Leucotoxic Activities of Staphylococcus aureus Strains Isolated from Cows, Ewes, and Goats with Mastitis: Importance of LukM/LukF′-PV Leukotoxin

Abstract

ABSTRACT Among the toxins that Staphylococcus aureus is able to secrete, bi-component toxins named leukotoxins target specifically leukocytes, mainly phagocytic cells. Isolates from cows, goats and ewes with mastitis were selected on the basis of the presence or not of the genes encoding the recently described LukM/LukF′-PV leukotoxin. Of the 128 isolates tested, 126 had moderate to high leukotoxic activity to bovine polymorphonuclear cells (PMN). The supernatants of lukM -positive isolates were much more leukotoxic than the supernatants of lukM -negative isolates: mean leukotoxic titers were 122 versus 20 and 581 versus 26 for isolates of bovine and caprine origin, respectively. Among lukM / lukF ′- PV positive isolates, those of caprine and ovine origins were more leukotoxic than were isolates of bovine origin ( P < 0.01). The two most abundant proteins in the culture supernatant of a highly toxic isolate were purified and identified as the two components of LukM (LukM and LukF′-PV) on the basis of their molecular mass, N-terminal amino acid sequence, and high synergistic activity. LukM/LukF′-PV induced the flattening of bovine PMN at a concentration as low as 3.6 ng/ml (0.1 nM). A higher concentration (18 ng/ml) was necessary to produce the same effect on caprine or ovine PMN. Affinity-purified antibodies to LukM or to LukF′-PV neutralized the leukotoxic effect of all the culture supernatants. They neutralized with the same efficiency the toxic activity of supernatants from lukM / lukF ′-PV positive or negative isolates. These results establish that LukM/LukF′-PV is very active on PMN of ruminants and suggest that this leukotoxin could be the most active leukotoxin produced by mastitis isolates. They prompt further studies to delineate the contribution of LukM/LukF′-PV to the pathogenesis of mastitis in ruminants and the protective effect of antibodies to this leukotoxin.