completely sterile showing no microbial growth.
This table also shows the permissible range, mean viable aerobic plate counts, the highest and the lowest viable aerobic counts per gram of different types of food examined.
Microbial contamination with reference to the site of collection was also determined (Table III).

The highest rate of contamination in dif­ferent type of foods was found in street side shops and vendors i.e. 70%, whereas in the middle and high class hotels and restaurants the rate of contamination was approximately 40%, which is again a high figure.

The microorganisms isolated from dif­ferent type of foods are given in Table IV.

The highest percentage after non pathogenic strains was of E. coli (14.9%). The coliform group and other microorganisms were 42.5% in the examined foods, Clostridium perfringens was present to the extent of 1.1% in the raw and cooked meat products only.
Streptococcus feacalis was found in 0.7% and Staphylococcus aureus was recovered from 1.8% of the food samples.
Bacillus cereus was incriminated in 0.3% of the examined foods. Salmonella spp. was isolated only from 2 samples (0.7%) one from milk and the other from raw meat. Fungus was mostly isolated from processed or dried foods and incidence was 12.3% of the examined foods.


Presence of microorganisms like Escherichia coli, faecal coliforms, Streptococcus faecalis and Staphylococcus aureus (relatively recently accepted as an indicator organism) in food and water pro­vides a useful indication that faecal contamination has occurred in the food1.
Foods that are eaten raw present a health hazard if they are contaminated with pathogenic microorganisms.11 Our results indicate that the highest percentage of contamination was mainly found in raw foods, raw meats and dry foods. It is an established fact that once food is subjected to the attack of microorganisms the micro­bial population increases within a few hours at normal temperature, because food is an ideal culture medium for multiplication of a variety of microorganisms. 12
The international Commission on Micro­biological Specifications for Foods (ICMSF) has recommended criteria for different foods (Table II). When these criteria were applied to the data of present study the bacterial numbers were very high. Viable bacterial counts suggest practice of inadequate hygienic measures, malhandling and unhygienic conditions of the retail shops. This is in agreement with another study. 13
Comparing the foods according to the site of collection it has been found that the highest contamination was recovered from the foods of street side shops and vendored foods, indicating more unhygienic conditions. But, the rate of contamination in the middle and high class hotels was also quite alarming (Table III).
It is stated that routine microbiological testing of foods is necessary for recommending effective sanitary control of hotels, markets and institutional canteens.5 It was also recommended that every catering service should carryout the microbiological quality control of its food routinely, in order to protect health of the con­sumer. 14
The percentage of Escherichia coli was highest among pathogenic strains (Table IV). Although the presence of E. cOil does not neces­sarily mean a pathogenic strain, but implies a certain risk as some serotypes are enteropathogenic and cause diarrhoea in infants and its presencein large numbers may cause diarrhea in adults3,15
Great majority of outbreaks of food poisoning in every country throughout the world were caused by enterotoxin of Strap hyiococcus aureus.16 In this study only 1.8% foods harboured this strain which is apparently a low incidence. This finding is in agreement with other studies12,17
Clostridium perfringens was present to the extent of 1.1% in meat and meat products. This is considered to be one of the most important causes of food poisoning. Spores of typical food poisoning strains and certain type c strains are heat resistant 18
Streptococczss faecalis was found in 0.7% food samples. These are preferred as indicators of feacal pollution, since they survive longer in water than coliform bacteria.1
Bacillus cereus was the third most common cause of bacterial food poisoning between 1960 and 1968 in Hungary. 19 However in our study only 0.3% of the foods were incriminated with Bacillus cereus.
Salnionellosis is usually food/waterborne infection and the number of organisms required to produce clinical infection depends on many factors. An extensive outbreak of Salmonella eastbourne food poisoning on North American Continent in 1974 involved chocolate candy containing well under one salmonella bacterium pergram of food20
Fungus was mostly isolated from processed or dried foods. The correlation between fungi and mycotoxins is, however, very poor. In many cases large number of mouldy foods have been examined for mycotoxins with a few or no posi­tive results1


1. Microbiological aspects of food hygiene. WHO Tech. Rep. 1976, 598: 6,8,50-5 2.
2. National Health Laboratory Guide notes on nutrition Islamabad, National Health Laboratories 1973;p. 61.
3. Rashid, F. Identification and characterization of enterotoxigenic E. coli isolated from infantile diarrhea cases and their culture sensitivity pattern. Islamabad, Quaid.e-Azam, University,1983; 24-25.
4. U.S. Department of Health, Education and Welfare Foodborne outbreak annual summary 1967-71 Public Health service, Centre of disease control, Atlanta, Georgia, Auther, 1972.
5. Bryan, F.L. Microbiological food hazards today based on epidemiological information. Food Technology, 1974;52:66.
6. National Institute of Health Food contamination study and control in Asia and far East Islamabad, National Institute of Health, 1984; p. 82.
7. SEAMIC Manual for the laboratory diagnosis of bacterial food poisoning and the assessment of the sanitary quality of food Japan, The South­east Asian Medical Information Centre Inter­national Medical Foundation, 1978; P. 70.
8. Coffins, C.H. and Lyne, P.M. Microbiological methods 4th ed. London, Butterworth, 1967; p.401.
9. Edwards, P.R. and Ewing, W.H. Identification of enterobacteriaceae. 3rd ed. Minneapalis, Burgess, 1972;p 258.
10. Wilson, G.S., Miles, A., and Parker, M.T. Topley and Wilson’s principles of bacteriology, virology and immunity. 7th ed. London, Butterworth, 1984; V.2. V.2.
11. WHO Expert Committee Microbiological aspects of food hygiene. WHO Tech. Rep. 1968; 399:42.
12. Begum, M. Bacteriological analysis of different foods to determine the fitness for human con­ sumption. JPMA., 1985; 35:79.
13. Munce, B.A. Microbial status of international airline food. Proceedings of world congress foodborne infections and intoxications. Berlin (West), Institute of Veterinary Medicine, 1980; p. 141.
14. Urn, V.S. and Jegathesan, M.A. Bacteriological study of some and non frozen foods. Southeast Asian J. Trop. Med. Public Health, 1977;8:37.
15. Pickering, L.K., Evans, D.J. and Munoz, 0. Prospective evaluation of enteropathogens in children with diarrhea in Houston and Mexico. J. Pediatr., 1979; 98: 383-388.
16. Bryan, F.L. Factors that contribute to outbreaks of food disease. J. Food: Prop., 1977 ;12: 311.
17. Donnelly, C.B., Black, L.A., and Lewis, K.H. Occurence of coagulase positive staphylococci in cheddar cheese. Appi. Microbiol., 1964; 12:311.
18. Cruickshank, R., Duguid, J., Marmion, B.P. and Swain, R.H.A. Medical microbiology, r2th ed. Edinburgh, Churchill livingstone, 1975; v.2; p. 473.
19. Ormay, L. and Novotony, T. The significance of Bacillus cereus food poisoning in Hungary Netherland, Proceeding of 6th International symposium in food microbiology, 1969.
20. Craven, P.C., Mackel, D.C., Baine, W.B., Baker, W.H., Gangarosa, E.J., Goldfield, M., Rosenfield, H.,Altman, R., Lachapeils, Q., Davies, J.W. and Swanson, R.C. International outbreak of Salmonella eastbourne infection traced to con­taminated chocolate. Lancet, 1975; 1: 78.

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