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Bacteriophage-resistant Staphylococcus aureus mutant confers broad immunity against staphylococcal infection in mice.

Capparelli R, Nocerino N, Lanzetta R, Silipo A, Amoresano A, Giangrande C, Becker K, Blaiotta G, Evidente A, Cimmino A, Iannaccone M, Parlato M, Medaglia C, Roperto S, Roperto F, Ramunno L, Iannelli D - PLoS ONE (2010)

Bottom Line: Acquisition of phage-resistance altered several properties of A172, causing reduced growth rate, under-expression of numerous genes and production of capsular polysaccharide.The same vaccine was also effective when administered as an aerosol.The above results demonstrate that selection for phage-resistance can facilitate bacterial vaccine preparation.

View Article: PubMed Central - PubMed

Affiliation: Faculty of Biotechnology, University of Naples, Portici, Naples, Italy.

ABSTRACT
In the presence of a bacteriophage (a bacteria-attacking virus) resistance is clearly beneficial to the bacteria. As expected in such conditions, resistant bacteria emerge rapidly. However, in the absence of the phage, resistant bacteria often display reduced fitness, compared to their sensitive counterparts. The present study explored the fitness cost associated with phage-resistance as an opportunity to isolate an attenuated strain of S. aureus. The phage-resistant strain A172 was isolated from the phage-sensitive strain A170 in the presence of the M(Sa) phage. Acquisition of phage-resistance altered several properties of A172, causing reduced growth rate, under-expression of numerous genes and production of capsular polysaccharide. In vivo, A172 modulated the transcription of the TNF-alpha, IFN-gamma and Il-1beta genes and, given intramuscularly, protected mice from a lethal dose of A170 (18/20). The heat-killed vaccine also afforded protection from heterologous methicillin-resistant S. aureus (MRSA) (8/10 mice) or vancomycin-intermediate S. aureus (VISA) (9/10 mice). The same vaccine was also effective when administered as an aerosol. Anti-A172 mouse antibodies, in the dose of 10 microl/mouse, protected the animals (10/10, in two independent experiments) from a lethal dose of A170. Consisting predominantly of the sugars glucose and galactose, the capsular polysaccharide of A172, given in the dose of 25 microg/mouse, also protected the mice (20/20) from a lethal dose of A170. The above results demonstrate that selection for phage-resistance can facilitate bacterial vaccine preparation.

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Expression levels of S. aureus A170 and A172 virulence factors.Acquisition of phage-resistance by A172 is accompanied by extensive alterations in virulence gene expression levels. Out of the 14 ORFs examined, 13 are significantly under-expressed, compared to the phage-sensitive strain A170. Bacteria were collected for transcriptional analysis during the exponential growth phase. SA0107 (spa, IgG binding protein A precursor); SA0112 (hypothetical protein, similar to cysteine synthase); SA0184 (hypothetical protein); SA039 (geh; glycerol ester hydrolase); SA1007 (α-haemolysin); SA1160 (nuc; thermonuclease); SA1898 (hypothetical protein, similar to SceD precursor); SA2097 (hypothetical protein, similar to secretory antigen precursor SsaA); SA2206 (sbi; IgG-binding protein SBI); SA2255 (oligopeptide transporter substrate binding protein); SA2406 (gbsA; glycine betaine aldehyde dehydrogenase gbsA); SA2459 (ica; N-glycosyltransferase); SAR216 (groEL; chaperonin GROEL); SAR2117 (groES; co-chaperonin GRES). ORFs numbers correspond to the S. aureus N315 genome sequence. Gene designations (when known) and proteins function are shown in parentheses.
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pone-0011720-g004: Expression levels of S. aureus A170 and A172 virulence factors.Acquisition of phage-resistance by A172 is accompanied by extensive alterations in virulence gene expression levels. Out of the 14 ORFs examined, 13 are significantly under-expressed, compared to the phage-sensitive strain A170. Bacteria were collected for transcriptional analysis during the exponential growth phase. SA0107 (spa, IgG binding protein A precursor); SA0112 (hypothetical protein, similar to cysteine synthase); SA0184 (hypothetical protein); SA039 (geh; glycerol ester hydrolase); SA1007 (α-haemolysin); SA1160 (nuc; thermonuclease); SA1898 (hypothetical protein, similar to SceD precursor); SA2097 (hypothetical protein, similar to secretory antigen precursor SsaA); SA2206 (sbi; IgG-binding protein SBI); SA2255 (oligopeptide transporter substrate binding protein); SA2406 (gbsA; glycine betaine aldehyde dehydrogenase gbsA); SA2459 (ica; N-glycosyltransferase); SAR216 (groEL; chaperonin GROEL); SAR2117 (groES; co-chaperonin GRES). ORFs numbers correspond to the S. aureus N315 genome sequence. Gene designations (when known) and proteins function are shown in parentheses.

Mentions: In liquid culture, compared to the parental strain, A172 formed colonies with the tendency to clump together starting at the early stationary phase (Figure 2A–B). Bacterial suspensions of A170 and A172 with the same density (OD600: 0.6) were assayed by plate count. In three independent experiments, A170 cultures contained about 3 fold as many CFU as the A172 cultures (3.4×107±6.7×106 vs 1.2×107±2.5×106; P 0.003). A172 displayed reduced growth rate during the exponential phase (Figure 2C) and reduced doubling time (A172: 45 min±2.8 min; A170: 30±2.5 min; P 0.002). Also, unlike A170, A172 produced capsular polysaccharide (Figure 3). Phage resistance altered the transcription of several virulence factors. Among 14 ORFs of A172 analyzed, 13 were significantly down-regulated compared to A170 (Figure 4). Down-regulated ORFs comprise 9 genes, which control virulence factors [25].


Bacteriophage-resistant Staphylococcus aureus mutant confers broad immunity against staphylococcal infection in mice.

Capparelli R, Nocerino N, Lanzetta R, Silipo A, Amoresano A, Giangrande C, Becker K, Blaiotta G, Evidente A, Cimmino A, Iannaccone M, Parlato M, Medaglia C, Roperto S, Roperto F, Ramunno L, Iannelli D - PLoS ONE (2010)

Expression levels of S. aureus A170 and A172 virulence factors.Acquisition of phage-resistance by A172 is accompanied by extensive alterations in virulence gene expression levels. Out of the 14 ORFs examined, 13 are significantly under-expressed, compared to the phage-sensitive strain A170. Bacteria were collected for transcriptional analysis during the exponential growth phase. SA0107 (spa, IgG binding protein A precursor); SA0112 (hypothetical protein, similar to cysteine synthase); SA0184 (hypothetical protein); SA039 (geh; glycerol ester hydrolase); SA1007 (α-haemolysin); SA1160 (nuc; thermonuclease); SA1898 (hypothetical protein, similar to SceD precursor); SA2097 (hypothetical protein, similar to secretory antigen precursor SsaA); SA2206 (sbi; IgG-binding protein SBI); SA2255 (oligopeptide transporter substrate binding protein); SA2406 (gbsA; glycine betaine aldehyde dehydrogenase gbsA); SA2459 (ica; N-glycosyltransferase); SAR216 (groEL; chaperonin GROEL); SAR2117 (groES; co-chaperonin GRES). ORFs numbers correspond to the S. aureus N315 genome sequence. Gene designations (when known) and proteins function are shown in parentheses.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2908692&req=5

pone-0011720-g004: Expression levels of S. aureus A170 and A172 virulence factors.Acquisition of phage-resistance by A172 is accompanied by extensive alterations in virulence gene expression levels. Out of the 14 ORFs examined, 13 are significantly under-expressed, compared to the phage-sensitive strain A170. Bacteria were collected for transcriptional analysis during the exponential growth phase. SA0107 (spa, IgG binding protein A precursor); SA0112 (hypothetical protein, similar to cysteine synthase); SA0184 (hypothetical protein); SA039 (geh; glycerol ester hydrolase); SA1007 (α-haemolysin); SA1160 (nuc; thermonuclease); SA1898 (hypothetical protein, similar to SceD precursor); SA2097 (hypothetical protein, similar to secretory antigen precursor SsaA); SA2206 (sbi; IgG-binding protein SBI); SA2255 (oligopeptide transporter substrate binding protein); SA2406 (gbsA; glycine betaine aldehyde dehydrogenase gbsA); SA2459 (ica; N-glycosyltransferase); SAR216 (groEL; chaperonin GROEL); SAR2117 (groES; co-chaperonin GRES). ORFs numbers correspond to the S. aureus N315 genome sequence. Gene designations (when known) and proteins function are shown in parentheses.
Mentions: In liquid culture, compared to the parental strain, A172 formed colonies with the tendency to clump together starting at the early stationary phase (Figure 2A–B). Bacterial suspensions of A170 and A172 with the same density (OD600: 0.6) were assayed by plate count. In three independent experiments, A170 cultures contained about 3 fold as many CFU as the A172 cultures (3.4×107±6.7×106 vs 1.2×107±2.5×106; P 0.003). A172 displayed reduced growth rate during the exponential phase (Figure 2C) and reduced doubling time (A172: 45 min±2.8 min; A170: 30±2.5 min; P 0.002). Also, unlike A170, A172 produced capsular polysaccharide (Figure 3). Phage resistance altered the transcription of several virulence factors. Among 14 ORFs of A172 analyzed, 13 were significantly down-regulated compared to A170 (Figure 4). Down-regulated ORFs comprise 9 genes, which control virulence factors [25].

Bottom Line: Acquisition of phage-resistance altered several properties of A172, causing reduced growth rate, under-expression of numerous genes and production of capsular polysaccharide.The same vaccine was also effective when administered as an aerosol.The above results demonstrate that selection for phage-resistance can facilitate bacterial vaccine preparation.

View Article: PubMed Central - PubMed

Affiliation: Faculty of Biotechnology, University of Naples, Portici, Naples, Italy.

ABSTRACT
In the presence of a bacteriophage (a bacteria-attacking virus) resistance is clearly beneficial to the bacteria. As expected in such conditions, resistant bacteria emerge rapidly. However, in the absence of the phage, resistant bacteria often display reduced fitness, compared to their sensitive counterparts. The present study explored the fitness cost associated with phage-resistance as an opportunity to isolate an attenuated strain of S. aureus. The phage-resistant strain A172 was isolated from the phage-sensitive strain A170 in the presence of the M(Sa) phage. Acquisition of phage-resistance altered several properties of A172, causing reduced growth rate, under-expression of numerous genes and production of capsular polysaccharide. In vivo, A172 modulated the transcription of the TNF-alpha, IFN-gamma and Il-1beta genes and, given intramuscularly, protected mice from a lethal dose of A170 (18/20). The heat-killed vaccine also afforded protection from heterologous methicillin-resistant S. aureus (MRSA) (8/10 mice) or vancomycin-intermediate S. aureus (VISA) (9/10 mice). The same vaccine was also effective when administered as an aerosol. Anti-A172 mouse antibodies, in the dose of 10 microl/mouse, protected the animals (10/10, in two independent experiments) from a lethal dose of A170. Consisting predominantly of the sugars glucose and galactose, the capsular polysaccharide of A172, given in the dose of 25 microg/mouse, also protected the mice (20/20) from a lethal dose of A170. The above results demonstrate that selection for phage-resistance can facilitate bacterial vaccine preparation.

Show MeSH
Related in: MedlinePlus