Recipient Organization
UNIVERSITY OF CALIFORNIA, DAVIS
410 MRAK HALL
DAVIS,CA 95616-8671
Performing Department
Population Health & Reproduction
Non Technical Summary
Coccidiosis in small ruminants is an important economic, animal health and welfare concern. Currently, producers and practitioners rely on oocyst count to strategically implementanti-coccidial therapy. Unfortunately, an oocyst count alone doesn't decifer thecoccidia that are likely to cause disease or even have an impact on production. This study will give us an opportunity to learn more about how management affects parasite burden and parasite population dynamics within the small ruminant gastrointestinal tract. The objectives of this project are to determine relationships between management practices, level of oocyst excretion (parasite burden), pathology of the intestines atnecropsy, and ecology of multi-species coccidia infections. An important outcome of this project is to develop a molecular-based test to be able to identify and quantify specific coccidian species in fecal samples. With this study we can lay a lot of ground work on developing management strategies to prevent coccidial infections and target treatment based on an informative diagnostic test.
Animal Health Component
50%
Research Effort Categories
Basic
25%
Applied
50%
Developmental
25%
Goals / Objectives
Coccidiosis causes clinical disease (severe diarrhea, anemia, ill thrift, chronic inflammatory bowel disease) and production losses predominately in young stock. Small ruminant producers in the US identify internal parasitism as a top health concern according to needs-assessment surveys conducted by NAHMS. Current strategies to control coccidiosis rely on oocyst counts, which count both pathogenic and non-pathogenic species, and may lead to overuse of anticoccidial treatments and consequnetly resistance.Development of tools to aid selective anticoccidial use is warranted given recent evidence for resistance of ovine Eimeria to the anticoccidial drug toltrazuril. While Eimeria spp. are host specific, understanding risk factors in both goats and sheep would provide a framework for strategic control methods and targeted management recommendations. The various Eimeria species affecting small ruminants have been identified using morphological differentiation of species, however this is time intensive and requires a high level of expertise that is not available for clinical samples at diagnostic laboratories. The development of a diagnostic platform that can identify the various species of Eimeria present in one fecal sample with a rapid turnaround could be practically applied by veterinarians and livestock owners to improve parasite management.Using validated and published quantitative and species-specific PCR methods for Eimeria in ruminants molecular quantification and species determination will be performed. In this study we aim to further adapt molecular methods for high throughput testing using multiplex qPCR and/or targeted sequencing.SPECIFIC OBJECTIVESDetermine fecal oocyst counts on small ruminant farms in northern California.Determine association between oocyst counts and farm factors (management practices including anticoccidial use, environmental and spatial factors) and presence and severity of clinical and histopathologic coccidiosis.Determine association between oocyst species causing infection and farm factors and presence and severity of clinical and histopathologic coccidiosis.Development of a high throughput platform for speciation of coccidia in clinical samples.
Project Methods
Aim 1: An on-farm survey will be conducted in three California regions: North Coast, Sacramento Valley, and the Sierra Foothills. These regions are served by the Davis CAHFS lab and locally by UCCE advisors, have diverse climates and production types, and represent 25% of all small ruminant farms and the animal inventory for California. We are targeting 20 farms on the coast, 10 farms in the valley, and 10 farms in the foothills. Farm inclusion criteria for participation are 1) more than 10 production age sheep or goats; and 2)raise young stock. Study participants will complete a short survey regarding housing, lamb and kid rearing practices, coccidia prevention or treatment strategies and history of morbidity or mortality associated with coccidiosis. Twelve fecal samples will be collected per farm from lambs/kids during peak oocyst shedding, between 2 and 5 months old. A total of 480 samples will be collected across the 3 regions based on expected 25% prevalence, herd sensitivity of 80% and specificity of 95%. The Five Point Check (FAMACHA score, body condition score, subcutaneous edema, and dag score/diarrhea), hematocrit, and total protein will be used as indicators of clinical severity of coccidia infection. Quantitative fecal analysis will be performed on fecal samples to quantify coccidia burden in oocysts/gram of feces. Total coccidia DNA will be extracted from oocysts recovered from positive samples and stored at -80C for Aim 3. Aim 2: Recruited farms from Aim 1 will be encouraged submit any severely sick or dead lambs/kids between 2 and 9 months old to CAHFS for necropsy. Animals submitted to CAHFS will have full necropsy performed (including histopathology of small intestine) and feces collected for quantitative fecal analysis and harvesting of DNA which will be stored at -80C for Aim 3. A scoring system will be developed to characterize severity of intestinal pathologic changes. Aim 3: A previously described quantitative PCR assay will be employed to determine the number of Eimeria oocysts present in fecal samples and validated with Modified McMaster's. Sequence data for the different species is available through NCBI and validated and published species-specific PCR assays targeting the 18S ribosomal RNA (rRNA) locus and ITS-1 region of Eimeria exist. The quantity and species composition of Eimeria oocysts will be determined by multiplex qPCR or targeted NGS of PCR amplicons (from validated and published PCR assays). The utility of targeting these loci to develop a high throughput approach utilizing either multiplex qPCR or targeted NGS for rapid quantitative determination of species will be evaluated. Platforms that will be investigated for NGS include iSeq (Illumina) and Minion (Oxford Nanpopore Technologies). The development of this assay will be used to determine the relative numbers of each coccidia species in the stored DNA from all collected samples.