, 2002; Hamamoto et al, 2004) The large size of the silkworm al

, 2002; Hamamoto et al., 2004). The large size of the silkworm allows for injection of quantitative amounts of samples into the hemolymph using syringes, a marked advantage over small invertebrate animals, including D. melanogaster and C. elegans (Kaito & Sekimizu, 2007; Kurokawa et al., 2007; Fujiyuki et al., 2010). The silkworm E7080 supplier can be maintained at 37 °C, the temperature at which

most pathogenic bacteria against humans show high virulence (Kaito et al., 2011a). Use of the silkworm model enabled us to identify S. aureus novel virulence genes, cvfA, cvfB, cvfC and sarZ, from hypothetical genes that are conserved among bacteria (Kaito et al., 2005, 2006; Matsumoto et al., 2007, 2010; Nagata et al., 2008; Ikuo et al., 2010). These genes contribute to virulence in mice and regulate the expression of hemolysins. Injection of α-hemolysin and β-hemolysin from S. aureus into silkworm hemolymph is lethal to silkworms (Hossain et al., 2006; Usui et al., 2009). α-Hemolysin and β-hemolysin contribute to S. aureus virulence Nutlin-3a chemical structure in mammals (O’Callaghan et al., 1997; Bubeck Wardenburg et al., 2007). Phenol-soluble modulins (PSMs) have recently been identified as cytolysins against erythrocytes and

neutrophils (Wang et al., 2007). Expression of these hemolysins is positively regulated by the agr locus (Novick, 2003; Queck et al., 2008). Our previous studies of the RN4220 strain transformed with the intact agr locus indicated that the agr locus contributes to S. aureus virulence in

silkworms (Kaito et al., 2005). These findings suggest that S. aureus possesses virulence factors that are not only specific for humans but also applicable Tangeritin to other invertebrates, and that the silkworm model is effective for the functional analysis of S. aureus virulence factors. The overall scope of S. aureus virulence factors that can be evaluated using the silkworm model, however, remains to be elucidated. In the present study, we evaluated virulence factors of S. aureus that have been characterized in mammalian infection models. Using mammalian systems, S. aureus genes encoding hemolysins and adhesins were identified to be involved in the infectious process (see Table 3 below). We constructed disruption mutants for these genes and compared their virulence in silkworms with that of the parent strain. We previously evaluated the virulence of the S. aureus RN4220 strain in silkworms (Kaito et al., 2005). The strain is constructed by mutagen treatment and contains previously unidentified mutations in the genome (Traber & Novick, 2006; Nair et al., 2011). For example, a point mutation in the agr locus, which positively regulates the expression of exotoxins, was discovered in the genome of RN4220, and results in decreased hemolysin production (Traber & Novick, 2006). Here, we used NCTC8325-4 as the parent strain. Escherichia coli JM109 was used as the host for plasmids.

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