an epidemiological study of listeriosis in dairy cattle

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addition to nalidixic acid. Lovett and others (1987) included cyclohexamide in the FDA (United States Food and Drug Administration) broth to inhibit yeast and moulds. This original formulation of Lovett and others (1987) was further developed by USDA (United States Department of Agriculture) to trace Listeria in meat samples (McClain and Lee 1988). In parallel to the development of selective enrichment broths, several selective agars were developed incorporating the same antibiotics. Gray and colleagues (1950) were again the first researchers to use selective agar by including potassium tellurate to inhibit Gram negative organisms. McBride and Girard (1960) developed a selective phenyl ethanol agar containing lithium chloride, glycine and blood. This was the only commercially available selective agar for a considerable time. A major problem was its inability to inhibit enterococci. Lee and McClain (1986) later modified McBride agar by removing glycine and blood, adding moxalactam and glycine anhydride and increasing the concentration of lithium chloride. Ralovich and colleagues (1971) also used trypaflavin and nalidixic acid in a serum based agar. The discovery that Listeria organisms were resistant to higher concentrations of lithium chloride led to development and modification of many agars and overcame the problem of enterococcal growth. Most of the agars currently used are a modification of McBride (McBride and Girard 1960) and the Oxford formulation (Curtis and others 1989a). In addition to antibiotics, one or more indicators for Listeria were also included in the medium such as aesculin in Oxford agar (Curtis and others 1989), blood in enhanced haemolysis agar (EHA, Cox and others 1991) and Fenlon Listeria agar (Fenlon 1985), mannitol and phenol red in PALCAM agar (van Netten and others 1989) and rhamnose in Modified Despeirres agar (Golden and other 1988). Although the selectivity of media was increased by incorporating antibiotics in the culture medium it was noticed that highly selective media could also inhibit the 131
growth of some strains of Listeria (Seeliger and Jones 1986, Curtis and other 1989b). To obtain maximum success in isolation of Listeria a three stage procedure was recommended (Curtis and Lee 1995); (a) resuscitation or pre-enrichment (selective or non-selective) (b) selective enrichment and (c) selective plating. This procedure has been used to isolate Listeria from food (Lewis and Cory 1991, McLain and Lee, USDA method, 1988), sewage (Watkins and Sleath 1981), clinical samples (Eld and others 1993) and environmental samples (Fenlon 1985, Husu 1990). The difficulty in isolating L. monocytogenes means that bacteriological methods are not always satisfactory when attempting to determine the exposure of animals to Listeria organisms (Gray and Killinger 1966) and serological techniques have been used as supplementary tools for this purpose. Specific ELISA tests, employing cell products involved in pathogenesis such as listeriolysin O, have been used in experimental studies. No large scale use of these ELISA assays, involving field studies has, yet been conducted (Berche and others 1990, Low and Donachie 1991, Low and others 1992b, Gholizadeh and others 1996). A pilot study involving a dairy herd was carried out to develop and standardise bacteriological and serological techniques which would be used in a longitudinal survey of L. monocytogenes infection. 5. 2. Materials and methods 132
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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
Page 97 and 98: a) Type of housing : Housing animal
Page 99 and 100: Herd sizes n Y N OR P value x Y N O
Page 101 and 102: of disease. Similarly storing feed
Page 103 and 104: a) Type of forages: Feeding maize s
Page 105 and 106: ) Use of bedding: Using straw beddi
Page 107 and 108: ) Source of forages: Home made gras
Page 109 and 110: 1.29-18.22) whereas straw bedding i
Page 111 and 112: 100 2 34 2.33 0.01 X 2 22 3.35 0.01
Page 113 and 114: a) Dung disposal: Storage of manure
Page 115 and 116: vaccination against Leptospirosis +
Page 117 and 118: suppression of the growth of Lister
Page 119 and 120: 1989, Sargison 1993). It is also po
Page 121 and 122: L. monocytogenes has been isolated
Page 123 and 124: association (Kirkwood 1988, Thrusfi
Page 125 and 126: The predictor variables that met ou
Page 127 and 128: Forage feeding Maize silage Hay Str
Page 129 and 130: indoor feeding fed ad libitum on th
Page 131 and 132: cases of Listeriosis in sheep 2.9 1
Page 133 and 134: and controlling moles in fields wer
Page 135 and 136: Cases of Listeriosis in beef cattle
Page 137 and 138: This model was similar to the Model
Page 139 and 140: Table 4. 9. The multivariate relati
Page 141 and 142: 5) Nervous signs: Cases of Listerio
Page 143 and 144: feeders, maize silage feeding in ri
Page 145 and 146: A contradictory finding was made ab
Page 147: selective enrichment broth, at refr
Page 151 and 152: 5. 2. 2. Bacteriology : a) Culture
Page 153 and 154: culturing negative LSEB (1/10w/v) s
Page 155 and 156: 6) Sugar tests: Listeria are capabl
Page 157 and 158: uffered saline (PBS), pH 7.2, centr
Page 159 and 160: Investigation of non-specific bindi
Page 161 and 162: and 0.94 for L. monocytogenes (17/1
Page 163 and 164: Detection limit of the method: A si
Page 165 and 166: Relationship between housing, silag
Page 167 and 168: 5 13 3 3 1 3 1 1 1 6 14 1 4 5 3 1 0
Page 169 and 170: Table 5. 11. The relationship betwe
Page 171 and 172: frequency of isolation of L. monocy
Page 173 and 174: CHAPTER 6 A study of the dynamic of
Page 175 and 176: 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
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) Environmental samples: Both L. in
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monocytogenes. By April the proport
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A total of 211 milking cows were ex
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100 % 90 80 70 60 50 40 30 20 10 0
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The number of new cases each month
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monocytogenes was also isolated fro
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Table 6. 18 The isolates, their ori
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Table 6. 19. The distribution of th
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On farm C pattern 1 was detected in
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Figure 6. 8. The repeatability of R
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Figure 6. 10. The distribution of s
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Figure 6. 12a. The RAPD pattern obt
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1988, Husu 1990) and from zero to a
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experimentally been shown that ther
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periods. However, on farms B and E
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A large number of L. monocytogenes
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Figure 6. 13. Animal-environment cy
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Some important farming practices we
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chance finding of L. monocytogenes
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Asahi, O., Hosoda, T. and Akuyama,
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Bourry, A. and Poutrel, B. (1996) B
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Chakraborty, T., Ebel, F., Wehland,
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delGarso, L. and Wallop, W. (1975)
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Eveland, W.C. (1963) Demonstration
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Fraser, J.A. and Sperber, W.H. (198
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Golden, D.A, Beuchat, L.R. and Brac
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humans. In Miller, A.J.,. Smith, J.
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Jones, F.S. and Little, R.B. (1934)
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Listeria innocua by oral inoculatio
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Low, J.C. and Donachie, W.A. (1997)
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McBride, M.E. and Girard, K.F. (196
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Nieman, R.E. and Lorber, B. (1980)
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Peters, M., Pohlenz, J., Jaton, K.,
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Bakteriologie, Mikrobiologie und Hy
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Seeliger, H.P.R. (1981) Apathogenic
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Tilney, L.G. and Portnoy, D.A. (198
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Walker, J.K., Morgan, J.H., McLauch
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APPENDIX 3 PREPARATION OF BACTERIOL
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Cool to 50 0 C Mix Solution A with
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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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