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Streptococcus iniae M-like protein contributes to virulence in fish and is a target for live attenuated vaccine development.

Locke JB, Aziz RK, Vicknair MR, Nizet V, Buchanan JT - PLoS ONE (2008)

Bottom Line: Our hybrid striped bass (HSB) and zebrafish models of infection revealed that M-like protein contributes significantly to S. iniae pathogenesis whereas C5a peptidase-like protein does not.Analysis of M-like protein and C5a peptidase through allelic replacement revealed that M-like protein plays a significant role in S. iniae virulence, and the Mga-like locus, which may regulate expression of this gene, has an unusual arrangement.The M-like protein mutant created in this research holds promise as live-attenuated vaccine.

View Article: PubMed Central - PubMed

Affiliation: Department of Pediatrics, University of California San Diego, La Jolla, California, United States of America.

ABSTRACT

Background: Streptococcus iniae is a significant pathogen in finfish aquaculture, though knowledge of virulence determinants is lacking. Through pyrosequencing of the S. iniae genome we have identified two gene homologues to classical surface-anchored streptococcal virulence factors: M-like protein (simA) and C5a peptidase (scpI).

Methodology/principal findings: S. iniae possesses a Mga-like locus containing simA and a divergently transcribed putative mga-like regulatory gene, mgx. In contrast to the Mga locus of group A Streptococcus (GAS, S. pyogenes), scpI is located distally in the chromosome. Comparative sequence analysis of the Mgx locus revealed only one significant variant, a strain with an insertion frameshift mutation in simA and a deletion mutation in a region downstream of mgx, generating an ORF which may encode a second putative mga-like gene, mgx2. Allelic exchange mutagenesis of simA and scpI was employed to investigate the potential role of these genes in S. iniae virulence. Our hybrid striped bass (HSB) and zebrafish models of infection revealed that M-like protein contributes significantly to S. iniae pathogenesis whereas C5a peptidase-like protein does not. Further, in vitro cell-based analyses indicate that SiMA, like other M family proteins, contributes to cellular adherence and invasion and provides resistance to phagocytic killing. Attenuation in our virulence models was also observed in the S. iniae isolate possessing a natural simA mutation. Vaccination of HSB with the Delta simA mutant provided 100% protection against subsequent challenge with a lethal dose of wild-type (WT) S. iniae after 1,400 degree days, and shows promise as a target for live attenuated vaccine development.

Conclusions/significance: Analysis of M-like protein and C5a peptidase through allelic replacement revealed that M-like protein plays a significant role in S. iniae virulence, and the Mga-like locus, which may regulate expression of this gene, has an unusual arrangement. The M-like protein mutant created in this research holds promise as live-attenuated vaccine.

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Nucleotide and ORF variability in the S. iniae Mga-like Mgx region.The S. iniae putative mga-like gene mgx and the putative tellurite resistance gene telX are highly conserved in strains K288, 9117, and 02161A. 02161A, however, has significant variation in the Mgx chromosomal region primarily due to four deletion or insertion sequences (A–D), two of which affect coding sequences. A 40 bp insertion/duplication (D) in the simA M-like protein gene splits it into two ORFs whose transcription and function is unknown. A 117 bp deletion (A) in the upstream mgx region generates a second putative mga-like gene, mgx2. In K288 and 9117 the mgx2 region is broken into two smaller ORFs. Similarity between adjacent strains is indicated as % nucleotide identity.
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pone-0002824-g003: Nucleotide and ORF variability in the S. iniae Mga-like Mgx region.The S. iniae putative mga-like gene mgx and the putative tellurite resistance gene telX are highly conserved in strains K288, 9117, and 02161A. 02161A, however, has significant variation in the Mgx chromosomal region primarily due to four deletion or insertion sequences (A–D), two of which affect coding sequences. A 40 bp insertion/duplication (D) in the simA M-like protein gene splits it into two ORFs whose transcription and function is unknown. A 117 bp deletion (A) in the upstream mgx region generates a second putative mga-like gene, mgx2. In K288 and 9117 the mgx2 region is broken into two smaller ORFs. Similarity between adjacent strains is indicated as % nucleotide identity.

Mentions: Our sequencing efforts, as well as the BCM-HGSC 9117 genome project, indicate the presence of a chromosomal region downstream from mgx which encodes two ORFs with high BLAST similarity to regions of Mgx and other Mga-like regulatory proteins, potentially representing an evolutionary distant duplication of a mga-like gene, whose function was lost through mutations over time (Fig. 3). Strain 02161A, however, through sequence variation in this region, including a 117 bp deletion, possesses a 1,326 bp ORF which may encode a second putative mga-like regulatory gene, mgx2 (Fig. 3). The 441 amino acid mgx2 gene product, Mgx2, has a predicted mass of 51.7 kDa and is most similar (tblastn) to the Mgx protein of S. iniae QMA0076 (42% identity, 58% positive) [29], the DmgB Mga-like protein of GCS strain Epi9 (32% identity, 52% positive) [31], and the Mga protein of GAS strain MGAS8232 (32% identity, 53% positive) [37].


Streptococcus iniae M-like protein contributes to virulence in fish and is a target for live attenuated vaccine development.

Locke JB, Aziz RK, Vicknair MR, Nizet V, Buchanan JT - PLoS ONE (2008)

Nucleotide and ORF variability in the S. iniae Mga-like Mgx region.The S. iniae putative mga-like gene mgx and the putative tellurite resistance gene telX are highly conserved in strains K288, 9117, and 02161A. 02161A, however, has significant variation in the Mgx chromosomal region primarily due to four deletion or insertion sequences (A–D), two of which affect coding sequences. A 40 bp insertion/duplication (D) in the simA M-like protein gene splits it into two ORFs whose transcription and function is unknown. A 117 bp deletion (A) in the upstream mgx region generates a second putative mga-like gene, mgx2. In K288 and 9117 the mgx2 region is broken into two smaller ORFs. Similarity between adjacent strains is indicated as % nucleotide identity.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2483786&req=5

pone-0002824-g003: Nucleotide and ORF variability in the S. iniae Mga-like Mgx region.The S. iniae putative mga-like gene mgx and the putative tellurite resistance gene telX are highly conserved in strains K288, 9117, and 02161A. 02161A, however, has significant variation in the Mgx chromosomal region primarily due to four deletion or insertion sequences (A–D), two of which affect coding sequences. A 40 bp insertion/duplication (D) in the simA M-like protein gene splits it into two ORFs whose transcription and function is unknown. A 117 bp deletion (A) in the upstream mgx region generates a second putative mga-like gene, mgx2. In K288 and 9117 the mgx2 region is broken into two smaller ORFs. Similarity between adjacent strains is indicated as % nucleotide identity.
Mentions: Our sequencing efforts, as well as the BCM-HGSC 9117 genome project, indicate the presence of a chromosomal region downstream from mgx which encodes two ORFs with high BLAST similarity to regions of Mgx and other Mga-like regulatory proteins, potentially representing an evolutionary distant duplication of a mga-like gene, whose function was lost through mutations over time (Fig. 3). Strain 02161A, however, through sequence variation in this region, including a 117 bp deletion, possesses a 1,326 bp ORF which may encode a second putative mga-like regulatory gene, mgx2 (Fig. 3). The 441 amino acid mgx2 gene product, Mgx2, has a predicted mass of 51.7 kDa and is most similar (tblastn) to the Mgx protein of S. iniae QMA0076 (42% identity, 58% positive) [29], the DmgB Mga-like protein of GCS strain Epi9 (32% identity, 52% positive) [31], and the Mga protein of GAS strain MGAS8232 (32% identity, 53% positive) [37].

Bottom Line: Our hybrid striped bass (HSB) and zebrafish models of infection revealed that M-like protein contributes significantly to S. iniae pathogenesis whereas C5a peptidase-like protein does not.Analysis of M-like protein and C5a peptidase through allelic replacement revealed that M-like protein plays a significant role in S. iniae virulence, and the Mga-like locus, which may regulate expression of this gene, has an unusual arrangement.The M-like protein mutant created in this research holds promise as live-attenuated vaccine.

View Article: PubMed Central - PubMed

Affiliation: Department of Pediatrics, University of California San Diego, La Jolla, California, United States of America.

ABSTRACT

Background: Streptococcus iniae is a significant pathogen in finfish aquaculture, though knowledge of virulence determinants is lacking. Through pyrosequencing of the S. iniae genome we have identified two gene homologues to classical surface-anchored streptococcal virulence factors: M-like protein (simA) and C5a peptidase (scpI).

Methodology/principal findings: S. iniae possesses a Mga-like locus containing simA and a divergently transcribed putative mga-like regulatory gene, mgx. In contrast to the Mga locus of group A Streptococcus (GAS, S. pyogenes), scpI is located distally in the chromosome. Comparative sequence analysis of the Mgx locus revealed only one significant variant, a strain with an insertion frameshift mutation in simA and a deletion mutation in a region downstream of mgx, generating an ORF which may encode a second putative mga-like gene, mgx2. Allelic exchange mutagenesis of simA and scpI was employed to investigate the potential role of these genes in S. iniae virulence. Our hybrid striped bass (HSB) and zebrafish models of infection revealed that M-like protein contributes significantly to S. iniae pathogenesis whereas C5a peptidase-like protein does not. Further, in vitro cell-based analyses indicate that SiMA, like other M family proteins, contributes to cellular adherence and invasion and provides resistance to phagocytic killing. Attenuation in our virulence models was also observed in the S. iniae isolate possessing a natural simA mutation. Vaccination of HSB with the Delta simA mutant provided 100% protection against subsequent challenge with a lethal dose of wild-type (WT) S. iniae after 1,400 degree days, and shows promise as a target for live attenuated vaccine development.

Conclusions/significance: Analysis of M-like protein and C5a peptidase through allelic replacement revealed that M-like protein plays a significant role in S. iniae virulence, and the Mga-like locus, which may regulate expression of this gene, has an unusual arrangement. The M-like protein mutant created in this research holds promise as live-attenuated vaccine.

Show MeSH
Related in: MedlinePlus