Assessment of Salmonella contamination using an antibody-ELISA test and a PCR technique in pigs at slaughter and on farm level

Salmonella spp. are among the most important food-associated infective agents of intestinal diseases in humans. In several European countries monitoring programmes have been established to estimate the risk of infection from pork, for example by testing for Salmonella antibodies in samples of meat juice taken from abattoirs. Producers are classified in three categories according to the percent of seropositive animals determined in the slaughter house. Such a monitoring programme is planned for Germany, as well. However, the potential risk from delivery of animals carrying or shedding Salmonella at the abattoir and for meat as food cannot be judged solely on the basis of antibody titres. Proof of actual contamination of animals in the herd with Salmonella is necessary, as it is unclear what measures should be taken to avoid cross-contaminations before and during slaughter when a large proportion of the pigs are seropositive. The purpose of the present investigations was to compare the Salmonella antibody status of pigs at the abattoir with the actual degree of colonisation of the animals‘ intestines (jejunum) and tonsils by Salmonella spp. in order better to judge the usefulness of serological test results for estimating the risk of contamination. At the same time this study was intended to test the usefulness of classifying fattening farms as free of Salmonella, slightly or seriously contaminated, on the basis of serological status, and to discuss possible sources of contamination at the farm. Samples (diaphragm pillar muscle, jejunum and adhering lymph nodes) were taken on twelve slaughter days from 383 pigs from 32 different fattening farms. On four of the twelve days (days 9 through 12), samples were additionally taken from the tonsils of 129 slaughter pigs. The order in which the animals were delivered and slaughtered was noted on every sampling day. The samples of the diaphragm pillar muscle were frozen for subsequent assessment of the meat juice in antibody ELISA. Tissue samples of intestine, lymph nodes and tonsils were pre-enriched in buffered peptone water in preparation for the detection of Salmonella spp. by PCR. A total of 7.04 % (27 animals) of the slaughtered pigs were tested seropositive; these originated from 19 % (6 fattening farms) of the 32 suppliers. (Sixteen animals came from farm F13, five from F11, two from F20, and one each from F15 and F27.) A single supplier, F13, was the source of 59 % (16 animals) of the ELISA-positive animals. Salmonella DNA was detected in jejunum samples with adhering lymph nodes in 63 (16.4 %) of the pigs slaughtered. One third of these (21 animals) were supplied by farm F13. Although 15.5 % of the 129 tonsil samples were found to be positive by PCR, only 1 animal came from pigs supplied by farm F13. Salmonella DNA was also isolated from the jejunum samples of about 40 % (11 animals) of the seropositive pigs. On the other hand, only circa 17.5 % of the jejunum samples found to be positive by PCR correlated with the ELISA test. Positive antibody ELISA and/or Salmonella spp. were found in the organs on only seven of the twelve sampling days. Strikingly, the time when animals from the various farms were delivered and slaughtered had a considerable influence on the frequency of Salmonella findings over the course of the day. When animals from farms with high levels of positive antibody titres and PCR findings in organ samples were slaughtered at the beginning of the day, there was clear evidence of Salmonella spp. in intestine and tonsil samples. But when animals from contaminated farms were slaughtered at the end of the day, Salmonella were found only sporadically in the pigs from other farms slaughtered earlier in the day. After classification of the suppliers on the basis of the results of the meat juice antibody ELISA (cut-off OD % > 40), closer investigation was undertaken of one farm each classified as category I or II (with between 0 and 20 % seropositive animals, and between 20 and 40 %, respectively). Three blood samples were taken at different times from twenty animals from newly stocked herds in each of these farms and tested for antibodies with ELISA. In addition, environmental samples were taken in the stalls (swabs of surfaces, feed samples), as were samples of faeces, which were analysed with traditional microbiological methods for the presence of Salmonella. In farm F11, 20 % of the blood sera collected on the first testing day were serologically positive, as were 25 % of the blood sera taken on days 2 and 3. In farm F13, on the other hand, there was no positive blood sample on day 1, and only 5 % were positive on days 2 and 3. This contradicts the prevalences determined by the meat juice samples for these farms, where the category I farm, F11, showed 9.4 % positive, and farm F13, category II, 29.1 %. All feed samples tested negative microbiologically, and only one collective faeces sample was positive (in farm F13). The results of this study show that the meat juice antibody ELISA is apparently only an insufficient indication of the actual contamination of the animals with Salmonella. More than twice as many pigs carrying Salmonella will be identified if the intestine (intestinal lymph nodes, material from the jejunum) are included in the investigation. These latent infections also increase the risk of unrecognised cross-contaminations at the abattoir when pigs afflicted with Salmonella are slaughtered at the beginning of the day. It is therefore necessary to know the pigs‘ antibody status at the production unit if at all possible. If this is not possible, then at least the animals‘ antibody status should be determined at the abattoir and linked to the suppliers. However, the findings also show that it is not enough to classify the fattening farms as to specific contamination categories solely on the basis of meat juice testing at the abattoir, because the antibody status of a herd can change dramatically every time the farm is restocked. This is particularly true when suppliers are changed frequently, and demonstrates the advantage of so-called integrated chains (with only one or a few, well-established suppliers). In farms not managed in accordance with the chain concept, animals should be tested in good time, at least a week before slaughter. This is essential for the recognition of risk and minimisation of contamination. Animals from units suspected of contamination should be slaughtered only at the end of the day, and the abattoir should be cleaned especially thoroughly and disinfected. It appears that close co-operation between producers, abbatoirs and public health authorities is necessary to break the chain of Salmonella contamination effectively. This is in the interest of animal owners and meat producers and will promote the health of both animals and consumers.