Pathogenic microorganisms causing bovine mastitis: Isolation, identification, and susceptibility to biosynthesized silver nanoparticles.

A.P. FERREYRA MAILLARD, M.N. GALLUCCI, P.R. DALMASSO ; M.S. PELLEGRINO

Córdoba

Congreso; XI Congreso Argentino de Microbiología General.; 2015

Bovine mastitis is one of the infectious diseases with the most significant adverse economic impact to milk producers, and it can be caused by a wide variety of factors. However, mastitis is nearly always caused by bacteria, usually Staphylococcus (S. aureus) and Streptococcus (S. agalactiae, S. uberis), which account for over 90% of pathogenic microorganisms. Also, Gram negative bacteria, such as Escherichia coli, Klebsiella spp. and Enterobacter spp. have been reported to cause a variety of symptoms of the disease. Currently, antibiotic therapy plays an important role in eliminating existing intramammary infections.Nevertheless, an inadequate application of these treatments leads to the development of antimicrobial resistance strains. In this scenario, nanotechnology provides opportunities for adjusting the biological properties of nanoparticles (NPs) to generate effective antimicrobials. The aim of the present work was to determine the effect of NPs against the main bovine mastitis pathogens. For this, a simple and economic method of biosynthesis of silver nanoparticles (AgNPs) from AgNO3 usingGeoffroea decorticans (chañar) aqueous leaf extract is presented. Green synthesis of the AgNPs was confirmed by UV-vis spectroscopy and they were characterized by transmission electron microscopy (TEM). Bacterial strains in milk samples collected from cows showing clinical signs of mastitis were isolated and identified for their biochemical properties (catalase test, Gram stain, CHROM agar) and by molecular techniques. For Staphylococci pathogens, a PCR-RFLP analysis of the groEL gene and the profiles generated using restriction enzymes (AluI, PvuII, and HindIII) were used to confirm our identification results obtained by the conventional biochemical method. Biochemical and molecular evidence allowed to identify 12 Staphylococcus strains, 7 S. aureus, 2 S. simulans, 1 S. haemolyticus, and 1 S. hycus (only one strain cannot be identified by molecular techniques), and 10 Streptococcus strains: 8 S. uberis, 1 S. dysgalactiae, and 1 S. bovis. The in vitro susceptibility ofthe strains identified against the biosynthesized AgNPs was evaluated using disc agar diffusion and microplate techniques. The results obtained showed an excellent antibacterial effect of AgNPs for all of the mastitis-causing pathogens tested to levels of concentration (pM) about 6 orders of magnitude lower than those reported for conventional antibiotics (around mM). Streptococcus strains were inhibited using AgNPs concentrations up to 20 times lower than those needed to inhibitStaphylococcus. In comparison, higher levels of AgNPs concentration were requires to inhibite S. aureus than those necessary for other Staphylococcus. Thus, the biosynthesized AgNPs may be used as a novel unconventional antimicrobial strategy for treatment of infectious mastitis in cow due to the broad spectrum of their antibacterial properties.