Spatiotemporal analysis of foot and mouth disease outbreaks in cattle and small ruminants in Türkiye between 2010 and 2019.

Determining the dynamics associated with foot-and-mouth disease (FMD) outbreaks is important for being able to develop effective strategic plans against the disease. In this direction, spatiotemporal analysis of FMD virus (FMDV) epidemic data that occurred in Türkiye between 2010 and 2019 was carried out. Spatiotemporal analysis was performed by the space-time scan statistic using data from a total of 7,796 FMD outbreaks. Standard deviational ellipse analysis (SDE) was performed to analyse the directional trend of FMD. Five, six, and three significant and high-risk clusters were identified by the space-time cluster analysis for serotypes A, O, and Asia-1, respectively. The SDE analysis indicated that direction of FMD transmission was northeast to southwest. A significant decrease in the number of outbreaks and cases were observed between 2014 and 2019 compared to 2010-2013 (p = 0.010). Most of the serotype A, serotype O, and serotype Asia-1 associated FMD outbreaks were observed during the dry season (April to September). Among FMD cases, cattle and small ruminants accounted for 80.75% (180,932 cases) and 19.25% (43,116 cases), respectively. Among the serotypes detected in the cases, the most frequently detected serotype was serotype O (50.84%), followed by serotypes A (35.67%) and Asia-1 (13.49%). The results obtained in this study may contribute to when and where control programs could be implemented more efficiently for the prevention and control of FMD. Developing risk-defined regional control plans by taking into account the current livestock production including uncontrolled animal movements in border regions, rural livestock, livestock trade between provinces are recommended.

Epidemiology of bluetongue virus infection among small ruminants in Turkey: Seroprevalence and associated risk factors.

Bluetongue (BT) is an endemic disease of small ruminants in Turkey, and it has substantial socio-economic impact at national level. To reduce this impact, vaccination has been used for the control of BT but sporadic outbreaks have been reported. Although sheep and goat farming plays an important role in rural communities, little is known about the BT epidemiological situation in small ruminants in Turkey. Therefore, this study aimed to estimate the seroprevalence of the bluetongue virus (BTV) and to identify the potential risk factors associated with BTV seropositivity in small ruminants. This study was conducted in the Antalya Province in the Mediterranean region of Turkey, from June 2018 to June 2019. A total of 1026 blood samples, from clinically healthy goats (n = 517) and sheep (n = 509), obtained from randomly selected unvaccinated flocks (n = 100) were tested for BTV anti-VP7 antibodies by using a competitive enzyme linked immunosorbent assay test. A questionnaire was administered to the flock owners to obtain data related to sampled flocks and animals. At the animal level, the true prevalence of BTV antibodies was 74.2% (n = 651/1026, 95% CI = 70.7-77.7) with 85.3% (n = 370/509, 95% CI = 80.6-89.9) seropositive sheep and 63.3% (n = 281/517, 95% CI = 58.2-68.4) seropositive goats. The true flock-level seroprevalence of BTV was higher in goats (100.0%, 95% CI = 92.8-100.0) than in sheep (98.8%, 95% CI = 86.6-100.0). The intra-flock seroprevalence within seropositive flocks varied between 36.4% and 100%, with a mean value of 85.5% and 61.9% in sheep and goat flocks, respectively. The logistic regression model revealed that odds of seropositivity for sheep were significantly higher in female animals (OR: 1.8, 95% CI = 1.1-2.9), animals older than 24 months old (OR: 5.8, 95% CI = 3.1-10.8), Pirlak breed (OR: 3.3, 95% CI = 1.1-10.0) and Merino breed (OR: 4.9, 95% CI = 1.6-14.9), whereas for goats, it was higher in female animals (OR: 1.7, 95% CI = 1.0-2.6), animals older than 24 months old (OR: 4.2, 95% CI = 2.7-6.6) and Hair breed (OR: 5.6, 95% CI = 2.8-10.9). The use of insecticides was identified as a protective factor. The present study revealed that BTV infection is widespread in sheep and goats in the Antalya Province. It is recommended to implement biosecurity measures in flocks and use insecticides to mitigate the spread of infection and contact between hosts and vectors.

Risk assessment and seroprevalence of bovine herpesvirus type 1 infection in dairy herds in the inner Aegean Region of Turkey.

This cross-sectional study was performed to investigate the seroprevalence and associated risk factors of bovine herpesvirus type 1 (BoHV-1) infection in dairy cattle herds in Afyonkarahisar province in the Aegean Region of Turkey. Blood samples were collected from 602 cattle from 56 unvaccinated dairy herds between May 2018 and June 2019. Animal and herd-level epidemiological information was collected with a questionnaire during blood collection. Specific antibodies against BoHV-1 and bovine viral diarrhea virus (BVDV) were detected by using a virus neutralization test and a commercial indirect ELISA kit, respectively. Univariable and multivariable logistic regression analyses were used to determine any association between categorical variables and BoHV-1 seropositivity. The animal-level and herd-level seroprevalences of BoHV-1 infection were determined to be 39.53% (95% confidence interval, CI: 35.71-43.50) and 73.21% (95% CI: 60.41-83.04), respectively. Within-herd prevalence was more than 50% in 34.14% of infected herds. Cattle age (odds ratio, OR= 2.34, 95% CI: 1.58-3.44), BVDV infection (OR= 7.74, 95% CI: 5.08-11.76), and the presence of goats in the herd (OR= 2.84, 95% CI: 1.91-4.19) were identified as risk factors for BoHV-1 seropositivity by the multivariable logistic regression model. This is the first study conducted in Turkey using two-layer sampling and logistic regression analyses to determine the herd-level and animal-level seroprevalence and associated risk factors of BoHV-1 infection.

Risk factors and genetic characterization of bovine respiratory syncytial virus in the inner Aegean Region, Turkey.

Bovine respiratory syncytial virus (BRSV) is one of the causative viral agents of the bovine respiratory disease complex. This study was conducted to determine the seropositivity and risk factors associated with BRSV infection and to evaluate the phylogenetic relatedness of the BRSVs in the inner Aegean region of Turkey. In this cross-sectional study, serum samples (n = 557) and nasal swabs (n = 21) were collected from cattle herds (n = 43) between February 2018 and March 2019. A commercial indirect-ELISA kit was used for the detection of antibodies in the sera samples. Reverse-transcriptase PCR was used to detect viral RNA in nasal swabs. Nasal samples were also examined for the detection of bovine parainfluenza-3, bovine viral diarrhoea virus, and bovine herpesvirus 1 by molecular detection methods. Genetic characterization of the local BRSV field isolates was conducted by sequencing attachment glycoprotein (G) gene segment. Epidemiological data on potential risk factors were collected from each sampled herd during blood collection. All herds had at least one seropositive animal. After adjustment for assay sensitivity and specificity, the overall true seropositivity was 58.48% (95% CI: 53.32-63.47). BRSV RNA was detected in 2 of the 21 nasal swabs, whereas other infectious agents were not detected in the investigated samples. Phylogenetic analysis showed that the field isolates of BRSV obtained in this study belonged to subgroup III, but they were located on separate branch from previously characterised Turkish subgroup III isolates. BRSV field strains from this study displayed 3 new amino acid substitutions (P89S, D115G, and S165L) in the G protein chains compared to other main reference BRSV isolates, demonstrating that BRSV is still evolving. Generalised estimating equation model showed that there were positive associations between BRSV infection, age (OR = 2.36, p = 0.001), herd size (OR = 10.32, p < 0.001), herd type (OR = 8.97, p < 0.001), a past history of respiratory disease (OR = 4.06, p < 0.001). The results of this study revealed that BRSV infection is common among cattle herds in the inner Aegean region of Turkey. The obtained epidemiological and genetic data on BRSV infection from this study could be beneficial for designing effective biosecurity practices and vaccination strategies.

Genomic characterization of pestiviruses isolated from bovine, ovine and caprine foetuses in Turkey: A potentially new genotype of Pestivirus I species.

This study was carried out to investigate the frequency and genetic diversity of pestiviruses in abortion cases in cattle and small ruminants in Turkey. During January 2012 and December 2017, a total of 2029 aborted foetuses (553 bovine foetuses, 1,388 sheep foetuses and 88 goat foetuses) were collected from different regions of Turkey. Real-time RT-PCR (RRT-PCR) assays were used to detect pestiviral RNA in aborted foetuses. To confirm the cause of abortion, pestivirus-positive foetuses were also examined for the presence of Brucella spp., Campylobacter spp., Chlamydophila abortus (C. abortus), akabane virus, bluetongue virus and Schmallenberg virus by molecular detection methods. Pestiviral RNA was detected in 61 (11%) of the 553 bovine foetuses, 124 (8.9%) of the 1,388 sheep foetuses and 3 (3.4%) of the 88 goat foetuses. Furthermore, C. abortus DNA was detected in 3 pestivirus-positive sheep foetuses, whereas other infectious agents were not detected in pestivirus-positive foetuses. Genetic characterization of the pestivirus RRT-PCR positive samples was conducted by sequencing 5' untranslated (5' UTR) and non-structural autoprotease (Npro ) genomic regions. A total of 68 sequences were obtained, and phylogenetic analyses revealed that all sequences belonged to BVDV-1, including 1b (8/68), 1f (2/68), 1l (4/68), 1r (10/68), Aydin-like pestivirus (20/68) and one unknown genotype (24/68). The 5' UTR and Npro sequences of this unknown genotype differed from pestiviruses previously described, providing evidence for the presence of an emerging genotype within the species Pestivirus I, tentatively named as 'Konya-like' pestivirus. 'Konya-like' pestivirus was the dominant genotype in sheep foetuses, whereas Aydin-like pestivirus was found to be the predominant genotype in bovine foetuses. To the best my knowledge, this is the first report of Aydin-like pestivirus infection in cattle. The information provided in this study contributes to the understanding the dissemination and evolution of pestiviruses and could be beneficial for developing more effective vaccines.

Comparative Evaluation of In-House and Commercial Real-Time PCR Methods for the Detection of the BK and JC Viruses in Clinical Samples.

Aim The two most common human polyomaviruses are the BK (BKV) and JC viruses (JCV). Diseases associated with polyomavirus usually occur in cases of severe cellular immunosuppression. BKV and JCV can cause many diseases, especially if they are reactivated in an immunosuppressed host. The aim of this study is to compare and evaluate the results of real-time polymerase chain reaction (PCR) methods targeting the small and large T gene regions of the viral genome, considering polymorphisms occurring in the viral genome of BKV and JCV. Materials and Methods Urinary specimens of 82 patients were taken from immunosuppressed patient and sent to molecular microbiology laboratory of Meram Medical Faculty. The small t gene was investigated using a commercial kit (LightMix, Roche) by real-time PCR method. Large T gene was investigated by using the optimized in-house real-time PCR method. Sequence analysis was accepted as the standard method. Results BKV positivity was detected in 9 samples and JCV positivity in 61 samples by real-time PCR method specific to small t gene region; BKV positivity in 21 samples and JCV positivity in 67 samples were determined by real-time PCR method specific to the large T gene region. Statistically, there was a significant difference for BKV, but not significant difference for JCV detection between the two methods. Conclusion Different polymorphisms in the target gene regions were responsible for the different outcomes obtained from this study. With this sensitivity and specificity, in-house PCR method which we used is a candidate for routine diagnosis.

Orf virus circulation in cattle in Turkey.

Orf virus (ORFV) causes contagious skin disease that mainly affects sheep and goats with zoonotic potential. However, there is not enough information about the association between ORFV and occurrence of skin disease in cattle. The present study describes outbreaks of ORFV infection in cattle in different provinces that are located in the Aegean, Central Anatolian and Mediterranean regions of Turkey. During the months of June and August 2017, vesicular fluid and scab samples were collected from cattle which had proliferative skin lesions. First, presence of lumpy skin disease virus (LSDV) and bovine herpesvirus 2 (BoHV-2, known as the causative agent of pseudo-lumpy skin disease) were investigated by real time PCR and PCR, respectively. Then, samples tested for the presence of parapoxviruses by PCR using primers specific to major envelope protein gene (B2L). Parapoxvirus DNA was detected in investigated samples whereas LSDV and BoHV-2 DNA were not detected. The analysis of the B2L gene sequences revealed that cattle were infected with ORFV. The isolates in the present study shared 100% sequence identity at the nucleotide and amino acid level when compared with previously characterised Turkish field ORFV isolates from goats in 2016. Results of the study show unusual infection of cattle with ORFV, and suggest that ORFV jumps the host species barrier from goats to cattle.

Detection of Peste des petits ruminants virus RNA in Culicoides imicola (Diptera: Ceratopogonidae) in Turkey.

An outbreak of Peste des petits ruminants (PPR) occurred in the Antalya Province in Turkey during  October  2015.  The  Antalya  Province  has  suitable  habitats  for  vectors.  There  is  no  information  available  on  the  role  of  Culicoides  spp.  in  the  transmission  of  Peste  des  petits  ruminants virus (PPRV). In this study we investigated the potential role of the Culicoides spp. in  the  transmission  of  PPRV.  Culicoides  were  trapped  throughout  middle  of  October  and  middle of December, 2015. A total of 12 pools of non-engorged females were analysed with real-time RT-PCR targeting the nucleocapsid (N) gene of the PPRV. PPRV RNA was detected in 7 of 12 Culicoides pools. These pools were negative for the bovine/ovine beta-actin mRNA. Culicoides spp. were identified to the species level by sequence analysis of the mitochondrial cytochrome  oxidase  subunit  I  gene.  The  species  of  Culicoides  found  PPRV  positive  was  Culicoides  imicola.  Molecular  characterization  of  field  isolates  from  recent  outbreaks  and  pools of midges that tested positive for PPRV suggests that PPRV replication might occur in Culicoides imicola, and it may have played a role in transmitting PPRV.

Molecular detection and genetic analysis of Akabane virus genogroup Ib in small ruminants in Turkey.

Akabane disease is a viral disease transmitted by biting midges and can cause teratogenic malformations in cattle, sheep and goats. Abortion outbreaks associated with arthrogryposis and cerebellar hypoplasia in two epidemiologically independent flocks were reported in the Mediterranean region of Turkey in 2015. Phylogenetic analysis based on nucleocapsid (N) gene sequences showed that the field isolates presented here belong to the genogroup Ib. The akabane virus (AKAV) field isolates analysed in this study displayed 6 new amino acid substitutions in N and non-structural protein chains compared with those of AKAV strains belonging to genogroup Ib. To the best of our knowledge, this is the first report on the presence of the AKAV genogroup Ib in Turkey.

Association of two clusters of Orf virus isolates in outbreaks of infection in goat in the Central Anatolian region of Turkey.

Orf virus (ORFV) is the etiological agent of contagious pustular dermatitis and can cause skin disease in sheep and goats. In this study, two outbreaks of ORFV infection in goats in the Central Anatolian region of Turkey were investigated. Samples were collected from 1- to 4-month-old kids (n = 9) in two different flocks in the Aksaray and Konya Provinces during the months of March and May 2016. The presence of ORFV in suspected samples was confirmed by PCR using primers specific to envelope gene (B2L). The analysis of the B2L gene sequences revealed that the nucleotide homology between the two isolates in the present study was 100%, whereas the similarity with Parapoxvirus isolates from different regions ranged from 83.6 to 99%. Phylogenetic analysis of the B2L gene revealed that there are two main clusters of ORFV isolates which were responsible for past outbreaks in Turkey. The information presented here will provide an insight into genetic diversity of field isolates of ORFV circulating in the Central Anatolian region of Turkey.

Serum Biochemistry of Lumpy Skin Disease Virus-Infected Cattle.

Lumpy skin disease is an economically important poxvirus disease of cattle. Vaccination is the main method of control but sporadic outbreaks have been reported in Turkey. This study was carried out to determine the changes in serum biochemical values of cattle naturally infected with lumpy skin disease virus (LSDV). For this study, blood samples in EDTA, serum samples, and nodular skin lesions were obtained from clinically infected animals (n = 15) whereas blood samples in EDTA and serum samples were collected from healthy animals (n = 15). A quantitative real-time PCR method was used to detect Capripoxvirus (CaPV) DNA in clinical samples. A real-time PCR high-resolution melt assay was performed to genotype CaPVs. Serum cardiac, hepatic, and renal damage markers and lipid metabolism products were measured by autoanalyzer. LSDV nucleic acid was detected in all samples which were obtained from clinically infected cattle. The results of serum biochemical analysis showed that aspartate aminotransferase, alkaline phosphatase, total protein, and creatinine concentrations were markedly increased in serum from infected animals. However, there were no significant differences in the other biochemical parameters evaluated. The results of the current study suggest that liver and kidney failures occur during LSDV infection. These findings may help in developing effective treatment strategies in LSDV infection.

Genetic characterization of peste des petits ruminants virus, Turkey, 2009-2013.

Peste des petits ruminants is an endemic disease of small ruminants in Turkey and vaccination has been the method of control but sporadic outbreaks have been reported. This study was carried out to characterize the local peste des petits ruminants virus (PPRV) by sequencing fusion (F) protein and nucleoprotein (N) gene segments and phylogenetic analysis, so as to focus on genetic variation in the field viruses. Samples were collected from sheep and goats clinically suspected of having PPRV infection in Central and Mediterranean regions of Turkey during 2009-2013. Phylogenetic analysis based on the F gene sequences showed that the field isolates in the present study belong to lineage 4 with other Middle East isolates. While N gene sequences revealed a different pattern, the field isolates in the present study clustered with previous Turkish isolates, which probably represents the true picture of molecular epidemiology for PPRV.

An 8-year longitudinal sero-epidemiological study of bovine leukaemia virus (BLV) infection in dairy cattle in Turkey and analysis of risk factors associated with BLV seropositivity.

Enzootic bovine leukosis (EBL) which is caused by bovine leukaemia virus (BLV) has an important economic impact on dairy herds due to reduced milk production and restrictions on livestock exports. This study was conducted to determine the BLV infection status in Central Anatolia Region of Turkey, an important milk production centre, and to examine the risk factors such as purchasing cattle, increasing cattle age, cattle breed and herd size associated with transmission of BLV infection. To estimate the rate of BLV infection, a survey for specific antibodies in 28,982 serum samples from animals belonging to 1116 different herds situated in Central Anatolia Region of Turkey were tested from January 2006 to December 2013. A generalized mixed linear model was used to evaluate the risk factors that influenced BLV seroprevalence. Antibodies against BLV were detected in 431 (2.28 %) of 18,822 Holstein and 29 (0.28 %) of 10,160 Brown Swiss cows. Among 1116 herds, 132 herds (11.82 %) had one or more positive animals. Also results of our study show that the prevalence of BLV infection increased from 2006 to 2011, and it tends to reduce with BLV control programme. Furthermore, we found positive associations between percentage of seropositive animal and increasing cattle age, herd size, cattle breed and purchased cattle. Age-specific prevalence showed that BLV prevalence increased with age. These factors should be taken into consideration for control of BLV infection.