| 1. |
Gagnon G,
Vadeboncoeur C,
Levesque RC,
Frenette M,
( 1992 ) Cloning, sequencing and expression in Escherichia coli of the ptsI gene encoding enzyme I of the phosphoenolpyruvate:sugar phosphotransferase transport system from Streptococcus salivarius. PMID : 1427100 : DOI : 10.1016/0378-1119(92)90163-j Abstract >>
We present the cloning and sequencing of the ptsI gene, encoding enzyme I (EI) of the phosphoenolpyruvate (PEP): sugar phosphotransferase (PTS) transport system from Streptococcus salivarius. The ptsI gene corresponds to an open reading frame of 1731 nucleotides, which translates into a putative 577-amino acid (aa) protein with a M(r) of 62,948 and a pI of 4.49. The EI was produced in Escherichia coli under the control of its own promoter located immediately upstream of ptsI, a situation never previously reported for any other gene coding for an EI. The deduced aa sequence of the S. salivarius EI shows a high degree of similarity with the E. coli EI and the EI moiety of the multiphosphoryl transfer protein from Rhodobacter capsulatus. The S. salivarius EI also shares a highly conserved aa cluster with a non-PTS protein, the maize pyruvate:orthophosphate dikinase. The conserved cluster is located in a domain which is hypothesized to be the PEP-binding site.
|
2. |
Frey N,
Nessler S,
Fieulaine S,
Vaillancourt K,
Frenette M,
Vadeboncoeur C,
( 2003 ) The HPr(Ser) kinase of Streptococcus salivarius: a hexameric bifunctional enzyme controlled by glycolytic intermediates and inorganic phosphate. PMID : 12855169 : DOI : 10.1016/S0378-1097(03)00429-4 Abstract >>
Phosphorylation of HPr, the small phosphocarrier protein of the phosphoenolpyruvate:sugar phosphotransferase system, on Ser46 by the HPr(Ser) kinase (HPrK/P) is a vital step in catabolite repression in Gram-positive bacteria. Streptococcus salivarius HPrK/P is reported to be a multimeric protein not regulated by metabolic intermediates. We re-evaluated the molecular mass of S. salivarius HPrK/P using sedimentation equilibrium ultracentrifugation, demonstrated that S. salivarius HPrK/P dephosphorylated HPr(Ser-P) and further characterised the effect of fructose 1,6-bisphosphate and other metabolic intermediates on enzyme activities. The molecular mass of S. salivarius HPrK/P was 201305 Da, suggesting that streptococcal HPrK/P was a hexameric protein. Fructose 1,6-bisphosphate poorly activated streptococcal HPrK/P but protected kinase activity against inhibition by inorganic phosphate and inhibited dephosphorylation of HPr(Ser-P). Phosphoenolpyruvate and 2-phosphoglycerate, but not fructose 1-P, fructose 6-P, and ribulose 1,5-bisphosphate, also protected kinase activity against inhibition by inorganic phosphate. Thus, unlike previous reports, we show that fructose 1,6-bisphosphate and other key glycolytic intermediates played a pivotal role as a modulator of streptococcal HPrK/P activities.
|
3. |
Yoshida Y,
Negishi M,
Amano A,
Oho T,
Nakano Y,
( 2003 ) Differences in the betaC-S lyase activities of viridans group streptococci. PMID : 12480520 : DOI : 10.1016/s0006-291x(02)02803-6 Abstract >>
betaC-S Lyase catalyzes the alpha,beta-elimination of L-cysteine to hydrogen sulfide, which is one of the main causes of oral malodor and is highly toxic to mammalian cells. We evaluated the capacity of six species of oral streptococci to produce hydrogen sulfide. The crude enzyme extract from Streptococcus anginosus had the greatest capacity. However, comparative analysis of amino acid sequences did not detect any meaningful differences in the S. anginosus betaC-S lyase. The capacity of S. anginosus purified betaC-S lyase to degrade L-cysteine was also extremely high, while its capacity to degrade L-cystathionine was unremarkable. These findings suggest that the extremely high capacity of S. anginosus to produce hydrogen sulfide is due to the unique characteristic of betaC-S lyase from that organism.
|
4. |
Teng LJ,
Hsueh PR,
Tsai JC,
Chen PW,
Hsu JC,
Lai HC,
Lee CN,
Ho SW,
( 2002 ) groESL sequence determination, phylogenetic analysis, and species differentiation for viridans group streptococci. PMID : 12202549 : DOI : 10.1128/jcm.40.9.3172-3178.2002 PMC : PMC130726 Abstract >>
The full-length sequences of the groESL genes (also known as cpn10/60) of Streptococcus anginosus, Streptococcus constellatus, Streptococcus gordonii, and Streptococcus sanguis and the near full-length sequence of the groESL genes of Streptococcus intermedius, Streptococcus bovis, Streptococcus mitis, Streptococcus mutans, Streptococcus oralis, and Streptococcus salivarius were determined. The lengths of the groES genes from the 10 species listed above ranged from 282 to 288 bp, and the full-length sequences of groEL determined for 4 species (S. anginosus, S. constellatus, S. gordonii, and S. sanguis) revealed that each was 1,623 bp. The intergenic region (spacer) between the groES and groEL genes varies in size (15 to 111 bp) and sequence between species. The variation of the groES sequences among the species tested was greater (62.1 to 95.1% nucleotide sequence identities) than that of the groEL sequences (77.2 to 95.2% nucleotide sequence identities). Phylogenetic analysis of the groES and groEL genes yielded evolutionary trees similar to the tree constructed by use of the 16S rRNA gene. The intraspecies variation of the spacer was minimal for clinical isolates of some species. The groESL sequence data provide an additional parameter for identification of viridans group streptococcal species.
|
5. |
Razonable RR,
Litzow MR,
Khaliq Y,
Piper KE,
Rouse MS,
Patel R,
( 2002 ) Bacteremia due to viridans group Streptococci with diminished susceptibility to Levofloxacin among neutropenic patients receiving levofloxacin prophylaxis. PMID : 12015693 : DOI : 10.1086/340352 Abstract >>
Despite the use of levofloxacin prophylaxis during the neutropenic period after autologous peripheral blood stem cell transplantation, viridans group (VG) streptococcal bacteremia developed in 6 (16.2%) of 37 patients who underwent transplantation between 1 January and 25 February 2001 at the Mayo Clinic in Rochester, Minnesota. All 6 patients presented with fever and mucositis after a mean of 4.5 days of neutropenia, and 3 developed septic shock. All 6 VG streptococcal isolates from these patients exhibited distinct patterns on pulsed-field gel electrophoresis. All isolates had diminished susceptibility to levofloxacin, 5 to gatifloxacin, and 4 to moxifloxacin. Quinolone resistance was associated with mutations in the quinolone resistance-determining region of GyrA and (for 1 isolate) of ParC. The use of levofloxacin may select VG streptococci with diminished susceptibility to levofloxacin and other quinolones with enhanced activity against gram-positive organisms and, therefore, may not be optimal for preventing VG streptococcal bacteremia in neutropenic patients.
|
6. |
Vaillancourt K,
Moineau S,
Frenette M,
Lessard C,
Vadeboncoeur C,
( 2002 ) Galactose and lactose genes from the galactose-positive bacterium Streptococcus salivarius and the phylogenetically related galactose-negative bacterium Streptococcus thermophilus: organization, sequence, transcription, and activity of the gal gene products. PMID : 11790749 : DOI : 10.1128/jb.184.3.785-793.2002 PMC : PMC139519 Abstract >>
Streptococcus salivarius is a lactose- and galactose-positive bacterium that is phylogenetically closely related to Streptococcus thermophilus, a bacterium that metabolizes lactose but not galactose. In this paper, we report a comparative characterization of the S. salivarius and S. thermophilus gal-lac gene clusters. The clusters have the same organization with the order galR (codes for a transcriptional regulator and is transcribed in the opposite direction), galK (galactokinase), galT (galactose-1-P uridylyltransferase), galE (UDP-glucose 4-epimerase), galM (galactose mutarotase), lacS (lactose transporter), and lacZ (beta-galactosidase). An analysis of the nucleotide sequence as well as Northern blotting and primer extension experiments revealed the presence of four promoters located upstream from galR, the gal operon, galM, and the lac operon of S. salivarius. Putative promoters with virtually identical nucleotide sequences were found at the same positions in the S. thermophilus gal-lac gene cluster. An additional putative internal promoter at the 3' end of galT was found in S. thermophilus but not in S. salivarius. The results clearly indicated that the gal-lac gene cluster was efficiently transcribed in both species. The Shine-Dalgarno sequences of galT and galE were identical in both species, whereas the ribosome binding site of S. thermophilus galK differed from that of S. salivarius by two nucleotides, suggesting that the S. thermophilus galK gene might be poorly translated. This was confirmed by measurements of enzyme activities.
|
7. |
Upton M,
Tagg JR,
Wescombe P,
Jenkinson HF,
( 2001 ) Intra- and interspecies signaling between Streptococcus salivarius and Streptococcus pyogenes mediated by SalA and SalA1 lantibiotic peptides. PMID : 11395456 : DOI : 10.1128/JB.183.13.3931-3938.2001 PMC : PMC95275 Abstract >>
Streptococcus salivarius 20P3 produces a 22-amino-acid residue lantibiotic, designated salivaricin A (SalA), that inhibits the growth of a range of streptococci, including all strains of Streptococcus pyogenes. Lantibiotic production is associated with the sal genetic locus comprising salA, the lantibiotic structural gene; salBCTX genes encoding peptide modification and export machinery proteins; and salYKR genes encoding a putative immunity protein and two-component sensor-regulator system. Insertional inactivation of salB in S. salivarius 20P3 resulted in abrogation of SalA peptide production, of immunity to SalA, and of salA transcription. Addition of exogenous SalA peptide to salB mutant cultures induced dose-dependent expression of salA mRNA (0.2 kb), demonstrating that SalA production was normally autoregulated. Inactivation of salR encoding the response regulator of the SalKR two-component system led to reduced production of, and immunity to, SalA. The sal genetic locus was also present in S. pyogenes SF370 (M type 1), but because of a deletion across the salBCT genes, the corresponding lantibiotic peptide, designated SalA1, was not produced. However, in S. pyogenes T11 (M type 4) the sal locus gene complement was apparently complete, and active SalA1 peptide was synthesized. Exogenously added SalA1 peptide from S. pyogenes T11 induced salA1 transcription in S. pyogenes SF370 and in an isogenic S. pyogenes T11 salB mutant and salA transcription in S. salivarius 20P3 salB. Thus, SalA and SalA1 are examples of streptococcal lantibiotics whose production is autoregulated. These peptides act as intra- and interspecies signaling molecules, modulating lantibiotic production and possibly influencing streptococcal population ecology in the oral cavity.
|
8. |
Mollerach M,
García E,
( 2000 ) The galU gene of Streptococcus pneumoniae that codes for a UDP-glucose pyrophosphorylase is highly polymorphic and suitable for molecular typing and phylogenetic studies. PMID : 11137293 : DOI : 10.1016/s0378-1119(00)00468-6 Abstract >>
The enzyme UTP-glucose-1-phosphate uridylyltransferase (UDP-glucose pyrophosphorylase, UDPG:PP) is synthesized by practically all organisms, although prokaryotic UDPG:PPs are evolutionarily unrelated to the eukaryotic counterparts. The primary structure of prokaryotic UDPG:PPs is well conserved, although little information exists on the polymorphism of the genes coding for these enzymes. It has been reported that the galU gene encoding the Streptococcus pneumoniae UDPG:PP is absolutely required for the synthesis of the capsular polysaccharide, a sine qua non prerequisite for virulence. A 594 bp fragment covering 66% of the galU gene from 37 pneumococcal isolates and the type strains of Streptococcus mitis, Streptococcus oralis, Streptococcus gordonii, Streptococcus sanguinis, Streptococcus salivarius, and Streptococcus sobrinus has been amplified by PCR and sequenced. Up to 21 different alleles were found in S. pneumoniae. They possess a mosaic-like structure and belong to, at least, two evolutionarily distinct families that show a sequence divergence of 15-20%. In spite of its marked polymorphism, phylogenetic relationships among pneumococcal strains deduced from the galU gene matched those previously established by using alternative approaches. Comparison of the pneumococcal galU alleles with those from other streptococci indicated the existence of a complex network of genetic interchange. The galU gene represents an informative marker to be used alone or in conjunction with other molecular typing methods.
|
9. |
Almirón-Roig E,
Mulholland F,
Gasson MJ,
Griffin AM,
( 2000 ) The complete cps gene cluster from Streptococcus thermophilus NCFB 2393 involved in the biosynthesis of a new exopolysaccharide. PMID : 11065358 : DOI : 10.1099/00221287-146-11-2793 Abstract >>
The cpsFGHIJKL genes from the cps cluster of Streptococcus thermophilus NCFB 2393 involved in the biosynthesis of EPS were identified, cloned and nucleotide sequenced. The complete cps cluster is contained on an approximately 11.2 kb chromosomal region which contains 12 ORFs, including the previously cloned cpsABCDE genes. Functions were assigned to some of the predicted gene products on the basis of homology to known sequences as follows: cpsK encodes a protein thought to be involved in the polymerization and export of the polysaccharide; cpsE, cpsF, cpsG, cpsH, cpsI and cpsJ encode putative sugar transferases. Two insertion sequences, IS1193 and ISS1, were identified within and flanking the 3' end of the cps cluster respectively. Analysis of the expression of the cpsE gene in Escherichia coli demonstrated that it encodes a glucose-1-phosphate transferase; the enzyme which catalyses the first step in EPS biosynthesis in S. thermophilus NCFB 2393.
|
10. |
Pelletier M,
Lortie LA,
( 2000 ) The gene encoding IIAB(Man)L in Streptococcus salivarius is part of a tetracistronic operon encoding a phosphoenolpyruvate: mannose/glucose phosphotransferase system. PMID : 10746771 : DOI : 10.1099/00221287-146-3-677 Abstract >>
Glucose and mannose are transported in streptococci by the mannose-PTS (phosphoenolpyruvate:mannose phosphotransferase system), which consists of a cytoplasmic IIAB protein, called IIAB(Man), and an uncharacterized membrane permease. This paper reports the characterization of the man operon encoding the specific components of the mannose-PTS of Streptococcus salivarius. The man operon was composed of four genes, manL, manM, manN and manO. These genes were transcribed from a canonical promoter (Pman) into a 3.6 kb polycistronic mRNA that contained a 5'-UTR (untranslated region). The predicted manL gene product encoded a 35.5 kDa protein and contained the amino acid sequences of the IIA and IIB phosphorylation sites already determined from purified S. salivarius IIAB(Man)L. Expression of manL in Escherichia coli generated a 35 kDa protein that reacted with anti-IIAB(Man)L antibodies. The predicted ManM protein had an estimated size of 27.2 kDa. ManM had similarity with IIC domains of the mannose-EII family, but did not possess the signature proposed for mannose-IIC proteins from Gram-negative bacteria. From multiple alignment analyses of sequences available in current databases, the following modified IIC(Man) signature is proposed: GX3G[DNH]X3G[LIVM]2XG2[STL][LT][EQ]. The deduced product of manN was a hydrophobic protein with a predicted molecular mass of 33.4 kDa. The ManN protein contained an amino acid sequence similar to the signature sequence of the IID domains of the mannose-EII family. manO encoded a 13.7 kDa protein. This gene was also transcribed as a monocistronic mRNA from a promoter located in the manN-manO intergenic region. A search of current databases revealed the presence of IIAB(Man)L, ManM, ManN and ManO orthologues in Streptococcus mutans, Streptococcus pyogenes, Streptococcus pneumoniae and Enterococcus faecalis. This work has elucidated the molecular structure of the mannose PTS in streptococci and enterococci, and demonstrated the presence of a putative regulatory protein (ManO) within the man operon.
|
11. |
Horz HP,
Meinelt A,
Houben B,
Conrads G,
( 2007 ) Distribution and persistence of probiotic Streptococcus salivarius K12 in the human oral cavity as determined by real-time quantitative polymerase chain reaction. PMID : 17311636 : DOI : 10.1111/j.1399-302X.2007.00334.x Abstract >>
The bacteriocin producer Streptococcus salivarius K12 is used as a probiotic targeting the oral cavity, so our study aimed to assess whether its dispersal and persistence could be monitored using real-time quantitative polymerase chain reaction. To this end, we designed polymerase chain reaction primers and a hybridization probe specifically targeting salA, which encodes for the prepropeptide of salivaricin A. Using a single individual as our subject, we administered four lozenges of K12 Throat Guard per day over 3 days, then measured salA gene levels for 16 different oral sites at six different intervals over 35 days. Four samples each from gingival sulci and from teeth all remained negative. In contrast, in saliva and at all mucosal membranes K12 was detected, but with varying amounts and time profiles. Relatively high salA gene copy numbers, calibrated on the basis of colony-forming units, were seen on the tongue (maximum 4.6 x 10(4)/swab at day 4), in stimulated saliva (2.4 x 10(4)/ml, day 4) and on buccal membranes (1.3 x 10(4)/swab, day 8). K12 was present on both sides of the pharynx but asymmetrically in both quantity and duration. In conclusion, we have developed a real-time quantitative-polymerase chain reaction for counting S. salivarius K12 at various sites in the oral cavity. In the individual studied, K12 could be detected at the mucosal membranes for as long as 3 weeks, but with steadily decreasing numbers after day 8. Thus, K12 may have the potential to control oral bacterial infections only when the uptake is repeated frequently.
|
12. |
Hyink O,
Wescombe PA,
Upton M,
Ragland N,
Burton JP,
Tagg JR,
( 2007 ) Salivaricin A2 and the novel lantibiotic salivaricin B are encoded at adjacent loci on a 190-kilobase transmissible megaplasmid in the oral probiotic strain Streptococcus salivarius K12. PMID : 17194838 : DOI : 10.1128/AEM.02265-06 PMC : PMC1828679 Abstract >>
The commercial probiotic Streptococcus salivarius strain K12 is the prototype of those S. salivarius strains that are the most strongly inhibitory in a standardized test of streptococcal bacteriocin production and has been shown to produce the 2,368-Da salivaricin A2 (SalA2) and the 2,740-Da salivaricin B (SboB) lantibiotics. The previously uncharacterized SboB belongs to the type AII class of lantibiotic bacteriocins and is encoded by an eight-gene cluster. The genetic loci encoding SalA2 and SboB in strain K12 have been fully characterized and are localized to nearly adjacent sites on pSsal-K12, a 190-kb megaplasmid. Of 61 strongly inhibitory strains of S. salivarius, 19 (31%) were positive for the sboB structural gene. All but one (strain NR) of these 19 strains were also positive for salA2, and in each of these cases of double positivity, the two loci were separated by fewer than 10 kb. This is the first report of a single streptococcus strain producing two distinct lantibiotics.
|
13. |
Patterson AJ,
Rincon MT,
Flint HJ,
Scott KP,
( 2007 ) Mosaic tetracycline resistance genes are widespread in human and animal fecal samples. PMID : 17178791 : DOI : 10.1128/AAC.00725-06 PMC : PMC1803118 Abstract >>
Mosaic tetracycline resistance genes comprising tet(O), tet(W), and tet(32) sequences were abundant in DNA extracted from pig and human fecal samples, accounting for 78% (50/64) and 46% (37/80) of genes amplified with a tet(O) primer set, respectively, in two samples. The nonmosaic tet(32) gene was isolated from a human saliva bacterium.
|
14. |
Delorme C,
Poyart C,
Ehrlich SD,
Renault P,
( 2007 ) Extent of horizontal gene transfer in evolution of Streptococci of the salivarius group. PMID : 17085557 : DOI : 10.1128/JB.01058-06 PMC : PMC1797340 Abstract >>
The phylogenetically closely related species Streptococcus salivarius and Streptococcus vestibularis are oral bacteria that are considered commensals, although they can also be found in human infections. The relationship between these two species and the relationship between strains isolated from carriers and strains responsible for invasive infections were investigated by multilocus sequence typing and additional sequence analysis. The clustering of several S. vestibularis alleles and the extent of genomic divergence at certain loci support the conclusion that S. salivarius and S. vestibularis are separate species. The level of sequence diversity in S. salivarius alleles is generally high, whereas that in S. vestibularis alleles is low at certain loci, indicating that the latter species might have evolved recently. Cluster analysis indicated that there has been genetic exchange between S. salivarius and S. vestibularis at three of the nine loci investigated. Horizontal gene transfer between streptococci belonging to the S. salivarius group and other oral streptococci was also detected at several loci. A high level of recombination in S. salivarius was revealed by allele index association and split decomposition sequence analyses. Commensal and infection-associated S. salivarius strains could not be distinguished by cluster analysis, suggesting that the pathogen isolates are opportunistic. Taken together, our results indicate that there is a high level of gene exchange that contributes to the evolution of two streptococcal species from the human oral cavity.
|
15. |
Itoh Y,
Kawamura Y,
Kasai H,
Shah MM,
Nhung PH,
Yamada M,
Sun X,
Koyana T,
Hayashi M,
Ohkusu K,
Ezaki T,
( 2006 ) dnaJ and gyrB gene sequence relationship among species and strains of genus Streptococcus. PMID : 16487673 : DOI : 10.1016/j.syapm.2005.12.003 Abstract >>
The dnaJ and gyrB nucleotide sequences were determined for members of the genus Streptococcus. The average similarity between the species tested was 76.4% (69.7-100%) for dnaJ and 75.9 (70.1-98.7%) for gyrB. These data indicated that the dnaJ and gyrB genes are more divergent and more discriminatory than the 16S rDNA gene. Furthermore, the variation in the dnaJ nucleotide sequences among the mitis group was greater than that of the gyrB nucleotide sequences, especially between Streptococcus pneumoniae and Streptococcus mitis. Subsequently, the high discrimination power of dnaJ within the mitis group was confirmed. Thus, we conclude that the dnaJ and gyrB genes are efficient alternative targets for the classification of the genus Streptococcus, and that dnaJ is suitable for phylogenetic analysis of closely related Streptococcus strains.
|
16. |
Wescombe PA,
Upton M,
Dierksen KP,
Ragland NL,
Sivabalan S,
Wirawan RE,
Inglis MA,
Moore CJ,
Walker GV,
Chilcott CN,
Jenkinson HF,
Tagg JR,
( 2006 ) Production of the lantibiotic salivaricin A and its variants by oral streptococci and use of a specific induction assay to detect their presence in human saliva. PMID : 16461700 : DOI : 10.1128/AEM.72.2.1459-1466.2006 PMC : PMC1392966 Abstract >>
Salivaricin A (SalA), the first Streptococcus salivarius lantibiotic to be characterized, appears to be inhibitory to most Streptococcus pyogenes strains. A variant of the SalA structural gene (salA1) is present in more than 90% of S. pyogenes strains, but only strains of M serotype 4 and T pattern 4 produce the biologically active peptide. The present study identifies four additional variants (salA2 to salA5) of the SalA structural gene and demonstrates that each of the corresponding inhibitory peptides (SalA2 to SalA5) is produced in vitro. These variants appear to be similar to SalA and SalA1 in their inhibitory activity against Micrococcus luteus and in their ability to act as inducers of SalA production. It had previously been shown that S. pyogenes strain SF370 had a deletion (of approximately 2.5 kb) in the salM and salT genes of the salA1 locus. In the present study, several additional characteristic deletions within the salA1 loci were identified. S. pyogenes strains of the same M serotype all share the same salA1 locus structure. Since S. salivarius is a predominant member of the normal oral flora of healthy humans, strains producing anti-S. pyogenes lantibiotics, such as SalA, may have excellent potential for use as oral probiotics. In the present study, we have used a highly specific SalA induction system to directly detect the presence of SalA in the saliva of humans who either naturally harbor populations of SalA-producing S. salivarius or who have been colonized with the SalA2-producing probiotic S. salivarius K12.
|
17. |
Hoshino T,
Fujiwara T,
Kilian M,
( 2005 ) Use of phylogenetic and phenotypic analyses to identify nonhemolytic streptococci isolated from bacteremic patients. PMID : 16333101 : DOI : 10.1128/JCM.43.12.6073-6085.2005 PMC : PMC1317212 Abstract >>
The aim of this study was to evaluate molecular and phenotypic methods for the identification of nonhemolytic streptococci. A collection of 148 strains consisting of 115 clinical isolates from cases of infective endocarditis, septicemia, and meningitis and 33 reference strains, including type strains of all relevant Streptococcus species, were examined. Identification was performed by phylogenetic analysis of nucleotide sequences of four housekeeping genes, ddl, gdh, rpoB, and sodA; by PCR analysis of the glucosyltransferase (gtf) gene; and by conventional phenotypic characterization and identification using two commercial kits, Rapid ID 32 STREP and STREPTOGRAM and the associated databases. A phylogenetic tree based on concatenated sequences of the four housekeeping genes allowed unequivocal differentiation of recognized species and was used as the reference. Analysis of single gene sequences revealed deviation clustering in eight strains (5.4%) due to homologous recombination with other species. This was particularly evident in S. sanguinis and in members of the anginosus group of streptococci. The rate of correct identification of the strains by both commercial identification kits was below 50% but varied significantly between species. The most significant problems were observed with S. mitis and S. oralis and 11 Streptococcus species described since 1991. Our data indicate that identification based on multilocus sequence analysis is optimal. As a more practical alternative we recommend identification based on sodA sequences with reference to a comprehensive set of sequences that is available for downloading from our server. An analysis of the species distribution of 107 nonhemolytic streptococci from bacteremic patients showed a predominance of S. oralis and S. anginosus with various underlying infections.
|
18. |
Duwat P,
Ehrlich SD,
Gruss A,
( 1992 ) A general method for cloning recA genes of gram-positive bacteria by polymerase chain reaction. PMID : 1629178 : DOI : 10.1128/jb.174.15.5171-5175.1992 PMC : PMC206342 Abstract >>
An internal fragment of the recA gene from eight gram-positive organisms has been amplified by using degenerate primers in a polymerase chain reaction. The internal 348- or 360-bp recA DNA segments from Bacillus subtilis, Clostridium acetobutylicum, Lactobacillus bulgaricus, Lactobacillus helveticus, Leuconostoc mesanteroides, Listeria monocytogenes, Staphylococcus aureus, and Streptococcus salivarus subsp. thermophilus were amplified, cloned, and sequenced. The G + C contents of the DNA from these species range from 28 to 52%. The sequences of the bacterial recA genes show strong relatedness. This method is particularly useful for the recovery of the recA genes of gram-positive bacteria and avoids the difficulties of using a genetic complementation test for cloning.
|
19. |
Hill JE,
Penny SL,
Crowell KG,
Goh SH,
Hemmingsen SM,
( 2004 ) cpnDB: a chaperonin sequence database. PMID : 15289485 : DOI : 10.1101/gr.2649204 PMC : PMC509277 Abstract >>
Type I chaperonins are molecular chaperones present in virtually all bacteria, some archaea and the plastids and mitochondria of eukaryotes. Sequences of cpn60 genes, encoding 60-kDa chaperonin protein subunits (CPN60, also known as GroEL or HSP60), are useful for phylogenetic studies and as targets for detection and identification of organisms. Conveniently, a 549-567-bp segment of the cpn60 coding region can be amplified with universal PCR primers. Here, we introduce cpnDB, a curated collection of cpn60 sequence data collected from public databases or generated by a network of collaborators exploiting the cpn60 target in clinical, phylogenetic, and microbial ecology studies. The growing database currently contains approximately 2000 records covering over 240 genera of bacteria, eukaryotes, and archaea. The database also contains over 60 sequences for the archaeal Type II chaperonin (thermosome, a homolog of eukaryotic cytoplasmic chaperonin) from 19 archaeal genera. As the largest curated collection of sequences available for a protein-encoding gene, cpnDB provides a resource for researchers interested in exploiting the power of cpn60 as a diagnostic or as a target for phylogenetic or microbial ecology studies, as well as those interested in broader subjects such as lateral gene transfer and codon usage. We built cpnDB from open source tools and it is available at http://cpndb.cbr.nrc.ca.
|
20. |
Drancourt M,
Roux V,
Fournier PE,
Raoult D,
( 2004 ) rpoB gene sequence-based identification of aerobic Gram-positive cocci of the genera Streptococcus, Enterococcus, Gemella, Abiotrophia, and Granulicatella. PMID : 14766807 : DOI : 10.1128/jcm.42.2.497-504.2004 PMC : PMC344509 Abstract >>
We developed a new molecular tool based on rpoB gene (encoding the beta subunit of RNA polymerase) sequencing to identify streptococci. We first sequenced the complete rpoB gene for Streptococcus anginosus, S. equinus, and Abiotrophia defectiva. Sequences were aligned with these of S. pyogenes, S. agalactiae, and S. pneumoniae available in GenBank. Using an in-house analysis program (SVARAP), we identified a 740-bp variable region surrounded by conserved, 20-bp zones and, by using these conserved zones as PCR primer targets, we amplified and sequenced this variable region in an additional 30 Streptococcus, Enterococcus, Gemella, Granulicatella, and Abiotrophia species. This region exhibited 71.2 to 99.3% interspecies homology. We therefore applied our identification system by PCR amplification and sequencing to a collection of 102 streptococci and 60 bacterial isolates belonging to other genera. Amplicons were obtained in streptococci and Bacillus cereus, and sequencing allowed us to make a correct identification of streptococci. Molecular signatures were determined for the discrimination of closely related species within the S. pneumoniae-S. oralis-S. mitis group and the S. agalactiae-S. difficile group. These signatures allowed us to design a S. pneumoniae-specific PCR and sequencing primer pair.
|
21. |
Lévesque C,
Vadeboncoeur C,
Frenette M,
( 2004 ) The csp operon of Streptococcus salivarius encodes two predicted cell-surface proteins, one of which, CspB, is associated with the fimbriae. PMID : 14702412 : DOI : 10.1099/mic.0.26592-0 Abstract >>
A Tn917 mutant library was generated to identify genes involved in the biogenesis of Streptococcus salivarius fimbriae. A fimbria-deficient mutant was isolated by negative selection using an immunomagnetic separation technique with specific anti-fimbriae polyclonal antibodies (pAbs). The transposon was inserted in an ORF, called orf176, which encoded a protein of unknown function. The transposon prevented the transcription of orf176 as well as two genes located downstream, which are designated cspA and cspB and which form the csp operon. Sequence analyses of CspA and CspB revealed that both proteins possessed the classic cell-wall-anchoring motif (LPXTG) of Gram-positive bacterial surface proteins. Recombinant CspA (rCspA) and CspB (rCspB) proteins were generated in Escherichia coli and used to produce pAbs. Immunolocalization experiments showed that anti-rCspB, but not anti-rCspA antibodies specifically recognized S. salivarius fimbriae. Our results suggested that the csp operon encoded predicted cell-surface proteins, one of which, CspB, was associated with the fimbriae.
|
22. |
Wescombe PA,
Upton M,
Renault P,
Wirawan RE,
Power D,
Burton JP,
Chilcott CN,
Tagg JR,
( 2011 ) Salivaricin 9, a new lantibiotic produced by Streptococcus salivarius. PMID : 21310787 : DOI : 10.1099/mic.0.044719-0 Abstract >>
Salivaricin 9 (Sal9) is a 2560 Da lantibiotic having just 46 % amino acid identity with its closest known homologue, the Streptococcus pyogenes lantibiotic SA-FF22. The Sal9 locus (designated siv) in Streptococcus salivarius strain 9 was partially sequenced and localized to an approximately 170 kb megaplasmid, which also harbours the locus for the lantibiotic salivaricin A4. The entire locus was fully characterized in the draft genome sequence of S. salivarius strain JIM8780 and shown to consist of eight genes, having the following putative functions: sivK, sensor kinase; sivR, response regulator; sivA, Sal9 precursor peptide; sivM, lantibiotic modification enzyme; sivT, ABC transporter involved in the export of Sal9 and concomitant cleavage of its leader peptide; and sivFEG, encoding lantibiotic self-immunity. Intriguingly, in contrast to strain 9, the siv locus was chromosomally located in strain JIM8780--the first lantibiotic locus shown not to be exclusively plasmid-associated in S. salivarius. Sal9-containing extracts specifically induced lantibiotic production in both strain 9 and strain JIM8780, indicating that Sal9 functions as a signal peptide for upregulation of its own biosynthesis. Screening representative strains of three streptococcal species (S. salivarius, S. pyogenes and S. mitis) for sivA indicated that it was present only in S. salivarius, with 12 of 28 tested S. salivarius positive. Since Sal9 was inhibitory to all tested S. pyogenes strains it appears to have potential as an important component of the bacteriocin armoury of S. salivarius probiotics intended to control S. pyogenes infections of the human oral cavity.
|
23. |
Itzek A,
Gillen CM,
Fulde M,
Friedrichs C,
Rodloff AC,
Chhatwal GS,
Nitsche-Schmitz DP,
( 2010 ) Contribution of plasminogen activation towards the pathogenic potential of oral streptococci. PMID : 21072208 : DOI : 10.1371/journal.pone.0013826 PMC : PMC2972214 Abstract >>
Oral streptococci are a heterogeneous group of human commensals, with a potential to cause serious infections. Activation of plasminogen has been shown to increase the virulence of typical human pathogenic streptococci such as S. pneumoniae. One important factor for plasminogen activation is the streptococcal �\-enolase. Here we report that plasminogen activation is also common in oral streptococci species involved in clinical infection and that it depends on the action of human plasminogen activators. The ability to activate plasminogen did not require full conservation of the internal plasminogen binding sequence motif FYDKERKVY of �\-enolase that was previously described as crucial for increased plasminogen binding, activation and virulence. Instead, experiments with recombinant �\-enolase variants indicate that the naturally occurring variations do not impair plasminogen binding. In spite of these variations in the internal plasminogen binding motif oral streptococci showed similar activation of plasminogen. We conclude that the pathomechanism of plasminogen activation is conserved in oral streptococci that cause infections in human. This may contribute to their opportunistic pathogenic character that is unfurled in certain niches.
|
24. |
Maeda Y,
Murayama M,
Goldsmith CE,
Coulter WA,
Mason C,
Millar BC,
Dooley JS,
Lowery CJ,
Matsuda M,
Rendall JC,
Elborn JS,
Moore JE,
( 2011 ) Molecular characterization and phylogenetic analysis of quinolone resistance-determining regions (QRDRs) of gyrA, gyrB, parC and parE gene loci in viridans group streptococci isolated from adult patients with cystic fibrosis. PMID : 21193474 : DOI : 10.1093/jac/dkq485 Abstract >>
Ciprofloxacin is the most frequently used member of the fluoroquinolones during initial eradication therapy of Pseudomonas aeruginosa, as well as during acute pulmonary exacerbations. However, its long-term effect on the susceptibility of the commensal flora within the cystic fibrosis (CF) airways has not yet been examined. The aim of this study was therefore to examine the consequence of oral ciprofloxacin usage on the resistance of the commensal viridans group streptococci (VGS), in terms of MICs and mutational analysis of the quinolone resistance-determining regions (QRDRs). The MICs of ciprofloxacin, efflux activities and amino acid substitutions in the QRDRs for 190 isolates of VGS, originating from the sputa of adult CF patients who had been exposed constantly to ciprofloxacin, were examined. VGS organisms included Streptococcus salivarius, Streptococcus mitis, Streptococcus sanguinis, Streptococcus oralis, Streptococcus parasanguinis, Streptococcus infantis, Streptococcus gordonii, Streptococcus anginosus, Streptococcus cristatus, Streptococcus australis and Streptococcus mutans. Ciprofloxacin susceptibility was determined by broth microdilution and QRDRs within the gyrA, gyrB, parC and parE gene loci were explored using sequence analysis. Twenty-seven (14.2%) streptococcal isolates were resistant to ciprofloxacin (MICs ?8 mg/L) and 21 (11.1%) had reduced susceptibility (MICs 4 mg/L). As a comparator, clinically non-significant and non-invasive VGS organisms were examined in 12 consecutive non-CF patients in the community, where no resistance to ciprofloxacin was observed. Five novel QRDR PCR assays were developed to elucidate mutations within the CF VGS population, where there were six positions, which corresponded to previously reported quinolone resistance responsible mutations, and eight novel potential QRDR resistance mutations. Double mutations in gyrA and parC/parE led to MICs of 16 to >64 mg/L, while single mutations in parC or parE resulted in MICs of 8-32 mg/L and 8 mg/L, respectively. The mean homologies of each species to Streptococcus pneumoniae R6 were: gyrA, 70.3%-95%; gyrB, 69.6%-96.2%; parC, 76.1%-94.8%; and parE, 70.7%-94.7%. The close relatives of S. pneumoniae, S. mitis and S. oralis, showed high similarity for all four genes (more than 86%). Treatment of P. aeruginosa with oral ciprofloxacin in patients with CF may concurrently reduce antibiotic susceptibility in the commensal VGS flora, where these organisms may potentially act as a reservoir of fluoroquinolone resistance gene determinants for newly acquired and antibiotic-susceptible pathogens, particularly the Streptococcus milleri group.
|
25. |
Shewmaker PL,
Gertz RE,
Kim CY,
de Fijter S,
DiOrio M,
Moore MR,
Beall BW,
( 2010 ) Streptococcus salivarius meningitis case strain traced to oral flora of anesthesiologist. PMID : 20504987 : DOI : 10.1128/JCM.00426-10 PMC : PMC2897516 Abstract >>
Two women in labor received intrapartum spinal anesthesia from the same anesthesiologist approximately 1 h apart. Within 15 h, both patients developed Streptococcus salivarius meningitis and one patient died. Blood and cerebrospinal fluid (CSF) samples from both patients and tongue swab specimens from the anesthesiologist yielded isolates of an indistinguishable S. salivarius strain.
|
26. |
Glazunova OO,
Raoult D,
Roux V,
( 2009 ) Partial sequence comparison of the rpoB, sodA, groEL and gyrB genes within the genus Streptococcus. PMID : 19620365 : DOI : 10.1099/ijs.0.005488-0 Abstract >>
Phylogenetic analysis and species identification of members of the genus Streptococcus were carried out using partial sequence comparison of the 16S rRNA gene (1468-1478 bp), rpoB, encoding the beta subunit of RNA polymerase (659-680 bp), sodA, encoding the manganese-dependent superoxide dismutase (435-462 bp), groEL, encoding the 60 kDa heat-shock protein (757 bp), and gyrB, encoding the Beta subunit of DNA gyrase (458-461 bp). For the first time, most species within the genus Streptococcus were represented in the study (65 strains, representing 58 species and nine subspecies). Phylogenies inferred from rpoB, sodA, gyrB and groEL sequence comparisons were more discriminative than those inferred from 16S rRNA gene sequence comparison, and showed common clusters. The minimal interspecies divergence was 0.3, 2.7, 0, 2.5 and 3.4 % for the 16S rRNA gene, rpoB, sodA, gyrB and groEL, respectively. In general, groEL partial gene sequence comparison represented the best tool for identifying species and subspecies and for phylogenetic analysis.
|
27. |
Glazunova OO,
Raoult D,
Roux V,
( 2010 ) Partial recN gene sequencing: a new tool for identification and phylogeny within the genus Streptococcus. PMID : 19880633 : DOI : 10.1099/ijs.0.018176-0 Abstract >>
Partial sequences of the recN gene (1249 bp), which encodes a recombination and repair protein, were analysed to determine the phylogenetic relationship and identification of streptococci. The partial sequences presented interspecies nucleotide similarity of 56.4-98.2 % and intersubspecies similarity of 89.8-98 %. The mean DNA sequence similarity of recN gene sequences (66.6 %) was found to be lower than those of the 16S rRNA gene (94.1 %), rpoB (84.6 %), sodA (74.8 %), groEL (78.1 %) and gyrB (73.2 %). Phylogenetically derived trees revealed six statistically supported groups: Streptococcus salivarius, S. equinus, S. hyovaginalis/S. pluranimalium/S. thoraltensis, S. pyogenes, S. mutans and S. suis. The 'mitis' group was not supported by a significant bootstrap value, but three statistically supported subgroups were noted: Streptococcus sanguinis/S. cristatus/S. sinensis, S. anginosus/S. intermedius/S. constellatus (the 'anginosus' subgroup) and S. mitis/S. infantis/S. peroris/S. oralis/S. oligofermentans/S. pneumoniae/S. pseudopneumoniae. The partial recN gene sequence comparison highlighted a high percentage of divergence between Streptococcus dysgalactiae subsp. dysgalactiae and S. dysgalactiae subsp. equisimilis. This observation is confirmed by other gene sequence comparisons (groEL, gyrB, rpoB and sodA). A high percentage of similarity was found between S. intermedius and S. constellatus after sequence comparison of the recN gene. To study the genetic diversity among the 'anginosus' subgroup, recN, groEL, sodA, gyrB and rpoB sequences were determined for 36 clinical isolates. The results that were obtained confirmed the high genetic diversity within this group of streptococci.
|
28. |
Pombert JF,
Sistek V,
Boissinot M,
Frenette M,
( 2009 ) Evolutionary relationships among salivarius streptococci as inferred from multilocus phylogenies based on 16S rRNA-encoding, recA, secA, and secY gene sequences. PMID : 19878555 : DOI : 10.1186/1471-2180-9-232 PMC : PMC2777182 Abstract >>
Streptococci are divided into six phylogenetic groups, i.e, anginosus, bovis, mitis, mutans, pyogenic, and salivarius, with the salivarius group consisting of only three distinct species. Two of these species, Streptococcus salivarius and Streptococcus vestibularis, are members of the normal human oral microflora whereas the third, Streptococcus thermophilus, is found in bovine milk. Given that S. salivarius and S. vestibularis share several physiological characteristics, in addition to inhabiting the same ecosystem, one would assume that they would be more closely related to each other than to S. thermophilus. However, the few phylogenetic trees published so far suggest that S. vestibularis is more closely related to S. thermophilus. To determine whether this phylogenetic relationship is genuine, we performed phylogenetic inferences derived from secA and secY, the general secretion housekeeping genes, recA, a gene from a separate genetic locus that encodes a major component of the homologous recombinational apparatus, and 16S rRNA-encoding gene sequences using other streptococcal species as outgroups. The maximum likelihood (ML) and maximum parsimony (MP) phylogenetic inferences derived from the secA and recA gene sequences provided strong support for the S. vestibularis/S. thermophilus sister-relationship, whereas 16S rRNA-encoding and secY-based analyses could not discriminate between alternate topologies. Phylogenetic analyses derived from the concatenation of these sequences unambiguously supported the close affiliation of S. vestibularis and S. thermophilus. Our results corroborated the sister-relationship between S. vestibularis and S. thermophilus and the concomitant early divergence of S. salivarius at the base of the salivarius lineage.
|
29. |
Tazumi A,
Maeda Y,
Goldsmith CE,
Coulter WA,
Mason C,
Millar BC,
McCalmont M,
Rendall J,
Elborn JS,
Matsuda M,
Moore JE,
( 2009 ) Molecular characterization of macrolide resistance determinants [erm(B) and mef(A)] in Streptococcus pneumoniae and viridans group streptococci (VGS) isolated from adult patients with cystic fibrosis (CF). PMID : 19584106 : DOI : 10.1093/jac/dkp213 Abstract >>
Although long-term use of azithromycin has shown a significant clinical improvement for patients with cystic fibrosis (CF), its long-term effect on the susceptibility of commensal flora within CF airways has not yet been examined. We therefore suggest that long-term use of azithromycin increases macrolide resistance in commensal streptococci. Erythromycin susceptibility in naturally colonizing viridans group streptococci (VGS) was characterized, as well as macrolide resistance gene determinants through sequence analysis, in pneumococci (n = 15) and VGS [n = 84; i.e. Streptococcus salivarius (n = 30), Streptococcus mitis (n = 17), Streptococcus sanguinis (n = 11), Streptococcus oralis (n = 10), Streptococcus parasanguinis (n = 6), Streptococcus gordonii (n = 3), Streptococcus infantis (n = 3), Streptococcus cristatus (n = 2), Streptococcus anginosus (n = 1) and Streptococcus australis (n = 1)] isolated from sputum from 24 adult CF patients, who were on oral azithromycin therapy for at least the previous 7 months. Almost three-quarters of isolates (74; 74.7%) were resistant to erythromycin, whilst a further 15 (15.2%) had reduced susceptibility, leaving only 10 (10.1%) isolates susceptible to erythromycin. The majority (89.8%) were not susceptible to erythromycin, as demonstrated by possession of the erm(B) gene in 25/99 (25.3%), the mef(A) gene in 1/99 (1.0%), the mef(E) gene in 75/99 (75.8%) and both erm(B) and mef(E) genes simultaneously in 11/99 (11.1%). These results indicate that genotypic resistance for macrolides is common in VGS in adult CF patients, with efflux being over three times more frequent. Long-term treatment with azithromycin in CF patients may reduce antibiotic susceptibility in commensal VGS, where these organisms may potentially act as a reservoir of macrolide resistance determinants for newly acquired and antibiotic-susceptible pathogens.
|
30. |
Santagati M,
Lupo A,
Scillato M,
Di Martino A,
Stefani S,
( 2009 ) Conjugal mobilization of the mega element carrying mef(E) from Streptococcus salivarius to Streptococcus pneumoniae. PMID : 19025575 : DOI : 10.1111/j.1574-6968.2008.01408.x Abstract >>
We report the isolation and characterization of an unusual strain of Streptococcus salivarius, 3C30, displaying both the macrolide-lincosamide-streptogramin B and the tetracycline resistance phenotypes. It harbours the mef(E), erm(B), and tet(M) genes carried by different genetic elements. The genetic element carrying mef(E), named mega, was investigated by long PCR and sequencing, while the presence of the Tn3872-like element, carrying tet(M) and erm(B), was demonstrated by sequencing of both the int-xis-Tn and the fragment between the two resistance genes. In strain 3C30 the mega element is 5388 bp in size and its nucleotide sequence is identical to that of the element described previously in S. salivarius, with the exception of a 912 bp deletion at the left end. The composite Tn3872-like element appeared to be nonconjugative while the mega element was transferred by conjugation to Streptococcus pneumoniae. It was, however, impossible to transfer it again from these transconjugants to other strains. In addition, only in the 3C30 strain did mega form circular structures, as identified by real-time PCR. In conclusion, we found a clinical strain of S. salivarius carrying both mega and Tn3872-like genetic elements. Mega is transferable by conjugation to S. pneumoniae but it is not transferable again from the transconjugants, suggesting a possible mobilization by recombinases of the coresident Tn3872-like transposon.
|
31. |
Warburton P,
Roberts AP,
Allan E,
Seville L,
Lancaster H,
Mullany P,
( 2009 ) Characterization of tet(32) genes from the oral metagenome. PMID : 18955517 : DOI : 10.1128/AAC.00788-08 PMC : PMC2612163 Abstract >>
tet(32) Was identified in three bacterial isolates and in metagenomic DNA from the human oral cavity. The regions immediately flanking the gene were found to have similarities to the mobile elements TnB1230 from Butyrivibrio fibrisolvens, ATE-3 from Arcanobacterium pyogenes, and CTn5 from Clostridium difficile.
|
32. |
Giffard PM,
Simpson CL,
Milward CP,
Jacques NA,
( 1991 ) Molecular characterization of a cluster of at least two glucosyltransferase genes in Streptococcus salivarius ATCC 25975. PMID : 1838391 : DOI : 10.1099/00221287-137-11-2577 Abstract >>
The oral micro-organism Streptococcus salivarius ATCC 25975 synthesizes extracellular glucosyltransferases (GTFs) which polymerize the glucose moiety of sucrose into glucan polymers. Two separate genes encoding the activities of a GTF-I (a GTF that synthesizes an insoluble product) and a GTF-S (a GTF that synthesizes soluble product) were cloned into bacteriophage lambda L47.1. The inserts in the lambda-clones were characterized by restriction mapping and Southern hybridization and were found to overlap, implying that the two genes lay very close to one another on the S. salivarius chromosome. Both genes were subcloned into phagemid vector pIBI30 where they were expressed at a high level. The GTF-I-encoding gene was named gtfJ and the GTF-S-encoding gene, gtfK. Nucleotide sequencing showed that gtfJ and most probably gtfK were closely related to the gtf genes of the mutans streptococci. Sequence alignment also indicated that gtfK lay very close to and downstream from gtfJ, and that both were transcribed in the same direction.
|
33. |
Chen YY,
Feng CW,
Chiu CF,
Burne RA,
( 2008 ) cadDX operon of Streptococcus salivarius 57.I. PMID : 18165364 : DOI : 10.1128/AEM.01878-07 PMC : PMC2258612 Abstract >>
A CadDX system that confers resistance to Cd(2+) and Zn(2+) was identified in Streptococcus salivarius 57.I. Unlike with other CadDX systems, the expression of the cad promoter was negatively regulated by CadX, and the repression was inducible by Cd(2+) and Zn(2+), similar to what was found for CadCA systems. The lower G+C content of the S. salivarius cadDX genes suggests acquisition by horizontal gene transfer.
|
34. |
Vadeboncoeur C,
Konishi Y,
Dumas F,
Gauthier L,
Frenette M,
( 1991 ) HPr polymorphism in oral streptococci is caused by the partial removal of the N-terminal methionine. PMID : 1799636 : DOI : 10.1016/0300-9084(91)90174-y Abstract >>
HPr is a small phosphocarrier protein of the bacterial phosphoenolpyruvate:sugar phosphotransferase system involved in the transport and phosphorylation of sugars. It has recently been reported that streptococci possess two forms of HPr having identical biochemical properties. In this communication, we show by N-terminal amino-acid sequencing and by ion-spray mass spectroscopy that these two forms differ by the presence or the absence of the N-terminal methionine.
|
35. |
Zhou M,
Yang Q,
Kudinha T,
Zhang L,
Xiao M,
Kong F,
Zhao Y,
Xu YC,
( 2016 ) Using Matrix-Assisted Laser Desorption Ionization-Time of Flight (MALDI-TOF) Complemented with Selected 16S rRNA and gyrB Genes Sequencing to Practically Identify Clinical Important Viridans Group Streptococci (VGS). PMID : 27617008 : DOI : 10.3389/fmicb.2016.01328 PMC : PMC5000867 Abstract >>
There are challenges in viridans group streptococci (VGS) identification especially for the mitis group. Few studies have investigated the performance of MALDI-TOF MS system in VGS identification. Using 16S rRNA gene and gyrB gene sequencing as a gold standard, the performance of two MALDI-TOF MS instruments in the identification of 181 VGS clinical isolates was studied. The Bruker Biotyper and Vitek MS IVD systems correctly identified 88.4% and 98.9% of the 181 isolates, respectively. The Vitek MS RUO system was the least reliable, only correctly identifying 38.7% of the isolates to species level with several misidentifications and invalid results. The Bruker Biotyper system was very unreliable in the identification of species within the mitis group. Among 22 non-pneumococci isolates (S. mitis/S. oralis/S. pseudopneumoniae), Biotyper misidentified 21 of them as S. pneumoniae leading to a low sensitivity and low positive predictive value in these species. In contrast, the Vitek MS IVD demonstrated a better resolution for pneumococci and non-pneumococci despite the inability to distinguish between S. mitis/S. oralis. For more accurate species-level identification, further improvements in the VGS spectra databases are needed. Based on MALDI-TOF analysis and selected 16S rRNA gene plus gyrB genes sequencing, we designed a practical VGS identification algorithm.
|
36. |
Naveen Kumar V,
van der Linden M,
Menon T,
Nitsche-Schmitz DP,
( 2014 ) Viridans and bovis group streptococci that cause infective endocarditis in two regions with contrasting epidemiology. PMID : 24220665 : DOI : 10.1016/j.ijmm.2013.10.004 Abstract >>
Viridans group (VGS) or bovis group streptococci (BGS) are the major causes for streptococcal infective endocarditis (IE). However, the causative isolates are not sufficiently characterized. Using multilocus sequence analysis we have examined VGS and BGS (VGS/BGS) isolates that caused IE in southern India and Germany, two distant geographic regions with a contrasting IE epidemiology. Other than in Germany, the majority of patients (68%) in Chennai, southern India had an underlying rheumatic heart disease (RHD). In accord with the high prevalence of RHD in the younger population and with the expansive age structure of India, the median age (24 years) of the VGS/BGS endocarditis patients was lower than in Germany (63 years), where RHD is rare and the age structure is contractive. Both in Germany and in southern India, the majority of cases were caused by mitis group streptococci, however, with considerable differences in the spectra of causative (sub)species. BGS endocarditis was more frequent in Germany. The spectrum of VGS/BGS that cause IE differs considerably between distant geographic regions in which different predisposing conditions prevail. Therefore, improved microbiological diagnosis in IE may facilitate determination of the optimal therapy.
|
37. |
Birri DJ,
Brede DA,
Nes IF,
( 2012 ) Salivaricin D, a novel intrinsically trypsin-resistant lantibiotic from Streptococcus salivarius 5M6c isolated from a healthy infant. PMID : 22101034 : DOI : 10.1128/AEM.06588-11 PMC : PMC3255740 Abstract >>
In this work, we purified and characterized a newly identified lantibiotic (salivaricin D) from Streptococcus salivarius 5M6c. Salivaricin D is a 34-amino-acid-residue peptide (3,467.55 Da); the locus of the gene encoding this peptide is a 16.5-kb DNA segment which contains genes encoding the precursor of two lantibiotics, two modification enzymes (dehydratase and cyclase), an ABC transporter, a serine-like protease, immunity proteins (lipoprotein and ABC transporters), a response regulator, and a sensor histidine kinase. The immunity gene (salI) was heterologously expressed in a sensitive indicator and provided significant protection against salivaricin D, confirming its immunity function. Salivaricin D is a naturally trypsin-resistant lantibiotic that is similar to nisin-like lantibiotics. It is a relatively broad-spectrum bacteriocin that inhibits members of many genera of Gram-positive bacteria, including the important human pathogens Streptococcus pyogenes and Streptococcus pneumoniae. Thus, Streptococcus salivarius 5M6c may be a potential biological agent for the control of oronasopharynx-colonizing streptococcal pathogens or may be used as a probiotic bacterium.
|
38. |
Srinivasan V,
Gertz RE,
Shewmaker PL,
Patrick S,
Chitnis AS,
O'Connell H,
Benowitz I,
Patel P,
Guh AY,
Noble-Wang J,
Turabelidze G,
Beall B,
( 2012 ) Using PCR-based detection and genotyping to trace Streptococcus salivarius meningitis outbreak strain to oral flora of radiology physician assistant. PMID : 22384169 : DOI : 10.1371/journal.pone.0032169 PMC : PMC3285201 Abstract >>
We recently investigated three cases of bacterial meningitis that were reported from a midwestern radiology clinic where facemasks were not worn during spinal injection of contrast agent during myelography procedures. Using pulsed field gel electrophoresis we linked a case strain of S. salivarius to an oral specimen of a radiology physician assistant (RPA). We also used a real-time PCR assay to detect S. salivarius DNA within a culture-negative cerebrospinal fluid (CSF) specimen. Here we extend this investigation through using a nested PCR/sequencing strategy to link the culture-negative CSF specimen to the case strain. We also provide validation of the real-time PCR assay used, demonstrating that it is not solely specific for Streptococcus salivarius, but is also highly sensitive for detection of the closely related oral species Streptococcus vestibularis. Through using multilocus sequence typing and 16S rDNA sequencing we further strengthen the link between the CSF case isolate and the RPA carriage isolate. We also demonstrate that the newly characterized strains from this study are distinct from previously characterized S. salivarius strains associated with carriage and meningitis.
|
39. |
Kitten T,
Munro CL,
Zollar NQ,
Lee SP,
Patel RD,
( 2012 ) Oral streptococcal bacteremia in hospitalized patients: taxonomic identification and clinical characterization. PMID : 22189124 : DOI : 10.1128/JCM.06438-11 PMC : PMC3295135 Abstract >>
Oral streptococci have been associated with systemic diseases, including infective endocarditis and neutropenic bacteremia. We analyzed 58 recent oral streptococcal bloodstream isolates, and we obtained clinical and demographic data for source patients. The sodA gene was found to be a better target than the 16S-23S rRNA internal transcribed spacer for DNA sequence-based species identification. Together, Streptococcus mitis and Streptococcus oralis were significantly more likely than the 12 combined remaining species to be isolated from neutropenic patients.
|
40. |
( 1996 ) The cpsABCDE genes involved in polysaccharide production in Streptococcus salivarius ssp. thermophilus strain NCBF 2393. PMID : 8996082 : DOI : 10.1016/s0378-1119(96)00405-2 Abstract >>
A 4074-bp EcoRI fragment of Streptococcus salivarius ssp. thermophilus (S. thermophilus) chromosomal DNA containing genes involved in exocellular polysaccharide (EPS) was identified and cloned. The nucleotide sequence of this fragment was determined and found to contain one partial and four complete open reading frames. These were designated cpsA, cpsB, cpsC, cpsD and cpsE and encoded proteins of > 130, 243, 230, 246 and 455 amino acids, respectively, that showed homology with the genes of the cps cluster, involved in polysaccharide biosynthesis, in Streptococcus pneumoniae Type 19F. The cpsA gene is predicted to encode a transcriptional regulator, while cpsC and cpsD are predicted to encode proteins involved in polysaccharide polymerization and export. The cpsE gene is likely to encode the phosphate-prenyl glycosyl-1-phosphate transferase catalyzing the first step in polysaccharide biosynthesis in S. thermophilus. Southern blot analysis revealed that cpsE is found only in polysaccharide producing strains of S. thermophilus.
|
41. |
( 1996 ) Streptococcus salivarius urease: genetic and biochemical characterization and expression in a dental plaque streptococcus. PMID : 8550211 : PMC : PMC173805 Abstract >>
The hydrolysis of urea by urease enzyme of oral bacteria is believed to have a major impact on oral microbial ecology and to be intimately involved in oral health and diseases. To begin to understand the biochemistry and genetics of oral ureolysis, a study of the urease of Streptococcus salivarius, a highly ureolytic organism which is present in large numbers on the soft tissues of the oral cavity, has been initiated. By using as a probe a 0.6-kpb internal fragment of the S. salivarius 57.I ureC gene, two clones from subgenomic libraries of S. salivarius 57.I in an Escherichia coli plasmid vector were identified. Nucleotide sequence analysis revealed the presence of one partial and six complete open reading frames which were most homologous to ureIAB-CEFGD of other ureolytic bacteria. Plasmid clones were generated to construct a complete gene cluster and used to transform E. coli and Streptococcus gordonii DL1, a nonureolytic, dental plaque microorganism. The recombinant organisms expressed high levels of urease activity when the growth medium was supplemented with NiCl2. The urease enzyme was purified from E. coli, and its biochemical properties were compared with those of the urease produced by S. salivarius and those of the urease produced by S. gordonii carrying the plasmid-borne ure genes. In all cases, the enzyme had a Km of 3.5 to 4.1 mM, a pH optimum near 7.0, and a temperature optimum near 60 degrees C. S. gordonii carrying the urease genes was then demonstrated to have a significant capacity to temper glycolytic acidification in vitro in the presence of concentrations of urea commonly found in the oral cavity. The ability to genetically engineer plaque bacteria that can modulate environmental pH through ureolysis will open the way to using recombinant ureolytic organisms to test hypotheses regarding the role of oral ureolysis in dental caries, calculus formation, and periodontal diseases. Such recombinant organisms may eventually prove useful for controlling dental caries by replacement therapy.
|
42. |
( 1993 ) The ftf gene encoding the cell-bound fructosyltransferase of Streptococcus salivarius ATCC 25975 is preceded by an insertion sequence and followed by FUR1 and clpP homologues. PMID : 8393068 : DOI : 10.1099/00221287-139-5-913 Abstract >>
Analysis of the region downstream of the ftf gene of Streptococcus salivarius led to the detection of two open reading frames (ORFs). The deduced amino acid sequences of these ORFs were homologous to proteins encoded by genes not previously described and/or sequenced in Gram-positive bacteria. The deduced amino acid sequence of the first of these (orf2) showed strong homology to the product of the FUR1 gene of Saccharomyces cerevisiae, which codes for a uracil phosphoribosyltransferase. The over-expression of the product of this gene appeared to be the source of the detrimental effect observed with phagemids carrying orf2 in Escherichia coli hosts. The deduced amino acid sequence of the second ORF (orf3) was homologous to the ClpP family of proteases. Examination of the upstream region of the ftf gene led to the discovery of a new insertion sequence-like element which has been designated IS1161.
|
43. |
( 1993 ) The cell-bound fructosyltransferase of Streptococcus salivarius: the carboxyl terminus specifies attachment in a Streptococcus gordonii model system. PMID : 8331080 : DOI : 10.1128/jb.175.14.4520-4527.1993 PMC : PMC204894 Abstract >>
The ftf gene, coding for the cell-bound beta-D-fructosyltransferase (FTF) of Streptococcus salivarius ATCC 25975, has been analyzed, and its deduced amino acid sequence has been compared with that of the secreted FTF of Streptococcus mutans and the levansucrases (SacBs) of Bacillus species. A unique proline-rich region detected at the C terminus of the FTF of S. salivarius preceded a hydrophobic terminal domain. This proline-rich region was shown to possess strong homology to the product of the prgC gene from pCF10 in Enterococcus faecalis, which encodes a pheromone-responsive protein of unknown function, as well as homology to the human proline-rich salivary protein PRP-4. A series of 3'-OH deletions of the S. salivarius ftf gene expressed in Streptococcus gordonii Challis LGR2 showed that the C terminus was required for cell surface attachment in this heterologous organism, as only the complete gene product was cell bound. This cell-bound activity was released in the presence of sucrose, suggesting that the mode of attachment and release of the S. salivarius FTF in S. gordonii was similar to that in its native host.
|
44. |
( 1993 ) Phosphotransferase system of Streptococcus salivarius: characterization of the ptsH gene and its product. PMID : 8294015 : DOI : 10.1016/0378-1119(93)90443-7 Abstract >>
The Streptococcus salivarius ptsH gene encoding histidine-containing phosphocarrier protein (HPr) of the phosphotransferase system (PTS) has been cloned, sequenced, and found to be part of a ptsH, ptsI operon. Upstream from ptsH, putative -35 and -10 boxes and a Shine-Dalgarno sequence highly similar to the Escherichia coli consensus regulatory elements were identified. A second promoter, located in the ptsH coding sequence was also observed and is sufficient for the expression of the S. salivarius ptsI gene, encoding enzyme I of the PTS in E. coli [Gagnon et al., Gene 121 (1992) 71-78]. The amino acid sequence of S. salivarius HPr, inferred from the ptsH sequence, shared identity varying between 37 and 76% with known HPr from other bacteria. Moreover, the S. salivarius HPr shared 78% identity with an HPr-like protein of Aspergillus fumigatus, a eukaroytic mold that does not possess a functional PTS. Expression analysis of S. salivarius HPr in E. coli demonstrated that (i) S. salivarius ptsH is expressed in E. coli under the control of its own promoter, (ii) S. salivarius HPr synthesized by E. coli is completely processed by methionine aminopeptidase, and (iii) S. salivarius HPr is phosphorylated in vivo by E. coli enzyme I. It was also observed that, in E. coli, the copy number of pUC18 bearing S. salivarius ptsH was reduced more than 25-fold, as compared to pUC18 without an insertion.
|
45. |
( 1993 ) Sequence of the gtfK gene of Streptococcus salivarius ATCC 25975 and evolution of the gtf genes of oral streptococci. PMID : 8371114 : DOI : 10.1099/00221287-139-7-1511 Abstract >>
Many strains of oral streptococci secrete glucosyltransferases (GTFs) that polymerize sucrose into glucans that form an integral part of the plaque matrix on the tooth surface. Recently, we reported the cloning of two closely linked GTF-encoding genes (gtfJ and gtfK) from Streptococcus salivarius ATCC 25975 as well as the sequence of gtfJ, which encodes a primer-dependent GTF that synthesizes an insoluble product (a GTF-I). In this communication we report the sequence of gtfK, which encodes a primer-dependent GTF that synthesizes a soluble product (a GTF-S), as well as the sequence of a small downstream open reading frame of unknown function. The deduced sequence of GtfK was compared with those of seven other streptococcal Gtfs and an unrooted phylogenetic tree constructed. This analysis suggested that Gtfs with similar product specificities do not form phylogenetic clusters and was consistent with currently accepted phylogenetic schemes. The tree was tested by constructing a series of 'sub-trees' from different blocks of the alignment. Evidence was obtained for recombination events involving gtfB and gtfC from S. mutans GS-5, gtfJ and gtfK from S. salivarius, as well as the gtfI genes from S. downei and S. sobrinus. The recombination events between gtfB and gtfC, and between the two gtfI genes, were confirmed by examining divergences at silent sites.
|
46. |
( 1993 ) Isolation and characterization of the lantibiotic salivaricin A and its structural gene salA from Streptococcus salivarius 20P3. PMID : 8357242 : PMC : PMC182229 Abstract >>
A bacteriocin-like inhibitory substance, salivaricin A, was purified from cultures of Streptococcus salivarius 20P3 and was shown by ion spray mass spectrometry to have a molecular mass of 2,315 +/- 1.1 Da. Amino acid composition analysis demonstrated the presence of lanthionine, indicating that salivaricin A may be a member of the lantibiotic class of antibiotic substances. The sequence of eight amino acids at the N terminus of the molecule was determined by Edman degradation, and mixed oligonucleotide probes based on part of this sequence (GSGWIA) were used to detect the salivaricin A structural gene. A 6.2-kb EcoRI fragment of chromosomal DNA from strain 20P3 that hybridized with the probes was cloned, and the hybridizing region was further localized to a 379-bp DraI-AluI fragment. Analysis of the nucleotide sequence of this fragment indicated that salivaricin A is synthesized as a 51-amino-acid prepeptide that is posttranslationally modified and cleaved to give a biologically active 22-residue peptide containing one lanthionine and two beta-methyllanthionine residues. The secondary structure of presalivaricin A was predicted to be similar to that of type A lantibiotics, with a hydrophilic alpha-helical leader sequence and a propeptide region with potential for beta-turn formation and a lack of alpha-helicity. The sequence around the cleavage site of presalivaricin A differed from that of other type A lantibiotics but was similar to that of several bacteriocin-like inhibitory substances produced by lactic acid bacteria.
|
47. |
Ohnishi Y,
Kubo S,
Ono Y,
Nozaki M,
Gonda Y,
Okano H,
Matsuya T,
Matsushiro A,
Morita T,
( 1995 ) Cloning and sequencing of the gene coding for dextranase from Streptococcus salivarius. PMID : 7737522 : DOI : 10.1016/0378-1119(95)00071-d Abstract >>
We cloned and sequenced the dextranase (Dex) (1,6-alpha-glucanhydrolase; EC 3.2.1.11)-encoding gene from Streptococcus salivarius (Ss) strain M-33. Recombinant clones from an Ss genomic library specifying Dex activity were identified as colonies surrounded by transparent halos on blue dextran plates. One of the clones had a 4.3-kb KpnI fragment containing the gene coding for an 826-amino-acid polypeptide with a molecular mass of 87.9 kDa, which corresponds well to that of native Dex from the Ss culture supernatant. There was no sequence homology between the gene encoding Ss Dex and the gene encoding dextran glucosidase of S. mutans, or between their protein products.
|
48. |
( 1994 ) Aminopeptidase N from Streptococcus salivarius subsp. thermophilus NCDO 573: purification and properties. PMID : 7989254 : Abstract >>
A 96 kDa aminopeptidase was purified from Streptococcus salivarius subsp. thermophilus NCDO 573. The enzyme had similar properties to aminopeptidases isolated from lactococci and lactobacilli and showed a high degree of N-terminal amino acid sequence homology to aminopeptidase N from Lactococcus lactis subsp. cremoris. It catalysed the hydrolysis of a range of aminoacyl 4-nitroanilides and 7-amido-4-methylcoumarin derivatives, dipeptides, tripeptides and oligopeptides. In common with aminopeptidases from other lactic acid bacteria, the enzyme from Strep. salivarius subsp. thermophilus showed highest activity with lysyl derivatives but was also very active with arginyl and leucyl derivatives. Relative activity with alanyl, phenylalanyl, tyrosyl, seryl and valyl derivatives was considerably lower and with glycyl, glutamyl and prolyl derivatives almost negligible. The aminopeptidase also catalysed the hydrolysis of dipeptides and tripeptides but mostly at rates much less than that with L-lysyl-4-nitroanilide and oligopeptides. The enzyme catalysed the successive hydrolysis of various amino acid residues from the N-terminus of several oligopeptides but it was unable to cleave peptide bonds on the N-terminal side of a proline residue.
|
49. |
( 1994 ) IS1139 from Streptococcus salivarius: identification and characterization of an insertion sequence-like element related to mobile DNA elements from gram-negative bacteria. PMID : 7991667 : DOI : 10.1006/plas.1994.1038 Abstract >>
An insertion sequence-like element, IS1139, was cloned and sequenced from Streptococcus salivarius ATCC 25975 chromosome. This insertion sequence-like element is 1168 bp long and is delimited by inverted repeats of 29 bp and by a duplicated sequence of 6 bp. This IS possesses an open reading frame that codes for a putative transposase of 339 amino acids which has, respectively, 94, 35, 33, and 30% amino-acid identity with the transposases of IS1161 from S. salivarius ATCC 25975, IS4351 from Bacteroides fragilis, IS30 from Escherichia coli, and IS1086 from Alcaligenes eutrophus. Sequence analysis revealed that these transposases may have evolved from a common ancestral gene. Southern hybridization of restriction endonuclease-digested genomic DNA from 21 strains of oral streptococci, using a probe specific to the transposase-encoding gene (tnpA), revealed that IS1139 is found in two strains of S. salivarius, ATCC 25975 and ATCC 13419, in eight and two copies, respectively.
|
50. |
Simpson CL,
Giffard PM,
Jacques NA,
( 1995 ) Streptococcus salivarius ATCC 25975 possesses at least two genes coding for primer-independent glucosyltransferases. PMID : 7822030 : PMC : PMC173040 Abstract >>
Fractionation of the culture medium showed that Streptococcus salivarius ATCC 25975 secreted a glucosyltransferase (Gtf) that was primer independent. On the basis of this observation, a gene library of S. salivarius chromosomal DNA cloned into lambda L47.1 was screened for a gene(s) coding for such an activity. As a result of this screening process, two new gtf genes, gtfL and gtfM, both of which coded for primer-independent Gtf activities, were isolated. GtfL produced an insoluble glucan that was refractory to digestion by the endo-(1-->6)-alpha-D-glucanase. of Chaetonium gracile, while GtfM produced a soluble glucan that was readily degraded by the glucanase. Comparison of the deduced amino acid sequences of gtfL and gtfM with 10 other available Gtf sequences allowed the relatedness of the conserved catalytic regions to be assessed. This analysis showed that the 12 enzymes did not form clusters based on their primer dependencies or on their product solubilities. Further analysis of the YG repeats in the C-terminal glucan-binding domains of GtfJ, GtfK, GtfL, and GtfM from S. salivarius showed that there was strong homology between a block of contiguous triplet YG repeats present in the four alleles. These blocks of YG repeats were coded for by a region of each gene that appeared to have arisen as a result of a recent duplication event(s).
|
51. |
Rul F,
Monnet V,
Gripon JC,
( 1994 ) Purification and characterization of a general aminopeptidase (St-PepN) from Streptococcus salivarius ssp. thermophilus CNRZ 302. PMID : 7836577 : DOI : 10.3168/jds.S0022-0302(94)77228-3 Abstract >>
A general aminopeptidase (St-PepN) was purified from an intracellular extract of Streptococcus salivarius ssp. thermophilus CNRZ 302 by ion-exchange chromatography and hydrophobic interaction chromatography. Gel electrophoresis of the purified enzyme in denaturing or nondenaturating conditions showed a single protein band. The enzyme is a monomer with a molecular mass of 97 kDa. Its activity is maximal at pH 7 and 36 degrees C and is completely abolished by CuCl2 and ZnCl2. The enzyme is strongly inhibited by metal-chelating reagents, such as EDTA and o-phenanthroline, which suggests that St-PepN is a metalloenzyme. The enzyme showed activity toward p-nitroanilide derivatives or dipeptides and tripeptides and showed a preference for hydrophobic or basic amino acids at the N-terminal position. Longer peptide chains, such as the B-chain of insulin, glucagon, or peptides generated by the hydrolysis of caseins, were degraded, too. The sequence of the first 21 residues of the mature enzyme was determined and showed high homology with that of the aminopeptidase PepN isolated from Lactococcus lactis ssp. cremoris Wg2. The properties of the enzyme are compared with those of corresponding enzymes of other species of lactic acid bacteria.
|
52. |
Jiang X,
Hall AB,
Arthur TD,
Plichta DR,
Covington CT,
Poyet M,
Crothers J,
Moses PL,
Tolonen AC,
Vlamakis H,
Alm EJ,
Xavier RJ,
( 2019 ) Invertible promoters mediate bacterial phase variation, antibiotic resistance, and host adaptation in the gut. PMID : 30630933 : DOI : 10.1126/science.aau5238 PMC : PMC6543533 Abstract >>
Phase variation, the reversible alternation between genetic states, enables infection by pathogens and colonization by commensals. However, the diversity of phase variation remains underexplored. We developed the PhaseFinder algorithm to quantify DNA inversion-mediated phase variation. A systematic search of 54,875 bacterial genomes identified 4686 intergenic invertible DNA regions (invertons), revealing an enrichment in host-associated bacteria. Invertons containing promoters often regulate extracellular products, underscoring the importance of surface diversity for gut colonization. We found invertons containing promoters regulating antibiotic resistance genes that shift to the ON orientation after antibiotic treatment in human metagenomic data and in vitro, thereby mitigating the cost of antibiotic resistance. We observed that the orientations of some invertons diverge after fecal microbiota transplant, potentially as a result of individual-specific selective forces.
|
53. |
Wyllie AL,
Pannekoek Y,
Bovenkerk S,
van Engelsdorp Gastelaars J,
Ferwerda B,
van de Beek D,
Sanders EAM,
Trzci?ski K,
van der Ende A,
( 2017 ) Sequencing of the variable region of rpsB to discriminate between Streptococcus pneumoniae and other streptococcal species. PMID : 28931649 : DOI : 10.1098/rsob.170074 PMC : PMC5627049 Abstract >>
The vast majority of streptococci colonizing the human upper respiratory tract are commensals, only sporadically implicated in disease. Of these, the most pathogenic is Mitis group member, Streptococcus pneumoniae Phenotypic and genetic similarities between streptococci can cause difficulties in species identification. Using ribosomal S2-gene sequences extracted from whole-genome sequences published from 501 streptococci, we developed a method to identify streptococcal species. We validated this method on non-pneumococcal isolates cultured from cases of severe streptococcal disease (n = 101) and from carriage (n = 103), and on non-typeable pneumococci from asymptomatic individuals (n = 17) and on whole-genome sequences of 1157 pneumococcal isolates from meningitis in the Netherlands. Following this, we tested 221 streptococcal isolates in molecular assays originally assumed specific for S. pneumoniae, targeting cpsA, lytA, piaB, ply, Spn9802, zmpC and capsule-type-specific genes. Cluster analysis of S2-sequences showed grouping according to species in line with published phylogenies of streptococcal core genomes. S2-typing convincingly distinguished pneumococci from non-pneumococcal species (99.2% sensitivity, 100% specificity). Molecular assays targeting regions of lytA and piaB were 100% specific for S. pneumoniae, whereas assays targeting cpsA, ply, Spn9802, zmpC and selected serotype-specific assays (but not capsular sequence typing) showed a lack of specificity. False positive results were over-represented in species associated with carriage, although no particular confounding signal was unique for carriage isolates.
|
54. |
( 1999 ) The HPr(Ser) kinase of Streptococcus salivarius: purification, properties, and cloning of the hprK gene. PMID : 9922231 : PMC : PMC93434 Abstract >>
In gram-positive bacteria, HPr, a protein of the phosphoenolpyruvate:sugar phosphotransferase system, is phosphorylated on a serine residue at position 46 by an ATP-dependent protein kinase. The HPr(Ser) kinase of Streptococcus salivarius ATCC 25975 was purified, and the encoding gene (hprK) was cloned by using a nucleotide probe designed from the N-terminal amino acid sequence. The predicted amino acid sequence of the S. salivarius enzyme showed 45% identity with the Bacillus subtilis enzyme, the conserved residues being located mainly in the C-terminal half of the protein. The predicted hprK gene product has a molecular mass of 34,440 Da and a pI of 5.6. These values agree well with those found experimentally by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate, molecular sieve chromatography in the presence of guanidine hydrochloride, and chromatofocusing using the purified protein. The native protein migrates on a Superdex 200 HR column as a 330,000-Da protein, suggesting that the HPr(Ser) kinase is a decamer. The enzyme requires Mg2+ for activity and functions optimally at pH 7.5. Unlike the enzyme from other gram-positive bacteria, the HPr(Ser) kinase from S. salivarius is not stimulated by FDP or other glycolytic intermediates. The enzyme is inhibited by inorganic phosphate, and its Kms for HPr and ATP are 31 microM and 1 mM, respectively.
|
55. |
( 1998 ) Transcriptional regulation of the Streptococcus salivarius 57.I urease operon. PMID : 9791132 : PMC : PMC107641 Abstract >>
The Streptococcus salivarius 57.I ure cluster was organized as an operon, beginning with ureI, followed by ureABC (structural genes) and ureEFGD (accessory genes). Northern analyses revealed transcripts encompassing structural genes and transcripts containing the entire operon. A sigma70-like promoter could be mapped 5' to ureI (PureI) by primer extension analysis. The intensity of the signal increased when cells were grown at an acidic pH and was further enhanced by excess carbohydrate. To determine the function(s) of two inverted repeats located 5' to PureI, transcriptional fusions of the full-length promoter region (PureI), or a deletion derivative (PureIDelta100), and a promoterless chloramphenicol acetyltransferase (CAT) gene were constructed and integrated into the chromosome to generate strains PureICAT and PureIDelta100CAT, respectively. CAT specific activities of PureICAT were repressed at pH 7.0 and induced at pH 5.5 and by excess carbohydrate. In PureIDelta100CAT, CAT activity was 60-fold higher than in PureICAT at pH 7.0 and pH induction was nearly eliminated, indicating that expression was negatively regulated. Thus, it was concluded that PureI was the predominant, regulated promoter and that regulation was governed by a mechanism differing markedly from other known mechanisms for bacterial urease expression.
|
56. |
( 2012 ) Use of tuf as a target for sequence-based identification of Gram-positive cocci of the genus Enterococcus, Streptococcus, coagulase-negative Staphylococcus, and Lactococcus. PMID : 23181410 : DOI : 10.1186/1476-0711-11-31 PMC : PMC3533577 Abstract >>
Accurate identification of isolates belonging to genus Enterococcus, Streptococcus, coagulase-negative Staphylococcus, and Lactococcus at the species level is necessary to provide a better understanding of their pathogenic potential, to aid in making clinical decisions, and to conduct epidemiologic investigations,especially when large blind samples must be analyzed. It is useful to simultaneously identify species in different genera using a single primer pair. We developed a primer pair based on the tuf gene (encoding elongation factor) sequence to identify 56 Gram-positive cocci isolates. The target sequences were amplified from all 56 samples. The sequencing results and the phylogenetic tree derived from the partial tuf gene sequences identified the isolates as three enterococcal species, two lactococcal species, two staphylococcal species, and six streptococcal species, as well as eight isolates that were novel species of the genus Streptococcus. Partial gene sequence analysis of the sodA, dnaK, and 16S RNA genes confirmed the results obtained by tuf gene sequencing. Based on the uniform amplification of the tuf gene from all samples and the ability to identify all isolates at both the genus and species levels, we conclude that the primer pair developed in this research provides a powerful tool for identifying these organisms in clinical laboratories where large blind samples are used.
|
57. |
( 1998 ) Identification of streptococci to species level by sequencing the gene encoding the manganese-dependent superoxide dismutase. PMID : 9431917 : PMC : PMC124804 Abstract >>
We have used a PCR assay based on the use of degenerate primers in order to characterize an internal fragment (sodA(int)) representing approximately 85% of the genes encoding the manganese-dependent superoxide dismutase in various streptococcal type strains (S. acidominimus, S. agalactiae, S. alactolyticus, S. anginosus, S. bovis, S. constellatus, S. canis, S. cricetus, S. downei, S. dysgalactiae, S. equi subsp. equi, S. equi subsp. zooepidemicus, S. equinus, S. gordonii, S. iniae, S. intermedius, S. mitis, S. mutans, S. oralis, S. parasanguis, S. pneumoniae, S. porcinus, S. pyogenes, S. salivarius, S. sanguis, S. sobrinus, S. suis, S. thermophilus, and S. vestibularis). Phylogenetic analysis of these sodA(int) fragments yields an evolutionary tree having a topology similar to that of the tree constructed with the 16S rRNA sequences. We have shown that clinical isolates could be identified by determining the positions of their sodA(int) fragments on the phylogenetic tree of the sodA(int) fragments of the type species. We propose this method for the characterization of strains that cannot be assigned to a species on the basis of their conventional phenotypic reactions.
|
331, Shih-Pin Rd., Hsinchu 30062, Taiwan
Phone: +886-3-5223191
E-mail: bcrcweb@firdi.org.tw
web maintainance: +886-3-5223191 ext 593
Copyright © 2018.BCRC All rights reserved.The duplication or use of information and data such as texts or images or any linkage the website at the "bcrc.firdi.org.tw" is only permitted with the indication of the source or with prior approval by the BCRC(Bioresource Collection and Research Center).