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Bacteriological Quality and Occurrence of Salmonella Species and Escherichia Coli o157:h7 in Roasted Rat (Arvicanthis niloticus) Meat

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Bacteriological Quality and Occurrence of Salmonella Species and Escherichia Coli o157:h7 in Roasted Rat (Arvicanthis niloticus) Meat.

ABSTRACT

Bacterial contamination of roasted ready-to-eat rat meat and occurrence of foodborne pathogens may be injurious to human health.

In Nigeria, the incidence and multidrug resistance of Salmonella spp. and E. coli O157:H7 in human cases is on the increase and these organisms have been found associated with meat and meat products in several studies.

A cross-sectional study was carried out to determine the bacteriological quality and occurrence of Salmonella spp. and Escherichia coli O157:H7 in roasted ready-to-eat rat (Arvicanthis niloticus) meat sold in Zaria, Nigeria.

A total of 384 samples were examined from four purposively selected districts in the study area; Samaru, Basawa, Jushi and Sabon Gari. Total aerobic plate count was determined using spread plate technique and bacteria load determined.

Isolates were screened for Escherichia coli O157:H7 and Salmonella using the conventional biochemical characterization, standardized micro-substrate (Microgen GN-ID A+B) detection kit for Gram negative bacteria and further confirmed using Rapid latex agglutination test for E. coli O157:H7 and PCR for Salmonella.

Agar disc diffusion method was used to evaluate the antibacterial resistance pattern; data obtained were interpreted using clinical laboratory standards institutes (CLSI).

Mean log and standard deviation of total aerobic plate counts (TAPC) were 10.01±0.27, 10.03±0.26, 10.10±0.29 and 9.9±0.29, from four sampling areas, Samaru, Sabon Gari, Jushi and Basawa.

The average TAPC ranged from 12 x 109 cfu.g-1 in Samaru to 15 x 109 cfu.g-1 in Jushi sampling districts. Salmonella was isolated from 2(0.5%) and E. coli O157:H7 isolated from 5 (1.3%) of the total 384 samples.

TABLE OF CONTENTS

Declaration———————————————————————————————————— i

Certification———————————————————————————————————- ii

Dedication———————————————————————————————————– iii

Acknowledgement————————————————————————————————- iv

Abstract————————————————————————————————————– vi

Table of contents————————————————————————————————- viii

List of Tables—————————————————————————————————— xiii

List of Figures—————————————————————————————————— xv

List of Plates——————————————————————————————————- xvi

List of Appendices———————————————————————————————- xvii

Abbreviations—————————————————————————————————- xviii

CHAPTER ONE: INTRODUCTION

1.1             Background information———————————————————————————- 1

1.2             Statement of Research Problem————————————————————————- 3

1.3             Justification————————————————————————————————– 4

1.4             Aims———————————————————————————————————- 5

1.5             Objectives————————————————————————————————— 6

1.6             Research questions—————————————————————————————- 6

CHAPTER TWO: LITERATURE REVIEW

  • Microbiology of meat————————————————————————————- 7
  • Bacterial contamination of meat———————————————————————— 7
  • Preparation of roasted rat meat————————————————————————– 9
  • Ready-to-eat foods—————————————————————————————— 9
  • The genus Salmonella———————————————————————————— 10
    • Historical perspective————————————————————————————- 10
    • Nomenclature———————————————————————————————– 11
    • Serovars—————————————————————————————————— 11
    • Morphology of Salmonella—————————————————————————— 12
    • Distribution and host range—————————————————————————— 13
    • Salmonella infection in humans————————————————————————- 14
    • Virulence factors of Salmonella————————————————————————- 15
      • Salmonella invasion (inv) genes———————————————————————- 15

2.5.7.2. Salmonella in meat and meat products in Nigeria———————————————– 15

  • Antibiotic resistance among Salmonella————————————————————- 16
    • Antibiotic resistant Salmonella in Nigeria———————————————————- 17
  • Detection of Salmonella———————————————————————————- 17
    • Culture based method——————————————————————————— 18

2.5.9.2 Control and prevention of Salmonella————————————————————– 22

  • History of Escherichia coli——————————————————————————– 22
    • Patho-types of Escherichia coli———————————————————————— 23
      • Enteroinvasive Escherichia coli——————————————————————– 23
      • Entero-haemorrhagic Escherichia coli————————————————————– 24
      • Enteropathogenic coli (EPEC)——————————————————————– 25
      • Enterotoxigenic coli (ETEC)———————————————————————- 26
      • Entero-aggregative coli—————————————————————————– 26
      • Diffusely adhering coli—————————————————————————– 27
    • Transmission of coli O157:H7———————————————————————- 27
    • Reservoir of coli O157:H7————————————————————————— 28
    • Clinical manifestation of EHEC O157: H7———————————————————- 29
    • Pathogenicity of Escherichia coli EHEC————————————————————- 30
    • Diagnosis of EHEC O157: H7 infection————————————————————- 30
    • Prevention and control of EHEC infection———————————————————– 31
    • Treatment of EHEC O157: H7————————————————————————- 32
    • Prognosis of EHEC O157: H7———————————————————————— 32
    • Antibiotic resistance of coli O157:H7———————————————————– 32
  • Arvicanthis niloticus (Nile grass rat)—————————————————————– 33
  • Polymerase chain reaction—————————————————————————– 34

CHAPTER THREE: MATERIALS AND METHODS————————————————— 35

  • Study Area————————————————————————————————– 35
  • Sample Size————————————————————————————————- 37
  • Study Design———————————————————————————————– 37
  • Questionnaire administration————————————————————————— 38
  • Sampling—————————————————————————————————- 38
  • Zoological Nomenclature——————————————————————————– 38
  • Laboratory Procedures———————————————————————————- 39
    • Non-Selective pre-enrichment————————————————————————- 42
    • Total aerobic plate count——————————————————————————- 42
      • Expression of total aerobic plate counts———————————————————— 42
    • Selective isolation of Salmonella——————————————————————— 43
      • Sub-culturing on nutrient agar plate—————————————————————– 43
    • Selective isolation of coli————————————————————————— 43
    • Conventional biochemical characterization of presumptive isolates————————– 43
    • Interpretation of conventional biochemical tests————————————————– 44

3.7.8   Isolation of E. coli O157:H7————————————————————————— 45

  • Bacterial characterization of presumptive isolates using the standardized micro- substrate detection kit for gram negative bacteria————————- 45
  • Rapid Latex Agglutination Test for Escherichia coli O157:H7——————————— 46
    • Test procedure for O157 (somatic antigen)———————————————————– 47
    • Test procedure for H7 (flagella antigen)————————————————————– 47
  • In vitro antibiotic susceptibility testing of isolates————————————————- 46
  • Detection of invA gene by PCR (Polymerase Chain Reaction) method Detection———- 48
    • DNA extraction of isolated Salmonella species—————————————————- 48
    • DNA extraction——————————————————————————————- 49
    • Primer set and PCR amplification——————————————————————— 49
    • Electrophoresis of PCR product———————————————————————- 50
  • Data Analyses——————————————————————————————— 50

CHAPTER FOUR: RESULTS——————————————————————————— 51

  • Zoological nomenclature——————————————————————————– 51
  • Total aerobic plate count——————————————————————————- 51
  • Frequency of isolation of Salmonella and coli O157:H7————————————- 51
    • Frequency of isolation of Salmonella—————————————————————– 51
    • Frequency of isolation of coli O157:H7——————————————————— 55

4.4. Biochemical Identification of isolates using the Standardized micro-substrate (Microgen GN-ID A+B) detection kit for gram negative bacteria————– 58

4.5     Results after further confirmatory tests using commercial Latex Agglutination Kits—61 4.6

Polymerase Chain Reaction (PCR)————————————————————-63

4.6.1 Virulence Marker of Salmonella Isolates of Polymerase Chain Reaction (PCR)———– 63

  • Antibacterial resistance———————————————————————————- 64
    • In vitro Susceptibilty of Salmonella Isolates to 13 Antimicrobial Agents——————— 64
    • In vitro Susceptibilty of coli O157:H7 Isolates to Antimicrobial Agents—————- 67
  • Demographic features of retailers and consumers of roasted ready-to-eat rat meat in Zaria 71
  • Practices of those that consume roasted ready-to-eat rat meat as regards consumption of roasted ready-to-eat rat meat in Zaria——————————— 71
  • Practices of retailers of roasted ready-to-eat rat meat as regards consumption of roasted ready-to-eat rat meat in Zaria———————– 71

CHAPTER FIVE: DISCUSSION—————————————————————————– 77

CHAPTER SIX: CONCLUSION AND RECOMMENDATIONS———————————— 88

6.1          Conclusion————————————————————————————————– 88

6.2          Recommendations—————————————————————————————- 89

INTRODUCTION

1.1 Background information

Meat refers to mostly skeletal muscles and associated fat but it may also refer to organs, including lungs, livers, skin, brains, bone marrow, kidneys, and a variety of other internal organs as well as blood. It is animal tissue used as food (Hammer, 1987).

The term “bush meat” stands for meat that has been sourced from wild animals for human consumption and could be consumed fresh, smoked, salted, or sun-dried (Gideon and Joseph, 2018).

Bush meat is central to the livelihood of many poor rural dwellers that consume and trade in it and the high demand in the urban regions means that the bush meat trade has become a lucrative business (ACET 2014; Gideon and Joseph, 2018).

Nearly 5 million tons of bush meats are being traded in Central and West Africa and in Nigeria about 25 percent of the population relies solely on resources of animal protein from bush meat (Wilkie and Carpenter, 1999; Fa et al., 2002; Gideon and Joseph, 2018).

Rodents are accepted as a popular source of protein and more than 71 genera and 89 species of hammerrodents, (mostly Hystricomorphs) have been consumed by man in the tropical world (Oyarekua and Ketiku, 2010).

REFERENCES

Aarestrup, F.M. (1995). Occurrence of Glycopeptides Resistance among Enterococcus faecium Isolates from Conventional and Ecological Poultry Farms. Journal of Microbial Drug Resistance. 1:255-257.
Abd El-Atty, N.S. and Meshref, A.M. (2007). Prevalence of Salmonella and E. coli O57 in some foods. BS. Veterinary Medicine Journal. 5th Scientific Conference, p. 73-78
Abolagba, O.J. and Iyeru, O.A., (1998). Study of Insect Pest infesting traditionally processed fish sold in Benin Metropolis. Nigeria. Nigeria Journal of Applied Science, Pp.25-29
Adetunji, V.O. and Isola, T.O. (2011). Enumeration of Listeria and enteric bacteria of public health significance on meat tables before and after sales of meat in Ibadan municipal abattoir, Nigeria. Pakistan Journal of Nutrition 10:224-228
Adriana, G., Tania, l.T., Valeria, R.O., Carla,R.T. and Marina, B.M. (2004). Molecular characterization of enteroinvasive Escherichia coli IPA genes by PCR-RFLP analysis. Brazilian journal of microbiology. 35:74-80
Agada, G.O.A., Abdullahi, I.O., Aminu, M., Odugbo, M., Chollom, S.C., Kumbish, P.R. and A. Okwori, A.E.J. (2014). Prevalence and Antibiotic Resistance Profile of Salmonella Isolates from Commercial Poultry and Poultry Farm-handlers in Jos, Plateau State, Nigeria. British Microbiology Research Journal. 4(4): 462-479

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