Influence of Temperature on the Physiology and Virulence of the Insect Pathogen Serratia sp. Strain SCBI

Abstract

The physiology of a newly recognized Serratia species, termed South African Caenorhabditis briggsae Isolate (SCBI), which is both a nematode mutualist and an insect pathogen, was investigated and compared to that of Serratia marcescens Db11, a broadhost-range pathogen. The two Serratia strains had comparable levels of virulence for Manduca sexta and similar cytotoxic activity patterns, but motility and lipase and hemolytic activities differed significantly between them. M embers of the genus Serratia are found widespread around the globe and are well known for their roles as insect pathogens (14). A newly recognized Serratia species, termed South African Caenorhabditis briggsae Isolate (SCBI), was identified following its isolation from the nematode C. briggsae KT0001 (2). These C. briggsae KT0001 nematodes were recovered from soil samples through Galleria mellonella bait traps in three provinces in South Africa (2). Serratia sp. strain SCBI is lethal to G. mellonella. When directly injected into the hemocoel in numbers of less than 1,000 CFU, larvae die within 72 h. A distinct characteristic of Serratia sp. SCBI is that it forms an apparent association with C. briggsae KT0001 as well as other Caenorhabditis nematodes, including Caenorhabditis elegans, which results in entomopathogenicity of the nematode (1, 2). This microbe-nematode association between Serratia sp. SCBI and C. briggsae may represent a potential emerging entomopathogenic association. Only two other Serratia species are known to use a nematode partner to establish an infection in an invertebrate host (40,43).Based on 16S phylogeny, Serratia sp. SCBI is closely related to S. marcescens Db11 (3), a reported pathogen of C. elegans (24,34,36). We have sequenced the entire Serratia sp. SCBI genome and have performed an analysis comparing it to the S. marcescens Db11 genome (F. Abebe-Akele, L. S. Tisa, V. Cooper, P. J. Hatcher, E. Abebe, and W. K. Thomas, unpublished data). S. marcescens Db11 is a well-known pathogen of vertebrates and invertebrates, including Caenorhabditis nematodes. Although their 16S RNA genes are 99% identical, genome-wide assessment (23) supports the idea that Serratia sp. SCBI and S. marcescens Db11 represent two distinct species. However, the high (about 80%) similarity at the 16S rRNA gene and within open reading frames indicates that Serratia sp. SCBI and S. marcescens Db11 share a close evolutionary relationship despite their distinct associations with Caenorhabditis nematodes. Since mutualistic associations have the potential to evolve from parasitic relationships (10), it is possible that Serratia sp. SCBI diverged from S. marcescens, making the leap from Caenorhabditis pathogen to a mutualistic relationship.Although the virulence factors of S. marcescens have been well studied (6,8,24,26,33), there have been no studies on the physiological properties of Serratia sp. SCBI and their contribution to pathogenesis. Due to their close evolutionary history, a comparative study of Serratia sp. SCBI and S. marcescens Db11 was cond...

Molecular Characterization of Protease Activity in Serratia sp. Strain SCBI and Its Importance in Cytotoxicity and Virulence

“…Cytotoxic activity of bacterial culture supernatant was determined on BGMK cells as described previously (4). Briefly, cell viability assays were carried out in 96-well flat-bottom plastic microplates.…”

“…Serratia sp. SCBI miniHimar RB1 mutants defective in protease activity were analyzed for their ability to kill thirdinstar M. sexta larvae using methods described previously (4). Briefly, overnight bacterial cultures of mutants were initially diluted to an OD 600 of 1.0 in LB medium.…”

“…Proteolytic activity was determined by a liquid azocasein assay described previously (4). Briefly, cultures were grown overnight at 28°C, which is the optimal temperature for protease activity in Serratia sp.…”

“…While Serratia sp. strain SCBI is nonpathogenic to Caenorhabditis nematodes, these bacteria are lethal to G. mellonella and the tobacco hornworm, Manduca sexta (4). When injected into the hemocoel of either species in numbers less than 1,000 CFU, larvae die within 72 h.…”

“…strain SCBI has protease, lipase, DNase, alkaline phosphatase, and chitinase activities, and is cytotoxic toward the mammalian Buffalo green monkey kidney (BGMK) cell line (4). The Serratia sp.…”