Toward harmonization of the European food hygiene/veterinary public health curriculum.

Prompted by developments in the agri-food industry and associated recent changes in European legislation, the responsibilities of veterinarians professionally active in veterinary public health (VPH), and particularly in food hygiene (FH), have increasingly shifted from the traditional end-product control toward longitudinally integrated safety assurance. This necessitates the restructuring of university training programs to provide starting competence in this area for veterinary graduates or a sub-population of them. To date, there are substantial differences in Europe in the way in which graduate programs in FH/VPH are structured and in the time allocated to this important curricular group of subjects. Having recognized this, the European Association of Establishments for Veterinary Education (EAEVE) recently instituted a working group to analyze the current situation, with a view to produce standard operating procedures allowing fair and transparent evaluations of universities/faculties constituting its membership and in concurrence with explicit European legislation on the professional qualifications deemed necessary for this veterinary discipline. This article summarizes the main conclusions and recommendations of the working group and seeks to contribute to the international efforts to optimize veterinary training in FH/VPH.

Survival of Toxoplasma gondii tachyzoites in simulated gastric fluid and cow's milk.

The protozoan parasite Toxoplasma gondii is one of the most important food-related pathogens worldwide. Besides contact to oocysts or ingestion of tissue cysts mainly by consumption of raw or undercooked meat from infected animals, raw milk is considered to be a risk factor and possible route of transmission for tachyzoites. This stage of the parasite is usually very sensitive to acidic pH and, therefore, considered unlikely to survive stomach passage. However, tachyzoites were shown to survive for several days in milk and there are also reports on transmission of toxoplasmosis via milk. Thus, the aim of the study was to examine retention of infectivity of tachyzoites in simulated gastric fluid (SGF) of different acidity and to elucidate whether addition of different shares of milk would affect survival of the parasites. Tachyzoites were exposed to SGF of pH 2.0 through 6.0 and their remaining infectivity was examined by cell culture. Furthermore, the impact on survival was investigated in different admixtures of milk to the SGF (25, 50, 75%) as well as in pure milk. Tachyzoites were shown to retain infectivity in SGF of pH 5.0 and 6.0 for at least 90min while they were more sensitive to lower pH values. Admixture of milk resulted in extension of survival. The results support the hypothesis of tachyzoites to survive stomach passage and their retention of infectivity.

Prevalences and transmission routes of Campylobacter spp. strains within multiple pig farms.

In this work, faecal samples were collected from 15 pig farms to determine the Campylobacter prevalences at different times during the rearing period and to visualize the exchange of strains among the pig population by genotyping specific isolates. All isolated strains were identified as C. coli. Whereas no Campylobacter were detectable in the faeces of piglets at the day of birth, the Campylobacter incidence rose within days to 32.8%. After transfer to the nursery unit the prevalence increased to 56.6%. Approximately two-thirds of the pigs remained C. coli shedders in the fattening unit. In contrast to most farms, one farm expressed a very low Campylobacter incidence during the whole rearing period. Amplified fragment length polymorphism (AFLP) analysis was performed on all C. coli isolates of one farm. Clonal strains were identified from the brood sows and their offsprings or neighbouring piglets. After moving to the nursery unit, new genotypes appeared in that pig group but the original C. coli strains largely remained within that group. C. coli genotypes, identified during the fattening period, replaced the previously isolated genotypes. Transportation to the abattoir had no significant influence on the shedding rate of C. coli. The detection rate before transportation was 79.1% and decreased slightly to 78.2% (n=330). Additionally, eleven of 1474 environmental samples from different sources of the pig farms were positive for C. coli. This study demonstrates the importance of pigs as a reservoir for C. coli. Maternal C. coli strains are the primary source of infection but non-related genotypes from different sources appear during the rearing period and these latter strains constitute largely the final C. coli flora.

Prevalence of Yersinia enterocolitica in fattening pigs.

The prevalence of pathogenic Yersinia enterocolitica in pig herds was monitored during six trials (at four different farrow-to-finisher farms). Samples were taken throughout the whole rearing period from birth of the piglets to the final fattening stage, and different samples were taken from these pigs during the slaughter process. Environmental samples also were evaluated to identify potential sources of on-farm infection. Y. enterocolitica was isolated using irgasan-ticarcillin-potassium chlorate broth enrichment and cefsulodin-irgasan-novobiocin agar culture. Colonies were identified using bio- and serotyping methods and by PCR assay. Pathogenic Y. enterocolitica were not isolated from fecal samples from piglets and weaners. The only fecal samples positive for Y. enterocolitica were obtained during the fattening stage. The prevalence of Y. enterocolitica in fattening pig herds ranged between 0 and 65.4%. Y. enterocolitica isolates were detected at the abattoir in 38.4% of the tonsils, in 3.8% of the ileocecal lymph nodes, on 0.3% of the carcass surfaces before chilling, and on 0% of the carcass surfaces after chilling. Almost all isolates belonged to bioserotype 4/O:3. Only one strain was identified as O:9. All isolates contained the ail gene. The yopT gene was found in 99.1% of the farm isolates but in only 76.6% of the isolates found at the abattoir from the corresponding carcasses. Although a direct link between porcine isolates and human infection has not been demonstrated, the similarity of the bioserotypes in infected pigs and humans and the presence of virulence factors in porcine isolates should encourage further studies to determine the risk of transmission of Y. enterocolitica to humans from pigs and pork products.

Growth of Salmonella enteritidis in yolk from eggs laid by immunized hens.

After hyperimmunization of laying hens with Salmonella enteritidis, antibodies can be found in egg yolks. This study was conducted to ascertain whether the growth of S. enteritidis would be suppressed in the presence of antibodies contained in egg yolk. Specifically pathogen-free (SPF)-laying hens were immunized with S. enteritidis; eggs were collected, the yolk was separated and the concentration of S. enteritidis antibodies was determined quantitatively by using the enzyme-linked immunosorbent assay (ELISA), the radial immunodiffusion and the bicinchoninic acid protein assay. Then, the yolk was inoculated with approximately 10, 100 or 1000 S. enteritidis cells/ml and incubated at 15, 20 and 30 degrees C for 0, 2, 6 and 24 h. The growth of organisms in each yolk was examined, and the generation times were calculated. The egg yolk from nonimmunized hens served as negative control. The highest level of antibody concentration was found in the hyperimmunized group. There was no difference in the generation times of S. enteritidis between the antibody-positive yolk and the negative yolk at the three different incubation temperatures. The results suggest that antibodies in the yolk do not influence the growth of S. enteritidis, even if the hens are highly immunized.

Impact of Sodium Chloride, Sucrose and Milk on Heat Stability of the Murine Norovirus and the MS2 Phage.

Until now, little is known about the influence of food additives on heat inactivation of noroviruses. Only a few studies have shown a protective or inhibiting effect on virus infectivity caused by the food matrix. Therefore, the aim of this study was to examine the influence of sodium chloride, sucrose and milk on heat stability of the surrogates murine norovirus (MNV) and MS2 phage at 60 °C for 1-5 min in PBS for MNV and for 5-120 min in suspension medium buffer for MS2 phage. Different concentrations of sodium chloride (5, 10 %) and sucrose (5, 50 %) were added to the respective buffers. In addition, commercially available milk with different fat concentrations (0.3, 1.5, 3.5 %) was investigated in this study. In general, a linear titre reduction for MNV and MS2 phage could be observed, except for the heat treatment of MNV in PBS with 50 % sucrose. A protective effect of PBS with 50 % sucrose and of the matrix milk on MNV could be concluded. All other tested conditions did not show any influence on virus inactivation. However, MS2 phage did show a higher heat resistance throughout the experiments compared to MNV. In future investigations, it should be tested, whether the achieved data may be considered in risk assessments of heat-treated food products with high concentrations of sugar. Furthermore, it should be clarified, whether these results can also be referred to complex food matrices.

Impact of Selected Factors on the Occurrence of Contact Dermatitis in Turkeys on Commercial Farms in Germany.

In a long term research project in Germany the influence of husbandry on the health of fattening turkeys (Study 1) as well as the influence of practiced rearing conditions on the health of turkey poults (Study 2) was examined in 24 farms and at the meat processing plant. In all examined rearing farms, litter samples for the determination of litter moisture were taken. This paper summarizes the results obtained by our working group from 2007 until 2012. The results elucidate the universal problem of foot pad dermatitis (FPD). Nearly 100% of the observed turkeys showed a clinically apparent FPD at the meat processing plant. Furthermore, skin lesions of the breast, especially breast buttons were diagnosed, particularly at the slaughterhouse. FPD was detected in the first week of the rearing phase. Prevalence and degree showed a progressive development up to the age of 22-35 days, whereas 63.3% of the poults had foot pad alterations. As even mild alterations in the foot pad condition can be indicators for suboptimal design of the rearing environment, especially high litter moisture, it is important to focus on the early rearing phase.

Salmonella status of pigs at slaughter--bacteriological and serological analysis.

Apart from Salmonella monitoring of pig herds during the period of growth to evaluate the efficacy of control programmes, monitoring at harvest level is of relevance to assess the Salmonella status of fattening pigs and the associated risk of introducing Salmonella organisms in the slaughter process. Samples from 1830 fattening pigs were gathered at slaughter. Ileocaecal lymph nodes, rectal and caecal content as well as tonsils were collected for bacteriological examinations, and a part of the diaphragm pillar muscle was taken to gain meat-juice for serological analysis. Salmonella spp. was recovered from 13.8% of all pigs examined. Salmonella Typhimurium and Derby were the dominating serovars. The highest detection rates were found in caecal content followed by ileocaecal lymph nodes. By analysing both organs nearly 90% of all Salmonella positive pigs could be identified. Serological examination revealed 9.6% of the pigs as positive using a cut-off value of OD % ≥ 40. Only one quarter of all Salmonella positive pigs showed also a positive serological result. A reduction of the cut-off value does not necessarily result in a higher compliance between bacteriologically and serologically positive slaughter pigs. Detection of antibodies is useful to verify whether pig herds were previously exposed to Salmonella organisms. However, the Salmonella status of pigs at time of slaughter and the associated risk of dissemination of Salmonella organisms can only be assessed by bacteriological examinations which should include both lymph nodes and caecal content.

Prevalence of specific IgG-antibodies against Toxoplasma gondii in domestic turkeys determined by kinetic ELISA based on recombinant GRA7 and GRA8.

The protozoan parasite Toxoplasma (T.) gondii is one of the most common zoonotic infectious agents worldwide. Besides its sexual reproduction in cats, T. gondii can also infect a wide spectrum of other warm-blooded animals. These include animals used for human consumption such as pigs or chickens. Nevertheless, the role of turkeys for the epidemiology of T. gondii infections has not been studied thoroughly. We have established a kinetic ELISA (KELA) for the detection of T. gondii-specific IgG antibodies in turkey serum samples. The test is based on the recombinant dense granule antigens GRA7 and GRA8. These proteins were used as an antigen mixture at a concentration of 0.13 µg per well. The overall sensitivity of the assay was between 92.6% and 100% and the specificity ranged from 78.1% to 100%, depending on the method used to calculate these parameters. Using this KELA we examined 1913 turkey serum samples from 14 turkey farms from different areas of Germany. From these sera, 387 produced a signal in the KELA, corresponding to a true seroprevalence of up to 20.2%. The seropositivity rate in individual fattening cycles at individual farms ranged from 0.0% to 77.1%, whereas the rates were highly variable within the individual farms and individual fattening cycles. Consequently, conditions of animal husbandry could not be associated with particular seroprevalence rates. Although seropositivity cannot be linked directly to infectious tissue cysts in the muscle tissue of commercially produced turkey meat, we state that there is a potential risk of being infected by consuming turkey meat products that were not heat treated.

[Examination of the prevalence of skin injuries in debeaked fattened turkeys]. / Untersuchungen zur Prävalenz von Hautverletzungen bei schnabelkupierten Mastputen.

Within the framework of a cooperative research project, turkey health as well as numerous aspects of animal welfare were examined in various intensive commercial farms with varying rearing forms. The present study demonstrates the prevalence of skin injuries concerning living animals as well as carcasses of slaughtered turkeys of both sexes. Although all turkeys were debeaked, prevalences of skin injuries at a value of 12.8% with male turkeys and 13.8% with female turkeys could be found at the age of 16 weeks. Within the scope of medically inspections skin injuries were mainly established in the head region, especially the snood as well as in the region of the back. By means of their clinical picture pecking wounds and scratch injuries could be distinguished. Skin lesion produced primary as a result of scratching the skin surface, could be enlarged by the animal itself or by conspecifics by beak pecking, especially after bleeding, in spite of debeaked upper beak. Injuries in the head region, especially of the snood, could be explained mainly as a result of pecking by conspecifics by reason of their clinical picture and were found mainly in male turkeys. Skin injuries in the region of the back, especially at the coxal tuberosity, were identified primarily as scratch marks. They were found mainly in female turkeys. A statistically significant correlation of the injury prevalence to particular husbandry parameters discussed in the literature as "predisposing" or "limiting" factors (e.g., population density, light regime or offer of employment material) could not be established in this study. At the meat inspection fresh scratching injuries at the chest and the hind legs were diagnosed most often. Especially fresh injuries of the carcass point to a misguided behaviour of the staff concerning animal handling during loading and transport which is not conform to animal welfare.

[Mycoplasma synoviae-associated egg-pole shell defects in laying hens]. / Mycoplasma synoviae-assoziierte Eischalenpoldefekte bei Legehennen.

Hens laying eggs with egg-pole shell defects (EPS) were examined in a clinical prospective study. 86 hens with EPS and 72 hens without EPS from three flocks were selected for this study. It could be proven serologically that hens with EPS had significant (p < 0.05) higher titers against Mycoplasma (M.) synoviae then hens without EPS. PCR tested cloacal swabs for M. synoviae were more frequently positive from hens with EPS (87%; n=72) then from hens without EPS (18%; n=13). Furthermore, M. synoviae could be cultivated from the oviduct of five hens with EPS. Additionally, M. synoviae-DNA was detectable in the albumen of nearly all eggs with EPS (n=48; 98%), contrary to the eggs without EPS (n=11; 26%). Ultrastructural investigation revealed that eggs with EPS showed considerable differences of the egg shell structure as well as the cross section dimension according to eggs without EPS. Due to the significantly more frequent detection of M. synoviae-DNA from the cloaca of chickens with EPS, is an involvement of M. synoviae in laying of EPS in the surveyed herds likely. Further infection experiments with the isolated M. synoviae were not perfomed, therefore about the causal pathogenic role of M. synoviae in the development of eggs with EPS in the surveyed herds can only be speculated.

[Examination of rearing standards and health status in turkeys in Germany]. / Erhebungen zur Haltung und Gesundheit bei Mastputen in Deutschland.

Within the framework of a cooperative research project, sponsored by the Federal Ministry of Food, Agriculture and Consumer Protection in Germany (BMELV), turkey health as well as numerous aspects of animal welfare in various intensive commercial farms with varying rearing forms were examined. For this purpose extensive documentation of flock management and health status information was conducted over the past two years at the farms. Additional information was ascertained from the carcasses at the slaughterhouse. The first results of this study, which look at flock management, will be presented here. Upon reviewing the questionnaires, it is clear that on one hand almost all farmers orientate there stocking rate upon the allowed limits, on the other hand, however, when considering national general requirements, which are very important parameters thatargely influence animal health (i. e. litter quality and care of sick/injured animals) farmer compliance lacks. Especially the litter quality, which, among other parameters, plays a very important role in food pad health, proved to be unsatisfactory. Unfortunately, no instruments are available that measure the litter quality, yet. The status of footpad health was a major problem in all the rearing systems. However, the prevalence of epithelial necrosis and deep skin lesions varied immensely among different flocks of the same age.

[Antibody response after immunization with a Salmonella Typhimurium live vaccine in dependence on the way of application]. / Antikörperantwort nach Impfung von Mastschweinen mit einem Salmonella-Typhimurium-Lebendimpfstoff in Abhängigkeit von der Applikationsform.

In Germany now, the recognition of Salmonella infections in pig herds is based on three different commercial tests detecting antibodies against Salmonella-derived lipopolysaccharide (LPS). However, a serious disadvantage of these tests, used so far, is the restricted detection of antibodies belonging predominantly to the immunoglobulin class g (IgG). Therefore, a new test was developed to detect three Ig classes (IgM, IgG and IgA). Different constellations between the three Ig classes allow the evaluation of the current infection status of each pig. Under field conditions, this was proved in three different vaccination trials using a commercial Salmonella Typhimurium live vaccine.

Influence of inoculation levels and processing parameters on the survival of Campylobacter jejuni in German style fermented turkey sausages.

This study investigated the influence of inoculum levels and manufacturing methods on the survival of Campylobacter (C.) jejuni in raw fermented turkey sausages. Sausages were prepared and inoculated with C. jejuni. After inoculation, these sausages were processed and ripened for 8 days. Samples were taken throughout the ripening process. The presence of C. jejuni was established bacteriologically. Additionally, lactic acid bacteria were enumerated, pH values and water activity were measured to verify the ripening process. To detect changes in genotype and verify the identity of the recovered clones, AFLP analysis was carried out on the re-isolated strains. Whereas no C. jejuni were detectable when inoculating the sausages with the lowest inoculum (0.08-0.44 log(10) cfu/g sausage emulsion), C. jejuni were detectable for 12-24h by enrichment when inoculated with approximately 2 log(10) cfu/g. After inoculation with 4 and 6 log(10) cfu/g respectively, C. jejuni were detectable without enrichment for 12-48 h and by enrichment for 144 h at the most. The greatest decrease of the C. jejuni population occurred during the first 4 h of ripening. Only a very high inoculum level allowed the survival of the organism during a fermentation process and during ripening to pose a potential risk for consumers. Lower initial Campylobacter inoculums will be eliminated during proper ripening of the sausages, if sufficient decrease in water activity and pH-value is ensured.

[Longitudinal study on the seroprevalence of Toxoplasma gondii infection in four German pig breeding and raising farms]. / Longitudinalstudie zur Seroprävalenz der Toxoplasma gondii-Infektion in vier deutschen Schweineaufzucht- und Mastbetrieben.

There are very few current data on the prevalence of Toxoplasma (T.) gondii in German pig farms. Consequently a reliable risk assessment of human Toxoplasmosis caused by ingesting raw or improperly cooked pork and pork products is not available. The aim of this study was to show current data on T. gondii prevalence in German pig farms. In four pig farms with different management systems (three conventional, one organic) 100 animals each were selected and tested for T. gondii antibodies. The test was done four times during the period from birth to slaughtering. In one farm 20 mother sows were tested additionally. The slaughtered pigs from conventional farms showed seroprevalences between 0 and 15.2% (mean value 5.6%). At the organic system T. gondii antibodies were not detected. All slaughtered seropositive pigs (6 months old) were tested negatively at the age of 9 weeks, but shortly after birth high titres of T. gondii antibodies had been detected in the same animals. Comparing the results gained in different seasons significantly more pigs were found to be infected during the autumn/winter than in the spring/summer period. In order to assess the current risk of Toxoplasmosis more pig farms should be tested. From the point of view of consumer protection the detection of highly infected pig herds is necessary.

[Distribution and genetic characterization of porcine Campylobacter coli isolates]. / Vorkommen und genetische Charakterisierung von porcinen Campylobacter coli-Isolaten.

The aim of this study was to evaluate the impact of the slaughter process on the Campylobacter (C.) coli prevalence on pig carcasses and finally pork. To detect C. spp., faecal samples, organ samples and surfaces of slaughter pigs were sampled. Additionally, various abattoir surfaces (n=208) and 227 pork and minced meat samples were included in our study. Whereas a high C. spp. prevalence (64.0%) was detectable in the faeces of slaughter pigs (all isolates were identified as C. coli), low detection rates were observed on pig carcasses after the slaughter process before the chilling period (21.1%). The impact of chilling reduced the detection rate of C spp. on pig carcasses even further to 0.8%. Only C. jejuni strains were isolated after the chilling process. Chilling and the associated drying of the skin are responsible for that massive reduction of C. spp prevalence. Significantly more C. spp. were isolated from livers compared to the corresponding carcasses. Only 5 out of 208 swab samples from different surfaces of the abattoir were C. coli positive. Bacteriological investigation could not detect any C. spp. strains from pork and minced pork meat. The low detection rates at the end of the slaughter and processing line indicate that pork may only play a minor role in the transmission of C. coli infections to humans. By genotyping C. coli-isolates from selected animals we were able to demonstrate three possible ways of contamination of the slaughter carcass surface. Genetically highly related strains were detectable on carcass surfaces of consecutively slaughtered animals. Faecal isolates and isolates from the carcass surface showed occasional high similarities. C. coli-genotypes from tonsils and genotypes from the corresponding slaughter carcasses formed a close cluster.

[Use of AFLP analysis for the molecular biological fine typing of Campylobacter jejuni strains from turkey flocks]. / Einsatz der AFLP-Analyse zur molekularbiologischen Feintypisierung von Campylobacter jejuni-Stämmen aus Putenbeständen.

One of the most promising genotyping methods for microorganisms is the AFLP (amplified fragment length polymorphism)-analysis, originally developed to genotype plants. The AFLP method has a high discriminatory power and an excellent reproducibility. Thus it is suitable for solving taxonomic and epidemiological questions. AFLP analysis was used to reveal the genetic diversity of Campylobacter (C.) jejuni strains from turkeys. The aim was to detect and genotype C. jejuni strains on individual turkeys throughout the complete slaughter process. A broad spectrum of genotypes was detectable on the turkey skin at the beginning of the slaughter line. At the end of the slaughter process-after chilling, only a diminished spectrum of genotypes was detectable by AFLP-analysis, suggesting, that slaughter specific environmental stressors may exert a selective pressure on the strain diversity. Our results proved, that AFLP analysis is an excellent tool for subtyping large numbers of Campylobacter strains.

A Rapid Test to Detect ST-producing Escherichia coli in Food.

Pasteurized milk, liquid egg, minced meat, and various salads were artificially contaminated with varying numbers of cells from six Escherichia coli ( E. coli ) strains able to produce heat-stable enterotoxins (ST). The ST-producing E. coli were detected by the following procedure within 24 h without isolation by cultivation. After enrichment of the food sample in GN broth (4 h at 37°C), the material was transferred to brain heart infusion broth, incubated (16-18 h at 37°C), centrifuged (20 min, 7000 g) and heated to 80°C for 10 min, the supernatant was tested with the infant mouse test. The sensitivity (= ratio of detectable E. coli per total microbial numbers in the food sample) of the test procedure was high even in many food samples with a considerable competitive microbial flora. The procedure was used to test 419 routine food samples. Enterotoxigenic bacteria were found in 7 samples of liquid egg and 4 samples of salad. The test is recommended as a rapid screening test in food control.