Pathogenicity assessment of a bovine viral diarrhea virus type 1l (BVDV-1l) strain in experimentally infected calves.

Bovine viral diarrhea is a widespread and economically important viral disease for livestock which can cause clinically diverse manifestations. The number of established BVDV subgenotypes has increased, not only the serological relationships of recently described subgenotypes but virulence and pathogenic characteristics have not yet been mostly elaborated. The dominant BVDV subgenotype in Turkiye was elaborated to be BVDV-1l, that involves more than half of field strains and there is no scientific data to identify the pathogenicity of this strain so far. This study investigated the pathogenicity of a selected field strain (TR-72) from subgenotype BVDV-1l. Experimental infection was implemented by intranasal inoculation of the strain TR-72 (10 ×105.5) to four young calves which were previously not vaccinated and were free both for BVDV antibodies and antigens. Clinical changes as well as blood parameters, body temperature, and viremia were monitored for 14 days. Only mild clinical signs associated with respiratory signs of BVDV infection were observed. Detected clinical signs included nasal discharge, conjunctivitis, cough, fatigue, high rectal temperature reaching 40.7 â„ƒ, and white blood cell counts depression started from the 2nd day and 40.4% decreased between the 12th and 14th days post-infection (poi). The presence of viremia was investigated by virus isolation, RT-PCR, and real-time RT-PCR from blood samples. The efficiency of experimental infection was established not only by observed clinical signs but also by virus isolation from blood leukocytes between the 5th and 8th days poi., virus detection was obtained by real-time PCR between the 3rd - 13th days poi. Besides, the recorded mild clinical signs, high fever, long duration of viremia , and high decrease in blood parameters obtained in this study, it was shown that the noncytopathogenic BVDV-1l strain TR-72 has a moderate virulence in naïve cattle.

Characterization of bovine rotavirus isolates from diarrheic calves in Türkiye.

BACKGROUND: Neonatal calf diarrhea, which is the most common cause in calf deaths, leads to significant economic losses in dairy farming around the world. Diarrhea develops due to infectious and non-infectious reasons. Group A Rotaviruses (RVA) are the leading and predisposing factor for acute neonatal gastroenteritis. METHODS AND RESULTS: In this study, 20 diarrheic fecal samples were collected from one farm in Balikesir province of Turkey. During virus isolation, a total of 2 stool samples were detected to produce cytopathogenic effects in MA-104 cell line. The two samples (RV-36, RV-38) were tested positive with antigen ELISA kits detecting RVA antigens. In order to detect the presence of rotavirus viral nucleic acid in cell supernatants, VP6 gene region-specific RT-PCR test was performed and the samples RV-36 and RV-38 were positive for RVA viral nucleic acid. By RT-PCR using genotype specific primers, both the isolates RV-36 and RV-38 formed amplicons compatible with G10 and P[11] genotypes of RVA. RVA nucleic acids segments were also visualized by poliacrilamide gel electrophoresis (PAGE) method. The phylogenetic tree constructed according to the VP6 gene region showed that these isolates were in the Rotavirus A group and in the I2 cluster same as other bovine and some human RVA isolates. CONCLUSION: Succesful isolation of RVA G10P[11] was echieved in the cattle farm. As rotaviruses play the most important role in the etiology of diarrhea in newborn calves respected genotype G10P[11] should be considered in selection of the vaccines applied to the dams. Those isolates can be further evaluated as vaccine candidate.

Recent strains of influenza D virus create a new genetic cluster for European strains.

Bovine respiratory diseases (BRD) are one of the significant health problems for cattle breeding industry. Influenza D virus (IDV) alone or in combination with other respiratory pathogens plays a role in BRD. According to the IDV-HEF gene region, phylogenetic analyzes revealed five lineages: D/OK, D/660, D/Yama2016, D/Yama2019, and D/CA2019, so far. In this study, despite no success in virus isolation, the presence of IDV was investigated by RT-PCR (partial HEF gene region) in 219 nasal swab samples collected from cattle with BRD between 2012 and 2021. The presence of IDV was demonstrated in two samples, and genome characterization data of the IDV sequences both in the partial and complete HEF gene regions showed that one of the obtained sequences (D/bovine/Turkey-Bursa/ET-138/2021) was in the lineage D/Yama2019 while the other (D/bovine/Turkey-Bursa/ET-130/2013) created a new lineage tentatively called D/Bursa2013 as including few partial IDV sequences reported in Europe. Two nucleotide substitutions (nt252A→G, nt299T→C) were typically characterized for the tentative lineage D/Bursa2013, one of which also leads to a unique amino acid change at position aa100 (V→A). When the amino acid differences between the lineages were evaluated, amino acid substitution changes were detected in four regions [aa12 (Alanine→Aspartic acid), aa19 (Glycine→Arginine), aa22 (Proline→Serine), and aa110 (Aspargine→Arginine)] of the D/Yama2019 lineage, unlike the other lineages. Considering the most common D/OK lineage in Europe, many nucleotide substitutions were shown between D/OK and D/Bursa2013. Accordingly, aminoacid substitutions were observed in aa27 (Threonine→Asparagine) and aa100 (Valine→Alanine) in the D/bovine/Turkey-Bursa/ET-138/2021 sequence. Study results describe the circulation of D/Yama2019 and D/Bursa2013 (new lineage) in Turkey. Expansion of new strains seems possible due to the high mutation rate of influenza viruses. It is important to understand the development of IDV with comprehensive characterization studies.

Inhibition of bovine and ovine capripoxviruses (Lumpy skin disease virus and Sheeppox virus) by ivermectin occurs at different stages of propagation in vitro.

Capripoxvirus diseases are listed as reportable diseases by World Organization for Animal Health (OIE). Lumpy skin disease virus (LSDV) and sheeppox virus (SPPV), which can only be distinguished by molecular analysis, cause moderately, severe, or sometimes fatal infections in cattle and sheep. Even though vaccines are the most effective way to control the infection, their effectiveness may decrease in some cases. Therefore, it is significant to explore antiviral drugs against these diseases along with the vaccine. This study aimed to investigate the antiviral efficiency of ivermectin (IVM) at different stages of in vitro replication of LSDV and SPPV. For this purpose, viral titers (TCID50/mL) of the viruses not treated with IVM (0.0 µM) and treated with non-cytotoxic concentrations of IVM (1.0 and 2.5 µM) were compared during a nine-day (216 h) post-infection period by viral titration assay. At 2.5 µM concentrations of IVM, the mean viral titer was significantly (P<0.05) reduced by approximately three logs for the replication stage of LSDV and SPPV. To evaluate the antiviral activity of IVM against LSDV and SPPV by treatment at the virus attachment and penetration stages, the titers of the virus either untreated or treated with 2,5 µM IVM were compared by virus titration assay. The number of infectious virions for LSDV and SPPV were decreased by 99.82% and 99.87% at the viral replication stage, 68.38% and 25.01% at the attachment stage, and 57.83% and 0.0% at the penetration stage, respectively. It was determined that ivermectin is statistically more effective on LSDV than SPPV at the virus attachment and penetration stages (P<0.05). This study found that the drug IVM can inhibit capripoxviruses, including LSDV and SPPV at various stages of the propagation. Moreover, this research predicted the in vitro antiviral ability of IVM against capripoxvirus infections for the first time.

First isolation and molecular characterization of pseudorabies virus detected in Turkey.

BACKGROUND: Pigs are the main host species for the pseudorabies virus. It causes fatal encephalitis in many species, including humans. This article aims to report the first clinical case of pseudorabies as well as isolation and molecular characterization of the virus from a hunting dog in Bursa province, Turkey. METHODS AND RESULTS: The dog shows clinical signs including pruritus and neurological signs such as stumbling and inability to stand up compatible with pseudorabies. The virus isolates were obtained from the supernatant of fresh tissue samples from the cerebellum, cornu ammonis, spleen, salivary gland, conjunctival swab, serum, and PBMC samples. The glycoprotein C region is targeted for viral DNA amplification. Pseudorabies virus genome detected both in fresh tissues and supernatants of third passage on Vero cells. The number of PCR positive samples was dramatically increased after cell culture inoculations. Genome sequencing of strain Bursa-10303, which was isolated from a non-endemic area, identified it to belong to clade A. CONCLUSIONS: This study confirms the possible presence of pseudorabies infection in the wildlife reservoirs in Turkey. Future studies may clarify the importance of the infection in Turkey region, where there is no prevalent pig production.

Ivermectin also inhibits the replication of bovine respiratory viruses (BRSV, BPIV-3, BoHV-1, BCoV and BVDV) in vitro.

Bovine respiratory disease (BRD) complex is an important viral infection that causes huge economic losses in cattle herds worldwide. However, there is no directly effective antiviral drug application against respiratory viral pathogens; generally, the metaphylactic antibacterial drug applications are used for BRD. Ivermectin (IVM) is currently used as a broad-spectrum anti-parasitic agent both for veterinary and human medicine on some occasions. Moreover, since it is identified as an inhibitor for importin α/ß-mediated nuclear localization signal (NLS), IVM is also reported to have antiviral potential against several RNA and DNA viruses. Since therapeutic use of IVM in COVID-19 cases has recently been postulated, the potential antiviral activity of IVM against bovine respiratory viruses including BRSV, BPIV-3, BoHV-1, BCoV and BVDV are evaluated in this study. For these purposes, virus titration assay was used to evaluate titers in viral harvest from infected cells treated with non-cytotoxic IVM concentrations (1, 2.5 and 5 µM) and compared to titers from non-treated infected cells. This study indicated that IVM inhibits the replication of BCoV, BVDV, BRSV, BPIV-3 and BoHV-1 in a dose-dependent manner in vitro as well as number of extracellular infectious virions. In addition, it was demonstrated that IVM has no clear effect on the attachment and penetration steps of the replication of the studied viruses. Finally, this study shows for the first time that IVM can inhibit infection of BRD-related viral agents namely BCoV, BPIV-3, BVDV, BRSV and BoHV-1 at the concentrations of 2.5 and 5 µM. Consequently, IVM, which is licensed for antiparasitic indications, also deserves to be evaluated as a broad-spectrum antiviral in BRD cases caused by viral pathogens.

Failure in dry period vaccination strategy for bovine viral diarrhea virus.

Bovine viral diarrhea is a common disease of cattle and has significant impact on animal welfare worldwide. There are fundamental approaches i.e. elimination of persistently infected animals, vaccination and biosecurity measures for effective control and eradication of BVD virus (BVDV). By this study, the presence of persistent infection with divergent BVDV subgenotype in the calves in a dairy herd having regular vaccination program was investigated. In the herd, vaccinated with a killed whole virion trivalent vaccine (composed of BVDV-1a) during the dry period of the cows, abortion cases were existed in the late autumn 2019. During herd screening by BVDV antigen-ELISA, 2 out of 300 dams were detected positive. Following, by ear notch-based BVDV antigen-ELISA, 30 calves were detected positive. Confirmation of persistent BVDV infection was performed 3 weeks later by testing with antigen-ELISA, where 8 of 9 selected newborn calves were positive for the second time. The entire antigen-ELISA positive samples were subjected to virus isolation on MDBK cell culture and identified as non-cytopathogenic pestiviruses by indirect immunoperoxidase assay. Presence of pestivirus RNA was detected in the 8 isolates by panpestivirus RT-PCR. Analysis of the 5'UTR regions revealed that BVDV-1 r circulate in the herd. Results of this study lead to questioning the efficiency of dry period vaccination strategy against BVDV. But otherwise, vaccination with BVDV-1a can be inefficient for complete protection against BVDV-1 r. Therefore, serological relationship between mentioned subgenotypes or protection by current vaccines against latest field isolates needs to be investigated before development of new BVDV vaccine candidates.