Eda ALTAN, Gilberto SABINO-SANTOS JR, Patricia PESAVENTO, Eric DELWART

The first discovery of Chaphamaparvovirus in sheep with encephalitis and anemia

Objective: Parvoviruses have been shown to exist in sheep since 1987. To this date there are only three reports pertaining to the existence parvoviruses in sheep. The first reported parvovirus study did not provide genomic information whereas the latter two belonged to the tetraparvovirus and copiparvovirus genera. This study focused on discovering the possible reasons of encephalitis and anemia in a dead sheep whose tissue samples were submitted to our laboratory. Methods: In the present study next-generation sequencing (NGS) was utilized. Nextera™ XT Sample Preparation Kit was used to generate a library for Illumina MiSeq using dual barcoding. An in-house pipeline was used for analyzing raw data generated from Miseq. Several software were used to create in housepipeline to trim sequences and de novo assembly. For the alignment genome and phylogenetic tree Geneious and MEGA X software were used. Results: A novel ovine chaphamaparvovirus and pestivirus D both were characterized simultaneously. Although chaphamaparvoviruses had had been reported in various animals, this is the first time they have been reported in sheep. PCR analyses confirmed the presence of chaphamaparvovirus in multiple tissues. The partial nonstructural protein (NS1) and the complete capsid proteins (VP1) protein sequences displayed the closest amino acid identity of 49% and 69%, respectively, to the proteins of a non-human primate chapparvovirus from Macaca fascicularis. Conclusion: This ovine parvovirus is the fourth parvovirus and the first chaphamaparvovirus reported in sheep. Both chaphamaparvovirus and pestivirus were shown to co-exist simultaneously in sheep. The role of this dual virus infection in the disease signs of this sheep remains to be determined. This study will shed light on future chaphamaparvovirus studies in sheep.

Ensefalitli ve anemili koyunda Chaphamaparvovirüs’ün ilk keşfi

Amaç: Parvovirüsler koyunlarda ilk olarak 1987 yılında tespit edilmiştir. Günümüze kadar koyunlarda parvovirüslerin varlığına ilişkin sadece üç rapor vardır. İlk bildirilen parvovirüs çalışması genomik bilgi sağlamazken, son ikisi tetraparvovirus ve copiparvovirus cinslerine aittir. Bu çalışmada doku örnekleri laboratuvarımıza gönderilen sebebi belli olmayan ensefalit ve anemi tespit edilen ölü bir koyunda olası nedenleri keşfetmeye odaklanılmıştır. Yöntem: Bu çalışmada yeni nesil dizileme (NGS) kullanılmıştır. Nextera™ XT Numune Hazırlama Kiti, ikili barkod kullanarak Illumina MiSeq platformu için bir kitaplık oluşturmada kullanıldı. Miseq’ten üretilen ham verileri analiz etmek için şirket içi bir data analiz altyapısı kullanıldı. Okuma dizilerini kırpmak ve de novo bağlama analizi için çeşitli yazılımlar kullanıldı. Genomun hizalama işlemi ve filogenetik ağaç için Geneious ve MEGA X yazılımından faydalanıldı. Bulgular: Çalışmada yeni bir küçükbaş hayvan chaphamaparvovirüsü ve pestivirus D aynı anda karakterize edildi. Chaphamaparvovirüsler çeşitli hayvanlarda bildirilmiş olmasına rağmen koyunlarda ilk kez rapor edilmektedir. PCR analizleri, birçok dokuda chaphamaparvovirus varlığını doğruladı. Kısmi yapısal olmayan protein (NS1) ve tam kapsid proteinleri (VP1) protein dizileri, insan olmayan bir primat Macaca fascicularis’te bulunan chapparvovirüsün proteinlerine sırasıyla %49 ve %69’luk en yakın amino asit özdeşliğini sergiledi. Sonuç: Bu küçükbaş hayvan parvovirüsü koyunlarda bildirilen dördüncü parvovirüs ve ilk chaphamaparvovirustur. Hem chaphamaparvovirus hem de pestivirüs’ün koyunlarda aynı anda birlikte var olduğu gösterilmiştir. Bu koyunun hastalık belirtilerinde bu ikili virüs enfeksiyonunun rolü henüz belirlenmemiştir. Bu çalışma ileride koyunlarda yapılacak olan chaphamaparvovirus çalışmalarına ışık tutacak niteliktedir.

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