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Journal of Animal Production Advances Isolation of Salmonella Species from Milk and Locally Processed Milk Products Traded for Human Consumption and Associated Risk Factors in Kanam, Plateau State, Nigeria Karshima N. S., Pam V. A., Bata S. I., Dung P. A. and Paman N. D. J Anim Prod Adv 2013, 3(3): 69-74 DOI: 10.5455/japa.20130330124355 Online version is available on: www.grjournals.com ISSN: 2251-7677 KARSHIMA ET AL. Original Article Isolation of Salmonella Species from Milk and Locally Processed Milk Products Traded For Human Consumption and Associated Risk Factors in Kanam, Plateau State, Nigeria 1 Karshima N. S., 2Pam V. A., 1Bata S. I., 1Dung P. A. and 1Paman N. D. 1 Department of Animal Health, Federal College of Animal Health and Production Technology Vom, Plateau State, Nigeria. 2 Department of Parasitology, National Veterinary Research Institution Vom, Plateau State, Nigeria. Abstract This study was conducted to determine the prevalence of Salmonella species in milk and locally processed milk products traded for human consumption and assess the risk factors associated with Salmonella milk contamination in the study area. The study utilized microbiological culture and isolation as well as questionnaire analysis. A total of 600 samples were studied, 200 each for fresh milk (madara), full creamed milk (kindirmo) and skimmed milk (nono). The study revealed an overall prevalence of 8.7% (52/600). About 0.2% (1/600), 3.0% (18/600), 1.0% (6/600), 2.0% (12/600) and 2.5% (15/600) of the overall prevalence were recorded in Dengi, Gargah, Kanam, Gagdi and Gidgid districts of the Local Government Area (LGA) respectively. This revealed a high level of statistical significance (P<0.05). Prevalence rates in relation to milk type were 6.4% (38/600), 1.5% (9/600) and 0.8% (5/600) for fresh milk, full creamed milk and skimmed milk respectively. This showed a high level of statistical significance (P<0.05). Based on sources of water used for milk processing, prevalence rates were 6.8% (41/600), 1.7% (10/600) and 0.2% (1/600) for stream water, well water and borehole water respectively. These differences were highly significant statistically (P<0.05). Risk factors analysis revealed that 31.3% (188/600) of the respondents were not aware of salmonellosis and 82.2% (493/600) were not aware of the risk of acquiring Salmonella via milk consumption. There were high association between the prevalence of Salmonella in milk and the risk of sucking milk directly from the udder of cows (P<0.00001, OR = 8.7448), the use of stream water in processing milk (P=0.0001, OR = 3.6733) and failure to wash the udder of cows before milking (P=0.0041, OR = 0.2072). This study has revealed the presence of Salmonella species in milk traded for human consumption and the risk factors associated with their contamination in Kanam Local Government Area of Plateau State, Nigeria. Keywords: Isolation, Salmonella, risk factors, milk, local milk products, human consumption. Corresponding author: Department of Animal Health, Federal College of Animal Health and Production Technology Vom, Plateau State, Nigeria. Received on: 09 Mar 2013 Revised on: 17 Feb 2013 Accepted on: 21 Mar 2013 Online Published on: 31 Mar 2013 69 J. Anim. Prod. Adv., 2013, 3(3):69-74 ISOLATION OF SALMONELLA SPECIES FROM MILK AND LOCALLY … Introduction Generally, milk has been described as a nearly perfect food because it contains the essential nutrients required by the body in appropriate proportions. However, it is a very efficient vehicle for bringing a large number of people into contact with potential microbial hazards (Jordan, 2007). The safety of dairy products with respect to foodborne diseases is a major global issue especially in the developing countries where production of milk and milk product takes place under poor hygienic, sanitary and Agricultural practices (Jordan, 2007). Bacterial organisms which may gain entry into milk can multiply and cause spoilage, rendering raw or processed milk unsuitable for human consumption due to rancidity, musty odour, or toxin production (Nanu et al., 2007). These organisms in milk indicate inadequate milking procedures, poor storage conditions, or unhygienic productions (Gran et al., 2003; Girmaud et al., 2009). The presence of pathogenic bacteria which may not be associated with spoilage of milk such as Salmonella spp, Brucella spp, Mycobacterium bovis, Listeria monocytogenes and Campylobacter jejuni can render raw and processed milk unsafe for human consumption (Nanu et al., 2007). Salmonellosis is the most common food-borne bacterial disease worldwide (Forshell and Wierup, 2006). There have been increased outbreaks of human salmonellosis in most parts of the world resulting from animal infections (Forshell and Wierup, 2006). The prevention and control of animal salmonellosis has become a global issue, as this has been established as the main source of outbreaks in humans (EFSA, 2006). Food-borne salmonellosis has remained a neglected zoonosis in Nigeria and other developing countries of the world, despite an upsurge of cases reported. Raw milk and milk products are increasingly becoming important sources of human infection with Salmonella. Food vehicles implicated in most food-borne disease outbreaks is often consumed or discarded before clinical symptoms develop in the exposed individual (Fontaine et al., 1980). The screening of milk and other dairy products against pathogenic organisms will play a vital role in curtailing human infection. This study 70 determined the prevalence and the risk factors associated with Salmonella contamination of milk and locally processed milk products in Kanam Local Government Area of Plateau State, Nigeria. Materials and Methods Study Area The study was conducted in Kanam Local Government Area of Plateau State, Nigeria. Kanam is located in the central zone of the state. The study covered five districts namely Dengi, Gargah, Kanam, Gagdi and Gidgid. The major occupation in this area is crop and livestock farming. Sample Collection A total of 600 milk samples were collected from five district of Kanam LGA of Plateau State, Nigeria. Milk samples were collected using systematic sampling technique. Using the sitting arrangement of milk sellers, the fifth milk sellers were sampled each day until 120 samples were collected from each of the five districts. A total of 100 ml of thoroughly mixed milk were purchased from milk sellers and transferred into sterilized, clean and dry sample bottles and transported in ice cool flash within 6 hours to the bacteriology laboratory of the National Veterinary Research Institute Vom, Nigeria for microbiological analysis. Isolation and Identification of Salmonella spp. The isolation of Salmonella spp. was carried out using the method described by Quinn et al. (2002). The samples were enriched in RapportVasilliadis broth. A sterile swab from raw or locally processed milk sample was inoculated into RapportVasilliadis broth and incubated at 37 OC for 24 hours. A loopful of Rapport-Vasilliadis broth was streaked onto xylose lysine desoxycholate agar and incubated for a further 24 hours at 37 OC. The colonies were examined for the characteristic pinkish colour of Salmonella with or without hydrogen sulphite. Suspected colonies were then subjected to indole, motility, oxidase, urease, citrate and sugar fermentation tests for identification of Salmonella spp. Results J. Anim. Prod. Adv., 2013, 3(3):69-74 KARSHIMA ET AL. A total of 600 samples were studied, 200 each for fresh milk (madara), full creamed milk (kindirmo) and skimmed milk (nono) as shown in Tables 1-3. The study revealed an overall prevalence of 8.7% (52/600) as shown in Tables 13. About 0.2% (1/600), 3.0% (18/600), 1.0% (6/600), 2.0% (12/600) and 2.5% (15/600) of the overall prevalence were recorded in Dengi, Gargah, Kanam, Gagdi and Gidgid districts of the Local Government Area (LGA). This revealed a high level of statistical significance (P<0.05) as shown in Table 1. Prevalence rates in relation to milk type were 6.4% (38/600), 1.5% (9/600) and 0.8% (5/600) for fresh milk, full creamed milk and skimmed milk respectively (Table 2). This showed a high level of statistical significance (P<0.05). Based on sources of water used for milk processing, prevalence rates were 6.8% (41/600), 1.7% (10/600) and 0.2% (1/600) for stream water, well water and borehole water respectively (Table 3). This difference was highly significant statistically (P<0.05). Risk factors analysis revealed that 31.3% (188/600) of the respondents were not aware of salmonellosis and 82.2% (493/600) were not aware of the risk of acquiring Salmonella via milk consumption. There were high association between the prevalence of Salmonella in milk and the risk of sucking milk directly from the udder of cows (P<0.00001, OR = 8.7448), the use of stream water in processing milk (P=0.0001, OR = 3.6733) and failure to wash the udder of cows before milking (P=0.0041, OR = 0.2072) as shown in Table 4. Table 1: District based prevalence of Salmonella isolates in milk traded for human consumption in Kanam LGA. Number of samples Number positive District specific Overall District examined for Salmonella prevalence (%) Prevalence (%) Dengi 120 1 0.83 0.2 Gargah 120 18 15.0 3.0 Kanam 120 6 5.0 1.0 Gagdi 120 12 10 2.0 Gidgid 120 15 12.5 2.5 Total 600 52 8.7 X2 = 2400.00 P < 0.00001, Degree of freedom = 16 Table 2: Prevalence of Salmonella isolates in milk traded for human consumption in Kanam LGA in relation to milk type Number of Number positive District specific Overall Milk type samples examined for Salmonella prevalence (%) Prevalence (%) Fresh milk (Madara) 200 38 19.0 6.4 Full creamed milk (Kindirmo) 200 9 4.5 1.5 Skimmed milk (Nono) 200 5 2.5 0.8 Total 600 52 X2 = 1200.00, P < 0.00001, Degree of freedom = 4 Table 3: Prevalence of Salmonella isolates in milk traded for human consumption in Kanam LGA in relation to water source. Number of samples Number positive District specific Overall Source of water examined for Salmonella prevalence (%) Prevalence (%) Stream 317 41 12.9 6.8 Well 204 10 4.9 1.7 Bore hole 79 1 1.3 0.2 Total 600 52 8.7 71 X2 = 1200.00, P < 0.00001, Degree of freedom = 4 J. Anim. Prod. Adv., 2013, 3(3):69-74 ISOLATION OF SALMONELLA SPECIES FROM MILK AND LOCALLY … Table 4: Analysis of risk factors associated with Salmonella contamination of milk traded for human consumption in Kanam LGA. Statistical significance Risk factor NQA R NR Prev (%) P Odd Ratio 95% CI Aware of salmonellosis 600 Yes 412 37 (8.89) 0.6858 1.1380 0.6082, 2.1290 No 188 15 (8.89) Aware that Salmonella can be acquired 600 Yes 107 12 (11.2) 0.3013 1.4305 0.7233, 2.8293 through milk consumption No 493 40 (8.11) Suck milk directly from the udder 600 Yes 302 46 (15.2) <0.00001 8.7448 3.6741, 20.814 No 298 6 (2.01) Use stream water to process milk 600 Yes 317 41 (12.9) 0.0001 3.6733 1.8493, 7.2962 No 283 11 (3.89) Forward milk from sick animals to the 600 Yes 211 18 (8.53) 0.9306 0.9738 0.5337, 1.7701 market No 389 34 (8.74) Wash udder before milking 600 Yes 128 3 (2.34) 0.0041 0.2072 0.0635, 0.6761 No 472 49 (10.4) Wash hands before milking 600 Yes 443 43 (9.71) 0.1283 1.7678 0.8411, 3.7155 No 157 9 (5.7) Discussion Prevalence of Salmonella in milk and locally processed milk products The microbial contamination of milk is multifactorial originating from sources like the air, feed, soil, faeces, grasses and the milking cow itself. Other possible sources of milk contamination include; utensils and water used in the collection and processing of milk (Coorevit et al., 2008). Salmonella contamination of milk (CDC, 2003; Van Kessel et al., 2004; Karns et al., 2005) and milk products (CDC, 1984; CDC, 1994) has been reported in several parts of world confirming its grouping as a neglected zoonotic pathogen. The occurrence of Salmonella in milk and milk products revealed by this study may not be unconnected with the underscored factors. The overall prevalence of 8.7% revealed by this study is lower than the 11.8% reported by Karns et al. (2005). However, it is on the higher side when compared with the 2.6% and 0.0% reported by Van Kessel et al. (2004) and Mhone et al. (2012) respectively. These variations may be explained by the differences in location, hygienic practices and the availability of potable water. The quality of Agricultural practices observed on the farms may 72 also cause variations in levels of contamination with Salmonella. Dengi district which is the most developed of the whole study area recorded the lowest prevalence rate. This may be explained by the availabilities of veterinary services and potable water used for milk procession. The use of water from the streams which are visited by humans, domestic animals and wildlife may be a possible reason for the higher prevalence rate observed by samples from Gargah and Gidgid. This is because these un-flowing streams are easily contaminated by visiting animals. The prevalence recorded by fresh milk samples was statistically higher than that from full creamed and skimmed milk. Both full creamed and skimmed milk are boiled during processing to destroy microbes. This may be the reason for the lower prevalence recorded by these sample types. The unboiled nature of fresh milk may be the probable reason for the higher prevalence observed. This suggests that the milk contamination might have arisen from the lactating cows, themselves, humans or unhygienic milking practices (Addis et al., 2011). Stream water is a source of water for both humans and animals in the rural settlements. Both domestic and wildlife visits such areas to drink water thereby contaminating the water with dirt J. Anim. Prod. Adv., 2013, 3(3):69-74 KARSHIMA ET AL. from their wool or defecation into the stream. For this reason stream water is believed to contain high levels of bacterial load. This is also the likely explanation for the higher prevalence recorded by milk samples processed using water from the stream. The lowest prevalence recorded by samples processed by borehole water may not be unconnected with the cleanliness of the water source used for milk processing. Risk factors associated with Salmonella contamination of milk There were high associations between the prevalence of Salmonella in milk and milk products and sucking milk directly from the udder of cows (P<0.00001, OR = 8.7448), the use of stream water for the procession of milk (P= 0.00001, OR = 3.6733) and lack of washing the udder of cows before milking (P= 0.0041, OR = 0.2072). These factors would have influenced the occurrence of Salmonella species in milk samples studied. Also 31.3% of respondents who were unaware of the infection and those who were unaware of the roles of water, dirty cows, sick animals and dirty hands in the contamination might have cause contamination of milk during processing. This lack of knowledge would have played a key role in the prevalence of Salmonella isolated from milk traded for human consumption in Kanam Local Government Area of Plateau State, Nigeria. 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