Challenge Dose Titration in a Mycobacterium bovis Infection Model in Goats.

Goats are natural hosts of Mycobacterium (M.) bovis, and affected herds can be the cause of significant economic losses. Similarites in disease course and lesions of M. bovis infections in goats and M. tuberculosis in humans make goats good models for human tuberculosis. The aim of this investigation was to characterize M. bovis challenge models in goats. For this, goats were endobronchially inoculated with three doses of M. bovis or culture medium. Clinical signs, shedding, and immune responses were monitored until 146 days post inoculation (dpi). At necropsy, lesions were examined by computed tomography, histology, and bacteriological culture. Infected goats did not develop clinical signs. M. bovis was cultured from feces, but never from nasal swabs. IGRAs were positive from 28 dpi onwards, antibodies at 140 dpi, and SICCT at 146 dpi. The increase in CD25+, IFN-γ+, and IFN-γ-releasing T-cell subpopulations was time-related, but not dose-dependent. All infected goats developed paucibacillary granulomas in the lungs and regional lymph nodes. M. bovis was regularly cultured. Dose-dependent effects included the size of pulmonary lesions, caverns, intestinal lesions, and early generalization in the high-dose group. In summary, reproducible challenge models with dose-dependent differences in lesions were established, which may serve for testing vaccines for veterinary or medical use.

Lethal Borna disease virus 1 infections of humans and animals - in-depth molecular epidemiology and phylogeography.

Borna disease virus 1 (BoDV-1) is the causative agent of Borna disease, a fatal neurologic disorder of domestic mammals and humans, resulting from spill-over infection from its natural reservoir host, the bicolored white-toothed shrew (Crocidura leucodon). The known BoDV-1-endemic area is remarkably restricted to parts of Germany, Austria, Switzerland and Liechtenstein. To gain comprehensive data on its occurrence, we analysed diagnostic material from suspected BoDV-1-induced encephalitis cases based on clinical and/or histopathological diagnosis. BoDV-1 infection was confirmed by RT-qPCR in 207 domestic mammals, 28 humans and seven wild shrews. Thereby, this study markedly raises the number of published laboratory-confirmed human BoDV-1 infections and provides a first comprehensive summary. Generation of 136 new BoDV-1 genome sequences from animals and humans facilitated an in-depth phylogeographic analysis, allowing for the definition of risk areas for zoonotic BoDV-1 transmission and facilitating the assessment of geographical infection sources. Consistent with the low mobility of its reservoir host, BoDV-1 sequences showed a remarkable geographic association, with individual phylogenetic clades occupying distinct areas. The closest genetic relatives of most human-derived BoDV-1 sequences were located at distances of less than 40 km, indicating that spill-over transmission from the natural reservoir usually occurs in the patient´s home region.

In turkeys, unlike chickens, the non-structural NS1 protein does not play a significant role in the replication and tissue tropism of the H7N1 avian influenza virus.

The economic losses caused by high pathogenicity (HP) avian influenza viruses (AIV) in the poultry industry worldwide are enormous. Although chickens and turkeys are closely related Galliformes, turkeys are thought to be a bridging host for the adaptation of AIV from wild birds to poultry because of their high susceptibility to AIV infections. HPAIV evolve from low pathogenicity (LP) AIV after circulation in poultry through mutations in different viral proteins, including the non-structural protein (NS1), a major interferon (IFN) antagonist of AIV. At present, it is largely unknown whether the virulence determinants of HPAIV are the same in turkeys and chickens. Previously, we showed that mutations in the NS1 of HPAIV H7N1 significantly reduced viral replication in chickens in vitro and in vivo. Here, we investigated the effect of NS1 on the replication and virulence of HPAIV H7N1 in turkeys after inoculation with recombinant H7N1 carrying a naturally truncated wild-type NS1 (with 224 amino-acid "aa" in length) or an extended NS1 with 230-aa similar to the LP H7N1 ancestor. There were no significant differences in multiple-cycle viral replication or in the efficiency of NS1 in blocking IFN induction in the cell culture. Similarly, all viruses were highly virulent in turkeys and replicated at similar levels in various organs and swabs collected from the inoculated turkeys. These results suggest that NS1 does not play a role in the virulence or replication of HPAIV H7N1 in turkeys and further indicate that the genetic determinants of HPAIV differ in these two closely related galliform species.

Deciphering bat influenza H18N11 infection dynamics in male Jamaican fruit bats on a single-cell level.

Jamaican fruit bats (Artibeus jamaicensis) naturally harbor a wide range of viruses of human relevance. These infections are typically mild in bats, suggesting unique features of their immune system. To better understand the immune response to viral infections in bats, we infected male Jamaican fruit bats with the bat-derived influenza A virus (IAV) H18N11. Using comparative single-cell RNA sequencing, we generated single-cell atlases of the Jamaican fruit bat intestine and mesentery. Gene expression profiling showed that H18N11 infection resulted in a moderate induction of interferon-stimulated genes and transcriptional activation of immune cells. H18N11 infection was predominant in various leukocytes, including macrophages, B cells, and NK/T cells. Confirming these findings, human leukocytes, particularly macrophages, were also susceptible to H18N11, highlighting the zoonotic potential of this bat-derived IAV. Our study provides insight into a natural virus-host relationship and thus serves as a fundamental resource for future in-depth characterization of bat immunology.

Automated quantification of avian influenza virus antigen in different organs.

As immunohistochemistry is valuable for determining tissue and cell tropism of avian influenza viruses (AIV), but time-consuming, an artificial intelligence-based workflow was developed to automate the AIV antigen quantification. Organ samples from experimental AIV infections including brain, heart, lung and spleen on one slide, and liver and kidney on another slide were stained for influenza A-matrixprotein and analyzed with QuPath: Random trees algorithms were trained to identify the organs on each slide, followed by threshold-based quantification of the immunoreactive area. The algorithms were trained and tested on two different slide sets, then retrained on both and validated on a third set. Except for the kidney, the best algorithms for organ selection correctly identified the largest proportion of the organ area. For most organs, the immunoreactive area assessed following organ selection was significantly and positively correlated to a manually assessed semiquantitative score. In the validation set, intravenously infected chickens showed a generally higher percentage of immunoreactive area than chickens infected oculonasally. Variability between the slide sets and a similar tissue texture of some organs limited the ability of the algorithms to select certain organs. Generally, suitable correlations of the immunoreactivity data results were achieved, facilitating high-throughput analysis of AIV tissue tropism.

Brachyspira in dogs: risk factors of shedding in central Germany and longitudinal study of an infected kennel.

BACKGROUND: Brachyspira (B.) pilosicoli is a zoonotic pathogen, able to infect different animal species such as pigs, poultry, and rodents, causing intestinal spirochetosis. An association of gastrointestinal clinical signs, such as diarrhea, with the isolation of B. pilosicoli from fecal samples or rectal swabs has not been proven in dogs. Other Brachyspira species commonly isolated from dogs, such as "B. canis" and "B. pulli", are considered commensals. This study investigated the occurrence of different Brachyspira species in rectal swabs and fecal samples in an independent canine cohort in central Germany. These included samples from shelter dogs, hunting dogs, and dogs presenting at regional small animal practices with various clinical signs. Data about the dogs, including potential risk factors for Brachyspira isolation, were obtained using a standardized questionnaire. The study also longitudinally investigated a colony of Beagle dogs for Brachyspira over 5 years. RESULTS: The rate of Brachyspira spp. isolation was 11% and included different Brachyspira species ("B. canis", "B. pulli", and B. pilosicoli). "B. canis" was detected in 18 dogs, whereas B. pilosicoli was only isolated from 1 dog in the independent cohort (not including the Beagle colony). Risk factors for shedding Brachyspira and "B. canis" were being less than 1 year of age and shelter origin. Gastrointestinal signs were not associated with the shedding of Brachyspira. B. pilosicoli and "B. canis" were isolated from several dogs of the same Beagle colony in 2017 and again in 2022, while Brachyspira was not isolated at multiple sampling time points in 2021. CONCLUSIONS: Shedding of B. pilosicoli in dogs appears to be uncommon in central Germany, suggesting a low risk of zoonotic transmission from dogs. Commensal status of "B. canis" and "B. pulli" is supported by the results of this study. Findings from the longitudinal investigation of the Beagle colony agree with an asymptomatic long-term colonization of dogs with "B. canis" and B. pilosicoli and suggest that introducing new animals in a pack can trigger an increased shedding of B. pilosicoli.

Analysis of gene and protein expression in the endometrium for validation of an ex vivo model of the equine uterus using PCR, digital and visual histopathology.

In the past, most research in equine reproduction has been performed in vivo but the use of in vitro and ex vivo models has recently increased. This study aimed to evaluate the functional stability of an ex vivo hemoperfused model for equine uteri with molecular characterization of marker genes and their proteins. In addition, the study validated the respective protein expression and the aptness of the software QuPath for identifying and scoring immunohistochemically stained equine endometrium. After collection, uteri (n = 12) were flushed with preservation solution, transported to the laboratory on ice, and perfused with autologous blood for 6 h. Cycle stage was determined by examination of the ovaries for presence of Graafian follicles or corpora lutea and analysis of plasma progesterone concentration (estrus: n = 4; diestrus: n = 4; anestrus: n = 4). Samples were obtained directly after slaughter, after transportation, and during perfusion (240, 300, 360 min). mRNA expression levels of progesterone (PGR), estrogen (ESR1) and oxytocin (OXTR) receptor as well as of MKI67 (marker of cell growth) and CASP3 (marker of apoptosis) were analyzed by RT-qPCR, and correlation to protein abundance was validated by immunohistochemical staining. Endometrial samples were analyzed by visual and computer-assisted evaluation of stained antigens via QuPath. For PGR, effects of the perfusion and cycle stage on expression were found (P < 0.05), while ESR1 was affected only by cycle stage (P < 0.05) and OXTR was unaffected by perfusion and cycle stage. MKI67 was lower after 360 min of perfusion as compared to samples collected before perfusion (P < 0.05). For CASP3, differences in gene expression were found after transport and samples taken after 240 min (P < 0.05). Immunohistochemical staining revealed effects of perfusion on stromal and glandular cells for steroid hormone receptors, but not for Ki-67 and active Caspase 3. OXTR was visualized in all layers of the endometrium and was unaffected by perfusion. Comparison of QuPath and visual analysis resulted in similar results. For most cell types and stained antigens, the correlation coefficient was r > 0.5. In conclusion, the isolated hemoperfused model of the equine uterus was successfully validated at the molecular level, demonstrating stability of key marker gene expression. The utility of computer-assisted immunohistochemical analysis of equine endometrial samples was also confirmed.

Cryptosporidium parvum infection alters the intestinal mucosa transcriptome in neonatal calves: implications for immune function.

One of the leading causes of infectious diarrhea in newborn calves is the apicomplexan protozoan Cryptosporidium parvum (C. parvum). However, little is known about its immunopathogenesis. Using next generation sequencing, this study investigated the immune transcriptional response to C. parvum infection in neonatal calves. Neonatal male Holstein-Friesian calves were either orally infected (N = 5) or not (CTRL group, N = 5) with C. parvum oocysts (gp60 subtype IIaA15G2R1) at day 1 of life and slaughtered on day 7 after infection. Total RNA was extracted from the jejunal mucosa for short read. Differentially expressed genes (DEGs) between infected and CTRL groups were assessed using DESeq2 at a false discovery rate < 0.05. Infection did not affect plasma immunohematological parameters, including neutrophil, lymphocyte, monocyte, leucocyte, thrombocyte, and erythrocyte counts as well as hematocrit and hemoglobin concentration on day 7 post infection. The immune-related DEGs were selected according to the UniProt immune system process database and were used for gene ontology (GO) and pathway enrichment analysis using Cytoscape (v3.9.1). Based on GO analysis, DEGs annotated to mucosal immunity, recognizing and presenting antigens, chemotaxis of neutrophils, eosinophils, natural killer cells, B and T cells mediated by signaling pathways including toll like receptors, interleukins, tumor necrosis factor, T cell receptor, and NF-KB were upregulated, while markers of macrophages chemotaxis and cytosolic pattern recognition were downregulated. This study provides a holistic snapshot of immune-related pathways induced by C. parvum in calves, including novel and detailed feedback and feedforward regulatory mechanisms establishing the crosstalk between innate and adaptive immune response in neonate calves, which could be utilized further to develop new therapeutic strategies.

[Zoonoses in endemic, free-ranging mammals]. / Zoonosen bei einheimischen Wildsäugetieren.

BACKGROUND: Zoonoses are diseases and infections that can be transmitted naturally between animals and humans. Direct and indirect contact of humans with wildlife occur during hunting activities, when diseased wildlife is found and treated, and in shared fields, forests, parks, gardens, and homes. Zoonoses can only be understood and controlled when ecosystems, animals, and humans are considered holistically. OBJECTIVE: This paper presents important zoonotic pathogens that are currently present in wild mammals as reservoirs in Germany. MATERIAL AND METHODS: The literature was searched to determine the prevalence of zoonotic pathogens currently occurring in wild mammals. RESULTS: Viral zoonotic agents currently present in free-ranging, mammalian animals in Germany as reservoirs of natural origin are bornaviruses, lyssaviruses, hepatitis E virus genotype 3, and Puumala orthohantavirus. Bacterial zoonotic agents beyond typical wound and foodborne pathogens include Brucella suis Biovar 2, Francisella tularensis ssp. holarctica, Leptospira interrogans sensu latu, Mycobacterium caprae, and Yersinia pseudotuberculosis. In particular, parasitic zoonotic agents common in wildlife are Alaria alata, Baylisascaris procyonis, Echinococcus multilocularis, Sacoptes scabei, and Trichinella spp. CONCLUSION: The presence of zoonotic infectious agents of risk groups 2 and 3 has to be regularly expected in numerous endemic wildlife species, especially canines, small bears, rodents, insectivores, and bats. Animal caretakers, hunters, veterinarians, and human health professionals should be aware of this risk and take protective measures appropriate to the situation.

Pyelonephritic end-stage kidney and ureterocutaneous fistula in a Shetland pony.

A 12-y-old Shetland Pony was presented with a mucus-secreting fistula in the right paralumbar fossa. Surgery was performed to unravel the origin of the fistula. The horse died under anesthesia and was forwarded to autopsy. The right kidney was markedly atrophic and fibrotic, consistent with unilateral end-stage kidney. The right ureter was markedly thickened, but with luminal continuity leading into the urinary bladder where a partial obstruction caused by nodular para-ureteral fat necrosis was evident. The lumen of the cutaneous fistula was continuous with the right ureter; therefore, we diagnosed the lesion as a ureterocutaneous fistula. Anomalies of the ureter are uncommon, and ureterocutaneous fistula formation in equids has not been reported previously to our knowledge.

Cryptosporidium parvumcompetes with the intestinal epithelial cells for glucose and impairs systemic glucose supply in neonatal calves.

Cryptosporidiosis is one of the main causes of diarrhea in children and young livestock. The interaction of the parasite with the intestinal host cells has not been characterized thoroughly yet but may be affected by the nutritional demand of the parasite. Hence, we aimed to investigate the impact of C. parvum infection on glucose metabolism in neonatal calves. Therefore, N = 5 neonatal calves were infected with C. parvum on the first day of life, whereas a control group was not (N = 5). The calves were monitored clinically for one week, and glucose absorption, turnover and oxidation were assessed using stable isotope labelled glucose. The transepithelial transport of glucose was measured using the Ussing chamber technique. Glucose transporters were quantified on gene and protein expression level using RT-qPCR and Western blot in the jejunum epithelium and brush border membrane preparations. Plasma glucose concentration and oral glucose absorption were decreased despite an increased electrogenic phlorizin sensitive transepithelial transport of glucose in infected calves. No difference in the gene or protein abundance of glucose transporters, but an enrichment of glucose transporter 2 in the brush border was observed in the infected calves. Furthermore, the mRNA for enzymes of the glycolysis pathway was increased indicating enhanced glucose oxidation in the infected gut. In summary, C. parvum infection modulates intestinal epithelial glucose absorption and metabolism. We assume that the metabolic competition of the parasite for glucose causes the host cells to upregulate their uptake mechanisms and metabolic machinery to compensate for the energy losses.

Comparison of systemic trimethoprim-sulfadimethoxine treatment and intrauterine ozone application as possible therapies for bacterial endometritis in equine practice.

Bacterial endometritis is one of the major problems in equine reproduction and usually treated with antimicrobial drugs. The study aimed to compare the effects of intrauterine ozone application and systemic antibiotic treatment (trimethoprim-sulfadimethoxine) on intrauterine bacterial growth and possible side effects on the endometrium in a clinical setting. Mares (n = 30) with signs of endometritis (positive uterine bacterial culture and cytological findings) were assigned randomly to different treatments: intrauterine insufflation of an ozone-air-mix (240 ml, 80 µg ozone/ml) twice at a 48 h-interval (Ozone; n = 10), systemic antibiotic therapy with trimethoprim-sulfadimethoxine (30 mg/kg, p.o., twice daily) for 5 days (TMS; n = 10), or intrauterine insufflation of air (240 ml, sterile-filtered) twice at a 48 h-interval (air; n = 10). Endometrial biopsy for histological examination was obtained before the treatment. Histological examination revealed no differences among groups. A control examination, including transrectal ultrasound, bacterial culture, cytological evaluation, and biopsy, was performed 7 days after the last treatment. Overall bacterial growth was reduced in every group after the treatment (p < 0.05), irrespective of the therapy [Ozone: 4/9 (positive culture after treatment/number of mares), TMS: 3/10 and Air: 6/10; p > 0.05]. However, Ozone and TMS (p < 0.05) were more effective in reducing growth of gram-negative bacteria as compared to Air (p > 0.05). No effects on the number of polymorphonuclear granulocytes (cytology) were observed (p > 0.05). In conclusion, trimethoprim-sulfadimethoxine and intrauterine ozone insufflation are safe treatment options for bacterial endometritis in mares but the efficacy of both treatments in reducing bacterial growth did not result in a complete absence of intrauterine bacterial growth.

[West-Nile-Virus infections in 12 horses in east-central Germany]. / West-Nil-Virus-Infektion bei 12 Pferden in Mitteldeutschland.

PURPOSE AND AIM: The presenting complaints, clinical signs, diagnostic evaluation, therapy, and outcome of 12 horses with clinically apparent West-Nile-Virus (WNV) infection are described. MATERIAL AND METHODS: Case series RESULTS: The adult horses (age 6-18 years, 7 mares, 5 geldings) from Saxony and Saxony-Anhalt were presented with various clinical histories between September 2018 and September 2020. All horses were presented in August or September and no horse was vaccinated against WNV. Fever as the most common general clinical sign was present in 8/12 horses. The most common neurological signs were muscle fasciculations (11/12 horses), ataxia (8/12 horses), hyperesthesia and head tilt (6/12 horses each). Diagnosis of WNV infection was confirmed by demonstrating IgM antibody and neutralizing antibody production in all horses; 2 euthanized horses also tested positive by PCR. Therapy was symptomatic and primarily included non-steroidal anti-inflammatories or dexamethasone as well as fluid therapy. Duration of hospitalization was 7.5 days on average. According to their owners, seven horses recovered completely, while information was missing for 2 horses. CONCLUSIONS AND CLINICAL RELEVANCE: In eastern-central Germany, WNV-encephalomyelitis must be considered a differential diagnosis for unvaccinated horses with acute neurologic disease occurring in summer and late summer. The reported clinical signs and the outcome of therapy are mostly congruent with reports from North America and other European countries.

A new S. suis serotype 3 infection model in pigs: lack of effect of buprenorphine treatment to reduce distress.

BACKGROUND: Streptoccocus suis (S. suis) is a major porcine pathogen causing meningitis, septicemia, arthritis and endocarditis. These diseases severely impair welfare of pigs. Experimental studies in pigs are important to better understand the pathogenesis and to identify protective antigens, as so far there is no vaccine available protecting against various serotypes (cps). Due to the severity of disease, application of appropriate refinement strategies in experimental S. suis infections is essential to reduce distress imposed on the piglets without jeopardizing the scientific output. The objectives of this study were to evaluate buprenorphine treatment as a refinement measure and serum cortisol levels as a distress read out parameter in a new S. suis cps3 infection model in pigs. RESULTS: Intravenous application of 2 × 108 CFU of S. suis cps3 (sly+, mrp+) to 6-week-old piglets led to severe morbidity in approximately 50% of the animals. Main pathological findings included suppurative meningoencephalitis and arthritis as well as fibrinosuppurative endocarditis. Buprenorphine treatment (0.05 mg/kg every 8 h) did not prevent signs of severe pain, high clinical scores, moderate to severe pathologies or high levels of serum cortisol in single severely affected piglets. Significant differences in the course of leukocytosis, induction of specific antibodies and bactericidal immunity were not recorded between groups with or w/o buprenorphine treatment. Of note, clinically unobtrusive piglets showed serum cortisol levels at 2 and 5 days post infectionem (dpi) comparable to the levels prior to infection with cps3. Cortisol levels in serum were significantly increased in piglets euthanized due to severe disease in comparison to clinically unobtrusive pigs. CONCLUSIONS: Different clinical courses and pathologies are induced after intravenous challenge of piglets with 2 × 108 CFU of this S. suis cps3 strain. The chosen protocol of buprenorphine application does not prevent severe distress in this infection model. Important parameters of the humoral immune response, such as the level of IgM binding to S. suis cps3, do not appear to be affected by buprenorphine treatment. Serum cortisol is a meaningful parameter to measure distress in piglets experimentally infected with S. suis and to evaluate refinement strategies. In this intravenous model, which includes close clinical monitoring and different humane endpoints, clinics and cortisol levels suggest convalescence in surviving piglets within 5 days following experimental infection.

Rift Valley Fever Virus Non-Structural Protein S Is Associated with Nuclear Translocation of Active Caspase-3 and Inclusion Body Formation.

Rift Valley fever phlebovirus (RVFV) causes Rift Valley fever (RVF), an emerging zoonotic disease that causes abortion storms and high mortality rates in young ruminants as well as severe or even lethal complications in a subset of human patients. This study investigates the pathomechanism of intranuclear inclusion body formation in severe RVF in a mouse model. Liver samples from immunocompetent mice infected with virulent RVFV 35/74, and immunodeficient knockout mice that lack interferon type I receptor expression and were infected with attenuated RVFV MP12 were compared to livers from uninfected controls using histopathology and immunohistochemistry for RVFV nucleoprotein, non-structural protein S (NSs) and pro-apoptotic active caspase-3. Histopathology of the livers showed virus-induced, severe hepatic necrosis in both mouse strains. However, immunohistochemistry and immunofluorescence revealed eosinophilic, comma-shaped, intranuclear inclusions and an intranuclear (co-)localization of RVFV NSs and active caspase-3 only in 35/74-infected immunocompetent mice, but not in MP12-infected immunodeficient mice. These results suggest that intranuclear accumulation of RVFV 35/74 NSs is involved in nuclear translocation of active caspase-3, and that nuclear NSs and active caspase-3 are involved in the formation of the light microscopically visible inclusion bodies.

Video Endoscopy-Guided Intrabronchial Spray Inoculation of Mycobacterium bovis in Goats and Comparative Assessment of Lung Lesions With Various Imaging Methods.

Bovine tuberculosis (bTB) not only poses a zoonotic threat to humans but also has a significant economic impact on livestock production in many areas of the world. Effective vaccines for humans, livestock, and wildlife are highly desirable to control tuberculosis. Suitable large animal models are indispensable for meaningful assessment of vaccine candidates. Here, we describe the refinement of an animal model for bTB in goats. Intrabronchial inoculation procedure via video-guided endoscopy in anesthetized animals, collection of lungs after intratracheal fixation in situ, and imaging of lungs by computed tomography (CT) were established in three goats using barium sulfate as surrogate inoculum. For subsequent infection experiments, four goats were infected with 4.7 × 102 colony-forming units of M. bovis by intrabronchial inoculation using video-guided endoscopy with spray catheters. Defined amounts of inoculum were deposited at five sites per lung. Four age-matched goats were mock-inoculated. None of the goats developed clinical signs until they were euthanized 5 months post infection, but simultaneous skin testing confirmed bTB infection in all goats inoculated with M. bovis. In tissues collected at necropsy, M. bovis was consistently re-isolated from granulomas in lymph nodes, draining the lungs of all the goats infected with M. bovis. Further dissemination was observed in one goat only. Pulmonary lesions were quantified by CT and digital 2D radiography (DR). CT revealed mineralized lesions in all the infected goats ranging from <5 mm to >10 mm in diameter. Small lesions <5 mm predominated. The DR failed to detect small lesions and to determine the exact location of lesions because of overlapping of pulmonary lobes. Relative volume of pulmonary lesions was low in three but high in one goat that also had extensive cavitation. CT lesions could be correlated to gross pathologic findings and histologic granuloma types in representative pulmonary lobes. In conclusion, video-guided intrabronchial inoculation with spray catheters, mimicking the natural way of infection, resulted in pulmonary infection of goats with M. bovis. CT, but not DR, presented as a highly sensitive method to quantify the extent of pulmonary lesions. This goat model of TB may serve as a model for testing TB vaccine efficacy.

Bone quality and composition are influenced by egg production, layer line, and oestradiol-17ß in laying hens.

Keel bone fractures are a serious animal welfare problem in laying hens. The aim of the current study was to assess the influence of egg production, oestradiol-17ß, and selection for high laying performance on bone quality. Hens of two layer lines differing in laying performance (WLA: 320 eggs per year, G11: 200 eggs per year) were allocated to four treatment groups. Group S received a deslorelin acetate implant that suppressed egg production. Group E received an implant with the sexual steroid oestradiol-17ß. Group SE received both implants and group C did not receive any implant. In the 63rd week of age, composition and characteristics of the tibiotarsi were assessed using histological analysis, three-point bending test, thermogravimetric analysis, infrared spectroscopy, and two-dimensional X-ray diffraction, respectively. Non-egg-laying hens showed a higher total bone area and a higher relative amount of cortical bone compared to egg-laying hens. Hens of layer line G11 showed a higher relative amount of medullary bone and a higher degree of mineralization of the cortical bone compared to hens of layer line WLA. These differences in bone composition may explain different susceptibility to keel bone fractures in non-egg-laying compared to egg-laying hens as well as in hens of layer lines differing in laying performance. The effect of exogenous oestradiol-17ß on bone parameters varied between the layer lines indicating a genetic influence on bone physiology and the way it can be modulated by hormone substitution.

Evaluation of an ex vivo model of the blood-perfused equine uterus.

Uterine pathologies are the most common causes of infertility in mares. This study aimed to establish an ex vivo blood-perfused model for equine uteri and investigate the possible effects of different cycle stages (estrus, diestrus and anestrus) on the applicability of the model. Uteri (n = 13) were collected at an abattoir, flushed with preservation solution, transported to the laboratory on ice, and isolated perfused with autologous blood for 6 h (n = 12). For negative control, one uterus was handled as described but left without perfusion for 6 h. The cycle stage was determined by examination of the ovaries for the presence of Graafian follicles or corpora lutea and analysis of plasma progesterone concentration (estrus: n = 4; diestrus: n = 4; anestrus: n = 4). Sonomicrometry crystals were implanted into the myometrium to record spontaneous contractions and the response to 0.5 IU oxytocin after 6 h of perfusion. Analyses of the arterial and venous perfusate were performed every hour to determine glucose consumption, lactate production, pH, lactate dehydrogenase activity (LDH), and potassium concentration (K+). Biopsy samples were obtained directly after slaughter, after transportation, after equilibration, after 4, 5, and 6 h of perfusion, and immediately after removal from the perfusion system. The uteri's glucose consumption and lactate production increased over time (p < 0.05), but no differences among cycle stages were detected. pH (arterial and venous) increased over time (p < 0.05). No changes for LDH were observed. K+ increased after 4 h of perfusion (p < 0.05), but was unaffected by the cycle stage. Spontaneous contractions were present in all perfused uteri, but myometrial activity in the negative control was limited to the 2nd hour of perfusion. No effects of cycle stage on contraction amplitude and duration after oxytocin administration were detected. The cycle stage did not affect frequency (except after 5 h of perfusion), amplitude, duration of contractions, or edema formation. Histology revealed congestion of endometrial capillaries after 4, 5, and 6 h perfusion time. In conclusion, the ex vivo model was capable of supporting the functionality of equine uteri for 6 h. However, viability and histomorphology of the endometrium appeared to be impaired after 4 h of perfusion. Effects of the cycle stage on the applicability of the model were absent.

NSG-Mice Reveal the Importance of a Functional Innate and Adaptive Immune Response to Overcome RVFV Infection.

Rift Valley fever (RVF) is a zoonotic disease caused by RVF Phlebovirus (RVFV). The RVFV MP-12 vaccine strain is known to exhibit residual virulence in the case of a deficient interferon type 1 response. The hypothesis of this study is that virus replication and severity of lesions induced by the MP-12 strain in immunocompromised mice depend on the specific function of the disturbed pathway. Therefore, 10 strains of mice with deficient innate immunity (B6-IFNARtmAgt, C.129S7(B6)-Ifngtm1Ts/J, B6-TLR3tm1Flv, B6-TLR7tm1Aki, NOD/ShiLtJ), helper T-cell- (CD4tm1Mak), cytotoxic T-cell- (CD8atm1Mak), B-cell- (Igh-Jtm1DhuN?+N2), combined T- and B-cell- (NU/J) and combined T-, B-, natural killer (NK) cell- and macrophage-mediated immunity (NOD.Cg-PrkdcscidIl2rgtm1WjI/SzJ (NSG) mice) were subcutaneously infected with RVFV MP-12. B6-IFNARtmAgt mice were the only strain to develop fatal disease due to RVFV-induced severe hepatocellular necrosis and apoptosis. Notably, no clinical disease and only mild multifocal hepatocellular necrosis and apoptosis were observed in NSG mice, while immunohistochemistry detected the RVFV antigen in the liver and the brain. No or low virus expression and no lesions were observed in the other mouse strains. Conclusively, the interferon type 1 response is essential for early control of RVFV replication and disease, whereas functional NK cells, macrophages and lymphocytes are essential for virus clearance.

Black rats (Rattus rattus) as potential reservoir hosts for Rift Valley fever phlebovirus: Experimental infection results in viral replication and shedding without clinical manifestation.

Rift Valley fever phlebovirus (RVFV) is an arthropod-borne virus that can cause severe disease in ruminants and humans. Epidemics occur mainly after heavy rainfall, which leads to a significant increase in the occurrence of RVFV-transmitting mosquitoes. During inter-epidemic periods, the virus is assumed to be maintained between mosquitoes, susceptible livestock and yet unknown wildlife. The widespread rodent Rattus rattus (black rat) has been suspected to be involved in RVFV maintenance. In order to elucidate its susceptibility and thus its possible role in the transmission cycle of the virus, an experimental infection study was performed. Black rats were subcutaneously infected with highly virulent RVFV strain 35/74 and euthanized on days 3, 14 and 28 post-infection. Additional black rats served as non-infected contact animals. The infected black rats showed high susceptibility to RVFV infection. Generation of RVFV-neutralizing antibodies was found, and the rats developed viraemias lasting up to 17 days. Viral RNA was found in tissues until the last day of the experiment. However, neither a clinical manifestation nor virus-induced histopathological lesions were observed in any rat. These findings indicate the persistence of RVFV in black rats without affecting the animals. In contact animals, no evidence of horizontal RVFV transmission was found, although the co-housed infected rats showed oral, rectal and conjunctival RVFV shedding. Results of this study point to an involvement of black rats in the RVFV transmission cycle, and further studies are needed to investigate their potential role in the maintenance of the virus.