The aim of the work was to develop a new, robust, portable diagnostic system based on the latest development of loop-mediated isothermal amplification (LAMP) technology for the detection of the main causative agent of contagious agalactia, an important disease of small ruminants. The integrated diagnostic system consists of a portable instrument and a kit, specific for the pathogen. The kit is ready to use and provides both the reagents for a rapid nucleic acid extraction from milk, and the amplification reagents using LAMP technology, The test showed a high sensitivity identifying only M.agalactiae DNA with a detection limit of less than 100 CFU / ml in milk samples, exceeding the traditional laboratory diagnostics for speed and handling The test also proved to be more tolerant to the inhibitory effect of milk components that affect amplification in the PCR. These preliminary results show that the LAMP system can be a practical and effective diagnostic tool for the diagnosis in the field of M.agalactiae pathogen.

Mycoplasma diseases of livestock such as contagious bovine pleuropneumonia, contagious caprine pleuropneumonia and contagious agalactia still represent major problems for animal health authorities worldwide. Many significant improvements have been seen as a result of research into these degenerate bacteria, mainly in the area of diagnosis, with the availability of specific, effective new molecular tools to detect small quantities of these pathogens (some of them highly fastidious, unculturable and often in mixed bacterial culture) directly from clinical samples. The objective of the present study was to identify and differentiate Mycoplasma species of sheep, goats and cattle by a new diagnostic test based on PCR of the 16S rRNA gene with specific primers and separation of the PCR product according to primary sequence using denaturing gradient gel electrophoresis (DGGE). Detection and identification results were compared with the ones obtained by sequence analysis. The development of a PCR DGGE offers advantages of a rapid identification of many Mycoplasma species for which no specific PCR is yet available, enabling the differentiation of animal Mycoplasma species. It represents a significant improvement from conventional culture and serological tests in term of specificity and time to perform.