Unlocking the role of a genital herpesvirus, otarine herpesvirus 1, in California sea lion cervical cancer

This research was funded by the Geoffrey Hughes Research Fellowship and The Marine Mammal Center.Urogenital carcinoma in California sea lions (Zalophus californianus) is the most common cancer of marine mammals. Primary tumors occur in the cervix, vagina, penis, or prepuce and aggressively metastasize resulting in death. This cancer has been strongly associated with a sexually transmitted herpesvirus, otarine herpesvirus 1 (OtHV1), but the virus has been detected in genital tracts of sea lions without cancer and a causative link has not been established. To determine if OtHV1 has a role in causing urogenital carcinoma we sequenced the viral genome, quantified viral load from cervical tissue from sea lions with (n = 95) and without (n = 163) urogenital carcinoma, and measured viral mRNA expression using in situ mRNA hybridization (Basescope®) to quantify and identify the location of OtHV1 mRNA expression. Of the 95 sea lions diagnosed with urogenital carcinoma, 100% were qPCR positive for OtHV1, and 36% of the sea lions with a normal cervix were positive for the virus. The non-cancer OtHV1 positive cases had significantly lower viral loads in their cervix compared to the cervices from sea lions with urogenital carcinoma. The OtHV1 genome had several genes similar to the known oncogenes, and RNA in situ hybridization demonstrated high OtHV1 mRNA expression within the carcinoma lesions but not in normal cervical epithelium. The high viral loads, high mRNA expression of OtHV1 in the cervical tumors, and the presence of suspected OtHV1 oncogenes support the hypothesis that OtHV1 plays a significant role in the development of sea lion urogenital carcinoma.Publisher PDFPeer reviewe

Efecto de la fragmentación y pérdida de hábitat sobre la abundancia de áfidos, parasitoides y parasitoidismo en cultivos de alfalfa experimentales

Memoria para optar al Título Profesional de Médico VeterinarioLa fragmentación del hábitat per se es la ruptura de un hábitat originalmente continuo en dos o más fragmentos, los que quedan separados por un hábitat diferente al original denominado matriz. Por otro lado, la pérdida de hábitat es la eliminación de hábitat, sin que necesariamente ocurra fragmentación. En la literatura estos dos términos usualmente se confunden. Por ello, en el presente estudio evaluamos sus efectos en conjunto y por separado sobre la abundancia de los áfidos Aphis craccivora y Therioaphis trifolii, sus parasitoides y el parasitoidismo en cultivos experimentales de alfalfa. Se sembraron 20 parches de alfalfa, 16 de los cuales fueron sometidos a dos niveles de fragmentación (4 ó 16 fragmentos) y dos niveles de pérdida de hábitat (55% ó 84%). Los 4 paisajes restantes no fueron sometidos a fragmentación y pérdida de hábitat y se utilizaron como control. Se analizaron los efectos inmediatamente luego de la fragmentación (empaquetamiento), en el corto (verano), mediano (otoño) y largo (primavera) plazo. Inmediatamente luego de la fragmentación no hubo un empaquetamiento ni de áfidos ni de parasitoides en los paisajes fragmentados. En general, en ningún periodo se encontró un efecto de la pérdida de hábitat o fragmentación per se sobre los áfidos y parasitoides. Sin embargo, la fragmentación y pérdida de hábitat en conjunto afectaron negativamente a A. craccivora y positivamente a T. trifolii respecto a un paisaje no perturbado. Adicionalmente, A. craccivora presentó mayores abundancias en los bordes que en los centros de los fragmentos en los paisajes con mayor fragmentación y pérdida de hábitat. La abundancia de parasitoides y el parasitoidismo no variaron significativamente entre paisajes, aunque la abundancia de parasitoides fue mayor en los bordes que en los centros de los fragmentos. Estos resultados sugieren que, independientemente de la fragmentación de un paisaje, los bordes de los cultivos deben ser considerados en el manejo de plagas

Identification of 3 novel herpesviruses in prosimians with lymphoproliferative disease

Although many studies have characterized catarrhine and platyrrhine primate herpesviruses, little is known about herpesviruses in prosimians. We aimed to identify and characterize herpesviruses in prosimians with proliferative lymphocytic disease. DNA was extracted from tissues of 9 gray mouse lemurs ( Microcebus murinus) and 3 pygmy slow lorises ( Nycticebus pygmaeus) with lymphoproliferative lesions, and we performed nested PCR and sequencing for detection of herpesviruses and polyomaviruses. We identified 3 novel herpesviruses and performed phylogenetic analyses to characterize their relationship with other herpesviruses. A gray mouse lemur herpesvirus clustered with other primate herpesviruses within the subfamily Betaherpesvirinae, just basal to the genus Cytomegalovirus. The other gray mouse lemur herpesvirus and the pygmy slow loris herpesvirus clustered within the subfamily Gammaherpesvirinae, although the relationships within the subfamily were less resolved. Quantitative PCR assays were developed for the 2 new gray mouse lemur viruses, providing specific, faster, less expensive, and quantitative detection tools. Further studies are needed to elucidate the relationship between the presence of these viruses and the severity or presence of lymphoproliferative lesions in prosimians

Novel application of single-cell next-generation sequencing for determination of intratumoral heterogeneity of canine osteosarcoma cell lines

Osteosarcoma (OSA) is a highly aggressive and metastatic neoplasm of both the canine and human patient and is the leading form of osseous neoplasia in both species worldwide. To gain deeper insight into the heterogeneous and genetically chaotic nature of OSA, we applied single-cell transcriptome (scRNA-seq) analysis to 4 canine OSA cell lines. This novel application of scRNA-seq technology to the canine genome required uploading the CanFam3.1 reference genome into an analysis pipeline (10X Genomics Cell Ranger); this methodology has not been reported previously in the canine species, to our knowledge. The scRNA-seq outputs were validated by comparing them to cDNA expression from reverse-transcription PCR (RT-PCR) and Sanger sequencing bulk analysis of 4 canine OSA cell lines (COS31, DOUG, POS, and HMPOS) for 11 genes implicated in the pathogenesis of canine OSA. The scRNA-seq outputs revealed the significant heterogeneity of gene transcription expression patterns within the cell lines investigated (COS31 and DOUG). The scRNA-seq data showed 10 distinct clusters of similarly shared transcriptomic expression patterns in COS31; 12 clusters were identified in DOUG. In addition, cRNA-seq analysis provided data for integration into the Qiagen Ingenuity Pathway Analysis software for canonical pathway analysis. Of the 81 distinct pathways identified within the clusters, 33 had been implicated in the pathogenesis of OSA, of which 18 had not been reported previously in canine OSA