Helminths in Invasive Raccoons (Procyon lotor) from Southwest Germany.

As hosts of numerous zoonotic pathogens, the role of raccoons needs to be considered in the One Health context. Raccoons progressively expand their range as invasive alien species in Europe. This study aimed to investigate the intestinal helminth fauna of raccoons in Baden-Wuerttemberg, Germany, as no such screening had ever been conducted there. In total, we obtained 102 animals from hunters in 2019 and 2020. Intestinal helminths were retrieved using the SSCT (segmented sedimentation and counting technique) and identified morphologically and by PCR-based Sanger sequencing. Fecal samples were assessed using the ELISA PetChekTM IP assay (IDEXX, Germany) and flotation technique. The artificial digestion method was employed for analyzing muscle tissue. We detected species of four nematode genera (Baylisascaris procyonis, Toxocara canis, Capillaria spp., and Trichuris spp.), three cestode genera (Atriotaenia cf. incisa/procyonis, Taenia martis, and Mesocestoides spp.), and three trematode genera (Isthmiophora hortensis/melis, Plagiorchis muris, and Brachylaima spp.). Echinococcus spp. and Trichinella spp. were not found. The invasive behavior and synanthropic habits of raccoons may increase the infection risk with these helminths in wildlife, domestic and zoo animals, and humans by serving as a connecting link. Therefore, it is crucial to initiate additional studies assessing these risks.

Gammaherpesvirus Infections in Cattle in Europe.

The genus Macavirus, subfamily Gammaherpesvirinae, comprises ungulate viruses that infect domestic and wild ruminants and swine. They cause asymptomatic latent infections in reservoir hosts and malignant catarrhal fever in susceptible species. Lung, spleen, bronchial lymph node, and tongue were collected from 448 cattle (348 necropsied, 100 slaughtered) in Switzerland, United Kingdom, Finland, Belgium, and Germany to determine their infection with bovine herpesvirus-6 (BoHV-6) and gammaherpesviruses of other ruminants, i.e., ovine herpesvirus-1 and -2, caprine herpesvirus-2, and bison lymphotropic herpesvirus, using quantitative PCR. Only BoHV-6 was detected, with an overall frequency of 32%, ranging between 22% and 42% in the different countries. Infection was detected across all ages, from one day after birth, and was positively correlated with age. There was no evidence of an association with specific disease processes. In positive animals, BoHV-6 was detected in all organs with high frequency, consistently in the lungs or spleen. Viral loads varied substantially. In BoHV-6-positive gravid cows, organs of fetuses tested negative for infection, indicating that the virus is not vertically transmitted. Our results confirm previous data indicating that BoHV-6 is a commensal of domestic cattle not associated with disease processes and confirm that infections with other macaviruses are rare and sporadic.

Identification of a novel clade of group A rotaviruses in fatally diseased domestic pigeons in Europe.

Rotaviruses are well-known causative agents of enteric disorders in humans and other mammals, but little is known about their virulence and pathogenic role in pigeons and other birds. Starting in summer 2017, a series of outbreaks of an acute disease with high mortalities was reported in domestic pigeons in Germany, Belgium and Denmark. The clinical picture was characterized by diarrhoea, vomiting, hepatic necrosis and sudden fatalities. From these severe outbreaks, we discovered several previously unknown group A rotavirus (RVA) lineages of genotype G18P[17]-I4-R4-C4-M4-A4-T4-N4-E19-H4, which were closely related but not identical to an RVA variant identified in cases of fatal hepatic necrosis in Australian pigeon lofts in 2016. Retrospective analysis demonstrated that the predecessors of the highly virulent variants have circulated in Europe since at least 2010. Our data indicate that reassortment and intercontinental spread has led to the emergence of novel RVA variants, which may constitute a major threat to animal welfare and health of domestic pigeon populations worldwide.

First isolation of EBLV-2 in Germany.

In Europe, rabies in bats is caused by European Bat Lyssavirus (EBLV) type 1 (EBLV-1) or type 2 (EBLV-2) which form two distinct genotypes (gt 5 and 6) within the genus Lyssavirus of the family of Rhadoviridae. Spill-over infections of EBLV in humans have caused fatal rabies encephalitis and highlighted the relevance of this wildlife disease for public health. The vast majority of the 831 European bat rabies cases reported between 1977 and 2006 were identified as EBLV-1. Only few virus isolates originating from Switzerland, The Netherlands and the United Kingdom were characterized as EBLV-2. Here we report the first EBLV-2 case detected in Germany in a Daubenton's bat (Myotis daubentonii) in August 2007. The bat showed clinical signs of disorders of the central nervous system and subsequently tested positive for rabies. The virus was isolated and characterized as EBLV-2 based on its antigen pattern and by nucleotide sequencing. Phylogenetic analysis indicated an association to EBLV-2 isolates from Switzerland which correlates with the origin of the bat close to the Swiss border.

Detection of Chlamydia suis from clinical specimens: comparison of PCR, antigen ELISA, and culture.

Cell culture is still widely regarded as the gold standard in chlamydial diagnosis despite its well-known limitations in terms of sensitivity. On the other hand, the polymerase chain reaction (PCR) has emerged as a promising alternative because of rapidity and high sensitivity. However, validation of methodologies is required before the issue of standardization can be addressed. In the present study, 109 clinical samples (organ tissue, nasal, and faecal swabs) from pigs experimentally infected with Chlamydia suis were examined by cell culture, nested PCR in the ompA gene region, and two different antigen enzyme-linked immunosorbent assays (ELISAs) in order to compare the diagnostic performance of these methods. Culture and PCR produced the highest proportion of concordant results (kappa coefficient 0.712). Among 99 samples, 34 were positive in both assays, 51 were negative in both assays, 12 culture-negatives were positive in PCR, and only 2 culture-positives were negative in PCR. Thus, the sensitivity and specificity of PCR vs. culture as standard were 94.4% and 81.0%, respectively, whereas the corresponding values for culture vs. PCR as standard were 73.9% and 96.2%, respectively. Both ELISA tests performed considerably weaker. The data underline the potential of PCR as a powerful detection method for chlamydiae.

Respiratory chlamydial infection based on experimental aerosol challenge of pigs with Chlamydia suis.

An experimental study of aerogeneous challenge in pigs was conducted in order to reveal characteristic features of porcine respiratory chlamydiosis. Eight conventionally raised pigs were exposed to a pathogenic strain of Chlamydia (C.) suis, four controls were mock infected. Besides pathological changes, the acute-phase and humoral immune responses, as well as the dissemination and transmission of the challenge strain was monitored in the course of infection. The data from clinical investigations, LPS-binding protein assay, antibody ELISAs, confocal laser scanning and light microscopy, immunohistochemical staining and PCR provided extensive evidence of the pathogenic potential of C. suis for the porcine respiratory system. This model appears suitable for further pathophysiological and immunological investigations of chlamydial respiratory infections and can also be recommended for studies of Chlamydia-associated infections of the human lung.

Presence of Clostridium difficile PCR ribotype clusters related to 033, 078 and 045 in diarrhoeic calves in Germany.

This study provides data on the distribution and relationship of C. difficile PCR ribotypes in diarrhoeic calves in Germany. C. difficile was isolated from 176 of 999 (17.6 %) faecal samples or swabs of diarrhoeic calves from 603 farms collected between January 2010 and August 2012 by eight federal laboratories of six states. Strains were assigned to 17 PCR ribotypes. PCR ribotypes 033 (57 %), 078 (17 %) and 045/FLI01 (closest match to 045 in the WEBRIBO database; 9 %) were found the most frequently. Nine per cent of all culture-positive tested animals shed more than one multiple locus variable number tandem repeat analysis (MLVA) or PCR ribotype. Eight PCR ribotypes with related profiles (including 033, 078 and 045/FLI01) representing 92 % of all isolates were grouped into three clusters. Molecular relatedness was supported by the absence of the MLVA locus A6Cd only in clustered strains and identical toxin gene profiles for strains within each cluster. Previously reported mulitilocus sequence typing analysis for PCR ribotypes that were also recovered in this study found identical sequence types and a tcdC deletion (Δ39 bp) for 033, 045, 078 and 126 (ST-11), confirming this clustering. A different geographical occurrence of PCR ribotypes was shown for cluster 033 (found more frequently in southern Germany) and 045 (found more frequently in northern Germany). This study showed that clusters of C. difficile PCR ribotypes related to 033, 078 and 045 are predominant in diarrhoeic calves in Germany. The high number of strains belonging to PCR ribotype 078 demonstrated that diarrhoeic calves are also potential reservoirs for human pathogenic C. difficile strains.

[Studies on the role of the red fox (Vulpes vulpes) as a potential definitive host of Neospora caninum]. / Untersuchungen zur Rolle des Rotfuchses (Vulpes vulpes) als möglicher Endwirt von Neospora caninum.

Neospora (N.) caninum is a protozoan parasite which is regarded as a major cause of abortion in cattle. Dogs and coyotes are definitive hosts of N. caninum which may shed environmentally resistant stages, oocysts, in their feces. Epidemiological studies in Germany showed that the presence of dogs increased the risk of a bovine herd to be N. caninum-positive in a bulk-milk ELISA test. However, there were also N. caninum-positive herds where dogs were not kept together with cattle.This leads to the question whether canids other than dogs, e.g., foxes, might be involved in the horizontal transmission of N. caninum. Therefore, the aim of our examinations in wild animals was to find out whether there are indications for a sylvatic cycle with foxes as definitive hosts and deer, roe deer and wild mice samples contained structures which resembled those of coccidian oocysts. In 13 of these 65 samples coccidian DNA was detected using a 18S rRNA gene based polymerase chain reaction (PCR).The examination of the 65 samples in a N. caninum-specific PCR revealed no positive result. Hammondia (H.) heydorni-DNA was detected in two samples. In addition, brain samples from 528 foxes, 224 wild mice, 16 deer and roe deer as well as from 1 wild boar were examined for the presence of N. caninum DNA by real time PCR. All samples tested negative by PCR. In conclusion, our study yielded no evidence indicating that the examined animals were part of a sylvatic cycle for N. caninum.

[An enzyme-linked immunosorbent assay (ELISA) for the detection of anti-chlamydial antibodies in pig sera]. / Enzyme-Linked Immunosorbent Assay (ELISA) zum Nachweis von Chlamydien- Antikörpern in Seren vom Schwein.

The evaluation of an enzyme-linked immunosorbent assay (ELISA) for the detection of antibodies against chlamydiae in pig sera is described. The most widely used serological test is the complement fixation test (CFT). The CFT has a lack of sensitivity and specificity because of low antibody titers and unspecific reactions. Eight conventionally raised pigs were exposed to a pathogenic strain of Chlamydia suis, four controls were mock infected. The immune responses was monitored by CFT and indirect ELISA. There was no agreement between CFT and ELISA data. These results were confirmed by a study with 191 sera from nine pig farms. As shown by ELISA and PCR chlamydiae are widespread in swine.

[Detection of goose haemorrhagic polyomavirus (GHPV) in flocks with haemorrhagic nephritis and enteritis of geese in southern Germany]. / Nachweis des Goose Haemorrhagic Polyomavirus (GHPV) in Beständen mit Hämorrhagischer Nephritis und Enteritis der Gänse in Süddeutschland.

Two flocks of geese in the South of Germany independently diseased on Haemorrhagic Nephritis and Enteritis of Geese (HNEG) at the age of 4 weeks. The flocks were approximately 300 km apart but had received goslings from the same hatchery. In both flocks the animals died within 12 hours mainly without showing clinical signs. Some of the first cases showed haemorrhagic typhlitis, whereas in later cases visceral gout was the main finding. In all cases, pathohistological examination generally showed necrosis of the tubular epithelium of the kidney. After a course of 5 weeks no new occurrences were seen. Death rates of 43.8% for the first flock and 29.2% for the second flock, respectively, were recorded. In both cases, the diagnosis HNEG was confirmed by the detection of the recently described Goose Haemorrhagic Polyomavirus (GHPV) using polymerase chain reaction.