Equine herpesvirus type 1 (EHV-1) is a common pathogen of horses that typically causes upper respiratory disease, however is also associated with late-term abortion, neonatal foal death and neurologic disease. Once a horse is infected, the virus concentrates to local lymphoid tissue, where it becomes latent. The virus can recrudesce during times of stress, which can lead to the initiation of devastating outbreaks. Some variants of EHV-1 have been associated with more severe disease outcomes. Appropriate outbreak management focuses on minimizing the movement of potentially exposed horses. This approach lacks a strategy for prevention at the level of latency largely due to a knowledge paucity in regards to carriage rate of latent EHV-1. Biosecurity decisions are also dependent on awaiting currently-available diagnostic testing that often take several days for results. Thus, our work has been focused on understanding the carriage rate of the latent virus in different geographic regions as well as improving diagnostic efficiency, both of which are essential for improving the management of EHV-1 disease. Loop mediated isothermal amplification (LAMP) is a method that amplifies nucleic acid rapidly at a constant temperature and is minimally affected by inhibitors that are often found in clinical samples. This procedure can be followed by multiple detection methods. A new, efficient sequencing method, called nanopore sequencing, has been developed in a handheld device, called MinION, that provides thorough output in a timely manner. When combined with LAMP, it has been referred to as LAMPore. The first objective of our work was to estimate the prevalence of latent EHV-1 and compare the frequency of each variant in the submandibular lymph nodes from horses in Virginia. Our second objective was to perform direct DNA sequencing of EHV-1 using the mobile MinION sequencer in combination with LAMP viral enrichment. Our findings demonstrated a low apparent prevalence of latent EHV-1 DNA in submandibular lymph nodes in this population of horses in Virginia as well as successful detection and identification of EHV-1 in equine nasal swab samples using LAMPore sequencing. / Doctor of Philosophy / Horses can develop disease from a virus called equine herpesvirus type 1 (EHV-1). Symptoms can vary from mild respiratory signs to the inability to rise leading to death or euthanasia. Horses transmit this virus to other nearby horses; however, the virus also becomes dormant once a horse is infected, meaning the virus is not infectious but is present within the animal. When the horse undergoes stress, such as during travel or competition, the virus can become active again, leading to the spread to other horses. This results in outbreaks, many of which are devastating to the equine industry. In order to minimize the risks of this virus spreading and causing disease, management is currently focused on minimizing the movement of horses that may have been exposed to the virus. There is little information regarding the number of horses that harbor the dormant virus and the current methods to detect the infectious virus can take multiple days for results. These limit decision-making during the management of an outbreak. Our work seeks to determine the number of horses in a region that harbor EHV-1 and also to test a new, efficient diagnostic method to identify the virus in samples from horses. Our findings showed a low number of horses in Virginia harbor dormant EHV-1 in the lymph nodes under their mandible, a common site of dormancy. Further, we found that our new method of detection was effective in identifying the virus in samples from nasal secretions from horse.

Made available in DSpace on 2014-08-20T14:37:54Z (GMT). No. of bitstreams: 1 tese_clairton_marcolongo_pereira_resumo.pdf: 10803 bytes, checksum: 3f7507f184a7adb5ceb1296595a35eca (MD5) Previous issue date: 2014-02-21 / This thesis is a study developed in period of 34 years about equine diseases diagnosed through out 34 municipalities of Southern Rio Grande do Sul, which includes de influence area of the Laboratório Regional de Diagnóstico of the Veterinary School of the Federal University of Pelotas. There is a general paper about the diseases diagnosed in this specie related by etiologic agent. This paper establishes the importance of dermatological diseases, including equine sarcoid and squamous cell carcinoma as a cause of economic losses in equines. It also demonstrates that leucoencephalomalacia, rabies, thromboembolism by Strongylus vulgaris and equine monocytic ehrlichiosis are the leading causes of death in horses in the region. Other three papers are presented: two related to the causes of abortion and the one about the epidemiology of pythiosis. These conditions are important causes of economic losses in the breeding of horses in the region. / Esta tese trata-se de um estudo das enfermidades de equinos diagnosticadas em 34 municípios da região Sul do Rio Grande do Sul, que compreende a área de influência do Laboratório Regional de Diagnóstico da Faculdade de Veterinária da Universidade Federal de Pelotas, em um período de 34 anos. É apresentado um trabalho geral das enfermidades diagnosticadas nesta espécie animal, relacionadas por agente etiológico tendo sido estabelecida a importância das doenças dermatológicas, dentre elas o sarcoide equino e o carcinoma de células escamosas como causas de prejuízos econômicos nesta espécie animal. Com este estudo foi possível, ainda, demonstrar que a leucoencefalomalacia, a raiva, o tromboembolismo por Strongylus vulgaris e a erliquiose monocítica são as principais causas de morte de equinos na região. São apresentados, também, dois trabalhos científicos referentes às causas de aborto e um à epidemiologia da pitiose, respectivamente, que são importantes causas de prejuízos econômicos na criação de equinos na região, identificadas neste estudo.