The novel polysaccharide deacetylase homologue Pdi contributes to virulence of the aquatic pathogen Streptococcus iniae

Abstract

The aquatic zoonotic pathogen Streptococcus iniae represents a threat to the worldwide aquaculture industry and poses a risk to humans who handle raw fish. Because little is known about the mechanisms of S. iniae pathogenesis or virulence factors, we established a highthroughput system combining whole-genome pyrosequencing and transposon mutagenesis that allowed us to identify virulence proteins, including Pdi, the polysaccharide deacetylase of S. iniae, that we describe here. Using bioinformatics tools, we identified a highly conserved signature motif in Pdi that is also conserved in the peptidoglycan deacetylase PgdA protein family. A Dpdi mutant was attenuated for virulence in the hybrid striped bass model and for survival in whole fish blood. Moreover, Pdi was found to promote bacterial resistance to lysozyme killing and the ability to adhere to and invade epithelial cells. On the other hand, there was no difference in the autolytic potential, resistance to oxidative killing or resistance to cationic antimicrobial peptides between S. iniae wild-type and Dpdi. In conclusion, we have demonstrated that pdi is involved in S. iniae adherence and invasion, lysozyme resistance and survival in fish blood, and have shown that pdi plays a role in the pathogenesis of S. iniae. Identification of Pdi and other S. iniae virulence proteins is a necessary initial step towards the development of appropriate preventive and therapeutic measures against diseases and economic losses caused by this pathogen. INTRODUCTIONIn the past few decades, the pathogen Streptococcus iniae has emerged as a major hindrance to aquaculture operations worldwide (Agnew & Barnes, 2007), causing economic losses measured in hundreds of millions of dollars annually. In addition, S. iniae has established itself as a zoonotic risk, especially in areas of the world that preferentially prepare and consume raw fish (Lau et al., 2003). To date, at least 25 human cases of invasive streptococcal infection attributed to S. iniae have been confirmed in the USA, Canada, China and Taiwan, many of which were in immunocompromised patients (Agnew & Barnes, 2007;Sun et al., 2007;Weinstein et al., 1997); since there is currently no prospective epidemiological surveillance for human S. iniae infections, the true number may be much higher (Facklam et al., 2005;Lau et al., 2006).In spite of the economic and human health risks that S. iniae presents, a fully assembled genome sequence for this emerging pathogen is not yet available, and very little is known about its disease mechanisms. To better understand the molecular genetic basis of virulence in this versatile pathogen, we created a library of randomly generated transposon mutants that we screened for virulence Abbreviations: AMP, antimicrobial peptide; HSB, hybrid striped bass, i.p., intraperitoneal; PIA, polysaccharide intercellular adhesin, WT, wild-type.3These authors contributed equally to this work.The GenBank/EMBL/DDBJ accession number for the pdi sequence of Streptococcus iniae is FJ664396.Four supplementary f...

Listeria monocytogenes Is Resistant to Lysozyme through the Regulation, Not the Acquisition, of Cell Wall-Modifying Enzymes

“…Therefore, exposure to blood is likely to occur during human listeriosis, where L. monocytogenes causes meningitis, abortion in pregnant women, and sepsis (11). The results of this and other studies suggest that the role of lysozyme in vivo is multifactorial (9,10,12,50,67) but that serum lysozyme is the major factor that protects against lysozyme-sensitive L. monocytogenes during i.v. infection.…”

“…S5A in the supplemental material). We next reasoned that the major bactericidal factor could be macrophages, neutrophils, or serum, since all have been previously implicated in the killing of lysozyme-sensitive bacteria (9,50,51). Upon the infection of bone marrow-derived macrophages, only a small defect was observed when the pgdA oatA pbpX triple mutant, the most lysozyme-sensitive strain, was used (see Fig.…”

The Bacillus subtilis Extracytoplasmic Function σ Factor σ V Is Induced by Lysozyme and Provides Resistance to Lysozyme

“…Staphylococcus aureus and Enterococcus faecalis both utilize O-acetyltransferases to acetylate the C6-OH group of MurNAc (4,19,27). Bacillus cereus, Bacillus anthracis, and several Streptococcus species encode PG deacetylases which remove the acetyl group from GlcNAc, which leads to increased lysozyme resistance (16,31,34,43).…”

“…Staphylococcus aureus and Enterococcus faecalis both utilize O-acetyltransferases to acetylate the C6-OH group of MurNAc (4,19,27). Bacillus cereus, Bacillus anthracis, and several Streptococcus species encode PG deacetylases which remove the acetyl group from GlcNAc, which leads to increased lysozyme resistance (16,31,34,43).In B. subtilis, several studies have identified genes which are regulated by V ; however, the signal(s) which induces V activity has not been identified (3, 47). We had previously found that only lysozyme was capable of inducing the expression of a sigV homolog, csfV, in Clostridium difficile (20).…”

The Streptococcus iniae Transcriptional Regulator CpsY Is Required for Protection from Neutrophil-Mediated Killing and Proper Growth In Vitro

“…S. iniae also contains a polysaccharide capsule that functions for protection from phagocytic clearance in whole blood (32,34,39). Furthermore, several genes not previously studied for their virulence traits have been shown to have critical roles during S. iniae infection of hybrid striped bass (HSB), including phosphoglucomutase (pgmA) (8) and a novel polysaccharide deacetylase (pdi) (38).…”