Poster communication in AHVLA International Conference 2011

September 15^th, 2011

Perez-Sancho M., Alvarez J., Garcia N., Martinez I., Garcia-Seco T., Gonzalez S., Goyache J. and Dominguez L.

Due to important limitations (high cost, biosafety facilities, etc.) related to bacteriological confirmation of infection by zoonotic agents such as Brucella spp., alternative tools for its direct detection in clinical samples have been developed in the last years. Among them, the LAMP (Loop-mediated isothermal amplification) technique has shown high sensitivity and specificity and can be performed without sophisticated equipment. LAMP could represent an alternative tool for Brucella spp. detection especially in developing countries where limited resources are usually available. In this context, a LAMP protocol was developed based on the IS711 target. Primers were designed using Primer ExplorerV4 software program from Eiken Chemical. Reaction mix (Bst pol, betaíne,…) and the amplification protocol was optimized as follows: first, its ability for detection of the main zoonotic Brucella species (B. suis, B. melitensis and B. abortus) was tested using bacterial DNA as template. Second, usefulness of the LAMP for direct detection of Brucella spp. was evaluated on different animal samples (lung, spleen, liver, stomach content) showing an overall sensitivity of 37.4-69.3% depending on type of sample. Our LAMP was designed based on the multiple insertion element IS711 in order to ensure a highly sensitivity of the assay, and the results show that the protocol assayed may be a useful ancillary tool for confirmation of brucellosis at a flock level, and could be carried out using only a water-bath. However, its sensitivity may be affected by type of sample so several samples should be used in order to ensure Brucella detection

	Servicio de Zoonosis Emergentes, de Baja Prevalencia y Agresivos Biológicos (NED). Centro de Vigilancia Sanitaria Veterinaria (VISAVET). Universidad Complutense (UCM).
	Departamento de Sanidad Animal. Facultad de Veterinaria. Universidad Complutense (UCM).