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904400.9904Impairment of novel non-coding small RNA00203 inhibits biofilm formation and reduces biofilm-specific antibiotic resistance in Acinetobacter baumannii. Small RNAs (sRNAs) are post-transcriptional regulators of many biological processes in bacteria, including biofilm formation and antibiotic resistance. The mechanisms by which sRNA regulates the biofilm-specific antibiotic resistance in Acinetobacter baumannii have not been reported to date. This study aimed to investigate the influence of sRNA00203 (53 nucleotides) on biofilm formation, antibiotic susceptibility, and expression of genes associated with biofilm formation and antibiotic resistance. The results showed that deletion of the sRNA00203-encoding gene decreased the biomass of biofilm by 85%. Deletion of the sRNA00203-encoding gene also reduced the minimum biofilm inhibitory concentrations for imipenem and ciprofloxacin 1024- and 128-fold, respectively. Knocking out of sRNA00203 significantly downregulated genes involved in biofilm matrix synthesis (pgaB), efflux pump production (novel00738), lipopolysaccharide biosynthesis (novel00626), preprotein translocase subunit (secA) and the CRP transcriptional regulator. Overall, the suppression of sRNA00203 in an A. baumannii ST1894 strain impaired biofilm formation and sensitized the biofilm cells to imipenem and ciprofloxacin. As sRNA00203 was found to be conserved in A. baumannii, a therapeutic strategy targeting sRNA00203 may be a potential solution for the treatment of biofilm-associated infections caused by A. baumannii. To the best of the authors' knowledge, this is the first study to show the impact of sRNA00203 on biofilm formation and biofilm-specific antibiotic resistance in A. baumannii.202337315907
248810.9903Antibiotic resistance, putative virulence factors and curli fimbrination among Cronobacter species. This study aimed to investigate antibiotic resistance and putative virulence factors among Cronobacter sakazakii isolated from powdered infant formula and other sources. The following 9 cultures (CR1-9) were collected from our culture collection: C. sakazakii and 3 Cronobacter species: C. sakazakii ATCC® 29544™, C. muytjensii ATCC® 51329™, C. turicensis E866 were used in this study. Isolates were subjected to antibiotic susceptibility and the following virulence factors (protease, DNase, haemolysin, gelatinase, motility and biofilm formation) using phenotypic methods. All the bacteria were able to form biofilm on agar at 37 °C and were resistant to ampicillin, erythromycin, fosfomycin and sulphamethoxazole. It was observed from this study that tested strains formed weak and strong biofilm with violet dry and rough (rdar), brown dry and rough (bdar), red mucoid and smooth (rmas) colony morphotypes on Congo red agar. Rdar expresses curli and fimbriae, while bdar expresses curli. Both biofilm colony morphotypes are commonly found in Enterobacteriaceae including Salmonella species. This study also reveals a new colony morphotypes in Cronobacter species. Conclusively, there was correlation between putative virulence factors and antibiotic resistance among the tested bacteria. Further study on virulence and antibiotic resistance genes is hereby encouraged.201931404630
880220.9897The Transcription Factor CsgD Contributes to Engineered Escherichia coli Resistance by Regulating Biofilm Formation and Stress Responses. The high cell density, immobilization and stability of biofilms are ideal characteristics for bacteria in resisting antibiotic therapy. CsgD is a transcription activating factor that regulates the synthesis of curly fimbriae and cellulose in Escherichia coli, thereby enhancing bacterial adhesion and promoting biofilm formation. To investigate the role of CsgD in biofilm formation and stress resistance in bacteria, the csgD deletion mutant ΔcsgD was successfully constructed from the engineered strain E. coli BL21(DE3) using the CRISPR/Cas9 gene-editing system. The results demonstrated that the biofilm of ΔcsgD decreased by 70.07% (p < 0.05). Additionally, the mobility and adhesion of ΔcsgD were inhibited due to the decrease in curly fimbriae and extracellular polymeric substances. Furthermore, ΔcsgD exhibited a significantly decreased resistance to acid, alkali and osmotic stress conditions (p < 0.05). RNA-Seq results revealed 491 differentially expressed genes between the parent strain and ΔcsgD, with enrichment primarily observed in metabolism-related processes as well as cell membrane structure and catalytic activity categories. Moreover, CsgD influenced the expression of biofilm and stress response genes pgaA, motB, fimA, fimC, iraP, ompA, osmC, sufE and elaB, indicating that the CsgD participated in the resistance of E. coli by regulating the expression of biofilm and stress response. In brief, the transcription factor CsgD plays a key role in the stress resistance of E. coli, and is a potential target for treating and controlling biofilm.202337761984
636430.9897Characterization of clumpy adhesion of Escherichia coli to human cells and associated factors influencing antibiotic sensitivity. Escherichia coli intestinal infection pathotypes are characterized by distinct adhesion patterns, including the recently described clumpy adhesion phenotype. Here, we identify and characterize the genetic factors contributing to the clumpy adhesion of E. coli strain 4972. In this strain, the transcriptome and proteome of adhered bacteria were found to be distinct from planktonic bacteria in the supernatant. A total of 622 genes in the transcriptome were differentially expressed in bacteria present in clumps relative to the planktonic bacteria. Seven genes targeted for disruption had variable distribution in different pathotypes and nonpathogenic E. coli, with the pilV and spnT genes being the least frequent or absent from most groups. Deletion (Δ) of five differentially expressed genes, flgH, ffp, pilV, spnT, and yggT, affected motility, adhesion, or antibiotic stress. ΔflgH exhibited 80% decrease and ΔyggT depicted 184% increase in adhesion, and upon complementation, adhesion was significantly reduced to 13%. ΔflgH lost motility and was regenerated when complemented, whereas Δffp had significantly increased motility, and reintroduction of the same gene reduced it to the wild-type level. The clumps produced by Δffp and ΔspnT were more resistant and protected the bacteria, with ΔspnT showing the best clump formation in terms of ampicillin stress protection. ΔyggT had the lowest tolerance to gentamicin, where the antibiotic stress completely eliminated the bacteria. Overall, we were able to investigate the influence of clump formation on cell surface adhesion and antimicrobial tolerance, with the contribution of several factors crucial to clump formation on susceptibility to the selected antibiotics. IMPORTANCE: The study explores a biofilm-like clumpy adhesion phenotype in Escherichia coli, along with various factors and implications for antibiotic susceptibility. The phenotype permitted the bacteria to survive the onslaught of high antibiotic concentrations. Profiles of the transcriptome and proteome allowed the differentiation between adhered bacteria in clumps and planktonic bacteria in the supernatant. The deletion mutants of genes differentially expressed between adhered and planktonic bacteria, i.e., flgH, ffp, pilV, spnT, and yggT, and respective complementations in trans cemented their roles in multiple capacities. ffp, an uncharacterized gene, is involved in motility and resistance to ampicillin in a clumpy state. The work also affirms for the first time the role of the yggT gene in adhesion and its involvement in susceptibility against another aminoglycoside antibiotic, i.e., gentamicin. Overall, the study contributes to the mechanisms of biofilm-like adhesion phenotype and understanding of the antimicrobial therapy failures and infections of E. coli.202438530058
906440.9897Bacillus subtilis var. natto increases the resistance of Caenorhabditis elegans to gram-positive bacteria. AIMS: This study aimed to investigate the effect of Bacillus subtilis var. natto on the susceptibility of the model host, Caenorhabditis elegans, to bacterial infection. METHODS AND RESULTS: Caenorhabditis elegans worms were fed with a standard food consisting of Escherichia coli OP50 strain (control) or B. subtilis (natto) during their larval stage. The worms were then infected with pathogenic bacteria. We analyzed their survival time and RNA sequencing-based transcriptome. Upon infection with Staphylococcus aureus and Enterococcus faecalis, the survival time of B. subtilis (natto)-fed worms was longer than that of the control. Transcriptome analyses showed upregulation of genes associated with innate immunity and defense response to gram-positive bacteria in B. subtilis (natto)-fed worms. CONCLUSIONS: Bacillus subtilis (natto) conferred an increased resistance of C. elegans to gram-positive bacteria. Our findings provided insights into the molecular mechanisms underlying B. subtilis (natto)-regulated host immunity and emphasized its probiotic properties for preventing and alleviating infections caused by gram-positive bacteria. SIGNIFICANCE AND IMPACT OF THE STUDY: To the best of our knowledge, this is the first study to show that B. subtilis (natto) confers specific resistance against gram-positive bacteria.202134157196
619650.9897New insights into the regulatory pathways associated with the activation of the stringent response in bacterial resistance to the PBP2-targeted antibiotics, mecillinam and OP0595/RG6080. OBJECTIVES: The diazabicyclooctane β-lactamase inhibitor OP0595 (RG6080) also acts as an antibiotic, targeting PBP2 in Enterobacteriaceae, but this activity is vulnerable to mutational resistance. We used WGS to investigate the basis of this resistance. METHODS: Twenty OP0595-selected mutants, comprising four derived from each of five different Escherichia coli strains, were sequenced on Illumina HiSeq. Reads from each mutant were mapped to the assembled genome of the corresponding parent. A variant-calling file generated with Samtools was parsed to determine genetic alterations. RESULTS: Besides OP0595, the mutants consistently showed decreased susceptibility to mecillinam, which likewise targets PBP2, and grew as stable round forms in the presence of subinhibitory concentrations of OP0595. Among the 20 mutants, 18 had alterations in genes encoding tRNA synthase and modification functions liable to induce expression of the RpoS sigma factor through activation of the stringent response or had mutations suppressing inactivators of RpoS or the stringent response signal-degrading enzyme, SpoT. TolB was inactivated in one mutant: this activates RcsBC regulation and was previously associated with mecillinam resistance. The mechanism of resistance remained unidentified in one mutant. Both the RpoS and RcsBC systems regulate genes of cell division, including ftsAQZ that can compensate for loss or inhibition of PBP2, allowing survival of the challenged bacteria as stable round forms, as seen. CONCLUSIONS: WGS identified the global stringent response signal, entailing induction of RpoS, as the main mediator of mutational resistance to OP0595 in E. coli.201627330062
901860.9896Transcriptome analysis of heat resistance regulated by quorum sensing system in Glaesserella parasuis. The ability of bacteria to resist heat shock allows them to adapt to different environments. In addition, heat shock resistance is known for their virulence. Our previous study showed that the AI-2/luxS quorum sensing system affects the growth characteristics, biofilm formation, and virulence of Glaesserella parasuis. The resistance of quorum sensing system deficient G. parasuis to heat shock was obviously weaker than that of wild type strain. However, the regulatory mechanism of this phenotype remains unclear. To illustrate the regulatory mechanism by which the quorum sensing system provides resistance to heat shock, the transcriptomes of wild type (GPS2), ΔluxS, and luxS complemented (C-luxS) strains were analyzed. Four hundred forty-four differentially expressed genes were identified in quorum sensing system deficient G. parasuis, which participated in multiple regulatory pathways. Furthermore, we found that G. parasuis regulates the expression of rseA, rpoE, rseB, degS, clpP, and htrA genes to resist heat shock via the quorum sensing system. We further confirmed that rseA and rpoE genes exerted an opposite regulatory effect on heat shock resistance. In conclusion, the findings of this study provide a novel insight into how the quorum sensing system affects the transcriptome of G. parasuis and regulates its heat shock resistance property.202236033895
844270.9896Staphylococcus epidermidis undergoes global changes in gene expression during biofilm maturation in platelet concentrates. BACKGROUND: Staphylococcus epidermidis forms surface-attached aggregates (biofilms) when grown in platelet concentrates (PCs). Comparative transcriptome analyses were undertaken to investigate differential gene expression of S. epidermidis biofilms grown in PCs. STUDY DESIGN AND METHODS: Two S. epidermidis strains isolated from human skin (AZ22 and AZ39) and one strain isolated from contaminated PCs (ST02) were grown in glucose-supplemented Trypticase Soy Broth (TSBg) and PCs. RNA was extracted and sequenced using Illumina HiSeq. Differential expression analysis was done using DESeq, and significantly differentially expressed genes (DEGs) were selected. DEGs were subjected to Kyoto encyclopedia of genes and genomes and Gene Ontology analyses. Differential gene expression was validated with quantitative reverse transcription-PCR. RESULTS: A total of 436, 442, and 384 genes were expressed in AZ22, AZ39, and ST02, respectively. DEG analysis showed that 170, 172, and 117 genes were upregulated in PCs in comparison to TSBg, whereas 120, 135, and 89 genes were downregulated (p < .05) in mature biofilms of AZ22, AZ39, and ST02, respectively. Twenty-seven DEGs were shared by all three strains. While 76 DEGs were shared by AZ22 and AZ39, only 34 and 21 DEGs were common between ST02, and AZ22 and AZ39, respectively. Significant transcriptional expression changes were observed in genes involved in platelet-bacteria interaction, biofilm formation, production of virulence factors, and resistance to antimicrobial peptides and antibiotics. CONCLUSION: Differential gene expression in S. epidermidis is triggered by the stressful PC storage environment. Upregulation of virulence and antimicrobial resistance genes could have clinical implications for transfusion patients.202133904608
872180.9894Chromium metabolism characteristics of coexpression of ChrA and ChrT gene. OBJECTIVE: Serratia sp. S2 is a wild strain with chromium resistance and reduction ability. Chromium(VI) metabolic-protein-coding gene ChrA and ChrT were cloned from Serratia sp. S2, and ligated with prokaryotic expression vectors pET-28a (+) and transformed into E. coli BL21 to construct ChrA, ChrT and ChrAT engineered bacteria. By studying the characteristics of Cr(VI) metabolism in engineered bacteria, the function and mechanism of the sole expression and coexpression of ChrA and ChrT genes were studied. METHODS: Using Serratia sp. S2 genome as template, ChrA and ChrT genes were amplified by PCR, and prokaryotic expression vectors was ligated to form the recombinant plasmid pET-28a (+)-ChrA, pET-28a (+)-ChrT and pET-28a (+)-ChrAT, and transformed into E. coli BL21 to construct ChrA, ChrT, ChrAT engineered bacteria. The growth curve, tolerance, and reduction of Cr(VI), the distribution of intracellular and extracellular Cr, activity of chromium reductase and intracellular oxidative stress in engineered bacteria were measured to explore the metabolic characteristics of Cr(VI) in ChrA, ChrT, ChrAT engineered bacteria. RESULTS: ChrA, ChrT and ChrAT engineered bacteria were successfully constructed by gene recombination technology. The tolerance to Cr(VI) was Serratia sp. S2 > ChrAT ≈ ChrA > ChrT > Control (P < 0.05), and the reduction ability to Cr(VI) was Serratia sp. S2 > ChrAT ≈ ChrT > ChrA (P < 0.05). The chromium distribution experiments confirmed that Cr(VI) and Cr(III) were the main valence states. Effect of electron donors on chromium reductase activity was NADPH > NADH > non-NAD(P)H (P < 0.05). The activity of chromium reductase increased significantly with NAD(P)H (P < 0.05). The Glutathione and NPSH (Non-protein Sulfhydryl) levels of ChrA, ChrAT engineered bacteria increased significantly (P < 0.05) under the condition of Cr(VI), but there was no significant difference in the indexes of ChrT engineered bacteria (P > 0.05). CONCLUSION: ChrAT engineered bacteria possesses resistance and reduction abilities of Cr(VI). ChrA protein endows the strain with the ability to resist Cr(VI). ChrT protein reduces Cr(VI) to Cr(III) by using NAD(P)H as electronic donor. The reduction process promotes the production of GSH, GSSG and NPSH to maintain the intracellular reduction state, which further improves the Cr(VI) tolerance and reduction ability of ChrAT engineered bacteria.202032768747
904090.9893Gene expression changes linked to antimicrobial resistance, oxidative stress, iron depletion and retained motility are observed when Burkholderia cenocepacia grows in cystic fibrosis sputum. BACKGROUND: Bacteria from the Burkholderia cepacia complex (Bcc) are the only group of cystic fibrosis (CF) respiratory pathogens that may cause death by an invasive infection known as cepacia syndrome. Their large genome (> 7000 genes) and multiple pathways encoding the same putative functions make virulence factor identification difficult in these bacteria. METHODS: A novel microarray was designed to the genome of Burkholderia cenocepacia J2315 and transcriptomics used to identify genes that were differentially regulated when the pathogen was grown in a CF sputum-based infection model. Sputum samples from CF individuals infected with the same B. cenocepacia strain as genome isolate were used, hence, other than a dilution into a minimal growth medium (used as the control condition), no further treatment of the sputum was carried out. RESULTS: A total of 723 coding sequences were significantly altered, with 287 upregulated and 436 downregulated; the microarray-observed expression was validated by quantitative PCR on five selected genes. B. cenocepacia genes with putative functions in antimicrobial resistance, iron uptake, protection against reactive oxygen and nitrogen species, secretion and motility were among the most altered in sputum. Novel upregulated genes included: a transmembrane ferric reductase (BCAL0270) implicated in iron metabolism, a novel protease (BCAL0849) that may play a role in host tissue destruction, an organic hydroperoxide resistance gene (BCAM2753), an oxidoreductase (BCAL1107) and a nitrite/sulfite reductase (BCAM1676) that may play roles in resistance to the host defenses. The assumptions of growth under iron-depletion and oxidative stress formulated from the microarray data were tested and confirmed by independent growth of B. cenocepacia under each respective environmental condition. CONCLUSION: Overall, our first full transcriptomic analysis of B. cenocepacia demonstrated the pathogen alters expression of over 10% of the 7176 genes within its genome when it grows in CF sputum. Novel genetic pathways involved in responses to antimicrobial resistance, oxidative stress, and iron metabolism were revealed by the microarray analysis. Virulence factors such as the cable pilus and Cenocepacia Pathogenicity Island were unaltered in expression. However, B. cenocepacia sustained or increased expression of motility-associated genes in sputum, maintaining a potentially invasive phenotype associated with cepacia syndrome.200818801206
8720100.9893Chromium resistance characteristics of Cr(VI) resistance genes ChrA and ChrB in Serratia sp. S2. OBJECTIVE: To find an efficient chromium (VI) resistance system, with a highly efficient, economical, safe, and environmentally friendly chromium-removing strain, ChrA, ChrB, and ChrAB fragments of the chromium (VI) resistance gene in Serratia sp. S2 were cloned, and their prokaryotic expression vectors were constructed and transformed into E. coli BL21. The anti-chromium (VI) capacity and characteristics of engineered bacteria, role of ChrA and ChrB genes in the anti-chromium (VI) processes, and the mechanism of chromium metabolism, were explored. METHODS: The PCR technique was used to amplify ChrA, ChrB, and ChrAB genes from the Serratia sp. S2 genome. ChrA, ChrB, and ChrAB genes were connected to the prokaryotic expression vector pET-28a and transferred into E. coli BL21 for prokaryotic expression. Cr-absorption and Cr-efflux ability of the engineered strains were determined. The effects of respiratory inhibitors and oxygenated anions on Cr-efflux of ChrA and ChrB engineered strains were explored. RESULTS: ChrA, ChrB, and ChrAB engineered strains were constructed successfully; there was no significant difference between the control strain and the ChrB engineered strain for Cr-metabolism (P > 0.05). Cr-absorption and Cr-efflux of ChrA and ChrAB engineered strains were significantly stronger than the control strain (P < 0.05). Oxyanions (sulfate and molybdate) and inhibitors (valinomycin and CN(-)) could significantly inhibit the Cr-efflux capacities of ChrA and ChrAB engineered strains (P < 0.05), while NADPH could significantly promote such capacities (P < 0.05). CONCLUSION: The Cr-transporter, encoded by ChrA gene, confer the ability to pump out intracellular Cr on ChrA and ChrAB engineered strains. The ChrB gene plays a positive regulatory role in ChrA gene regulation. The Cr-metabolism ability of the ChrAB engineered strain is stronger than the ChrA engineered strain. ChrA and ChrAB genes in the Cr-resistance system may involve a variety of mechanisms, such as sulfate ion channel and respiratory chain electron transfer.201829655157
578110.9893Characterization of radiation-resistance mechanism in Spirosoma montaniterrae DY10(T) in terms of transcriptional regulatory system. To respond to the external environmental changes for survival, bacteria regulates expression of a number of genes including transcription factors (TFs). To characterize complex biological phenomena, a biological system-level approach is necessary. Here we utilized six computational biology methods to infer regulatory network and to characterize underlying biologically mechanisms relevant to radiation-resistance. In particular, we inferred gene regulatory network (GRN) and operons of radiation-resistance bacterium Spirosoma montaniterrae DY10[Formula: see text] and identified the major regulators for radiation-resistance. Our results showed that DNA repair and reactive oxygen species (ROS) scavenging mechanisms are key processes and Crp/Fnr family transcriptional regulator works as a master regulatory TF in early response to radiation.202336959250
628120.9893Resistance to bismuth among gram-negative bacteria is dependent upon iron and its uptake. Bismuth antimicrobial action is poorly understood. Many trivalent metals possess antibacterial activity, especially under low iron conditions. Protection of bacteria from the deleterious effects of bismuth and other trivalent metals was demonstrated in iron-fortified media. Near-equimolar quantities of Fe3+ neutralized the growth-inhibitory effects of 250 microM Bi3+. Resistance to bismuth action also depended on the production of virulence-related siderophores. Escherichia coli, Aeromonas hydrophila or Pseudomonas aeruginosa producing aerobactin, amonabactin or pyoverdin respectively, were most resistant to Bi3+. Enterochelin or pyochelin producers were less resistant to Bi3+, but more resistant than strains lacking siderophores. Purified pyoverdin restored Bi3+ resistance in a mutant lacking this siderophore, but not in one lacking the pyoverdin receptor. Bismuth-treated bacteria exhibited unique outer membrane proteins, similar in size to iron-repressible proteins. Thus, resistance to the inhibitory action of Bi3+ among Gram-negative bacteria is inversely related to iron concentration and strongly dependent on iron transport mechanisms. The data suggest that bismuth action is largely a nonspecific, competitive interference with iron-transport, related primarily to atomic valence Furthermore, resistance to Bi3+ among bacteria is predictive of virulence.19969023650
8887130.9892RpoE is a Putative Antibiotic Resistance Regulator of Salmonella enteric Serovar Typhi. Bacterial antimicrobial resistance has been associated with the up regulation of genes encoding efflux pumps and the down regulation of genes encoding outer membrane proteins (OMPs). Gene expression in bacteria is primarily initiated by sigma factors (σ factors) such as RpoE, which plays an important role in responding to many environmental stresses. Here, we report the first observation that RpoE serves as an antibiotic resistance regulator in Salmonella enteric serovar Typhi (S. Typhi). In this study, we found that the rpoE mutant (ΔrpoE) of S. Typhi GIFU10007 has elevated resistance to several antimicrobial agents, including β-lactams, quinolones, and aminoglycosides. Genomic DNA microarray analysis was used to investigate the differential gene expression profiles between a wild type and rpoE mutant in response to ampicillin. The results showed that a total of 57 genes displayed differential expression (two-fold increase or decrease) in ΔrpoE versus the wild-type strain. The expressions of two outer membrane protein genes, ompF and ompC, were significantly down-regulated in ΔrpoE (six and seven-fold lower in comparison to wild-type strain) and RamA, a member of the efflux pump AraC/XylS family, was up-regulated about four-fold in the ΔrpoE. Our results suggest RpoE is a potential antimicrobial regulator in S. Typhi, controlling both the down regulation of the OMP genes and up-regulating the efflux system.201626742769
5181140.9892Differential Expression of fimH, ihf, upaB, and upaH Genes in Biofilms- and Suspension-Grown Bacteria From Samples of Different Uropathogenic Strains of Escherichia coli. Uropathogenic Escherichia coli (UPEC) strains are the main bacteria that cause urinary tract infections (UTIs). UPEC are a significant public health hazard due to their high proliferation, antibiotic resistance, and infection recurrence. The ability to form biofilms is a mechanism of antibiotic resistance, which requires the expression of different genes such as fimH, ihf, upaB, and upaH. Despite the relevance of biofilm formation in bacterial pathogenicity, differences in the expression level of these genes among bacterial growth conditions have been little studied. Here, we have characterized the expression of fimH, ihf, upaB, and upaH genes in biofilms and suspension-grown bacteria of different E. coli strains. These included the UPEC CFT073, the multidrug-resistant strain CDC-AR-0346, and clinical isolates obtained from UTI patients. The expression of fimH, ihf, upaB, and upaH was markedly heterogeneous in clinical isolates, both in terms of transcript levels and response to suspension or biofilm conditions. That expression pattern was distinct from the one in UPEC CFT073, where upaB and upaH were upregulated and ihf and fimH were slightly downregulated in biofilm. In conclusion, the data presented here show that the pattern of biofilm-associated genes in the clinical isolates from UTI patients is not fully related to the reference strain of UPEC CFT073. However, analysis of a larger number of samples is required.202439703715
8883150.9892Expression of Meiothermus ruber luxS in E. coli alters the antibiotic susceptibility and biofilm formation. Quorum sensing (QS) and signal molecules used for interspecies communication are well defined in mesophiles, but there is still a plethora of microorganisms in which existence and mechanisms of QS need to be explored, thermophiles being among them. In silico analysis has revealed the presence of autoinducer-2 (AI-2) class of QS signaling molecules in thermophiles, synthesized by LuxS (AI-2 synthase), though the functions of this system are not known. In this study, LuxS of Meiothermus ruber was used for understanding the mechanism and functions of AI-2 based QS among thermophilic bacteria. The luxS gene of M. ruber was expressed in luxS(-) deletion mutant of Escherichia coli. Complementation of luxS resulted in significant AI-2 activity, enhanced biofilm formation, and antibiotic susceptibility. Transcriptome analysis showed significant differential expression of 204 genes between the luxS-complemented and luxS(-) deletion mutant of E. coli. Majority of the genes regulated by luxS belonged to efflux pumps. This elucidation may contribute towards finding novel alternatives against incessant antibiotic resistance in bacteria.Key Points• Expression of luxS in luxS(-)E. coli resulted in increase in biofilm index. • Reduction in the MIC of antibiotics was observed after complementation of luxS. • Downregulation of efflux pump genes was observed after complementation of luxS. • Transcriptome analysis showed that 204 genes were differentially regulated significantly.202032215705
6373160.9892Antibiotic resistance and multidrug-resistant efflux pumps expression in lactic acid bacteria isolated from pozol, a nonalcoholic Mayan maize fermented beverage. Pozol is a handcrafted nonalcoholic Mayan beverage produced by the spontaneous fermentation of maize dough by lactic acid bacteria. Lactic acid bacteria (LAB) are carriers of chromosomal encoded multidrug-resistant efflux pumps genes that can be transferred to pathogens and/or confer resistance to compounds released during the fermentation process causing food spoiling. The aim of this study was to evaluate the antibiotic sensibility and the transcriptional expression of ABC-type efflux pumps in LAB isolated from pozol that contributes to multidrug resistance. Analysis of LAB and Staphylococcus (S.) aureus ATCC 29213 and ATCC 6538 control strains to antibiotic susceptibility, minimal inhibitory concentration (MIC), and minimal bactericidal concentration (MBC) to ethidium bromide were based in "standard methods" whereas the ethidium bromide efflux assay was done by fluorometric assay. Transcriptional expression of efflux pumps was analyzed by RT-PCR. LAB showed antibiotic multiresistance profiles, moreover, Lactococcus (L.) lactis and Lactobacillus (L.) plantarum displayed higher ethidium bromide efflux phenotype than S. aureus control strains. Ethidium bromide resistance and ethidium bromide efflux phenotypes were unrelated with the overexpression of lmrD in L. lactics, or the underexpression of lmrA in L. plantarum and norA in S. aureus. These findings suggest that, moreover, the analyzed efflux pumps genes, other unknown redundant mechanisms may underlie the antibiotic resistance and the ethidium bromide efflux phenotype in L. lactis and L. plantarum. Phenotypic and molecular drug multiresistance assessment in LAB may improve a better selection of the fermentation starter cultures used in pozol, and to control the antibiotic resistance widespread and food spoiling for health safety.201627247772
6060170.9891Safety Evaluation and Colonisation Abilities of Four Lactic Acid Bacteria as Future Probiotics. The study evaluated the safety and colonisation properties of four lactic acid bacteria (LAB), by determining their cell hydrophobicity and aggregation abilities. In addition, the presence of virulence and resistance genes was assayed in these probiotic candidates. Lactobacillus reuteri ZJ625, Lactobacillus reuteri VB4, Lactobacillus salivarius ZJ614 and Streptococcus salivarius NBRC13956 were tested for cell surface hydrophobicity abilities against xylene, chloroform and ethyl acetate. The isolates were also tested for auto-aggregation and co-aggregation abilities; the optical densities of cell growth were measured after 1, 2, 3 and 4 h of experimental set-up. DNA was extracted from all the four isolates and amplified using PCR with specific primers to detect virulence genes of adhesion collagen protein (ace) and aggregation substances (agg and asa); also, resistance genes of Vancomycin vanA, Vancomycin vanC1 and Vancomycin vanC2/3 were assayed in the four isolates. The isolates showed high hydrophobicity to all solvents: xylene (78-84%), chloroform (68-75%) and ethyl acetate (52-60%). High auto- and co-aggregations ranging from 60 to 70% and from 45 to 56% respectively were observed in the isolates after 4 h of incubation at 37 °C. Some of the tested isolates showed the presence of virulence and resistance genes; however, this does not indicate that these genes are unsafe because their transmission and expression abilities are unknown. Therefore, in this study, the isolates studied are considered safe for use as future probiotics, as revealed from results presented, which generally represents the scanned safety evaluations of the isolates as promising probiotics.201929881953
6372180.9891Sensitizing multi drug resistant Staphylococcus aureus isolated from surgical site infections to antimicrobials by efflux pump inhibitors. BACKGROUND: Staphylococcus aureus is a common hospital acquired infections pathogen. Multidrug-resistant Methicillin-resistant Staphylococcus aureus represents a major problem in Egyptian hospitals. The over-expression of efflux pumps is a main cause of multidrug resistance. The discovery of efflux pump inhibitors may help fight multidrug resistance by sensitizing bacteria to antibiotics. This study aimed to investigate the role of efflux pumps in multidrug resistance. METHODS: Twenty multidrug resistant S. aureus isolates were selected. Efflux pumps were screened by ethidium bromide agar cartwheel method and polymerase chain reaction. The efflux pump inhibition by seven agents was tested by ethidium bromide agar cartwheel method and the effect on sensitivity to selected antimicrobials was investigated by broth microdilution method. RESULTS: Seventy percent of isolates showed strong efflux activity, while 30% showed intermediate activity. The efflux genes mdeA, norB, norC, norA and sepA were found to play the major role in efflux, while genes mepA, smr and qacA/B had a minor role. Verapamil and metformin showed significant efflux inhibition and increased the sensitivity to tested antimicrobials, while vildagliptin, atorvastatin, domperidone, mebeverine and nifuroxazide showed no effect. CONCLUSION: Efflux pumps are involved in multidrug resistance in Staphylococcus aureus. Efflux pump inhibitors could increase the sensitivity to antimicrobials.202034394224
6076190.9891Isolation and identification of mucin-degrading bacteria originated from human faeces and their potential probiotic efficacy according to host-microbiome enterotype. AIM: Mucin-degrading bacteria are known to be beneficial for gut health. We aimed to isolate human-derived mucin-degrading bacteria and identify potential probiotic characteristics and their effects on the bacterial community and short-chain fatty acid (SCFA) production according to three different enterotypes of the host. METHODS AND RESULTS: Bacteria with mucin decomposition ability from human faeces were isolated and identified by 16S rRNA sequencing and MALDI-TOF. Heat resistance, acid resistance, antibiotic resistance, and antibacterial activity were analysed in the selected bacteria. Their adhesion capability to the Caco-2 cell was determined by scanning electron microscopy. Their ability to alter the bacterial community and SCFA production of the isolated bacteria was investigated in three enterotypes. The three isolated strains were Bifidobacterium(Bif.) animalis SPM01 (CP001606.1, 99%), Bif. longum SPM02 (NR_043437.1, 99%), and Limosilactobacillus(L.) reuteri SPM03 (CP000705.1, 99%) deposited in Korean Collection for Type Culture (KCTC-18958P). Among them, Bif. animalis exhibited the highest mucin degrading ability. They exhibited strong resistance to acidic conditions, moderate resistance to heat, and the ability to adhere tightly to Caco-2 cells. Three isolated mucin-degrading bacteria incubation increased Lactobacillus in the faecal bacteria from Bacteroides and Prevotella enterotypes. However, only L. reuteri elevated Lactobacillus in the faecal bacteria from the Ruminococcus enterotype. B. longum and B. animalis increased the α-diversity in the Ruminococcus enterotype, while their incubation with other intestinal types decreased the α-diversity. Bifidobacterium animalis and L. reuteri increased the butyric acid level in faecal bacteria from the Prevotella enterotype, and L. reuteri elevated the acetic acid level in those from the Ruminococcus enterotype. However, the overall SCFA changes were minimal. CONCLUSIONS: The isolated mucin-degrading bacteria act as probiotics and modulate gut microbiota and SCFA production differently according to the host's enterotypes. SIGNIFICANCE AND IMPACT OF STUDY: Probiotics need to be personalized according to the enterotypes in clinical application.202235365862