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Comparison of pathogenic Salmonella spp. detection from porcine oral fluids, over-the-shoe booties, and fecal samples

The goal of this project was to compare the accuracy of oral-fluids and over-the-shoe booties to the current gold standard of fecal sampling for identification of Salmonella in pigs.

Monday 4 June 2018 (1 years 6 months 1 days ago)
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Salmonellosis in pigs, which is typically seen during the weaning or finishing phase, may result in clinical disease or in establishment of a carrier state. In a carrier state, Salmonella is shed intermittently in the feces, creating a public health concern as a result of potential pork contamination. Accurate and timely detection of Salmonella is vital to the initiation of appropriate treatment protocols and prevention strategies. Veterinarians commonly question which sample type is best for herd level surveillance of Salmonella in swine herds. Oral fluids, currently widely used for the detection of PRRSV and SIV, are quick and inexpensive to collect. Over-the- shoe booties, while easy to collect, are not currently used as frequently for surveillance of pathogens in the swine industry. Fecal samples are the most commonly used sample for detection of many viruses and bacteria in swine but are more time consuming to collect than the previous two sample types. The goal of this project was to compare the accuracy of oral-fluids and over-the-shoe booties to the current gold standard of fecal sampling for identification of Salmonella using pigs experimentally inoculated with various serotypes of Salmonella.

Five-week old pigs were experimentally infected with 109 CFU of Salmonella via oral gavage and tonsil swab using isolates obtained from clinical samples submitted to the Iowa State University Veterinary Diagnostic Laboratory. Twenty pigs were infected with Salmonella Typhimurium and 20 with Salmonella I 4,[5],12:i:- (2 separate groups with 5 pens, 4 pigs in each pen). Rectal temperatures were taken on all pigs daily until 7 days post-inoculation (PI) then biweekly for the remaining 3 weeks. Samples were collected on days 2, 4, 7, 14, 21 and 28 post-inoculation. Over-the-shoe booties were collected during pen walk-through at each time point, with one pair of booties worn through all pens per infecting serotype. Oral fluids were collected by hanging a rope in each pen of 4 pigs for 30-60 minutes. Fecal samples were collected via digital rectal palpation of each pig. Samples were stored at -80°C from the time of collection until quantification of Salmonella was performed. Salmonella quantification was completed using the standard plate count method on Xylose-Lysine-Tergitol-4 (XLT4) agar. All samples were also enriched in Buffered Peptone Water for 18-24 hours and were then plated to Brilliant Green (BG) agar and XLT4 agar. A subset of colonies for each sample characteristic of Salmonella, black on XLT4 agar and red on BG agar, were confirmed as Salmonella using MAL- DI-TOF-MS.

The median temperatures of pigs peaked on day two post-inoculation at 39.3ºC and returned to near baseline levels by day five PI at 38.5 ºC. Average Salmonella quantities in the feces peaked at day two PI at 8.04 × 104 CFU/mL. Salmonella was still detectable in the feces from enrichment at 28 days PI. From oral fluids, average Salmonella quantities peaked at day 3 PI at 5.6 × 102 CFU/ mL. Salmonella was no longer detectable from oral fluids past day seven PI. Over-the-shoe booties were only sporadically positive throughout the testing period, with the highest average level of Salmonella detected being only 100 CFU/mL at day 4 PI. Salmonella was only detectable from the booties through day four PI. Selection of the most appropriate sample type is important for proper surveillance of pathogens. Over-the-shoe booties proved to be an unacceptable sample type to detect Salmonella given that booties positive for Salmonella were not reflective of the level of disease nor were they consistently positive. Oral fluids would be an acceptable sample type for detection but only during the acute phase of a Salmonella outbreak when Salmonella levels are the highest. Enrichment of those oral fluid samples would be recommended to ensure there is adequate Salmonella for detection through culture.

Although feces is the most time-consuming to collect, it should remain the gold standard for Salmonella detection as the fecal samples were most reflective of the level of disease as indicated by the rectal temperatures in addition to having the highest levels of Salmonella for the longest amount of time post-infection.

S. Naberhaus, A. Krull, B. Arruda, P. Arruda, D. Magstadt, F. Matias Ferreyra, Honorato Gatto, H. Meiroz de Souza Almeida, A. Kreuder. Comparison of pathogenic Salmonella spp. detection from porcine oral fluids, over-the-shoe booties, and fecal samples. 49 th Annual Meeting of the American Association of Swine Veterinarians (March 3-6, 2018)

Article Comments

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05-Jun-2018Peter vd WolfPeter vd WolfUnfortunately I do not agree with the authors of this article. Our experience is that sock samples are a very sensitive method to establish whether or not pigs are shedding Salmonella in their feaces. If wanted / necessary this can be combined with dust samples, which I would recommend when sampling hallways/corridors. We published our results on the ESPHM in 2017 in Prague, of which I copy the text here below.
P. van der Wolf1, K. Koenders1, V. Gotter2
1 IDT-Biologika, Benelux, Breda, Netherlands
2 IDT-Biologika, International, Dessau, Germany
Salmonella on farm can be detected by different types of samples: sock, swab and fecal. Here we describe the results of 281 samples collected at 15 different sow farms.
From corridors only a sock and dust sample were taken. In a compartment, a pooled fecal sample (mixed feces from the floor of each pen, max 50 ml total) was taken additionally. For each set of samples, new gloves and overshoes were used to prevent cross-contamination. For socks we used non-sterile gauze tissue and for the dust samples we used Swiffer® pads. All samples were analyzed via a standard bacteriological technique for Salmonella and suspicious colonies were identified with respect to serotype.
In total, 42% of the samples were positive for Salmonella, of which 58% was S. Typhimurium (ST). Out of 117 socks 54 were Salmonella positive (30 were ST); of 89 dust samples 43 were positive (25 were ST) and of 66 fecal samples 17 were positive (10 were ST). Relative number of ST out of Salmonella positive samples was comparable for all sample types. Dust tended to be more often positive than socks in corridors (54% vs. 43%). Socks tended to be more often positive in compartments (49% vs. 44%). Within compartments, fecal samples were always negative if the sock was negative. Only in two samples the fecal sample was positive but the dust sample was negative (however in those cases the sock was positive as well). Dust was positive in 4 compartments where the sock was negative and the sock was positive in 7 compartments where the dust was negative.
We conclude that sock and dust samples are good matrixes to detect Salmonella in sow herds. Taking a pooled fecal sample, when taking a sock and dust sample from the same compartment, has no added value."
08-Jun-2018IPigVetIPigVetI also believe that the authors are too definite in their conclusions. What this study shows very well is that depending on what kind of question you want to answer you have to use different types of samples and sampling techniques. There is no such thing as "one size fits all" in diagnostics.
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