Investigation of canine chaphamaparvovirus, canine bufavirus, and canine adenovirus in dogs with diarrhea: First report of novel canine bufavirus in Turkey.

Viral enteritis is a significant cause of death among dogs younger than 6 months. In this study, the presence of canine chaphamaparvovirus (CaChPV), canine bufavirus (CBuV), and canine adenovirus (CAdV) was investigated in 62 diarrheal dogs previously tested for other viral pathogens (canine parvovirus type 2, canine coronavirus, and canine circovirus). CBuV was detected in two dogs (3.22%) and CaChPV in one dog (1.61%). One dog tested positive for three parvoviruses (CPV-2b, CBuV, and CaChPV). All dogs tested negative to CAdV-1/CAdV-2. A long genome fragment of one of the two identified CBuVs and of the CaChPV was obtained and analyzed. New Turkish CBuVs had high identity rates (96%-98% nt; 97%-98% aa) with some Italian CBuV strains (CaBuV/9AS/2005/ITA and CaBuV/35/2016/ITA). The phylogenetic analysis powerfully demonstrated that these viruses belonged to a novel genotype (genotype 2). A part of the genome ChPV-TR-2021-19 revealed high identity rates (> 98% nt and > 99% aa) with some Canadian CaChPV strains (NWT-W88 and NWT-W171) and the Italian CaChPV strain Te/37OVUD/2019/IT. This study is the first report on the detection of CBuV-2 and the concomitant presence of three canine parvoviruses in Turkey. The obtained data will contribute to the molecular epidemiology and the role in the etiology of enteric disease of new parvoviruses.

Predominance and first complete genomic characterization of canine parvovirus 2b in Turkey.

Viral enteritis is a significant threat to domestic dogs. The two primary pathogens that cause viral enteritis in dogs are canine coronavirus (CCoV) and canine parvovirus (CPV). In this study, we investigated the occurrence of CPV-2, CCoV, and canine circovirus coinfection by characterizing circulating subtypes of CPV-2 in faecal samples from symptomatic dogs admitted to veterinary clinics located in Ankara, Elazig, Kayseri, and Kocaeli provinces of Turkey, between 2019 and 2022. Virus detection by PCR and RT-PCR revealed that CPV-2 was present in 48 (77.4%) samples, and no other agents were detected. Based on the occurrence of the codon GAT at positions 1276 to 1278 (coding for aspartate at residue 426) of VP2, all CPV-2 isolates were confirmed to be of the CPV-2b subtype. The complete genome sequences of two CPV-2b isolates showed a high degree of similarity to and phylogenetic clustering with Australian and East Asian strains/isolates. The predominant CPV strain circulating in the three different regions of Turkey was found to be a CPV-2b strain containing the amino acid substitutions at Y324I and T440A, which commonly contribute to immune escape. This is the first report of complete genomic analysis of CPV-2 isolates circulating in symptomatic domestic dogs in Turkey. The evolution of CPV-2 has raised questions about the efficacy of current vaccination regimes and highlights the importance of monitoring the emergence and spread of new CPV-2 variants.

First report of Serotype-1 Marek's disease virus (MDV-1) with oncogenic form in backyard turkeys in Turkey: a molecular analysis study.

BACKGROUND: Marek's disease (MD) is a lymphoproliferative disease caused by Gallid alphaherpesvirus 2 (GaHV-2, MDV-1), which primarily affects chickens. However, the virus is also able to induce tumors and polyneuritis in turkeys, albeit less frequently than in chickens. RESULTS: This is the first study in Turkey reporting the molecular characterization of a MDV-1 strain detected in a flock of backyard turkeys exhibiting visceral lymphoma. Here, MEQ, vIL-8, pp38 and 132-bp tandem repeat regions, which are frequently preferred in the pathotyping of MDV-1, were examined. It was determined that the MEQ gene of MDV-1/TR-21/turkey strain obtained in the present study encoded 339 amino acids (1020 nt) and had four proline-rich repeat regions (PPPP). Based on the nucleotide sequence of the MEQ gene of the MDV-1/TR-21/turkey strain, a phylogenetic tree was created using the MEGA-X software with the Maximum Likelihood Method (in 1000 replicates). Our strain was highly identical (> 99.8) to the Italian/Ck/625/16, Polish (Polen5) and some Turkish (Layer-GaHV-2-02-TR-2017, Tr/MDV-1/19) MDV-1 strains. Also, nt and aa sequences of the MEQ gene of our strain were 99.1 and 99.41% identical to another Turkish strain (MDV/Tur/2019) originated from chickens. Sequence analysis of pp38 and vIL-8 genes also supported the above finding. The identity ratios of nucleotide and amino acid sequences of vIL-8 and pp38 genes of MDV-1/TR-21/turkey strain were 99.64-100% and 99.79-100%, respectively, when compared with those of the Polish strain. According to 132-bp tandem repeat PCR results, the MDV-1/TR-21/turkey strain had five copies. CONCLUSIONS: These results suggested that the MDV-1/TR-21/turkey strain obtained from backyard turkeys can be either very virulent or very virulent plus pathotype, though experimental inoculation is required for precise pathotyping.

First detection of feline bocaparvovirus 2 and feline chaphamaparvovirus in healthy cats in Turkey.

The pet cat's population and the number of viruses that infect them are increasing worldwide. Recently, feline chaphamaparvovirus (FeChPV, also called fechavirus) and feline bocaparvovirus (FBoV) infections, which are novel parvovirus species, have been reported in cats from different geographic regions. Here, we investigated FBoV 1-3 and FeChPVs in healthy cats in Turkey using PCR, where nuclear phosphoprotein 1 (NP1) is targeted for FBoV and NP for FeChPV. For this purpose, oropharygeal swabs were obtained from 70 healthy cats with different housing status from June 15 to December 1, 2020. After PCR screening tests, six out of 70 cats (5/47 shelter cats; 1/23 domestic cats) were found to be positive for FBOV, while two were positive for FeChPV (1/47 shelter cats; 1/23 domestic cats). No cat was found in which both viruses were detected. The nucleotide (nt) sequence comparison in the 310 base pair (bp) NP gene of the two FeChPVs identified in this study shared a high identity with each other (95.0% nt and 99% aa identities) and with previously reported FeChPVs (92.4-97.1% nt and 98.1-99.0% aa identities), including 313R/2019/ITA, 49E/2019/ITA, VRI_849, 284R/2019/ITA, and IDEXX-1. Here, the near-full length (1489 nt, 495 amino acids-aa) of the VP2 gene of the FechaV/Tur-2020/68 isolate obtained from the study was also sequenced. The nt and aa identity ratio of this isolate with other FeChPVs was 98.0-98.5%-96-96.5%, respectively. Sequences of the 465 bp NP1 gene of the six Turkish FBoV strains shared high identities with each other (99.6-100% nt and 99.3-100% aa identities) and with those of FBoV-2 strains (97.8-99.1% nt and 98.0-100% aa identities), including 16SY0701, 17CC0505-BoV2, HFXA-6, and POR1. All FBoVs detected in this study were classified as genotype 2, similar to the study conducted in Japan and Portugal. Here, the NS1 (partial), NP1, VP1 and VP2 gene of the FBoV-2/TUR/2020-14 strain obtained from the study were also sequenced and the nt and aa sequences showed high identities to the above-mentioned FBoV-2 strain/isolates (> 96%, except for the aa ratio of strain 16SY0701). In conclusion, this study shows that FBoV and FeChPV are present in healthy cats in Turkey, and these viruses can be detected from oropharyngeal swabs. Our findings contribute to further investigation of the prevalence, genotype distribution, and genetic diversity of Turkish FBoVs and FeChPVs, adding to the molecular epidemiology of FBoV and FeChPVs worldwide.

Short-Term Humoral Immune Response of the pcDNA4-G Plasmid Expressing the Bovine Ephemeral Fever Virus G Gene in BALB/c Mice.

Bovine ephemeral fever (BEF) is an arthropod­borne viral disease characterised by a short­term clinical expression that can lead to significant losses in high­yielding cattle and water buffaloes. In this study, we aimed to generate a recombinant plasmid expressing the glycoprotein (G) of the BEF virus (BEFV) and to stimulate a humoral immune response to this protein in BALB / c mice immunised with the recombinant plasmid. Expression of the encoded protein was demonstrated by western blotting and immunoperoxidase tests. The suitable plasmids were intramuscularly administered to BALB/c mice on days 0, 14 and 21. The antibody response in the immunised mice was measured by a plaque reduction neutralization test (PRNT) and enzyme­linked immunosorbent assay (ELISA). According to BEFV ELISA, only two of the seven animals in these groups exceeded the cut­off value. A significant difference was observed in the mean OD values at 450 nm absorbance in the pcDNA4­G­immunised group when compared with those in the plasmid control group at 30 days (p < 0.05). According to PRNT50 results, a 1:20 (p < 0.05) antibody response was obtained at 30 days in pcDNA4­G (100 µg)­immunised mice, whereas this ratio was 1:80 (p < 0.001) in BEFV­immunised mice (1,000 PFU/0.5 ml). We conclude that the humoral immune response was stimulated in experimental mice immunised with the recombinant plasmid. However, disappointingly, the antibody response was markedly low in pcDNA4­G­immunised mice.

Detection and molecular characterisation of feline viruses from swab samples.

Feline calicivirus (FCV), feline alphaherpesvirus 1 (FHV-1) and feline panleukopenia virus (FPLV) as well as retroviral agents such as feline leukaemia virus (FeLV) and feline immunodeficiency virus (FIV) are important viral pathogens of cats. The aim of this study was to detect and characterise FHV-1, FPLV, FeLV, FIV and feline foamy virus (FFV) in oropharyngeal, nasal and conjunctival swabs from 93 cats that had been screened for FCV previously. We wanted to determine the possible risk factors for infection with these viruses. The prevalence was found to be 12.9% for FHV-1 and 9.7% for FPLV. FIV was detected only in two samples and FeLV in one sample, whereas the presence of FFV was not demonstrated in any of the clinical samples. The statistical analysis of the results showed that breed, age, health status, and lifestyle are important predisposing factors to FHV-1 (P < 0.05). For FPLV, only clinically unhealthy animals were found to be at risk (P < 0.001). Sequence analysis revealed that the two FIV-positive samples in this study contained different (A and B) subtypes of the virus. This is the first report on the occurrence of subtype A FIV in Turkey.

Detection and molecular characterization of a highly oncogenic Marek's disease virus from vaccinated hens in Turkey.

Marek's disease (MD) is a highly contagious neoplastic disease of chickens associated with economic losses, often due to visceral lymphomas. The etiological agent is MD virus serotype 1 (MDV-1), also called Gallid alphaherpesvirus 2 (GaHV-2). Despite intensive vaccination, MDV is constantly evolving and maintaining its presence in the world. The aim of this study was to genetically analyze a highly oncogenic MDV/Tur/2019 strain obtained from a poultry farm in Turkey's Elazig province in 2019. Genes associated with viral pathogenicity and oncogenicity Marek's EcoRI-Q-encoded protein (MEQ), phosphoprotein-38 (pp38), and viral interleukin 8 (vIL-8) were selected for this purpose. The vIL-8 nucleotide sequence showed high similarity (100% identity) to some European (EU-1, Polen 5) and Asian (03 India, GADVASU-M2) MDV strains. The pp38 nucleotide sequence showed high similarity (100% identity) to some American (CU-2, JM/102W, RB1B) and European (MD70/13, ATE2539) MDV strains. There were no disrupted four-proline molecules (PPPP) within the transactivation domain of the MEQ. However, according to phylogenetic results, the MDV/Tur/2019 strain was included in cluster 2a alongside European MDV strains (Polish, Hungarian, Italian) with very virulent and very virulent plus pathotypes. In conclusion, we believe that the MDV/Tur/2019 strain obtained from turkey herpesvirus (HVT)-vaccinated chickens has a very virulent or very virulent plus pathotype. Although this result provides some clues regarding the virulence of this strain, in vivo studies are needed to achieve exact pathotyping. Further, combination of HVT with the CVI988 strain should be used for vaccination to provide the best protection, as highly pathogenic MDV strains can break sterile immunity against the HVT vaccine. Keywords: GaHV-2; Marek's disease; oncogenes; Turkey.

Interpretation of SARS-CoV-2 behaviour on different substrates and denaturation of virions using ethanol: an atomic force microscopy study.

Coronavirus (SARS-CoV-2) is a respiratory infection virus that was first detected in Wuhan, China. The virus causes COVID-19 disease and the outbreak was recognised as a pandemic by the World Health Organization (WHO) in March 2020. SARS-CoV-2 virion was first imaged using cryo-electron microscopy by the Chinese Center for Disease Control and Prevention (CDC). Atomic Force Microscopy is a unique technique that can allow imaging of biomolecules under different conditions. In this work, we used Atomic Force Microscopy to characterize SARS-CoV-2 on tissue culture polystyrene (TCPS) and glass coverslip surfaces. We isolated SARS-CoV-2 and drop casted it on coverslip glass and tissue culture polystyrene surfaces. We analyzed height profiles, density, and aggregation behavior of the virion on glass and polystyrene surfaces. We observed the coffee ring effect on the drop casted samples and close packing of virions near the coffee rings on both surfaces with relatively higher virion distribution on the tissue culture polystyrene (TCPS) substrates. We compare virion agglomeration on the two types of surfaces. Finally, we applied ethanol disinfectant to virions on the surface to visualize the effect of ethanol and image the ultrastructure of SARS-CoV-2.

Complete genome analysis of highly pathogenic bovine ephemeral fever virus isolated in Turkey in 2012.

Relatively high prevalence and mortality rates of bovine ephemeral fever (BEF) have been reported in recent epidemics in some countries, including Turkey, when compared with previous outbreaks. A limited number of complete genome sequences of BEF virus (BEFV) are available in the GenBank Database. In this study, the complete genome of highly pathogenic BEFV isolated during an outbreak in Turkey in 2012 was analyzed for genetic characterization. The complete genome of the Turkish BEFV isolate was amplified by reverse transcription-polymerase chain reaction (RT-PCR) and sequenced. It was found that the complete genome of the Turkish BEFV isolate was 14,901 nt in length. The complete genome sequence obtained from the study showed 91-92% identity at nucleotide level to Australian (BB7721) and Chinese (Bovine/China/Henan1/2012) BEFV isolates. Phylogenetic analysis of the glycoprotein gene of the Turkish BEFV isolate also showed that Turkish isolates were closely related to Israeli isolates. Because of the limited number of complete BEFV genome sequences, the results from this study will be useful for understanding the global molecular epidemiology and geodynamics of BEF.