Bacterial contaminants of salad vegetables in Abuja Municipal Area Council, Nigeria

Tags: salad vegetables, Lettuce, Federal Capital Territory, Nigeria, leafy green vegetables, heat treatment, Bacteriological quality, vegetables, Malaysian Journal of Microbiology, Ibrahim Kolo Department of Microbiology, Human Virology and Biotechnology, Pseudomonas aeruginosa, Abuja Municipal Area Council, Nigeria, Nigeria Aboh, pathogenic microorganism, bacterial contamination, National Institute for Pharmaceutical Research and Development, Carrots Cucumber, pathogenic microorganisms, Federal Capital Territory, green leafy vegetables, samples, public health concern, Salmonella sp, Shigella spp, urinary tract infections, Salmonella spp, Klebsiella spp, urinary tract, Sampling Lugbe Lugbe Lugbe Wuse Wuse, Cucumber, Viable Count, Carrots, total viable count, Lugbe
Content: Malaysian Journal of Microbiology, Vol 7(2) 2011, pp. 111-114
SHORT COMMUNICATION
Bacterial contaminants of salad vegetables in Abuja Municipal Area Council, Nigeria
Aboh Mercy Itohan*, Oladosu Peters and Ibrahim Kolo
Department of Microbiology, Human Virology and Biotechnology, National Institute for Pharmaceutical Research and Development, Idu. P.M.B 21 Garki, Abuja, Nigeria. E-mail: [email protected] Received 18 March 2010; received in revised form 8 September 2010; accepted 17 September 2010 _______________________________________________________________________________________________ ABSTRACT
Salad vegetables are essential part of people's diet all around the world. They are usually consumed raw and often without heat treatment or thorough washing; hence have been known to serve as vehicles for the transmission of pathogenic microorganism associated with human diseases. Fresh samples of lettuce, carrot and cucumber collected from different markets and vendors in Abuja Municipal Area Council, Federal Capital Territory, Nigeria were evaluated for bacterial loads using spread plate agar dilution method. Bacterial loads ranged from 1.6 x 106 to 2.9 x 108 cfu/g. Escherichia coli, Klebsiella and Enterobacter were amongst the coliforms (lactose fermenters), while Proteus, Pseudomonas aeruginosa, Salmonella and Shigella were non-lactose fermenters associated with the samples. Staphylococcus aureus was isolated from majority of the samples.
Keywords: salad vegetables, heat treatment, coliforms, pathogenic, bacterial load, agar dilution _______________________________________________________________________________________________
INTRODUCTION Vegetable salad is a very common food accompaniment in Nigeria. The vegetables that usually make up this recipe include tomatoes, cucumber, carrots, green chili, cabbage and lettuce. They are sold in almost every market, and can be seen hawked around by traders. Fruits and vegetables have been identified as significant sources of pathogens and chemical contaminants (Uzeh et al., 2009). As a result, environmental and food microbiologists have continued to identify and suggest control measures for hazards at all stages in the supply chain (Johngen, 2005). Khan et al. (1992) reported that bacterial contamination results from various unsanitary cultivation and marketing practices. In another study, Tambekar et al. (2006) reported that bacterial contamination of salad vegetables was linked to the fact that they are usually consumed without any heat treatment. These vegetables can become contaminated with pathogenic microorganisms during harvesting, through human handling, harvesting equipments, transport containers, wild and domestic animals. Pathogens from the human and animal reservoir as well as other environmental pathogens can be found at the time of consumption. Although spoilage bacteria, yeasts and mould dominate the micro flora on raw fruits and vegetable, the occasional presence of pathogenic bacteria, parasites and viruses capable of causing human infections has also been documented (Hassan et al., 2006). Coliforms are facultative anaerobic Gram negative rods belonging to the family Enterobacteriacaea. They are
known contaminants of food and water, causing various intestinal and extra-intestinal infections such as urinary, central nervous system and Respiratory Tract Infections (John, 2007). The presence of E. coli, Enterobacter sp., Salmonella sp., Shigella sp and Pseudomonas aeruginosa, has been reported in salad vegetables (Khan et al., 1992; Tambekar, 2006). Mehmet and Aydin (2008), also reported the presence of E. coli in some green leafy vegetables. Due to the favourable climatic condition for cultivation of salad vegetables, as well as the cultural practice of dwellers in Northern Nigeria, the consumption rate of these vegetables is higher than in other regions in Nigeria. This study was therefore designed to assess the bacteriological profile of some salad vegetables sold in Abuja Municipal Area Council (AMAC), Federal Capital Territory (FCT), Nigeria. MATERIALS AND METHODS Collection of samples A total of 15 samples of carrots (5), cucumber (5) and lettuce (5) were collected in sterile polythene bags from different markets and vendors in AMAC, Federal Capital Territory, Nigeria as shown in Table 1. Isolation and identification of bacteria by spread plate method A 25 g of vegetable samples were weighed, rinsed in a 250 mL beaker containing 100 mL of sterile distilled water
*Corresponding author
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and rinsed water samples were diluted 1:10-2 and 1:10-4. 0.1 mL of each dilution was spread on MacConkey agar (Oxoid, England) and the plates were incubated at 37 °C for 24 h for isolation of bacteria (Khan et al., 1992). Total viable counts were determined by counting both red and non-red colonies growing on the plates. Based on their morphological characteristics, red and non-red colonies were selected from each MacConkey agar plate for Gram staining, biochemical tests namely: IMViC (Indole, Methyl Red, Voges-Proskauer, citrate), urease, oxidase, catalase, triple sugar iron; and sub-culture on differential media (eosin methylene blue agar at 37 °C and 44 °C for 24­48 h; and mannitol salt agar, and cetrimide agar at 37 °C), nutrient agar (Holt et al., 1994). RESULTS Total viable counts The salad vegetables showed a wide variation in total viable count ranging from 1.6 x 106 to 2.9 x 108 cfu/g at 37 °C (Table1). Cultural characteristics and biochemical Identification of isolated strains A total of 22 bacterial types were isolated. Non-red colonies from MacConkey plates that grew with golden yellow on mannitol salt agar and were Gram positive, coagulase-positive and catalase-positive were taken as Staphylococcus aureus; non-red colonies from MacConkey plates that grew with greenish pigment on nutrient and on cetrimide agar, and were Gram negative, indole-negative, oxidase positive and coagulase-negative were taken as P. aeruginosa; non-red colonies from MacConkey plates that were Gram-negative, indolenegative, methyl red­positive, Voges-Proskauer-negative,
citrate-negative, acidic butt, alkaline slant with no blackening on TSI slant were taken as Shigella spp., nonred colonies from MacConkey plates that were Gram negative, indole-negative, methyl red­positive, VogesProskauer-negative, citrate-positive, acidic butt and alkaline slant with blackening on TSI slant, ureasenegative and colourless colonies with black center on SS agar were taken as Salmonella spp., non-red colonies from MacConkey plates that were Gram negative, indolenegative, methyl red­positive, Voges-Proskauer-negative, citrate-positive, acidic butt and alkaline slant with blackening on TSI slant and urease-positive were taken as Proteus spp. Red colonies from MacConkey plates that grew with greenish metallic sheen on EMB agar and were Gram negative, indole-positive, methyl red-positive, Voges-Proskauer-negative and citrate-negative were taken as E. coli; red mucoid colonies from MacConkey plates that were Gram negative, indole-negative, methyl red-negative, Voges-Proskauer-positive and citratepositive were taken as Klebsiella spp.; red colonies from MacConkey plates that were Gram negative, indolenegative, methyl red-negative, Voges-Proskauer-positive, citrate-positive and urease-positive were taken as Enterobacter spp. The genera of the bacteria isolated and the percentage occurrences are S. aureus (46.7%), Klebsiella spp. (26.7%), Enterobacter spp. (20.0%), Proteus spp. (13.3%), and P. aeruginosa (13.3%), E. coli (6.7%), Shigella spp. (6.7%) and Salmonella spp. (6.7%). DISCUSSION Freshly consumed vegetables especially those used in salad mixtures, have been implicated in Food poisoning and thus hazardous to the health of the consumers. This could be linked to the fact that most of these vegetables are consumed without being subjected to thermal process or even thorough washing (Lund, 1992).
Table 1: Total viable count of salad vegetables.
Sampling No 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13. 14. 15.
Date of Sampling 27/04/09 27/04/09 27/04/09 04/05/09 04/05/09 04/05/09 10/05/09 10/05/09 10/05/09 18/05/09 19/05/09 18/05/09 24/05/09 24/05/09 24/05/09
Site of Sampling Lugbe Lugbe Lugbe Wuse Wuse Wuse Garki Garki Garki Karimo Karimo Karimo Kuchingoro Kuchingoro Kuchingoro
Type of Sampling Lettuce Carrots Cucumber Lettuce Carrots Cucumber Lettuce Carrots Cucumber Lettuce Carrots Cucumber Lettuce Carrots Cucumber
Total Viable Count (cfu/g) 8.6 x 107SD 8.49x106 4.4 x 107 SD 5.66x106 2.4 x 107 SD 2.83x106 2.9 x 108 SD 1.70x107 4.6 x 107 SD 2.83x106 7.6 x 106 SD 5.65x105 6.4 x 107 SD 1.13x105 2.4 x 107 SD 0 1.8 x 107 SD 2.83x106 8.4 x 107 SD 5.66x106 1.0 x 108 SD 0 1.5 x 107 SD 1.13x106 1.2 x 108 SD 5.66x106 1.6 x 108 SD 0 1.6 x 106 SD 5.66x105
Key: SD = standard deviation
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Table 2: Presence of pathogenic bacterial flora on vegetables.
Vegetables
No. of samples
Proteus spp.
E. coli
Enterobacter spp.
P. aeruginosa
S. aureus
Salmonella spp.
Shigella spp.
Lettuce
5
1
1
1
Carrots
5
1
-
1
Cucumber
5
-
-
1
Total
15
2
1
3
%
13.3 6.7
20
-
3
1
-
1
2
-
-
1
2
-
1
2
7
1
1
13.3
46.7
6.7
6.7
Klebsiell a spp. 2 1 1 4 26.7
In this study, the total viable count was done on MacConkey agar (Oxoid, England) by spread plate count method (Khan et al., 1992). Among the carrots analyzed, samples sourced from Kuchingoro had the highest total viable count of 1.6 x 108 cfu/g. This was higher than the total viable count of 5.7 x 106 cfu/g for carrots samples reported by Uzeh et al. (2009). Hall et al. (1967) suggested that a limit of 10 cfu/g should be standard with market raw food. Carrots are usually harvested from the soil hence can become contaminated by pathogenic organisms in soil. Among the three different salad vegetables analyzed, cucumber samples had the lowest bacterial load. The highest total viable count for cucumber samples was gotten from samples sourced from Lugbe with a load of 2.4 x 107 cfu/g while those sourced from Kuchingoro had the lowest load of 1.6 x 106 cfu/g. This result was comparable to the bacterial load of 1.9 x 106 cfu/g reported by Abdullahi et al. (2010). Unlike carrots, cucumber fruits are rarely contaminated by soil pathogens as they do not come in contact with soil. Contamination with these pathogens could be due to poor hygiene practices by handlers. Lettuce samples sourced from Kuchingoro had the highest coliform load of 2.9 x 108 cfu/g. This was higher than a load of 6.9 x 106 cfu/g reported for lettuce samples by Mehmet et al. (2008). The high bacterial load in lettuce can be attributed to the large surface area of the leaves suitable for water contact, making them susceptible to bacterial contamination. Ercolani et al. (1976) reported an average count for total coliform and faecal coliform of 5.95 x 104 and 6.13 x 102 cfu/g for lettuce respectively. The presence of Enterobacter spp., Klebsiella spp. and S. aureus is observed in all samples of salad vegetables in which S. aureus was predominant (Table 2). The detection of S. aureus is of serious public health importance because of its ability to cause a wide range of infections especially food- borne intoxication (Tambekar et al., 2006). Contamination with S. aureus has been linked to carriage in nasal passages of food handlers or by infected workers. The presence of S. aureus and some Gram negative rods have been reported to contaminate some salad vegetables such as carrots, cucumber, tomato and radishes (Beuchat, 1995). The presence of E. coli in the salad vegetables analyzed is indicative of faecal contamination. E. coli are part of the normal flora of the human intestines. Some
strains of E. coli have been linked to diahorrea, gastroenteritis and Urinary tract infections (Hassan et al., 2006). Klebsiella spp. is second only to E. coli as a urinary tract pathogen. It is well known in the environment and can be cultured from soil, water and vegetables when consumed raw as in salads. E. coli were also isolated from some leafy green vegetables (Ibrahim, 1996). Khan et al. (1992) also isolated E. coli, Klebsiella spp. and Enterobacter spp. from salad vegetables. In a study done by Tambekar et al. (1995), E. coli was found to be predominant on some salad vegetables which included coriander followed by carrot, radish, spinach, fenugreek and cucumber. Salmonella spp. and Shigella spp. are non-lactose fermenters usually associated with water contamination. Contamination with these organisms could arise from washing vegetables with contaminated water or handling of vegetables by infected workers. In this study, Salmonella sp. was isolated from lettuce. Its presence n food is of serious concern to safety. According to the WHO (2002), effect of microbiological hazards such as Salmonella on food safety is now a major public health concern worldwide. Ibrahim (1996) isolated Salmonella spp. from lettuce, cucumber and parsley. Salmonella spp. has also been isolated from salad vegetables in waakye a street food in Ghana. In this study, Salmonella spp. was present only in lettuce samples. Shigella spp. has been frequently found in salads and dairy products. It is a principal agent of bacterial dysentery. Pseudomonas spp. is a prominent inhabitant of soil and water. The organism is responsible for diseases of vegetables like angular leaf spot of cucumber (Uzeh et al., 2009). Its presence in salad vegetable is also of public health concern as it has been implicated in several infections. CONCLUSION In conclusion, the high bacterial load and presence of these organisms especially E. coli in the salad vegetable samples could serve as an indicator for the need to promote awareness about the possible health hazards that could be due to poor handling of these vegetables. There is therefore, the need for regulatory bodies to ensure that microbiological standards are established and practiced by farmers and marketers for the handling and distribution of salad vegetables.
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Mal. J. Microbiol. Vol 7(2) 2011, 111-114 REFERENCES Abdullahi, I. O. and Abdulkareem, S. (2010). Bacteriological quality of some ready to eat vegetables as retailed and consumed in SabonGari, Zaria, Nigeria Bayero. Journal of Pure and Applied Sciences 3, 173-175. Beuchat, C. R. (1995). Pathogenic microorganisms associated with fresh produces. Journal of Food Protection 59, 204-216. Ercolani, G. L. (1976). Bacteriological quality assessment of fresh marketed lettuce and fennel. Applied Environmental Microbiology 31, 847-852. Hall, H. E., Brown, D. F. and Lewis, K. H. (1967). Examination of market foods for coliform organisms. Applied Microbiology 15, 1062-1069. Hasan, A., Utku, O. and Koray, K. (2006). Determination of total aerobic and indicator bacteria on some raw eaten vegetables from wholesalers in Ankara, Turkey. International Journal of Hygiene and EnvironMental Health 209, 197-201. Holt, J. G., Kreyg, H. R. A., Standly, J. T. and Williams, S. T. (1994). Bergey's Manual of Determinative Bacteriology. 9th edn. The Williams and Wilken Company, Baltimore, MD, USA. Ibrahim, S. A. (1996). Microbiological studies on some salad vegetable in local markets. Journal of King Saud University 8, 99-106. John, R. W. (2007). The Enterobacteriacaea basic properties. Department of Pathology Northwestern University, FeinberG School of Medicine. Jongen, W. (2005). Improving the safety of fresh fruits and vegetables. Wageningen University, THE NETHERLANDS. Khan, M. R., Saha, M. L. and Kibria, A. M. (1992). A bacteriological profile of salad vegetables in Bangladesh with special reference to coliforms. Applied Microbiology 14, 88-90. Lund, B.M. (1992). Ecosystems in vegetable foods. Journal of Applied Bacteriology 73, 115-135. Mehmet, E. E. and Aydin, V. (2008). Investigation of the microbial quality of some leafy green vegetables. Journal of Food Technology 6, 285288. Tambekar, D. H. and Mundhada, R. H. (2006). Bacteriological quality of salad vegetables sold in Amravati city, India. Journal of biological sciences 6, 28­30. Uzeh, R. E., Alade, F. A. and Bankole, M. (2009). The bacterial quality of pre-packed mixed vegetable in some retail outlets in Lagos, Nigeria. African Journal of Food Science 3, 270272. WHO. (2002). WHO Global Strategy for Food Safety: Safer Food for Better Health. Food Safety issues, WHO Geneva. 114
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