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Antibiotics (Basel, Switzerland) Jun 2023The subgroup of viridans group streptococci are important human pathogens. We previously showed that a substantial portion of strains (>25%) are 'destined' to develop...
The subgroup of viridans group streptococci are important human pathogens. We previously showed that a substantial portion of strains (>25%) are 'destined' to develop rapid, high-level, and stable daptomycin (DAP) resistance (DAP-R) during DAP exposures in vitro. Such DAP-R is often accompanied by perturbations in distinct membrane phenotypes and metabolic pathways. The current study evaluated two bloodstream isolates, 73 and 205. Strain 73 developed stable, high-level DAP-R (minimum inhibitory concentration [MIC] > 256 µg/mL) within 2 days of in vitro DAP passage ("high level" DAP-R [HLDR]). In contrast, strain 205 evolved low-level and unstable DAP-R (MIC = 8 µg/mL) under the same exposure conditions in vitro ("non-HLDR"). Comparing the parental 73 vs. 73-D2 (HLDR) strain-pair, we observed the 73-D2 had the following major differences: (i) altered cell membrane (CM) phospholipid profiles, featuring the disappearance of phosphatidylglycerol (PG) and cardiolipin (CL), with accumulation of the PG-CL pathway precursor, phosphatidic acid (PA); (ii) enhanced CM fluidity; (iii) increased DAP surface binding; (iv) reduced growth rates; (v) decreased glucose utilization and lactate accumulation; and (vi) increased enzymatic activity within the glycolytic (i.e., lactate dehydrogenase [LDH]) and lipid biosynthetic (glycerol-3-phosphate dehydrogenase [GPDH]) pathways. In contrast, the 205 (non-HLDR) strain-pair did not show these same phenotypic or metabolic changes over the 2-day DAP exposure. WGS analyses confirmed the presence of mutations in genes involved in the above glycolytic and phospholipid biosynthetic pathways in the 73-D2 passage variant. These data suggest that strains which are 'destined' to rapidly develop HLDR do so via a conserved cadre of genotypic, membrane phenotypic, and metabolic adaptations.
PubMed: 37508179
DOI: 10.3390/antibiotics12071083 -
Journal of Oral Microbiology 2017is a new member of the Mitis group and is associated with infective endocarditis. Low and high virulent variants have been described. A search was made in the national...
is a new member of the Mitis group and is associated with infective endocarditis. Low and high virulent variants have been described. A search was made in the national reference collection of endocarditis isolates for -like strains by sequencing housekeeping genes (16S rRNA-gene, ). The strains were further profiled by polymerase chain reaction (PCR) targeting a choice of virulence genes (-like, -like, ). To study the prevalence and abundance of in the saliva and on the mucosal membranes of 35 healthy adults, PCRs detecting two variants of the 16S operon and virulence genes were applied. Among the endocarditis isolates, eight strains (all -negative and former ) holding the specific 16S motif were found, but the pattern of genes related to high virulence found in the type strain could not be detected in any of these strains. A close phylogenetic proximity between and was observed, with intersectional hybrid strains formed. This was supported by concatenated housekeeping sequences, DNA-DNA hybridization, pathogenomic profiling, and multidimensional scaling. In the oral samples, could be detected frequently, especially in the most common operon variant, but none of the type strain-related virulence factors were found. Low virulent -like strains can be found frequently and in high prevalence (66%) and abundance (12.5%) in the oral cavity of healthy adults. In strain collections, they are among the formerly known -negative . Highly virulent strains seem to be uncommon. Though closely related, and can be separated by the presence or absence of and dextran production. Hybrids of both species can be found. The variable arsenal of virulence genes found in this study emphasizes the genetic plasticity of Mitis group streptococci.
PubMed: 28473881
DOI: 10.1080/20002297.2017.1307079 -
Antibiotics (Basel, Switzerland) Apr 2021infections are increasingly prevalent in specific populations, including neutropenic cancer and endocarditis patients. strains have a propensity to evolve rapid,...
infections are increasingly prevalent in specific populations, including neutropenic cancer and endocarditis patients. strains have a propensity to evolve rapid, high-level and durable resistance to daptomycin (DAP-R) in vitro and in vivo, although the mechanism(s) involved remain incompletely defined. We examined mechanisms of DAP-R versus cross-resistance to cationic host defense peptides (HDPs), using an isogenic strain-pair: (i) DAP-susceptible (DAP-S) parental 351-WT (DAP MIC = 0.5 µg/mL), and its (ii) DAP-R variant 351-D10 (DAP MIC > 256 µg/mL). DAP binding was quantified by flow cytometry, in-parallel with temporal (1-4 h) killing by either DAP or comparative prototypic cationic HDPs (hNP-1; LL-37). Multicolor flow cytometry was used to determine kinetic cell responses associated with resistance or susceptibility to these molecules. While overall DAP binding was similar between strains, a significant subpopulation of 351-D10 cells hyper-accumulated DAP (>2-4-fold vs. 351-WT). Further, both DAP and hNP-1 induced cell membrane (CM) hyper-polarization in 351-WT, corresponding to significantly greater temporal DAP-killing (vs. 351-D10). No strain-specific differences in CM permeabilization, lipid turnover or regulated cell death were observed post-exposure to DAP, hNP-1 or LL-37. Thus, the adaptive energetics of the CM appear coupled to the outcomes of interactions of S. with DAP and selected HDPs. In contrast, altered CM permeabilization, proposed as a major mechanism of action of both DAP and HDPs, did not differentiate DAP-S vs. DAP-R phenotypes in this strain-pair.
PubMed: 33918000
DOI: 10.3390/antibiotics10040404 -
Frontiers in Cellular and Infection... 2019subspecies is explored as an anti-cariogenic probiotic. Here, subjecting freshly stimulated saliva samples of 35 healthy volunteers, six epidemiologically unrelated...
subspecies is explored as an anti-cariogenic probiotic. Here, subjecting freshly stimulated saliva samples of 35 healthy volunteers, six epidemiologically unrelated and two related strains were isolated (prevalence around 20%) applying a newly developed three-step procedure. Furthermore, the probiotic strain 7746 (AB-Dentisanium®) was tested under a variety of environmental conditions for its inhibitory effect on six , two , 15 other oral or intestinal streptococci, 15 strains, and six representatives of other species including periodontopathogens. All except one of the strains were inhibited by 7746 colonies or culture supernatant concentrate but only if either the test cell number was low or the producer or its bacteriocin concentration, respectively, was high. OMI 332, OMI 315, OMI 335, OMI 238, and the intestinal OMI 339 were not inhibited, while the other 10 streptococcal strains (especially OMI 334 and intestinal OMI 326) showed a certain degree of inhibition. From the panel of other bacterial species only was slightly inhibited. With the exception of OMI 285 and OMI 291 that possessed a 7746 bacteriocin-like gene cluster, all strains and especially type strain 7747 were strongly inhibited by 7746. In conclusion, probiotic strain 7746 might antagonize the initiation and progression of dental caries by reducing if not too abundant. strains inhibit each other, but strains with similar bacteriocin-related gene clusters, including immunity genes, are able to co-exist due to cross-resistance. In addition, development of resistance and adaptation to 7746-bacteriocins was observed during our study and needs attention. Hence, mechanisms underlying such processes need to be further investigated using omics-approaches. On the manufacturing level, probiotic strains should be continuously tested for function. Further clinical studies investigating inhibition of by AB-Dentisanium® are required that should also monitor the impact on the oral microbiome composition including resident strains.
Topics: Aggregatibacter actinomycetemcomitans; Anti-Bacterial Agents; Antibiosis; Bacterial Typing Techniques; Bacteriocins; Carrier State; Healthy Volunteers; Prevalence; Probiotics; Streptococcal Infections; Streptococcus mutans; Streptococcus oralis; Streptococcus sobrinus
PubMed: 31041198
DOI: 10.3389/fcimb.2019.00110 -
Cells May 2020Human gingival epithelial cells (HGEps) and fibroblasts (HGFs) are the main cell types in peri-implant soft tissue. HGEps are constantly exposed to bacteria, but HGFs...
Human gingival epithelial cells (HGEps) and fibroblasts (HGFs) are the main cell types in peri-implant soft tissue. HGEps are constantly exposed to bacteria, but HGFs are protected by connective tissue as long as the mucosa-implant seal is intact. is one of the commensal bacteria, is highly abundant at healthy implant sites, and might modulate soft tissue cells-as has been described for other streptococci. We have therefore investigated the effects of the biofilm on HGEps and HGFs. HGEps or HGFs were grown separately on titanium disks and responded to challenge with biofilm. HGFs were severely damaged after 4 h, exhibiting transcriptional inflammatory and stress responses. In contrast, challenge with only induced a mild transcriptional inflammatory response in HGEps, without cellular damage. HGFs were more susceptible to the biofilm than HGEps. The pro-inflammatory interleukin 6 (IL-6) was attenuated in HGFs, as was interleukin 8 (CXCL8) in HGEps. This indicates that can actively protect tissue. In conclusion, commensal biofilms can promote homeostatic tissue protection, but only if the implant-mucosa interface is intact and HGFs are not directly exposed.
Topics: Biofilms; Cell Adhesion; Cell Shape; Cell Survival; Cytokines; Epithelial Cells; Fibroblasts; Gingiva; Humans; Inflammation Mediators; Prostheses and Implants; Streptococcus oralis; Transcription, Genetic; Up-Regulation
PubMed: 32429151
DOI: 10.3390/cells9051226 -
Bioengineered Dec 2021The correlation between oral bacteria and dental implants failure has been reported. However, the effect and mechanism of bacteria during dental implants is unclear. In...
The correlation between oral bacteria and dental implants failure has been reported. However, the effect and mechanism of bacteria during dental implants is unclear. In this study, we explored key genes and candidate gene clusters in human gingival fibroblasts (HGF) cells in response to biofilm through weighted gene co-expression network analysis (WGCNA) and differential genes analysis using gene expression matrix, GSE134481, downloaded from the Gene Expression Omnibus (GEO) database. We obtained 325 genes in the module significantly associated with infection and 113 differentially expressed genes (DEGs) in the biofilm; 62 DEGs indicated significant correlation with injury. Multiple immune pathways, such as the tumor necrosis factor (TNF) signaling pathway, were considerably enriched. We obtained a candidate genes cluster containing 12 genes - , and ; we observed 5 candidate hub genes associated with infection - , and . The fraction of macrophage M0 cells was significantly increased in biofilm treatment compared with control; expression of and was significantly positively correlated with macrophage M0 cells. Our findings present a fierce inflammation changes in the transcript level of HGF in response to .
Topics: Biofilms; Fibroblasts; Gene Expression Regulation; Gene Regulatory Networks; Gingiva; Humans; Macrophages; Protein Interaction Maps; Streptococcus oralis
PubMed: 33781179
DOI: 10.1080/21655979.2021.1902697 -
Antimicrobial Agents and Chemotherapy May 2017Synthesis and integrity of the cytoplasmic membrane are fundamental to cellular life. Experimental evolution studies have hinted at unique physiology in the...
Synthesis and integrity of the cytoplasmic membrane are fundamental to cellular life. Experimental evolution studies have hinted at unique physiology in the Gram-positive bacteria and These organisms commonly cause bacteremia and infectious endocarditis (IE) but are rarely investigated in mechanistic studies of physiology and evolution. Unlike in other Gram-positive pathogens, high-level (MIC ≥ 256 μg/ml) daptomycin resistance rapidly emerges in and after a single drug exposure. In this study, we found that inactivating mutations in are associated with high-level daptomycin resistance in and IE isolates. This is surprising given that is an essential gene for life in commonly studied model organisms. CdsA is the enzyme responsible for the synthesis of CDP-diacylglycerol, a key intermediate for the biosynthesis of all major phospholipids in prokaryotes and most anionic phospholipids in eukaryotes. Lipidomic analysis by liquid chromatography-mass spectrometry (LC-MS) showed that daptomycin-resistant strains have an accumulation of phosphatidic acid and completely lack phosphatidylglycerol and cardiolipin, two major anionic phospholipids in wild-type strains, confirming the loss of function of CdsA in the daptomycin-resistant strains. To our knowledge, these daptomycin-resistant streptococci represent the first model organisms whose viability is CdsA independent. The distinct membrane compositions resulting from the inactivation of not only provide novel insights into the mechanisms of daptomycin resistance but also offer unique opportunities to study the physiological functions of major anionic phospholipids in bacteria.
Topics: Anti-Bacterial Agents; Cardiolipins; Cytidine Diphosphate Diglycerides; Daptomycin; Drug Resistance, Bacterial; Humans; Membrane Lipids; Microbial Sensitivity Tests; Nucleotidyltransferases; Phosphatidic Acids; Phosphatidylglycerols; Phospholipids; Streptococcal Infections; Streptococcus mitis; Streptococcus oralis
PubMed: 28223392
DOI: 10.1128/AAC.02552-16 -
BMC Microbiology Feb 2021In caries, low pH drives selection and enrichment of acidogenic and aciduric bacteria in oral biofilms, and development of acid tolerance in early colonizers is thought...
BACKGROUND
In caries, low pH drives selection and enrichment of acidogenic and aciduric bacteria in oral biofilms, and development of acid tolerance in early colonizers is thought to play a key role in this shift. Since previous studies have focussed on planktonic cells, the effect of biofilm growth as well as the role of a salivary pellicle on this process is largely unknown. We explored acid tolerance and acid tolerance response (ATR) induction in biofilm cells of both clinical and laboratory strains of three oral streptococcal species (Streptococcus gordonii, Streptococcus oralis and Streptococcus mutans) as well as two oral species of Actinomyces (A. naeslundii and A. odontolyticus) and examined the role of salivary proteins in acid tolerance development.
METHODS
Biofilms were formed on surfaces in Ibidi® mini flow cells with or without a coating of salivary proteins and acid tolerance assessed by exposing them to a challenge known to kill non-acid tolerant cells (pH 3.5 for 30 min) followed by staining with LIVE/DEAD BacLight and confocal scanning laser microscopy. The ability to induce an ATR was assessed by exposing the biofilms to an adaptation pH (pH 5.5) for 2 hours prior to the low pH challenge.
RESULTS
Biofilm formation significantly increased acid tolerance in all the clinical streptococcal strains (P < 0.05) whereas the laboratory strains varied in their response. In biofilms, S. oralis was much more acid tolerant than S. gordonii or S. mutans. A. naeslundii showed a significant increase in acid tolerance in biofilms compared to planktonic cells (P < 0.001) which was not seen for A. odontolyticus. All strains except S. oralis induced an ATR after pre-exposure to pH 5.5 (P < 0.05). The presence of a salivary pellicle enhanced both acid tolerance development and ATR induction in S. gordonii biofilms (P < 0.05) but did not affect the other bacteria to the same extent.
CONCLUSIONS
These findings suggest that factors such as surface contact, the presence of a salivary pellicle and sensing of environmental pH can contribute to the development of high levels of acid tolerance amongst early colonizers in oral biofilms which may be important in the initiation of caries.
Topics: Acids; Adaptation, Physiological; Biofilms; Humans; Hydrogen-Ion Concentration; Mouth; Salivary Proteins and Peptides; Streptococcal Infections; Streptococcus
PubMed: 33583397
DOI: 10.1186/s12866-021-02089-2 -
Frontiers in Oral Health 2023While oral mirobial dysbiosis due to tobacco smoking has been studied thoroughly, there is limited data on the effect of waterpipe smoking on the oral microbiome. This...
BACKGROUND
While oral mirobial dysbiosis due to tobacco smoking has been studied thoroughly, there is limited data on the effect of waterpipe smoking on the oral microbiome. This study aims to compare the salivary microbiome between waterpipe smokers and non-smokers.
MATERIALS AND METHODS
Unstimulated saliva samples were collected from 60 participants, 30 smokers and 30 non-smokers in Kuala Lumpur and Klang Valley, Malaysia. DNA extraction was performed using the Qiagen DNA mini kit, and the 16S rRNA bacterial gene was amplified and sequenced using the Illumina MiSeq platform. Sequencing reads were processed using DADA2, and the alpha and beta diversity of the bacterial community was assessed. Significantly differentiated taxa were identified using LEfSe analysis, while differentially expressed pathways were identified using MaAsLin2.
RESULTS
A significant compositional change (beta diversity) was detected between the two groups (PERMANOVA < 0.05). Specifically, the levels of phylum Firmicutes and genus were elevated in smokers, whereas phylum Proteobacteria and genus were depleted compared to non-smokers. At the species level, and were enriched in smokers. We observed significant differences in the abundance of thirty-seven microbial metabolic pathways between waterpipe smokers and non-smokers. The microbial pathways enriched in smokers were those implicated in polymer degradation and amino acid metabolism.
CONCLUSION
The taxonomic and metabolic profile of the salivary microbiome in waterpipe smokers compared to healthy controls exhibited a paradigm shift, thus, implying an alteration in the homeostatic balance of the oral cavity posing unique challenges for oral health.
PubMed: 38024144
DOI: 10.3389/froh.2023.1275717 -
Frontiers in Microbiology 2018is a highly impactful bacterial pathogen on a global scale. The principal pneumococcal virulence factor and target of effective vaccines is its polysaccharide capsule,...
is a highly impactful bacterial pathogen on a global scale. The principal pneumococcal virulence factor and target of effective vaccines is its polysaccharide capsule, of which there are many structurally distinct forms. Here, we describe four distinct strains of three Mitis group commensal species (, , and ) recovered from upper respiratory tract specimens from adults in Kenya and the United States that were PCR-positive for the pneumococcal serotype 5 specific gene, . For each of the four strains, the 15 genes comprising the capsular polysaccharide biosynthetic gene cluster () shared the same order found in serotype 5 pneumococci, and each of the serotype 5-specific genes from the serotype 5 pneumococcal reference strain shared 76-99% sequence identity with the non-pneumococcal counterparts. Double-diffusion experiments demonstrated specific reactivity of the non-pneumococcal strains with pneumococcal serotype 5 typing sera. Antiserum raised against strain KE67013 specifically reacted with serotype 5 pneumococci for a positive Quellung reaction and stimulated serotype 5 specific opsonophagocytic killing of pneumococci. Four additional commensal strains, identified using PCR serotyping assays on pharyngeal specimens, revealed loci highly homologous to those of pneumococci of serotypes 12F, 15A, 18C, and 33F. These data, in particular the species and strain diversity shown for serotype 5, highlight the existence of a broad non-pneumococcal species reservoir in the upper respiratory tract for the expression of capsular polysaccharides that are structurally related or identical to those corresponding to epidemiologically significant serotypes. Very little is known about the genetic and antigenic capsular diversity among the vast array of commensal streptococcal strains that represent multiple diverse species. The discovery of serotype 5 strains within three different commensal species suggests that extensive capsular serologic overlap exists between pneumococci and other members of the diverse Mitis group. These findings may have implications for our current understanding of naturally acquired immunity to and pneumococcal serotype distributions in different global regions. Further characterization of commensal strains carrying homologs of serotype-specific genes previously thought to be specific for pneumococci of known serotypes may shed light on the evolution of these important loci.
PubMed: 30671034
DOI: 10.3389/fmicb.2018.03199