Staphylococcus aureus is an important pathogen, with methicillin-resistant (MRSA) and multi-drug resistant strains becoming increasingly prevalent in both human and veterinary clinics. S. aureus causing bovine mastitis yields high annual losses to the dairy industry. Conventional treatment of mastitis by broad range antibiotics is often not successful and may contribute to development of antibiotic resistance. Bacteriophage endolysins present a promising new source of antimicrobials. The endolysin of prophage ΦSH2 of Staphylococcus haemolyticus strain JCSC1435 (ΦSH2 lysin) is a peptidoglycan hydrolase consisting of two catalytic domains (CHAP and amidase) and an SH3b cell wall binding domain. In this work, we demonstrated its lytic activity against live staphylococcal cells and investigated the contribution of each functional module to bacterial lysis by testing a series of deletion constructs in zymograms and turbidity reduction assays. The CHAP domain exhibited three-fold higher activity than the full length protein and optimum activity in physiological saline. This activity was further enhanced by the presence of bivalent calcium ions. The SH3b domain was shown to be required for full activity of the complete ΦSH2 lysin. The full length enzyme and the CHAP domain showed activity against multiple staphylococcal strains, including MRSA strains, mastitis isolates, and CoNS.

Monoclonal antibodies (MAb) were produced against two B. pseudomallei high-purified membrane proteins with Mr 29 kDa (p29) and 45 kDa (p45). Monoclonal antibodies from culture supernatant fluids of 4F2 and 1G11 clones showed specific interaction with protein moiety of p29 both Burkholderia pseudomallei and Burkholderia mallei in ELISA and Western blotting. However, MAb of 3G4 clone were bound to the LPS-protein structures of these microbial cells. Analysis of interaction of Mabs from 4F2 and 1G11 clones with antigens of different lysates of pathogenic cells confirmed high specificity of these antibodies to p29 membrane protein of B. pseudomallei and B. mallei.