Can pooled pen floor samples be used for diagnosing Escherichia coli F18 positive diarrhoeic nursery pigs?

Transcription

1 Can pooled pen floor samples be used for diagnosing Escherichia coli F18 positive diarrhoeic nursery pigs? Slide 1 Nicolai Weber DVM, Ph.D STUDENT, Resident ECPHM Department of Large Animal Sciences, University of Copenhagen

2 Ph.D project: Intestinal Health Development of diagnostic decision tool for treatment of diarrhoea in grower pigs Supervisors: Jens Peter Nielsen (Professor, DVM, PhD, Dipl. ECPHM) Christian Fink Hansen (Professor, MSc Animal Science, PhD) Ken Steen Pedersen (Affiliated Associate Professor, CEO Ø-Vet A/S, DVM, PhD, Dipl. ECPHM) Slide 2

3 Background Slide 3

4 Slide 4 Background Quantitive PCR E. coli F4 E. coli F18 Lawsonia intracellularis Brachyspira pilosicoli Solid verification of L. intracellularis Association between faecal load of lawsonia intracellularis and pathological findings of proliferative enteropathy in pigs with diarrhoea Pedersen et. al 2012 BMC VETERINARY RESEARCH Pooling of porcine fecal samples for quantification of Lawsonia intracellularis by realtime polymerase chain reaction Pedersen et. al 2014 JOURNAL OF VETERINARY DIAGNOSTIC INVESTIGATION

5 Background % pens 100% 80% 60% 40% 20% 0% Distribution of pathogens detected Day 14 Day 21 Day 28 Day 35 Days after weaning B. pilosicoli L. intracellularis E. coli F18 E. coli F4 Detection of pathogens in pen floor by qpcr in non-medicated pigs from three Danish herds Slide 5

6 Background Slide 6

7 Study design Pig samples Pen samples 3 nursery facilities In pens with >25 % diarrhoea 3 diarrheic pigs per pen 1 floorsample per pen Slide 7

8 Study design Laboratory work: Pig samples: Culture + PCR (F4, F5, F6, F18, F41, STa, STb, LT, VT2e) Pen floor samples: Culture + PCR + qpcr (F4, F18, LI, PILO) Slide 8

9 Study design culture Department of Large Animal Sciences Golden standard culture qpcr Slide 9

10 Classification of E. coli Definition: Virulent E.coli = fibriae positive and toxin positive Slide 10

11 Results Sampling size 86 pigs housed in 31 pens 380 E. coli isolates Slide 11

12 Results Culture + RT-PCR: 89 virulent E. coli all F18 positive 22/86 sampled pigs in 12/31 pens 13/31 sampled pens Slide 12

13 Results qpcr: E.coli F18 gens detected in 16/31 sampled pens Mean excretion of positive samples = CFU/g faeces Slide 13 Detectionlimit = 1.5 x 10 3 CFU/g faeces

14 Results CULTURE-PEN vs CULTURE-PIG Detection of virulent E. coli in pen floor samples Detection of virulent E. coli in pig samples + - Total Total Slide 14

15 Results qpcr-pen vs CULTURE-PIG Detection of E. coli gens in pen floor samples Detection of virulent E. coli in pig samples + - Total Total Slide 15

16 Results Method Se CL95% Sp CL95% Culture qpcr Slide 16

17 Results Method Se CL95% Sp CL95% Culture qpcr Slide 17

18 Results 3 qpcr-pen 2 vs CULTURE-PIG 1 Detection of E. coli gens in 0 pen floor samples Pens with disagreement Department of Large Animal Sciences Detection of virulent E. coli in pig samples + - Total Total Slide 18 qpcr results (CFU pr. gram faeces)

19 Conclusion 10 of 31 pens virulent E. coli isolated in pen and pig Similar sensitivity when using culture of qpcr from pen floor samples Low sample size needs verifications in large scale study Slide 19

20 Can pooled pen floor samples be used for diagnosing Escherichia coli F18 positive diarrhoeic nursery pigs? Yes! Slide 20

21 Resistance profiling from pen floor samples? Table2 Antimicrobial class Agreementof resistance classification Pens with agreement Sulphamethoxazole 10/10 Trimethoprim 10/10 Ampicillin 10/10 Tetracycline 10/10 Streptomycin 10/10 Spectinomycin 8/10 Slide 21

22 Acknowledgments Centre for Herd-oriented, Education, Research and Development SEGES-VSP Elisabeth Okholm Nielsen Svend Haugegaard DTU-VET Charlotte Kristiane Hjulsager Student helpers Christian Møller, Martin Rasmussen, Thomas Kusk Funding for this project was provided by Danish Pig Levy Fund Slide 22