Sequestration of membrane cholesterol by cholesterol-binding proteins inhibits SARS-CoV-2 entry into Vero E6 cells.

Membrane lipids and proteins form dynamic domains crucial for physiological and pathophysiological processes, including viral infection. Many plasma membrane proteins, residing within membrane domains enriched with cholesterol (CHOL) and sphingomyelin (SM), serve as receptors for attachment and entry of viruses into the host cell. Among these, human coronaviruses, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), use proteins associated with membrane domains for initial binding and internalization. We hypothesized that the interaction of lipid-binding proteins with CHOL in plasma membrane could sequestrate lipids and thus affect the efficiency of virus entry into host cells, preventing the initial steps of viral infection. We have prepared CHOL-binding proteins with high affinities for lipids in the plasma membrane of mammalian cells. Binding of the perfringolysin O domain four (D4) and its variant D4E458L to membrane CHOL impaired the internalization of the receptor-binding domain of the SARS-CoV-2 spike protein and the pseudovirus complemented with the SARS-CoV-2 spike protein. SARS-CoV-2 replication in Vero E6 cells was also decreased. Overall, our results demonstrate that the integrity of CHOL-rich membrane domains and the accessibility of CHOL in the membrane play an essential role in SARS-CoV-2 cell entry.

SARS-CoV-2 molecular epidemiology in Slovenia, January to September 2021.

BackgroundSequencing of SARS-CoV-2 PCR-positive samples was introduced in Slovenia in January 2021. Our surveillance programme comprised three complementary schemes: (A) non-targeted sequencing of at least 10% of samples, (B) sequencing of samples positive after PCR screening for variants of concern (VOC) and (C) sequencing as per epidemiological indication.AimWe present the analysis of cumulative data of the non-targeted surveillance of SARS-CoV-2 and variant-dependent growth kinetics for the five most common variants in Slovenia for the first 9 months of 2021.MethodsSARS-CoV-2 PCR-positive samples, from January to September 2021, were selected for sequencing according to the national surveillance plan. Growth kinetics studies were done on Vero E6 cells.ResultsAltogether 15,175 genomes were sequenced and 64 variants were detected, of which three successively prevailed. Variant B.1.258.17 was detected in ca 80% of samples in January and was replaced, within 9 weeks, by the Alpha variant. The number of cases decreased substantially during the summer of 2021. However, the introduction of the Delta variant caused a fourth wave and completely outcompeted other variants. Other VOC were only detected in small numbers. Infection of Vero E6 cells showed higher replication rates for the variants Alpha and Delta, compared with B.1.258.17, B.1.258, and B.1.1.70, which dominated in Slovenia before the introduction of the Alpha and Delta variants.ConclusionInformation on SARS-CoV-2 variant diversity provided context to the epidemiological data of PCR-positive cases, contributed to control of the initial spread of known VOC and influenced epidemiological measures.

TXM peptides inhibit SARS-CoV-2 infection, syncytia formation, and lower inflammatory consequences.

After three years of the SARS-CoV-2 pandemic, the search and availability of relatively low-cost benchtop therapeutics for people not at high risk for a severe disease are still ongoing. Although vaccines and new SARS-CoV-2 variants reduce the death toll, the long COVID-19 along with neurologic symptoms can develop and persist even after a mild initial infection. Reinfections, which further increase the risk of sequelae in multiple organ systems as well as the risk of death, continue to require caution. The spike protein of SARS-CoV-2 is an important target for both vaccines and therapeutics. The presence of disulfide bonds in the receptor binding domain (RBD) of the spike protein is essential for its binding to the human ACE2 receptor and cell entry. Here, we demonstrate that thiol-reducing peptides based on the active site of oxidoreductase thioredoxin 1, called thioredoxin mimetic (TXM) peptides, can prevent syncytia formation, SARS-CoV-2 entry into cells, and infection in a mouse model. We also show that TXM peptides inhibit the redox-sensitive HIV pseudotyped viral cell entry. These results support disulfide targeting as a common therapeutic strategy for treating infections caused by viruses using redox-sensitive fusion. Furthermore, TXM peptides exert anti-inflammatory properties by lowering the activation of NF-κB and IRF signaling pathways, mitogen-activated protein kinases (MAPKs) and lipopolysaccharide (LPS)-induced cytokines in mice. The antioxidant and anti-inflammatory effects of the TXM peptides, which also cross the blood-brain barrier, in combination with prevention of viral infections, may provide a beneficial clinical strategy to lower viral infections and mitigate severe consequences of COVID-19.

Rehoming and Other Refinements and Replacement in Procedures Using Golden Hamsters in SARS-CoV-2 Vaccine Research.

Effective vaccines are needed to fight the COVID-19 pandemic. Forty golden hamsters were inoculated with two promising vaccine candidates and eighteen animals were used in pilot trials with viral challenge. ELISA assays were performed to determine endpoint serum titres for specific antibodies and virus neutralisation tests were used to evaluate the efficacy of antibodies. All tests with serum from vaccinated hamsters were negative even after booster vaccinations and changes in vaccination protocol. We concluded that antibodies did not have sufficient neutralising properties. Refinements were observed at all steps, and the in vitro method (virus neutralisation test) presented a replacement measure and ultimately lead to a reduction in the total number of animals used in the project. The institutional animal welfare officer and institutional designated veterinarian approved the reuse or rehoming of the surplus animals. Simple socialization procedures were performed and ultimately 19 animals were rehomed, and feedback was collected. Recently, FELASA published recommendations for rehoming of animals used for scientific and educational purposes, with species-specific guidelines, including mice, rats, and rabbits. Based on our positive experience and feedback from adopters, we concluded that the rehoming of rodents, including hamsters, is not only possible, but highly recommended.

Prevalence of pigeon circovirus infections in feral pigeons in Ljubljana, Slovenia.

Pigeon circovirus (PiCV) was detected by real-time PCR in cloacal swabs, pharyngeal swabs, and serum samples taken from 74 feral pigeons (Columba livia var. domestica) that were caught at various locations in the city of Ljubljana, Slovenia. PiCV infections were detected in the majority of the tested birds. The highest (74.3%) detection rate was observed in the cloacal swabs and the lowest (31.1%) in serum samples. PiCV DNA was more readily detected in the cloacal swabs, pharyngeal swabs, and serum samples of birds younger than 1 yr. Molecular analysis of partial open reading frame V1 sequences showed that PiCV strains detected in feral pigeons share high nucleotide and amino acid sequence identities with PiCV strains detected in ornamental, racing, meat, and feral pigeons.

Surveillance of influenza A viruses in wild birds in Slovenia from 2006 to 2010.

Within the framework of the surveillance program for the early detection of H5 and H7 subtypes of avian influenza (AI) viruses, samples from 2547 wild birds of different species that were collected between 2006 and 2010 were examined by PCR-based methods. AI viruses of various subtypes were detected in 4.4% of birds from four different orders: Anseriformes, Ciconiiformes, Charadriiformes, and Pelecaniformes. Highly pathogenic avian influenza (HPAI) H5N1 viruses were detected only in 2006. HPAI H5N1 virus was confirmed in 1.9% of birds from four different species. Comparison of nucleotide sequences of the H5N1 hemagglutinin gene indicated that two different HPAI H5N1 viruses from the European-Middle Eastern-African clade 1 had been introduced into Slovenia, despite the relatively short duration of the HPAI outbreak. Low pathogenic avian influenza (LPAI) viruses were detected in 2.5% of birds during a 5-yr period. The subtypes H1, H2, H3, H4, H5, H7N7, H8, H10, H11, and H13N6 were determined in 18 out of 64 cases. The highest prevalence (81%) of LPAI viruses, including the H5 subtype, were found in birds sampled as a part of the "active" surveillance system.

Effects of Coenzyme Q10 Supplementation on Oxidative Stress Markers, Inflammatory Markers, Lymphocyte Subpopulations, and Clinical Status in Dogs with Myxomatous Mitral Valve Disease.

Scarce data exist on the effects of coenzyme Q10 (CoQ10) supplementation in dogs with myxomatous mitral valve disease (MMVD). The purpose of this study was to investigate the effect of CoQ10 supplementation on oxidative stress markers (glutathione peroxidase, F2-isoprostanes), markers of inflammation (tumor necrosis factor-α, TNF soluble receptor II, leucocytes, and their subtypes), lymphocyte subpopulations (T helper and cytotoxic T lymphocytes, including activated T lymphocytes, and B lymphocytes), and echocardiographic and clinical parameters in dogs with MMVD. In this randomized, controlled, double-blind, longitudinal study, 43 MMVD dogs in stages ACVIM (American College of Veterinary Internal Medicine classification) B2 and ACVIM C and D (congestive heart failure (CHF)) received water-soluble coenzyme Q10 (100 mg twice daily) or placebo for 3 months, and 12 non-supplemented healthy dogs served as controls. All parameters were measured before and after supplementation in MMVD dogs and once in healthy dogs. CoQ10 supplementation had a positive impact on neutrophil percentage, lymphocyte percentage, and lymphocyte concentration in our cohort of dogs with CHF (ACVIM C and D). Conclusion: CoQ10 as an oral supplement may have benefits in terms of decreasing inflammation in dogs with MMVD and CHF.

Effect of intrabronchial administration of autologous adipose-derived mesenchymal stem cells on severe equine asthma.

BACKGROUND: Severe equine asthma (SEA) is a common chronic respiratory disease and a significant health and well-being problem in horses. Current therapeutic strategies improve pulmonary function and clinical signs in some horses, but in the long-term, return to full athletic function appears to be rare. The aim of this study was to assess the safety and the effect of intrabronchial administration of adipose-derived mesenchymal stem cells (AD-MSC) on pulmonary inflammatory and clinical parameters in horses with SEA. METHODS: This was a randomized controlled trial. Twenty adult horses diagnosed with SEA were randomly divided into two groups (n = 10), and treated either with a single intrabronchial application of autologous AD-MSC or oral dexamethasone for three weeks. A targeted clinical examination with determination of clinical score, maximal change in pleural pressure during the breathing cycle, and an endoscopic examination of the airways were performed at baseline and three weeks after treatment. Bronchoalveolar lavage fluid was analyzed cytologically, and IL-1ß, IL-4, IL-8, IL-17, TNFα and IFNγ mRNA and protein concentrations were measured at baseline and three weeks. The horses were then monitored over one year for recurrence of SEA. A non-inferiority analysis and a linear mixed-effects model were performed to assess differences between treatments. RESULTS: The non-inferiority of AD-MSC treatment was not established. However, AD-MSC administration significantly ameliorated the clinical score (P = 0.01), decreased the expression of IL-17 mRNA (P = 0.05) and IL-1ß (P ≤ 0.001), IL-4 (P ≤ 0.001), TNFα (P = 0.02) protein levels, and had a positive long-term effect on SEA-associated clinical signs (P = 0.02). CONCLUSIONS: Intrabronchial administration of AD-MSC had limited short-term anti-inflammatory effects but improved the clinical signs of SEA at one year.

Circulation of infectious bronchitis virus strains from Italy 02 and QX genotypes in Slovenia between 2007 and 2009.

Fourteen infectious bronchitis viruses (IBVs) detected in broiler, broiler breeder, and layer flocks in Slovenia between 2007 and 2009 were molecularly analyzed by reverse transcription-PCR and direct sequencing of the S1 gene. Phylogenetic analyses based on partial S1 gene sequences revealed that seven strains were classified as the Italy 02 genotype, a genetic group of IBV that has not previously been detected in Slovenia. Another seven strains were classified as the QX genotype. The results of phylogenetic analyses and epidemiologic investigations revealed that the genetic classification correlates with the geographic origins of the IBV strains. Greater genetic variability (maximum nucleotide and amino acid divergences were 4.8% and 9.9%, respectively) was observed within the Slovenian strains from the Italy 02 genotype detected between 2007 and 2009 than within strains from the QX genotype isolated in 2008 and 2009 (1.2% and 1.3%, respectively). The Slovenian strains classified as the Italy 02 genotype form a separate genetic cluster. These strains shared five specific amino acid substitutions that became fixed during the period of surveillance. Strains from the QX genotype that shared one specific amino acid substitution also form a separate genetic cluster.

Field efficacy of different vaccines against infectious bursal disease in broiler flocks.

A field study was performed to determine the efficacy of three commercially available vaccines against infectious bursal disease (IBD) in commercial broilers raised in a high IBD virus (IBDV) risk area. Live attenuated intermediate and intermediate plus vaccines were used in four flocks. Birds were vaccinated orally at the estimated vaccination time. Three broiler flocks were vaccinated subcutaneously with a turkey herpesvirus (HVT)-IBD vector vaccine at one day old. Evaluation of the efficacy of different vaccines was focused on humoral immune response, bursa/body weight (B/Bw) ratio, molecular detection of IBDV in ileocaecal tonsils and bursa of Fabricius, and production parameters. The serological results showed that although the uptake of all three vaccine strains was confirmed in the lymphoid organs, no significant antibody response to vaccination was detected in flocks vaccinated with intermediate and intermediate plus vaccines. A significant increase in antibody titres detected in flocks vaccinated with the vector vaccine indicated its ability to induce an immune response in birds with a high level of maternally derived antibodies. Observations obtained in this field trial did not confirm the expected reduction of the B/Bw ratio in flocks vaccinated with less attenuated vaccines. No significant differences were observed between birds vaccinated with the vector vaccine and those immunised with the intermediate plus vaccine. Very virulent IBDV was confirmed in the flock vaccinated with the intermediate vaccine. The infection induced reduced B/Bw and moderate mortality but did not affect the production parameters. Field infection was not detected in broilers vaccinated with the intermediate plus vaccine and the vector vaccine.

Detection of Laryngotracheitis Virus in Poultry Flocks with Respiratory Disorders in Slovenia.

Infectious laryngotracheitis (ILT) is an acute, highly contagious infectious disease of the upper respiratory tract in chickens and other poultry species that causes significant economic losses in countries worldwide. Between 2017 and 2019, seven outbreaks of mild to severe respiratory disorders with high suspicion of ILT occurred in commercial and backyard poultry flocks in Slovenia. In all submissions, infection with ILT virus (ILTV) was confirmed by PCR, which is the first report of ILT in Slovenia. Circulating ILT strains were characterized by the sequence and phylogenetic analysis of two fragments of the ICP4 gene. Four strains-three detected in non-vaccinated flocks and one in a flock vaccinated against ILT-were identical or very similar to the chicken embryo-origin live virus vaccines, and the other three were closely related to Russian, Chinese, Australian, and American field strains and to tissue culture origin vaccine strains. As in other diseases, coinfections with other respiratory pathogens in confirmed ILT cases may cause a more severe condition and prolong the course of the disease. In our study, coinfections with Mycoplasma synoviae (7/7 tested flocks), infectious bronchitis virus (5/5 tested flocks), Mycoplasma gallisepticum (4/7 tested flocks), Ornithobacterium rhinotracheale (3/4 tested flocks), and avian pox virus (1/2 tested flocks) were confirmed, indicating the importance of these pathogens in the occurrence of ILT infections.

DETECTION OF HERPESVIRUSES IN PASSERINE BIRDS CAPTURED DURING AUTUMN MIGRATION IN SLOVENIA.

Herpesviruses (HVs) were detected by PCR in the cloacal swabs of 0.76% (4/525) clinically healthy free-living passerine birds from 32 different species captured in mist nets in Slovenia during the 2014 and 2017 autumn migrations. Herpesviruses were detected in the Eurasian Blackcap (Sylvia atricapilla), the Common Blackbird (Turdus merula), and the Eurasian Blue Tit (Cyanistes caeruleus). Phylogenetic analysis of partial DNA polymerase gene nucleotide sequences of the HV strains showed a distant relationship with other alphaherpesviruses of birds. In the phylogenetic tree, the HVs detected were clustered together with HV detected in Sulphur-crested Cockatoo and Neotropic Cormorants, as well as with known HVs such as gallid HV1, psittacid HV1 and HV2, and passerine HV1. Different sequences of HVs with relatively low identity were detected in our study, suggesting that different HVs were circulating in passerines sampled during the autumn migration in Slovenia.

Health Status and Stress in Different Categories of Racing Pigeons.

The influence of different stress parameters in racing pigeon flocks, such as the presence of diseases and environmental conditions at the time of the races, were described. A total of 96 racing pigeons from 4 pigeon flocks were examined, and health monitoring was carried out. No helminth eggs and coccidia were found. Trichomonas sp. was confirmed in subclinical form. Paramyxoviruses and avian influenza viruses were not confirmed, but circovirus infections were confirmed in all flocks. Chlamydia psittaci was confirmed in one flock. Blood samples were collected, and HI antibody titers against paramyxoviruses before and 25 days after vaccination were determined. To improve the conditions during racing and the welfare of the pigeons, critical points were studied with regard to stress factors during the active training season. Serum corticosterone levels were measured in the blood serum of four different categories of pigeons from each flock. Corticosterone levels were almost twice as high in pigeons from the category that were active throughout the racing season, including medium- and long-distance racing, compared to the other three categories that were not racing actively. Within five hours of the finish of a race, the average serum corticosterone level was 59.4 nmol/L in the most physically active category. The average serum corticosterone level in this category remained at 37.5 nmol/L one month after the last race.

Evaluation of Welfare in Commercial Turkey Flocks of Both Sexes Using the Transect Walk Method.

The study was conducted between March and September 2019 in six meat-type turkey flocks with similar management standard procedures using the transect walk method. The concept of the method is based on visual observation of the birds while slowly walking across the entire farm in predetermined transects. Each flock was evaluated at three different times during the fattening cycle: at 3 to 4, 12 to 13, and 19 to 20 weeks of age, and total number of males and females that were immobile or lame, had visible head, vent, or back wounds, were small, featherless, dirty, or sick, had pendulous crop, or showed aggression toward birds or humans were recorded. At each visit, NH3 and CO2 were measured within the facilities. In the first assessment, the most frequently observed welfare indicators were small size (0.87%) and immobility (0.08%). Males showed a significantly higher prevalence of small size (p < 0.01), sickness (p < 0.05), and dirtiness (p < 0.1) compared to females. In the second assessment, the most common findings in both sexes were dirtiness (1.65%) and poor feather condition (1.06%), followed by immobility (0.28%). Males were significantly dirtier (p < 0.001), had more immobile birds (p < 0.01) and birds with vent wounds (p < 0.1), but had fewer sick birds (p < 0.05). In the last assessment, an increase in immobile, lame, sick, and dead birds was recorded, indicating an increase in health problems. Higher CO2 (3000 and 4433 ppm) and NH3 (40 and 27.6 ppm) values were noted only at the first assessment in two facilities. Further analyses showed that slightly elevated NH3 and CO2 levels did not influence the occurrence of welfare indicators. This study is the first description of the welfare of commercial turkey flocks in Slovenia.

Molecular analysis of infectious bronchitis viruses isolated in Slovenia between 1990 and 2005: a retrospective study.

Fifteen infectious bronchitis viruses (IBV) isolated from broiler and broiler breeder flocks in Slovenia between 1990 and 2005 were molecularly characterised. IBV strains were divided into four genotypes by the analysis of the S1 gene region. Four strains belonged to the Massachusetts genotype, one strain was placed into the QX genotype, one strain formed a cluster together with the B1648 strain and nine strains were classified into the 624/I genotype. Nine Slovenian strains of the 624/I genotype formed two subgroups independently of the time of isolation and the geographical origin. Phylogenetic analysis of the partial N gene sequences revealed lower sequence variability and different clustering of the Slovenian IBV. Fourteen strains were grouped together with the strains H120 and D1466. One strain formed a cluster with the strain 793/B.

Molecular characterization of avian paramyxovirus type 1 (Newcastle disease) viruses isolated from pigeons between 2000 and 2008 in Slovenia.

Fourteen avian paramyxovirus type 1 (APMV-1; Newcastle disease) viruses isolated from dead free-living and domestic pigeons in Slovenia between 2000 and 2008 were analyzed by a molecular characterization of a part of the fusion protein gene, which included the region encoding the fusion protein cleavage site. Phylogenetic analysis indicated that the Slovene pigeon paramyxovirus type 1 (PPMV-1) viruses do not cluster together but instead are divided into two groups--4bi and 4bii--of sublineage 4b. Nine Slovenian strains were placed in group 4bii. Five other strains clustered together with PPMV-1 from group 4bi. The sequence of the fusion protein cleavage site of all Slovenian strains was typical for pathogenic APMV-1. The 112RRQKRF117 motif was present in the strains from group 4bii, whereas strains from group 4bi displayed the 112GRQKRF117 motif.

Loop-mediated isothermal amplification: rapid molecular detection of virulence genes associated with avian pathogenic Escherichia coli in poultry.

Infections with pathogenic Escherichia coli can lead to different animal- and human-associated diseases. E. coli infections are common in intensive poultry farming, and important economic losses can be expected during infections with avian pathogenic E. coli (APEC) strains followed by colibacillosis. Loop-mediated isothermal amplification (LAMP) assays were developed for rapid detection of 3 APEC-associated virulence genes: sitA, traT, and ompT. All 3 LAMP assays are shown to be specific, repeatable, and reproducible. High sensitivities of the assays are shown, where as few as 1,000 bacterial cells/mL can be detected in different matrices. On-site applicability of this LAMP method is demonstrated through testing of different sample types, from animal swabs and tissues, and from environmental samples collected from 6 commercial poultry farms. All 3 virulence genes were detected at high rates (above 85%) in samples from layer and broiler chickens with clinical signs and, interestingly, high prevalence of those genes was detected also in samples collected from clinically healthy broiler flock (above 75%) while lower prevalence was observed in remaining 3 clinically healthy chicken flocks (less than 75%). Importantly, these virulence genes were detected in almost all of the air samples from 11 randomly selected poultry houses, indicating air as an important route of E. coli spread. Three LAMP assays that target APEC-associated virulence genes are shown to be sensitive and robust and are therefore applicable for rapid on-site testing of various sample types, from animal swabs to air. This on-site LAMP testing protocol offers rapid diagnostics, with results obtained in <35 min, and it can be applied to other important microorganisms to allow the required prompt measures to be taken.

Molecular characterisation of infectious bursal disease viruses isolated in recent acute outbreaks in Slovenia.

In 2004 and then in 2006 several outbreaks of infectious bursal disease (IBD) were reported in broiler and broiler breeder flocks in Slovenia. In this report ten recently emerged IBD viruses (IBDV) were characterised by sequence analysis of the VP2 hypervariable region and compared to previous Slovene IBDV strains from 1995/1996 and to some representative serotype 1 IBDV strains of different pathotypes. On the basis of nucleotide and amino acid identities, phylogenetic analyses and the presence of very virulent IBDV (vvIBDV) conserved amino acid substitutions, all Slovene isolates from recent outbreaks were identified as vvIBDV. Although some unique nucleotide exchanges and amino acid substitutions have been observed, the results of this study indicated that recent vvIBDV isolates are closely related with those from outbreaks in the 1990s. However, acute IBD has not been reported in commercial flocks in Slovenia for some years. This could lead to the conclusion that poor biosecurity and relaxed vaccination could be responsible for the re-emergence of vvIBDV.

Detection and Phylogenetic Analysis of Herpesviruses Detected in Wild Owls in Slovenia.

Herpesvirus (HV) was detected using PCR in the organs of eight of 55 wild owls (14.5%) from seven species that were found dead in various locations in Slovenia between 1995 and 2015. HV was detected in three species: the Eurasian eagle owl (Bubo bubo), Ural owl (Strix uralensis), and long-eared owl (Asio otus). Phylogenetic analysis of partial DNA polymerase gene nucleotide sequences showed that the detected HVs are similar to the avian and mammal alphaherpesviruses. Two sequences were very similar to known bird HV sequences. One sequence was identical to the columbid herpesvirus 1 (CoHV1) sequence, and the other was very similar to the gallid herpesvirus 2 (GaHV2) sequence. The phylogenetic tree revealed a lower similarity of the other six analyzed Slovenian sequences with the sequences of alphaherpesviruses of birds and mammals. This is the first study to report the detection of different HVs in owls.

Detección y análisis filogenético de herpesvirus detectados en búhos silvestres en Eslovenia Se detectaron virus herpes mediante PCR en los órganos de ocho de 55 búhos silvestres (14.5%) pertenecientes a siete especies y que se encontraron muertos en varios lugares de Eslovenia entre los años 1995 y 2015. Se detectaron virus herpes en tres especies: el búho real (Bubo bubo), cárabo uralense (Strix uralensis) y el búho chico (Asio otus). El análisis filogenético de las secuencias de nucleótidos parciales del gene de la polimerasa de ADN mostró que los virus herpes detectados son similares a los alphaherpesvirus aviares y de mamíferos. Dos secuencias fueron muy similares a las secuencias de virus herpes de aves conocidas. Una secuencia fue idéntica a la secuencia del herpesvirus 1 de palomas (CoHV1) y otra fue muy similar a la secuencia del herpesvirus 2 del pollo (GaHV2). El árbol filogenético reveló menores similitudes entre las otras seis secuencias analizadas de Eslovenia con las secuencias de alphaherpesvirus de aves y de mamíferos. Este es el primer estudio que informa la detección de diferentes virus herpes en búhos.

Trichomonosis outbreak in a flock of canaries (Serinus canaria f. domestica) caused by a finch epidemic strain of Trichomonas gallinae.

In the present paper, an outbreak of trichomonosis in a flock of 15 breeding pairs of canaries is described. Trichomonosis was diagnosed on characteristic clinical signs, microscopic examination of crop/esophageal swabs, gross pathology and histopathology. Trichomonads were successfully grown in culture media and were characterized by multi-locus sequence typing. The three genomic loci ITS1-5.8S-ITS2, 18S rRNA and Fe-hydrogenase were analyzed. Molecular characterization confirmed the finch trichomonosis strain, identical to the strain that caused emerging disease in free-living passerine birds in Europe. Flock treatment with metronidazole (200mg/L) in drinking water for 5days was partially effective. After individual treatment with oral application of metronidazole (20mg/kg SID) for 5days no further clinical signs were observed in the flock over next 30 months.