an epidemiological study of listeriosis in dairy cattle

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etween increased anti-LLO antibodies and faecal excretion has also been suggested in an experimental infection of L. monocytogenes (Mietinnen and others 1991). The sources of infection on these farms were investigated using RAPD test. One criticism of RAPD is its lack of reproducibility (Wernars and others 1996). The RAPD technique used in this study had been validated in another laboratory (O’Donoghue and others 1995). The use of 6 primers allowed internal validation and the experiments showed that the technique was repeatable. Similar results were reported by other researchers (Mazurier and Wernars 1992, MacGowan and others 1993, O’Donoghue and others 1995). Alternative typing methods such as serotyping, ribotyping, restriction enzyme analysis (REA) multilocus enzyme electrophoresis (MEE) and pulsed field gel electrophoresis (PFGE), have been used for L. monocytogenes (Ralovich 1993) and a comparison with RAPD has also been made (Boerlin and others 1995, Louie and others 1996). RAPD was compared with serotyping, ribotyping, MEE and REA by Boerlin and others (1995). They acknowledged the superiority of RAPD. RAPD was also compared with ribotyping and PFGE. RAPD and PFGE were found to be the most discriminating typing methods and their ability to differentiate between strains was comparable (Louie and others 1996). However PFGE may take several days, it is costly and sophisticated laboratory equipment is required to perform it. RAPD, on the other hand, is cheap, rapid (results are obtained within several hours) and easy to perform (O’Donoghue and others 1995, Louie and others 1996). The wide diversity of L. monocytogenes found on five study farms makes it difficult to link environmental strains with faecal strains. Because of the lack of understanding of the population dynamics of strains of L. monocytogenes it is also not known if temporal distribution of strains occurs (Low and Donachie 1997). This was also the case in this study. 212
A large number of L. monocytogenes isolates were obtained from the environment and animals. It was impossible to examine all the isolates because of time and financial constraints. However the limited results obtained in this study provide some interesting information. A total of 12 “strains” were identified but on individual farms this was limited to 5 or 6 different “strains”. The maximum number in environmental samples was 4 and in faecal samples it was 6. This diversity within the strains of L. monocytogenes is in accordance with the results reported by other researchers (Boerlin and Piffaretti 1991, Fenlon and others 1996, Wiedmann and others 1996). In interpreting the difference between the isolates made at different times care has to be taken because of the potential influence of the sampling procedure. Competition between the strains invariably takes place during the enrichment procedure and the selection of only 5 colonies from a plate means that only “strains” present at a prevalence of 50% will be detected. However, given these constraints the results suggest that the predominant faecal isolates may vary with time. It is possible that the epidemic curves observed for L. monocytogenes are composite results from a large number of “strains” related epidemic curves. If this is the case then any potential vaccine against L. monocytogenes infection would need to incorporate immunogens from a number of different “strains”. The isolates from milk in the bulk tank on three farms (farm A, B and D) revealed that each farm had different patterns; the pattern 4 on farm A, 1 on farm B and 5 on farm D. Except for the pattern 4, this pattern 4 was not detected in any other samples. Both pattern 1 and 5 were also detected in the environmental and faecal isolates on these farms. This may suggest that milk was contaminated with the organism either by animals excreting it in milk or the organism got into the tank via environmental contamination where the refrigeration temperature allowed the organism 213
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AN EPIDEMIOLOGICAL STUDY OF LISTERI
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In the second part of the study a l
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I would like to thank to Metin Oztu
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DECLARATION I declare that apart fr
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2. 2. Materials and Methods 47 2. 2
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4. 4. Discussion 125 CHAPTER 5 A pi
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CHAPTER 1 LIST OF TABLES AND FIGURE
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Table 3. 11. The relationship betwe
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CHAPTER 5 Table 5. 1. Date of visit
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Figure 6. 4. The monthly faecal exc
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Listeralla hepatolytica. In the sam
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Listeriolysin O (Fernandaz-Garayzab
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L. monocytogenes was the only recog
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Table 1. 2. Serovar distribution of
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Serotyping is based on the identifi
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electrophoresis. Different strains
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and others (1995) came to the concl
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animals suggests that L. monocytoge
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Immunity reducing Septicaemia facto
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monocytogenes and a prerequisite to
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a variable number of cases of encep
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The relationship between age and Li
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a reservoir from which it spreads t
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1. 8. Clinical signs and pathology
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is seldom clinical illness in the d
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Bacterial culture is widely used fo
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Differential diagnosis: Listerial m
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Since elimination of L. monocytogen
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first description, it was only 1980
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and meat products have resulted in
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infectious dose for the infection i
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a longitudinal study involving five
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in Britain. In this part of the stu
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groups were excluded from analysis.
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The proportions of farms with cases
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Table 2. 3. The proportion of anima
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percentage 50 45 40 35 30 25 20 15
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Milking cows (%) Rep. heifers (%) D
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Abortion 274 (57.8%) 28 (21.9%) Ner
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with others (Kanuk and Berenson 197
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occurred in replacement heifers and
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CHAPTER 3 The relationship between
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(ii) Cases in milking cows: Farms r
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To assess the relationship between
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) Duration of housing: If animals w
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d) Presence of moles: Farmers were
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listeriosis. Maize silage feeding a
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found between the duration of feedi
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a) Type of housing : Housing animal
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Herd sizes n Y N OR P value x Y N O
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of disease. Similarly storing feed
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a) Type of forages: Feeding maize s
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) Use of bedding: Using straw beddi
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) Source of forages: Home made gras
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1.29-18.22) whereas straw bedding i
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100 2 34 2.33 0.01 X 2 22 3.35 0.01
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a) Dung disposal: Storage of manure
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vaccination against Leptospirosis +
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suppression of the growth of Lister
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1989, Sargison 1993). It is also po
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L. monocytogenes has been isolated
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association (Kirkwood 1988, Thrusfi
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The predictor variables that met ou
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Forage feeding Maize silage Hay Str
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indoor feeding fed ad libitum on th
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cases of Listeriosis in sheep 2.9 1
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and controlling moles in fields wer
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Cases of Listeriosis in beef cattle
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This model was similar to the Model
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Table 4. 9. The multivariate relati
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5) Nervous signs: Cases of Listerio
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feeders, maize silage feeding in ri
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A contradictory finding was made ab
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selective enrichment broth, at refr
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growth of some strains of Listeria
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5. 2. 2. Bacteriology : a) Culture
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culturing negative LSEB (1/10w/v) s
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6) Sugar tests: Listeria are capabl
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uffered saline (PBS), pH 7.2, centr
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Investigation of non-specific bindi
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and 0.94 for L. monocytogenes (17/1
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Detection limit of the method: A si
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Relationship between housing, silag
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5 13 3 3 1 3 1 1 1 6 14 1 4 5 3 1 0
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Table 5. 11. The relationship betwe
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frequency of isolation of L. monocy
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CHAPTER 6 A study of the dynamic of
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There were no sample size calculati
Page 177 and 178: 4 of the farms had their silage ana
Page 179 and 180: a) Faecal samples: Faecal samples w
Page 181 and 182: • • • 1 5 9 6. 2. 5. Sample p
Page 183 and 184: A total of 944 isolates of L. monoc
Page 185 and 186: UK), 0.5μM DNA primer, 1 unit Supe
Page 187 and 188: entered and left the herd over the
Page 189 and 190: Table 6. 7. Monthly frequency of ex
Page 191 and 192: Table 6. 8. The relationship betwee
Page 193 and 194: (MA) mean age, (n) number, (R) rang
Page 195 and 196: frequency of excretion (P
Page 197 and 198: There was no evidence from this far
Page 199 and 200: ) Environmental samples: Both L. in
Page 201 and 202: monocytogenes. By April the proport
Page 203 and 204: A total of 211 milking cows were ex
Page 205 and 206: 100 % 90 80 70 60 50 40 30 20 10 0
Page 207 and 208: The number of new cases each month
Page 209 and 210: monocytogenes was also isolated fro
Page 211 and 212: Table 6. 18 The isolates, their ori
Page 213 and 214: Table 6. 19. The distribution of th
Page 215 and 216: On farm C pattern 1 was detected in
Page 217 and 218: Figure 6. 8. The repeatability of R
Page 219 and 220: Figure 6. 10. The distribution of s
Page 221 and 222: Figure 6. 12a. The RAPD pattern obt
Page 223 and 224: 1988, Husu 1990) and from zero to a
Page 225 and 226: experimentally been shown that ther
Page 227: periods. However, on farms B and E
Page 231 and 232: Figure 6. 13. Animal-environment cy
Page 233 and 234: Some important farming practices we
Page 235 and 236: chance finding of L. monocytogenes
Page 237 and 238: Asahi, O., Hosoda, T. and Akuyama,
Page 239 and 240: Bourry, A. and Poutrel, B. (1996) B
Page 241 and 242: Chakraborty, T., Ebel, F., Wehland,
Page 243 and 244: delGarso, L. and Wallop, W. (1975)
Page 245 and 246: Eveland, W.C. (1963) Demonstration
Page 247 and 248: Fraser, J.A. and Sperber, W.H. (198
Page 249 and 250: Golden, D.A, Beuchat, L.R. and Brac
Page 251 and 252: humans. In Miller, A.J.,. Smith, J.
Page 253 and 254: Jones, F.S. and Little, R.B. (1934)
Page 255 and 256: Listeria innocua by oral inoculatio
Page 257 and 258: Low, J.C. and Donachie, W.A. (1997)
Page 259 and 260: McBride, M.E. and Girard, K.F. (196
Page 261 and 262: Nieman, R.E. and Lorber, B. (1980)
Page 263 and 264: Peters, M., Pohlenz, J., Jaton, K.,
Page 265 and 266: Bakteriologie, Mikrobiologie und Hy
Page 267 and 268: Seeliger, H.P.R. (1981) Apathogenic
Page 269 and 270: Tilney, L.G. and Portnoy, D.A. (198
Page 271 and 272: Walker, J.K., Morgan, J.H., McLauch
Page 273 and 274: APPENDIX 3 PREPARATION OF BACTERIOL
Page 275 and 276: Cool to 50 0 C Mix Solution A with
Page 277 and 278: Phosphate buffered saline (PBS; 10x
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APPENDIX 2 THE OVERALL RESULTS OF U
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Grass silage Y N OR (95% CL) p Valu
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storage Y N OR (95% CL) p Value cla
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source home made 38 182 0.6 (0.1-16
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source home made 34 345 0.8 (0.3-2.
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source barley 27 251 0.93(0.6-1.6)
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source potatoes 2 11 1.4 (0.-7.7) 0
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in a covered barn 73 595 0.8 (0.4-1
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sawdust 0 16 0.0 (0.0-2.73) 0.3 str
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APPENDIX 5 THE QUESTIONNAIRE AND TH
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Abbreviations: 1 first blood sampli
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B 113 1 114 1 115 1 116 1 117 1 118
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B 104 1 104 2 104 3 105 1 105 2 105
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B 20 1 20 2 20 3 21 1 21 2 21 3 19
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0.169 1.157 1.134 1.467 1.356 0.145
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Plate 4 Layout and ODs B 73 1 73 2
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UNIVERSITY OF BRISTOL LISTERIOSIS I
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PART C : HERD SIZE :--BETWEEN JULY
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(please circle one for each) 3. Wha
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etween which months were your milki
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not stored beneath the slats compos
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STUDY OF LISTERIOSIS IN DAIRY CATTL
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STUDY OF LISTERIOSIS IN DAIRY CATTL
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Now I would like to get some detail
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Root crops : ______________________
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In this section I will be asking qu
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33. Which days of the week bulk tan
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-what was used to treat? -what was
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an epidemiological study of listeriosis in dairy cattle

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