Detection of immunological markers in wildlife: a case study in Phyllotis darwini (Cricetidae) in Chile
5:15 a 5:30 pm, Centro de Estudios Científicos
Esperanza Beltrami1,2*, Claudio Henríquez3, Camilo Tomckowiack1, Claudio Verdugo4,5, Pablo Beldoménico6, Josefina Gutiérrez4,5, Javiera Delgado1, Gerardo Acosta-Jamett1,5*
- Instituto de Medicina Preventiva Veterinaria, Facultad de Ciencias Veterinarias, Universidad Austral de Chile, Valdivia, Chile; 2. Escuela de Graduados, Facultad de Ciencias Veterinarias, Universidad Austral de Chile, Valdivia, Chile; 3. Instituto de Farmacología y Morfofisiología, Facultad de Ciencias Veterinarias, Universidad Austral de Chile, Valdivia, Chile; 4. Instituto de Patología Animal, Facultad de Ciencias Veterinarias, Universidad Austral de Chile, Valdivia, Chile; 5. Programa de Investigación Aplicada en Fauna Silvestre, Facultad de Ciencias Veterinarias, Universidad Austral de Chile, Valdivia, Chile; 6. Laboratorio de Ecología de Enfermedades, Instituto de Ciencias Veterinarias del Litoral (ICiVet-Litoral), Universidad Nacional del Litoral (UNL) – Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), Santa Fe, Argentina.
Ponente: Esperanza Beltrami, firstname.lastname@example.org
Studying immune responses in wildlife species is challenging, due to a great lack of background knowledge about genetic and physiological aspects of wild species, and the need to adapt some laboratory techniques from their domestic counterparts. We studied immunological markers that represent defence mechanisms in a wild small mammal, Phyllotis darwini, captured in a semi-arid area of the Coquimbo region of Chile. We focused on detecting GATA3, IFNG, TBX21 and IL10, as well as potential internal control genes. We show technical steps and discuss the complexities for undertaking this research. From peripheral blood samples of live P. darwini, we extracted mRNA and RT-qPCR was performed to measure gene expression. This gene expression was standardized under a housekeeping gene to measure DCt. Specific primers were designed based on reference sequences of the genes and considering conserved segments from several species by a multiple alignment analysis. Gene expression was detected by real-time PCR and products sequenced. From a total of 40 blood samples of P. darwini, GATA3 amplified in 27.5% of the samples, TBX21 in 60% and the housekeeping, YWHAZ in 87.5% of the samples. We failed to detect IFNG, IL10, and the housekeeping SDHA in blood samples with any of the designed primers. Average DCt was higher for GATA3 (6.8) than for TBX21 (3.9), indicating a higher expression of the later which is associated to an inflammatory and Th1 immune response. This is the first study that reports immunological markers sequences from a South American wild mammal. The immunological markers identified here represent phenotypic subsets of lymphocytes with specific defence functions against infection. Future quantification of the expression levels of these genes will allow us to better understand how immune phenotypes are allocated under different parasite types and how responses vary across contrasting environmental scenarios. Fondecyt 1180119, 1211190, PhD grant 21171018, 21201700.