Karsten Fehlhaber

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.

Migration of Salmonella enteritidis from the albumen into the egg yolk.

Migration of Salmonella enteritidis from the albumen into the egg yolk was investigated in 860 eggs. After artificial contamination of the albumen with different doses of S. enteritidis phage-type 4 the migration and the effect of temperature on the migration process during a storage period of up to 4 weeks were studied. The experiments showed that the first cells can be detected in the yolk within a few days. The process will be relatively rare during chilled storage if the albumen is only slightly contaminated. The migration rate was positively correlated with the level of contamination, the storage temperatures and the age of the eggs.

The importance of Campylobacter coli in human campylobacteriosis: prevalence and genetic characterization

The primary aim of this study was to detect and genotype Campylobacter coli strains from humans over a period of one year from November 2002 to October 2003. Campylobacter spp. were isolated from patients with symptoms of enteric infection. Amplified fragment length polymorphism (AFLP) analysis was used to identify the genetic diversity of C. coli strains by cluster analysis. A total of 18.6% of all Campylobacter isolates were identified as C. coli. These data show, that C. coli is indeed of importance to human campylobacter infections. Heterogeneous patterns were detectable among the human C. coli pool by AFLP analysis suggesting different sources of infection. A continuous seasonal shift of genotypes was detectable.

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.

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.

Effects of pH, Sodium Chloride, and Curing Salt on the Infectivity of Toxoplasma gondii Tissue Cysts

Toxoplasma gondii is one of the most common zoonotic parasites in the world. The parasite causes no or mild symptoms in immunocompetent humans. However, a high potential hazard exists for seronegative pregnant women and immunocompromised patients. The consumption of meat containing tissue cysts or oocyst-contaminated vegetables and fruits or the handling of cat feces poses a high risk of infection with T. gondii. It is known that raw minced meat, raw fresh sausages, and locally produced raw meat products are possible causes of T. gondii infection. The infectivity of T. gondii tissue cysts in meat products depends, among other factors, on the pH and the salt concentration. Therefore, the impact of these two factors on the tissue cysts was examined. For this purpose, dissected musculature and brain from experimentally infected mice (donor mice) were placed in a cell culture medium (RPMI 1640). The medium was adjusted to different pH values (pH 5, 6, and 7) with lactic acid and to different salt concentrations (2.0, 2.5, and 3.0%) with sodium chloride (NaCl) or nitrite-enriched curing salt (NCS) for the various tests. After storage at 4°C for different time periods, the materials were fed to bioassay mice. Later, the brains were examined for presence of T. gondii to assess the infectivity. The data show that T. gondii tissue cysts have a high pH tolerance. Cysts were infectious in the muscle for up to 26 days (pH 5). In contrast to their tolerance to pH, cysts were very sensitive to salt. Muscle cysts survived at an NaCl concentration of up to 2.0% only, and for no longer than 8 days. At NaCl concentrations of 2.5 and 3.0%, the cysts lost their infectivity after 1 day. When NCS instead of NaCl was used under the same conditions, T. gondii muscle cysts retained infectivity for only 4 days at 2.0%. Consequently, NCS (NaCl plus 0.5% nitrite) has a stronger effect on T. gondii cysts than does common table salt. Sausages produced with low NaCl concentration and short contact times pose a potential risk for susceptible individuals.

Typing of Salmonella enterica serovar Infantis isolates from 51 outbreaks in Germany between 1974 and 2009 by a novel phage-typing scheme

We developed a new phage-typing method and evaluated its application in combination with XbaI macrorestriction analysis by pulsed-field gel electrophoresis (PFGE) as a useful tool for the long-term epidemiology of Salmonella enterica serovar Infantis. In this study, we investigated 1008 S. Infantis isolates recovered from humans, various animal species and food products from 1973 to 2009. The typing scheme is based on 17 typing phages, defining 61 different patterns within the strain collection. The experiments showed that phage typing is a reliable method for differentiation of outbreaks and sporadic clinical cases as well as for elucidation of chains of transmission. The combined analysis of phage typing and PFGE revealed the existence of epidemic clones with a high stability over time like PT29/XB27 which was identified in nosocomial salmonellosis, community outbreaks as well as in broiler chickens from 2002 to 2009.

Experimental Toxoplasma gondii oocyst infections in turkeys (Meleagris gallopavo)

Toxoplasma (T.) gondii is a protozoan parasite with a broad range of intermediate hosts. Humans are often infected by ingestion of tissue cysts in raw or undercooked meat or meat products. Turkeys as food-producing animals can also serve as intermediate hosts. The aim of the present study was to investigate occurrence and predilection sites of T. gondii infection in turkeys after oral infection with oocysts. Experimental infections with different doses of T. gondii oocysts were performed in 36 turkeys to mimic natural infection. Systemic distribution of parasitic stages was investigated by screening 14 different tissues including the edible tissues heart, liver, thigh, breast and drumstick muscle. Parasite detection was based on a conventional nested polymerase chain reaction (PCR). Animals were sacrificed 6-12 weeks after infection. Results demonstrated parasite spreading over the whole organism after oral infection by oocysts. Most frequently affected tissues were brain (47.2% of all brains were positive for T. gondii) and thigh muscle (25.0% positive samples). Other muscles were regularly T. gondii-positive, all other sampled tissues were positive at least once. Thus, edible tissues are one of the predilection sites of T. gondii in turkeys which renders raw or undercooked turkey meat a potential risk for parasite transmission to humans. Data were compared to results from previous parenteral turkey infections with tachyzoites. With the exception of brain, liver and breast muscle affection, no significant differences were observed between both infection routes. Both infection models could be used for research purposes with certain advantages and disadvantages.

Examinations on the prevalence of footpad lesions and breast skin lesions in B.U.T. Big 6 fattening turkeys in Germany. Part II: Prevalence of breast skin lesions (breast buttons and breast blisters)

This project involves the prevalence of breast skin alterations (breast buttons, blisters, purulent bursitis) in male and female fattened turkeys (British United Turkeys Big 6) from conventional intensive farming. In this case, 24 turkey farms across Germany and 11,860 turkeys (5,740 toms, 6,120 hens) were clinically examined, and 16,200 birds (8,400 hens, 7,800 toms) were evaluated on slaughter lines at 7 turkey abattoirs. Each turkey farm included examination of at least 3 consecutive fattening cycles. Especially in the meat examinations, breast skin alterations were a frequent discovery, particularly in the form of breast buttons. The turkey toms exhibited a prevalence of 27.15%, and the toms were affected significantly more frequently (P < 0.001) by these variations than the turkey hens (7.77%). It is possible that the higher BW of the male specimens leads to increased lying times and is therefore connected with more severe strain on the breast region. The increased fattening period of the turkey toms (slaughtering at wk 21 to 22 of life) vs. the female specimens (slaughtering at wk 15 to 17 of life) must be discussed as a cause. Breast blisters and bursitis sternalis were discovered with a prevalence of 7.36 and 1.24% in toms and more seldom in hens, at 0.30% and 0.15%. Large differences in prevalence were partially recognizable between individual farms as well as between the 3 fattening cycles on a farm. Breast buttons and breast blisters are pathological skin variations for which the etiology is multifactorial. Clear correlations between the prevalence of breast skin alterations and the manner of rearing the birds were not able to be determined. An improvement in this situation, which is considered very problematic from the perspective of bird protection and economics, may be achieved only via cooperation to achieve optimized rearing methods, management, and breeding practices.