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Assessment exposure to Escherichia coli among Halal cattle abattoir workers in Malaysia.

INTRODUCTION

An abattoir is a premise approved and registered by the controlling authority for hygienic slaughtering and inspection of animals, processing and effective preservation and storage of meat products for human consumption [1]. It may become an area or place for propagation and growth of pathogenic microbes. Dissemination of the microbes directly depends on the hygienic and sanitary precautions taken by the abattoir industry. Culpable microbes which are mostly found to contaminate and cause meat spoilage and its products are bacteria, yeast and moulds. Butchers and work men introduced the organisms directly or indirectly during animal processing. Indirect sources for meat contamination include; water and air in the dressing, cooling, and cutting rooms or tables and even the environment [2]. High ambient temperature of the tropical region coupled with insufficiency of portable water and poor handling practices by the butchers predisposes meat and the resulting products to considerable microbial contamination which may lead to rapid deterioration and even poisoning [3]. Bacteria such as Salmonella spp, Escherichia coli and Clostridium spp are of public health concern worldwide for their role in contamination of meat and its product as well as their role in food-borne disease transmission. Serotypes of these organisms have also been severally reported to be resistant to many antimicrobials. [4][5][6][3][7].

Many occupational zoonotic diseases of different sources and origins are encountered among abattoir workers who handled the slaughter of different species animals for human consumption. Zoonoses are described as those diseases and infections which are naturally transmitted from animals to humans. The slaughtering process involved various stages; slaughtering, hide removal, evisceration, deboning and cutting. Each of the processes requires proper handling of the animal as set by the guidelines for operating abattoirs. If precautions are not taken, abattoir workers constitute a major group at risk of occupational zoonosis. With technological advancement, close contact with the animals is significantly reduced. However, in developing countries where such facilities are unavailable, close contact that exists between the abattoir workers and animals/tissue of animals during slaughtering or processing may pose occupational hazards. Although Clinical signs of the zoonotic diseases may be observed easily. Confirmation of disease requires standard laboratory diagnostic techniques such as microbiological, immunological and molecular [8].

Zoonotic diseases represent about 70% of the number of emerging infectious diseases in recent time [9]. There are over 300 zoonotic diseases of diverse etiologies which cause high morbidity and mortality [10]. Zoonotic diseases occur in both sexes, in all age groups, in all seasons, in all climatic zones and in rural and urban settings [10][11]. High incidences of the diseases are linked with increased demand for meat and meat product by continuous growth of human population and human contact with animals become unprecedented. Movement of animals across international boundaries to complement the local supply can increase the risk of zoonotic diseases especially from endemic zones [12]. Zoonotic infections can be transmitted via various routes [13]. However, direct contact appeared to be the most common route of entry for the pathogen in the employees working in abattoirs [14][15].

Enterohemorrhagic Escherichia coli (EHEC) are one of the zoonotic pathogens that can be transmitted to the abattoir workers. It has been identified as a major cause of serious illness and mortality in food borne outbreaks that involved different variety of foods [16]. It was reported that an E. coli 0157:H7 strain that was involved in an outbreak of Hemorrhagic Colitis in the United States produced a toxin named Shiga toxin [17]. Similar report suggested that Shiga Toxin Producing E.coli (STEC) were epidemiologically associated with Hemolytic Uremic Syndrome HUS [18]. Generic E.coli can be a nonpathogenic or nontoxic member of the normal microflora in humans and other animals. However, virulence genes assimilated through various methods in the environment have conferred different forms of pathogenicity to strains of E.coli. World Health Organization (WHO) reported that illness caused by the consumption of contaminated foods is one of the most frequent health problems in the modern world [19]. Meat falls into the above because of its daily consumption to satisfy the protein need of the human population. Public health burden is increased due to hospitalization and cause decline in human productivity, which invariably could result in substantial decline economically. The most frequently implicated source of E. coli 0157:H7 outbreaks is still ground beef accounting for 75% of E.coli 0157:H7 outbreaks [20]. Dairy products and undercooked minced beef can be directly contaminated by cattle feces during either milking or slaughtering processes by the workers [21]. Results from a study of 90 outbreaks that were microbiologically confirmed in the UK, Ireland, Denmark, Norway, Finland, USA, Canada, and Japan, occurring between 1982, and 2006, indicated that the source of transmission was food in 42.2% of the outbreaks, dairy products in 12.2%, animal contact in 7.8%, water in 6.7%, environmental in 2.2%, and unknown in 28.9% [22]. This research aims at finding potential risk of exposure to zoonotic E.coli abattoir workers and possible source contamination.

MATERIALS AND METHODS

Study Location:

Malaysia consists of thirteen (13) States. Six (6) States were randomly selected for the assessment; Pahang, Selangor, Terengganu, Negeri Sembilan, Melaka and Perak. Figure 1 shows the map of Peninsular Malaysia and the Thirteen States.

Sampling:

Population:

Butchers form government Halal cattle abattoir.

Sampling Frame; all butchers working in government Halal cattle abattoir.

Sample Collection:

Hands Swab:

A total of One hundred and sixty five (156) samples were collected. Sterile cotton swab which contained transport medium (MEUS, Italy) were used for sample collection. Samples from hands of abattoir workers were taken before and after work. All samples will be kept in cooler box and transported to Applied Microbiology Laboratory Universiti Putra Malaysia.

Carcass and Fecal Samples:

A sum of sixty (130) cattle carcass and fecal samples were collected for the research. Sterile swabs sticks with transport medium were used to collect carcass samples by rubbing the shoulder to the thigh in a zigzag manner as describe by McEvoy et al., 2003 with slight modification. For fecal samples, cotton swabs were loaded with each of the samples by dipping the tip end of the cotton swab into the faeces. The swab sticks were stored in transport medium and then placed into an Ice pack for transportation to the laboratory before the bacteriological analysis. The bacteriological analysis of all the samples was conducted within 2 hours of sample collection or kept refrigerated at 4[degrees]C but not beyond 24 hours (Abdullahi et al., 2005).

Bacteriological Analysis:

All samples were subjected to Bacteriological analysis. The samples from hands of abattoir workers were inoculated onto Chromo cult[c] agar and CT-SMAC (Merck, Germany) agar for isolation of Escherichia coli O157:H7 and other enterobacteriaceae.

Laboratory Procedures:

The swabbed samples were appropriately inoculated onto Chromocult Agar[R] (Merck, Germany). Colonies with dark blue to violate coloration counted after incubation at 37[degrees]C for 24 h. All isolates that showed dark blue coloration from swabs samples on Chromocult Agar[R] were characterized biochemically based on methods previously described Cheesebrough [23]. Isolates identified biochemically as E.coli were further screened on Cefixime Tellurite Sorbitol MacConkey agar (Merck, Germany) by incubation for 24 hrs at 37[degrees]C. E.coli O157:H7 appeared colorless, while Non-O157H7 appeared pink [24]. Colonies that appeared colorless (non-Sorbitol fermenters) on CT-SMAC were presumptively identified as E.coli O157:H7 and were preserved on nutrient agar slant for confirmation using Slide agglutination test. Isolates that were colorless were serotyped using Serotest[R] for E.coli O157:H7 (S&A Lab., Thailand), a polyclonal antibody produced for serological identification based on agglutination method. A drop of serum was placed onto the test area and a drop of saline onto the control area of a clean glass slide. Using a platinum wire loop, a portion of the growth was transferred onto drop of serum and mixed. Another loopful from the growth was also mixed with the normal saline (control). The glass slide was tilted back and forth for one minute. Agglutination was observed and recorded according to manufacturer's instruction.

RESULTS AND DISCUSSION

The results have shown that no Escherichia coli 0157:H7 was isolated on the hands of abattoir workers before and after work. However, a total prevalence a prevalence of 9.7% was recorded for all samples during work. Only two abattoirs were found to have E.coli 0157:H7 on the hands of abattoir workers while working Kuala Pilah (67%) and Ipoh (20%) as shown in Table 1. For non-O157:H7, total prevalence of 33.3% during work and 13% after work were obtained. The occurrence of the bacteria during work may be link with contact with the intestinal content or hide of the animals. None of the abattoir workers had non O157:H7 E.coli on his hands before work. Perhaps, absence of the bacteria before work may arise from hand washing preformed prior to the animal processing. High prevalence were recorded in sample taken during work from Tampin, Jasin and Kemaman (100% each) while low prevalence where observed in Shah Alam, Banting and Ipoh (20% each). Tan et al., 2013 have reported the prevalence of E.coli among food handlers to be 71.76%, 71.76% and 68.24% for hands swabs taken before, during and after work [25]. Though, lower prevalence (24%) was reported by Mayada et al., 2014[26]. Other pathogenic bacteria isolated from the swab samples are Salmonella entritidis and Citrobacterfreundii. The prevalence of the bacteria range from 20-100% during working hours while after work it was 50-100% as shown in Table 3 and Fig.3. Citrobacter freundii showed higher prevalence range (60-100%) during work among the abattoir workers. Lower prevalence was recorded (20-40%) after work compared with S.entritidis.

The total prevalence of E.coli 0157:H7 (11%) in the carcass samples was lower than that reported by Son et al., 1998 (36%) and Sahilah, 1997 (76%) from retailed meats in Malaysia. However, individual prevalence in some abattoirs tends to be higher than the reported figure. On the other hand, the total prevalence of Non-0157:H7 E.coli was higher (49%) compared to that of 0157:H7 (11%) but within the range (22.6-80%) reported by Adzitey , 2011. Other bacteria isolated from the abattoir are shown in table 5. Notable among them were Salmonella entritidis and Citrobacter freundii. They are pathogenic and therefore of medical concern. Total prevalence of Escherichia coli O157:H7 and non-O157:H7 were 10% and 81% respectively.

Low level of prevalence in the abattoir compared with retail meats can be linked with different forms of contamination that can be encountered during transportation of the beef meat. In addition, meat handlers in the market, due to inadequate knowledge in meat handling may increase the level of contamination. Inanimate objects such as knife, cutting tables and air are some of the potential sources of contamination. Most the Abattoirs visited allowed non-government workers to work there and such practice may immensely contribute to the high prevalence of E.coli in some of the abattoirs as indicated in the results.

Conclusion:

Workers harboring non-O157:H7 may serve as a potential source of contamination and infections. Presence of E.coli O157:H7 during work will be of public health concern due to cross contamination from hands of workers to the cattle carcass. The possible source of contamination of workers hands and cattle carcass is the fecal matter collected in the abattoirs according to our findings.

ARTICLE INFO

Article history:

Received 28 September 201?

Accepted 15 November 2015

Available online 24 November 2015

ACKNOWLEGEMENT

The Authors will like to acknowledge the contribution made by the following organizations; Division of Veterinary Services, Ministry of Agriculture and Agro-Based Industry, Malaysia, Faculty of Medicine and Health Sciences (Postgraduate Study Grant), Gombe State University Nigeria, Tertiary Education Fund (TETFUND) Nigeria.

Abbreviations:

EHEC-Enteroheamorrhagic Escherichia coli

WHO-World Health Organization

STEC-Shiga Toxin-producing Escherichia coli

CT-SMAC-Cefixime Tellurite Sorbitol MacConkey Agar

Competing Interest:

The authors declare that they have no competing interest

Author Contributions:

Shamsul BMT--Chairman Supervisory Committee for the research

Adamu MT--Sample collection and laboratory analysis

Desa MN--Member Supervisory Committee and Head of laboratory analysis

Khairani--Bejo S-Member Supervisory Committee. Provided the laboratory procedures

Irwan Syah MDY-Sample Collection and Interpreter

REFERENCES

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[25] Tan, S.L., H.Y. Lee, Abu F. Bakar, M.S. Abdul Karim, Y. Rukayadi and N.A. Mahyudin, 2013. Microbiological quality on food handlers' hands at primary schools in Hulu Langat District, Malaysia. International Food Research Journal. 20(d): 2973-2977.

[26] Mayada, G., H.L.G. Helmut and T. Herbert, 2014. Occurrence of Enterobacteriaceae in Raw Meat and in Human Samples from Egyptian Retail Sellers. International Scholarly Research Notices, Article ID 565671, http://dx.doi.org/10.H55/2014/565671.

(1) Adamu Muhammed T., (2) Shamsul Bahri M. Tamrin, (3) Desa Mohd N., (4) Khairani-Bejo S. and (5) Irwan Syah MDY

(1) Department of Environmental and occupational Health, Faculty of Medicine and Health Sciences, Universiti Putra Malaysia, Serdang, Selangor, Malaysia

(1) Department of Microbiology, Faculty of Science, Gombe State University, PMB 127, Gombe, Nigeria

(2) Department of Environmental and occupational Health, Faculty of Medicine and Health Sciences, Universiti Putra Malaysia, 43400, Serdang, Selangor, Malaysia.

(3) Department of Biomedical Science, Faculty of Medicine and Health Sciences, Universiti Putra Malaysia, 43400, Serdang, Selangor, Malaysia.

(4) Department of Resource Management Consumer Studies, Universiti Putra Malaysia, 43400, Serdang, Selangor, Malaysia.

Corresponding Author: Adamu Muhammed T., Department of Environmental and occupational Health, Faculty of Medicine and Health Sciences, Universiti Putra Malaysia, 43400, Serdang, Selangor, Malaysia.

Tel: +601136556174

Table 1: Prevalence of E.coli O157:H7 on Hands
of Workes Based on Location.

Location       Before work   During work   After work

Sha Alam           0%            0%            0%
Banting            0%            0%            0%
Senawang           0%            0%            0%
Kuala Pilah        0%            67%           0%
Tampin             0%            0%            0%
Jasin              0%            0%            0%
Ipoh               0%            20%           0%
Teluk Intan        0%            0%            0%
Kuantan            0%            0%            0%
Kemaman            0%            0%            0%
Dungun             0%            0%            0%

Total prevalence of E.coli O157:H7 on Hands swab- Before
work = 0% During Work = 9.7%, After work = 0%

Table 2: Prevalence of non-O157:H7 on hands of workers.

Location       Before work   During work   After work

Sha Alam           0%            20%           0%
Banting            0%            20%           0%
Senawang           0%            0%            0%
Kuala Pilah        0%            67%          33%
Tampin             0%           100%           0%
Jasin              0%           100%          100%
Ipoh               0%            20%          20%
Teluk Intan        0%            0%           20%
Kuantan            0%            0%            0%
Kemaman            0%           100%           0%
Dungun             0%            0%            0%

Total prevalence of E.coli (Non-O157:H7) on Hands swab- Before
work=0%, During Work= 35.5%, After work=13%

Table 3: Prevalence of S.entritidis on hand swab.

Location       Before work   During work   After work

Sha Alam           0%            20%          60%
Banting            0%            40%           0%
Senawang           0%            0%           60%
Kuala Pilah        0%           100%          67%
Tampin             0%           100%          100%
Jasin              0%           100%          100%
Ipoh               0%            20%           0%
Teluk Intan        0%            0%           50%
Kuantan            0%            0%           100%
Kemaman            0%           100%           0%
Dungun             0%           100%           0%

Table 4: Prevalence of C. freundii on hand swab.

Location       Before work   During work   After work

Sha Alam           0%            0%           40%
Banting            0%            60%           0%
Senawang           0%            0%           20%
Kuala Pilah        0%            67%          33%
Tampin             0%            0%            0%
Jasin              0%            0%            0%
Ipoh               0%            0%            0%
Teluk Intan        0%            0%            0%
Kuantan            0%            0%            0%
Kemaman            0%           100%           0%
Dungun             0%           100%           0%

Table 5: Prevalence of E.coli O157:H7 and Non-O157:H7
on Cattle Carcass Based On Location.

Location       O157:H7   NON-O157:H7

Sha Alam         0%          60%
Banting          0%          67%
Senawang         0%          16%
Kuala Pilah      50%         50%
Tampin           0%          0%
Alor Gajah        -           -
Jasin            0%          67%
Ipoh             25%         0%
Teluk Intan      50%         50%
Kuantan          0%          50%
Kemaman          0%          0%
Dungun           0%          67%

Total prevalence of E.coli O157:H7 and Non-O157:H7
on Carcasses- E.coli O157:H7= 11%, Non-O157:H7= 49%

Table 6: Prevalence of Salmonella enteritidis and
Citrobacter freundii in selected abattoirs.

Location     S.enteritidis    Gfreundii

Sha Alam          80%            80%
Banting           67%            33%
Senawang           0%            0%
K. Pilah          50%            50%
Tamping           50%            0%
Jasing            33%            33%
Ipoh              50%            0%
T.Intan            0%            0%
Kuantan            0%            0%
Kemaman           50%            0%
Dungun            67%            67%

Total Prevale     32%

Table 6: Prevalence of E.Coli O157:H7 and Non-O157:H7
in fecal samples Based On Location.

Location       O157:H7   NON-O157:H7

Sha Alam         0%          83%
Banting          0%          70%
Senawang         0%          80%
Kuala Pilah      60%         80%
Tampin           0%          50%
Alor Gajah        -           -
Jasin            0%          70%
Ipoh             20%         83%
Teluk Intan      40%         75%
Kuantan          0%          90%
Kemaman          0%          50%
Dungun           0%          50%

Total prevalence of E.coli O157:H7 and Non-O157:H7
in fecal samples- E.coli O157:H7= 10%, Non-O157:H7= 81%
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Author:Adamu, Muhammed T.; Shamsul, Bahri M. Tamrin; Desa, Mohd N.; Khairani-Bejo, S.; Irwan Syah, Mdy
Publication:Advances in Environmental Biology
Article Type:Report
Geographic Code:9MALA
Date:Nov 1, 2015
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