Biofilm Formation Related Genes Research Articles

Surfactin and fengycin contribute differentially to the biological activity of Bacillus subtilis NCD-2 against cotton verticillium wilt

Bacillus subtilis strain NCD-2 is a promising biocontrol agent for soil borne plant diseases. A previous study indicated that strain NCD-2 could produce surfactin and fengycin simultaneously. In this study, the exact roles of surfactin and fengycin in strain NCD-2 against cotton verticillium wilt were determined by knockout of the surfactin synthetase gene srfAA and the fengycin synthetase gene fenC. Compared to the wild-type strain NCD-2, the fengycin-deficient mutant ΔFenC decreased the antifungal ability toward Verticillium dahliae, but the surfactin-deficient mutant ΔSrfAA mainly decreased the swarming and biofilm formation abilities of strain NCD-2. Comparatively, the fengycin and surfactin double mutant ΔFenC/SrfAA decreased the antifungal activity and swarming and biofilm formation abilities to more significant levels. The colonization abilities of the wild type strain NCD-2 and its derivative mutants were compared in the cotton rhizosphere. Compared to the wild-type strain NCD-2, ΔSrfAA and ΔFenC/SrfAA exhibited the poorest colonization ability. RT-qPCR results confirmed that knockout of the srfAA and fenC genes decreased the expression of biofilm formation-related genes. The biocontrol abilities of the strains against cotton verticillium wilt were evaluated in a greenhouse. All mutants decreased the biocontrol abilities against cotton verticillium wilt, while ΔFenC/SrfAA showed the poorest biocontrol ability, followed by ΔFenC. Taken together, fengycin was the main antifungal active compound, and surfactin affected root colonization. Both the antifungal ability and root colonization of strain NCD-2 played important roles in suppressing cotton verticillium wilt.

Inhibition of Biofilm Formation and Virulence Factors of Cariogenic Oral Pathogen Streptococcus mutans by Shikimic Acid.

ABSTRACTStreptococcus mutans is known as an important oral pathogen causing dental caries, a widespread oral infectious disease. S. mutans synthesize exopolysaccharide (EPS) using glucosyltransferases (Gtfs), resulting in biofilm formation on the tooth surface. Bacterial cells in the biofilms become strongly resistant to a harsh environment, such as antibiotics and host defense mechanisms, making biofilm-based infections difficult to eliminate. Discovering novel antibiofilm agents, especially from natural products, helps to develop effective strategies against this kind of diseases. The present study investigated the inhibitory effect of shikimic acid (SA), one abundant compound derived from Illicium verum extract, on the biofilm formation of S. mutans. We found SA can reduce the EPS synthesized by this oral pathogen and modulate the transcription of biofilm formation related genes, leading to fewer bacterial cells in its biofilm. SA also interacted with cell membrane and membrane proteins, causing damage to bacterial cells. Ex vivo testing of biofilm formation on bovine teeth showed SA strongly decreased the number of S. mutans cells and the number of EPS accumulated on dental enamel surfaces. Moreover, SA exhibits almost no toxicity to human oral cells evaluated by in vitro biocompatibility assay. In conclusion, shikimic acid exhibits remarkable antibiofilm activity against S. mutans and has the potential to be further developed as a novel anticaries agent.IMPORTANCE Natural products are an important and cost-effective source for screening antimicrobial agents. Here, we identified one compound, shikimic acid, from Illicium verum extract, exhibiting antimicrobial activity against S. mutans proliferation. It also inhibits biofilm formation of this bacteria through decreasing Gtf expression and EPS synthesis. Furthermore, this compound exhibits no significant cytotoxicity at its MIC against S. mutans, providing evidence for its clinical application.

Synergistic Effect of Combination of a Temoporfin-Based Photodynamic Therapy with Potassium Iodide or Antibacterial Agents on Oral Disease Pathogens In Vitro.

5, 10, 15, 20-Tetrakis(3-hydroxyphenyl)chlorin (temoporfin) is a photosensitizer used in photodynamic therapy for oral cancer and periodontal disease treatment. This study determined the minimum inhibitory concentrations (MICs) and minimum bactericidal concentrations (MBCs) of temoporfin. Additionally, the combination of potassium iodide (KI) or antimicrobial agents in oral pathogens under hypoxic or normoxic conditions were determined. We also evaluated the biofilm removal effect and detected the expressions of the antibiotic resistance-related genes and biofilm formation-related genes of methicillin-resistant staphylococcus aureus (MRSA). The results provided reveal that the combination of the temoporfin and KI had a synergistic effect of reducing the MICs and MBCs of Lactobacillus acidophilus and Lactobacillus paracasei under normoxic and hypoxic conditions due to increasing H2O2 production. Temoporfin increased the biofilm removal of Aggregatibacter actinomycetemcomitans, Enterococcus faecalis, and Staphylococcus aureus under normoxic condition, and it reduced the antibiotic resistance-related genes expression of MRSA. The combination of temoporfin with ampicillin or chlorhexidine significantly enhanced the bactericidal effect on MRSA. This study provides a potential application of temoporfin on the clinical side against oral pathogens and the prevention of oral diseases.

Celastrol mitigates staphyloxanthin biosynthesis and biofilm formation in Staphylococcus aureus via targeting key regulators of virulence; in vitro and in vivo approach

BackgroundStaphylococcus aureus is a leading cause of human infections. The spread of antibiotic-resistant staphylococci has driven the search for novel strategies to supersede antibiotics use. Thus, targeting bacterial virulence rather than viability could be a possible alternative.ResultsThe influence of celastrol on staphyloxanthin (STX) biosynthesis, biofilm formation, antibiotic susceptibility and host pathogenesis in S. aureus has been investigated. Celastrol efficiently reduced STX biosynthesis in S. aureus. Liquid chromatography-mass spectrometry (LC–MS) and molecular docking revealed that celastrol inhibits STX biosynthesis through its effect on CrtM. Quantitative measurement of STX intermediates showed a significant pigment inhibition via interference of celastrol with CrtM and accumulation of its substrate, farnesyl diphosphate. Importantly, celastrol-treated S. aureus was more sensitive to environmental stresses and human blood killing than untreated bacteria. Similarly, inhibition of STX upon celastrol treatment rendered S. aureus more susceptible to membrane targeting antibiotics. In addition to its anti-pigment capability, celastrol exhibits significant anti-biofilm activity against S. aureus as indicated by crystal violet assay and microscopy. Celastrol-treated cells showed deficient exopolysaccharide production and cell hydrophobicity. Moreover, celastrol markedly synergized the action of conventional antibiotics against S. aureus and reduced bacterial pathogenesis in vivo using mice infection model. These findings were further validated using qRT-PCR, demonstrating that celastrol could alter the expression of STX biosynthesis genes as well as biofilm formation related genes and bacterial virulence.ConclusionsCelastrol is a novel anti-virulent agent against S. aureus suggesting, a prospective therapeutic role for celastrol as a multi-targeted anti-pathogenic agent.

Staphylococcus aureus biofilm inhibition by high voltage prick electrostatic field (HVPEF) and the mechanism investigation

The study was to investigate the inhibitory effect and mechanism of high voltage prick electrostatic field (HVPEF) on Staphylococcus aureus biofilms. Results showed that HVPEF effectively inactivated 24-h and 48-h established S. aureus biofilms, and the effect was verified on different food-contact materials. Confocal laser scanning microscopy and scanning electron microscopy analysis suggested that HVPEF disintegrated the established biofilms by killing the embedded bacteria, but it hardly reduced the bacteria adhesion. HVPEF also effectively inhibit the formation of S. aureus biofilms, the effects varied with electric voltage, treatment time and biofilm culture conditions. The direct effect of HVPEF on planktonic S. aureus was a possible mode of biofilm formation inhibition. HVPEF also suppressed biofilm formation by reducing the release of key compositions of extracellular polymeric substance, including extracellular DNA (eDNA), protein and polysaccharide intercellular adhesion (PIA), and regulating the expression of biofilm formation related genes (icaA, ebh, cidA, sarA, icaR and sigB). We propose HVPEF as a novel method to inhibit bacteria biofilm, based on the results, HVPEF has positive effects to prevent biofilm-associated contamination of S. aureus.

A Novel LysR Family Factor STM0859 is Associated with The Responses of Salmonella Typhimurium to Environmental Stress and Biofilm Formation.

Salmonella enterica subsp. enterica serovar Typhimurium (ST) is an intracellularly parasitic bacterium. This zoonotic pathogen causes food poisoning and thus imposes a severe threat to food safety. Here, to understand the regulatory roles of the novel transcription factor STM0859 on the response of ST to environmental stress and biofilm formation, the STM0859 gene-deficient strain and the complementation strain ΔSTM0859/STM0859 were generated, respectively. Then, its capacity of responding to environmental stresses and biofilm (BF) formation ability under different stresses, including acid, alkali, high salt, cholate, and oxidative stresses was tested. We further analyzed the interaction between the STM0859 protein and the promoter of the acid stress response-related gene rcsB by performing an electrophoresis mobility shift assay (EMSA). The results showed that acid resistance and BF formation capacities of ST-ΔSTM0859 strain were significantly weaker, as compared with those of Salmonella Typhimurium SL1344 (ST-SL1344) wild strain (p < 0.01). Quantitative qRT-PCR analysis showed that the expression levels of acid stress and BF formation-related genes, rcsB and rpoS, of ST-ΔSTM0859 strain were significantly reduced at the transcription levels, while the transcription levels of these genes were fully restored in complementation strain ST-ΔSTM0859/STM0859. The results of EMSA showed that STM0859 was capable of binding the promoter DNA fragments of the rcsB gene, suggesting that STM0859 can promote the transcription of the rcsB gene through interaction with its promoter, thereby exerting an indirectly regulatory role on the adaptive responses to acid stress and BF formation of ST. This study provided new insights into the regulatory mechanisms of the LysR family factors on the tolerances of ST under adverse environmental stresses.

Role of agglutinin-like sequence protein 3 (Als3) in the structure and antifungal resistance of Candida albicans biofilms.

Agglutinin-like sequence protein 3 (Als3) is a cell surface glycoprotein of Candida albicans that plays essential roles in the processes of adherence and biofilm formation in vitro. In this study, we focused on the contribution of Als3 to the structure and drug susceptibility of biofilms. The C. albicans wild-type (WT) strain DAY185, the als3Δ/Δ null strain and the als3Δ/Δ + pALS3 complemented strain were used. Colony-forming unit enumeration, crystal violet and cell surface hydrophobicity assays, scanning electron microscopy and confocal laser scanning microscopy coupled with analyses using COMSTAT software were performed to evaluate the biomass and architecture of the biofilms. The detailed architectural analysis showed a significant variation in the biofilm parameters of the als3Δ/Δ biofilms compared with those of the WT biofilms. Fluconazole, miconazole and amphotericin B were selected as the antifungal agents for the antimycotic susceptibility test, and increased susceptibility was found with the ALS3 deletion biofilms. A quantitative real-time polymerase chain reaction analysis showed downregulation of biofilm formation-related genes (ALS1, EFG1, HWP1 and CSH1) and drug resistance-related genes (ERG11, CDR1, CDR2 and MDR1) in the als3Δ/Δ biofilms. We concluded that Als3 contributes to biofilm formation by changing the biofilm architecture and is involved in the antifungal resistance of C. albicans biofilms.

Antisense yycG modulates the susceptibility of Staphylococcus aureus to hydrogen peroxide via the sarA

BackgroundThe infectious pathogen Staphylococcus aureus (S. aureus) is primarily associated with osteomyelitis. Hydrogen peroxide drainage is an effective antimicrobial treatment that has been adopted to combat S. aureus infections. Previous investigations have indicated that the antisense RNA (asRNA) strategy negatively modulates S. aureus YycFG TCS, and it significantly disrupts biofilm formation. However, the effects of the antisense yycG RNA (ASyycG) strategy on the susceptibility of biofilm-producing S. aureus to hydrogen peroxide and the mechanisms underlying this effect have not been elucidated to date.ResultsOverexpression of ASyycG inhibited the transcription of biofilm formation-related genes, including sarA and icaA. Additionally, the CFU counts and the live bacterial ratios of ASyycG biofilm-producing S. aureus treated with H2O2 were notably reduced across the groups. Notably, the predicted promoter regions of the sarA and icaA genes were directly regulated by YycF.ConclusionsASyycG was observed to sensitize biofilm-producing S. aureus to H2O2 intervention synergistically via the sarA and thus may represent a supplementary strategy for managing osteomyelitis. However, future in-depth studies should attempt to replicate our findings in animal models, such as the rat osteomyelitis model.

Diclazuril Inhibits Biofilm Formation and Hemolysis of Staphylococcus aureus.

Biofilm formation and hemolysis induced by Staphylococcus aureus are closely related to pathogenicity. However, no drugs exist to inhibit biofilm formation or hemolysis induced by S.aureus in clinical practice. This study found diclazuril had antibacterial action against S.aureus with minimum inhibitory concentrations (MICs) at 50 μM for both methicillin-sensitive S.aureus (MSSA) and methicillin-resistant S.aureus (MRSA). Diclazuril (at 1/4× or 1/8× MICs) significantly inhibited biofilm formation of S.aureus under static or flow-based conditions and also inhibited hemolysis induced by S.aureus. The RNA levels of transcriptional regulatory genes (agrA, agrC, luxS, sarA, sigB, saeR, saeS), biofilm formation-related genes (aur, bap, ccpA, cidA, clfA, clfB, fnbA, fnbB, icaA, icaB, sasG), and virulence-related genes (hla, hlb, hld, hlg, lukDE, lukpvl-S, spa, sbi, alpha-3 PSM, beta PSM, coa) of S.aureus were decreased when treated by diclazuril (at 1/4× MIC) for 4 h. The diclazuril nonsensitive clones of S.aureus were selected in vitro by induction of wildtype strains for about 90 days under the pressure of diclazuril. Mutations in the possible target genes of diclazuril against S.aureus were detected by whole-genome sequencing. This study indicated that there were three amino acid mutations in the diclazuril nonsensitive clone of S.aureus, two of which were located in genes with known function (SMC-Scp complex subunit ScpB and glyceraldehyde-3-phosphate dehydrogenase 1, respectively) and one in a gene with unknown function (hypothetical protein). Diclazuril showed a strong inhibition effect on planktonic cells and biofilm formation of S.aureus with the overexpression of the scpB gene.

Roles of ompA of Citrobacter werkmanii in bacterial growth, biocide resistance, biofilm formation and swimming motility

The genus Citrobacter is commonly found in environmental and industrial settings, some members of which have been used for bioremediation of heavy metals owing to the absorption ability of their biofilms. Although our previous studies have found that the outer membrane protein A (OmpA) contributes to the process of Citrobacter werkmanii biofilm formation, the underlying mechanisms remain elusive. Therefore, we deleted ompA from the genome of C. werkmanii and investigated its phenotypes in comparison to the wild type strain (WT) and the complementary strain using biochemical and molecular techniques including RNA-Seq. Our results demonstrated that the deletion of ompA led to an increase in biofilm formation on both polystyrene and glass surfaces due to upregulation of some biofilm formation related genes. Meanwhile, swimming ability, which is mediated by activation of flagellar assembly genes, was increased on semi-solid plates in the ∆ompA strain when compared with WT. Additionally, inactivation of ompA also caused increased 1,2-benzisothiazolin-3-one (BIT) resistance, differential responses to Ca2+ stress, curli protein expression and cellulose production. Finally, ∆ompA caused differential expression of a total of 1470 genes when compared with WT, of which 146 were upregulated and 1324 were downregulated. These genes were classified into different Gene Ontology (GO) and KEGG pathways. In summary, ompA in C. werkmanii contributes to a variety of biological functions and may act as a target site to modulate biofilm formation. KEY POINTS: • ompA is a negative regulator for biofilm formation by C. werkmanii. • ompA inhibits swimming motility of C. werkmanii. • ompA deletion causes different expression profiles in C. werkmanii.

Cadmium sulfide nanoparticle biomineralization and biofilm formation mediate cadmium resistance of the deep-sea bacterium Pseudoalteromonas sp. MT33b.

Cadmium (Cd) is a common toxic heavy metal in the environment, and bacteria have evolved different strategies to deal with Cd toxicity. Here, a bacterium designated Pseudoalteromonas sp. MT33b possessing strong Cd resistance was isolated from the Mariana Trench sediments. Supplement of cysteine significantly increased bacterial Cd resistance and removal rate. Biofilm formation was demonstrated to play a positive role toward bacterial Cd resistance. Transcriptome analysis showed the supplement of cysteine effectively prevented Cd2+ from entering bacterial cells, promoted saccharide metabolism and thereby facilitating energy production, which consists well with bacterial growth trend analysed under the same conditions. Notably, the expressions of many biofilm formation related genes including flagellar assembly, signal transduction, bacterial secretion and TonB-dependent transfer system were significantly upregulated when facing Cd stress, indicating their important roles in determining bacterial biofilm formation and enhancing Cd resistance. Overall, this study indicates the formation of insoluble CdS precipitates and massive biofilm is the major strategy adopted by Pseudoalteromonas sp. MT33b to eliminate Cd stress. Our results provide a good model to investigate how heavy metals impact biofilm formation in the deep-sea ecosystems, which may facilitate a deeper understanding of microbial environmental adaptability and better utilization of these microbes for bioremediation purposes in the future.

Decreased biofilm formation by planktonic cells of Listeria monocytogenes in the presence of sodium hypochlorite

The objective of this study was to determine if the adaptation at planktonic stage to subinhibitory concentrations (SIC) of sodium hypochlorite (NaOCl) could modulate the biofilm forming ability of five Listeria monocytogenes strains V7, Scott A, FSL-N1-227, FSL F6-154 and ATCC 19116 representing serotypes 1/2a, 4b and 4c. Biofilm formation by NaOCl nonadapted and adapted L. monocytogenes planktonic cells was measured in the presence or absence of SIC of NaOCl. The biofilm formation ability of NaOCl nonadapted and adapted L. monocyotgenes planktonic cells was reduced only in the presence of NaOCl (P < 0.05). Scanning electron microscopy revealed that the continuous exposure of NaOCl induced morphological changes in the L. monocytogenes biofilm structure and reduced its attachment to polystyrene surface. The qRT-PCR results also showed that the subinhibitory NaOCl reduced biofilm formation related gene expression such as motility and quorum sensing signals (P < 0.05). These findings indicate that subinhibitory NaOCl can reduce the ability of L. monocytogenes planktonic cells to form biofilms on polystyrene surface.

Housefly Phormicin inhibits Staphylococcus aureus and MRSA by disrupting biofilm formation and altering gene expression in vitro and in vivo

The increasing drug resistance of pathogenic bacteria is a crisis that threatens public health. Antimicrobial peptides (AMPs) have been suggested to be potentially effective alternatives to solve this problem. Here, we tested housefly Phormicin-derived peptides for effects on Staphylococcus aureus and methicillin-resistant Staphylococcus aureus (MRSA) infections in vitro and in vivo. A decreased bacterial load of MRSA was observed in the mouse scald model after treatment with Phormicin and in the positive control group (vancomycin). A mouse scrape model indicated that Phormicin helps the host fight drug-resistant MRSA infections. The protective effect of Phormicin on MRSA was confirmed in the Hermetia illucens larvae model. Phormicin also disrupted the formation of S. aureus and MRSA biofilms. Furthermore, this effect coincided with the downregulation of biofilm formation-related gene expression (agrC, sigB, RNAIII, altA, rbf, hla, hld, geh and psmɑ). Notably, virulence genes and several regulatory factors were also altered by Phormicin treatment. Based on these findings, housefly Phormicin helps the host inhibit MRSA infection through effects on biofilm formation and related gene networks. Therefore, housefly Phormicin potential represents a candidate agent for clinical MRSA chemotherapy.

Inhibition of enteropathogenic Escherichia coli biofilm formation by DNA aptamer.

Enteropathogenic Escherichia coli (EPEC) is a bioagent that causes diarrhea through the formation of biofilm. The recalcitrant of EPEC to the current conventional antibiotic treatment has grown a big concern in a way to find effective alternative inhibitors. Aptamers have been demonstrated to show the ability to kill the pathogenic bacteria through inhibition of biofilm formation. Therefore, this study aimed to investigate antibiofilm activities of six types of aptamers against EPEC K1.1 which was isolated from patients with diarrhea. Environmental conditions such as temperatures and pH which impacted on biofilm formation of EPEC K1.1 and also biofilm inhibition of aptamer on EPEC K1.1 were performed by counting the crystal violet formation in 96-well polystyrene microplates at OD570. The motility examination combined with qPCR were applied to prove the mechanism of aptamers inhibition on biofilm by targeting essential genes that involve biofilm formation. The result showed that by applying cut off value at 0.399, aptamer SELEX 10 Colony 5 exhibited the highest biofilm inhibition against EPEC K1.1 with an absorbance value of 0.126. Further analysis showed that this aptamer also was able to reduce the motility diameter of EPEC K1.1. The effect of this aptamer on EPEC K1.1 motility was confirmed by qPCR where the mRNA level of motB, csgA and lsrA gene reduced significantly compared to the untreated group. Aptamer SELEX 10 Colony 5 was able to inhibit biofilm formation through interfering the motility ability of EPEC K1.1 and also by reducing the mRNA level of biofilm formation-related genes. This study provides evidences that aptamer is effective and promising for both antibiofilm of EPEC K1.1 and alternative treatment of diarrhea.

Sodium New Houttuyfonate Inhibits Candida albicans Biofilm Formation by Inhibiting the Ras1-cAMP-Efg1 Pathway Revealed by RNA-seq.

Here, we aim to investigate the antifungal effect and mechanism of action of sodium new houttuyfonate (SNH) against Candida albicans. Microdilution analysis results showed that SNH possesses potent inhibitory activity against C. albicans SC5314, with a MIC80 of 256 μg/mL. Furthermore, we found that SNH can effectively inhibit the initial adhesion of C. albicans. Inverted microscopy, crystal violet staining, scanning electron microscopy and confocal laser scanning microscopy results showed that morphological changes during the transition from yeast to hypha and the biofilm formation of C. albicans are repressed by SNH treatment. We also found that SNH can effectively inhibit the biofilm formation of clinical C. albicans strains (Z103, Z3044, Z1402, and Z1407) and SNH in combination with fluconazole, berberine chloride, caspofungin and itraconazole antifungal agents can synergistically inhibit the biofilm formation of C. albicans. Eukaryotic transcriptome sequencing and qRT-PCR results showed that SNH treatment resulted in significantly down-regulated expression in several biofilm formation related genes in the Ras1-cAMP-Efg1 pathway (ALS1, ALA1, ALS3, EAP1, RAS1, EFG1, HWP1, and TEC1) and significantly up-regulated expression in yeast form-associated genes (YWP1 and RHD1). We also found that SNH can effectively reduce the production of key messenger cAMP in the Ras1-cAMP-Efg1 pathway. Furthermore, using Galleria mellonella as an in vivo model we found that SNH can effectively treat C. albicans infection in vivo. Our presented results suggest that SNH exhibits potential antibiofilm effects related to inhibiting the Ras1-cAMP-Efg1 pathway in the biofilm formation of C. albicans.

Genotype, biofilm formation ability and specific gene transcripts characteristics of endodontic Enterococcus faecalis under glucose deprivation condition

ObjectiveTo study the relationship between the specific gene and biofilm formation ability of seven wild type Enterococcus faecalis (E. faecalis) under glucose deprivation conditions. DesignWild type E. faecalis (3RC, 5RC, 25RC, 31RC, 33RC, 37RC, 58RC) extracted from the teeth with persistent apical periodontitis were cultured under glucose deprivation conditions and then resequenced. The biofilm formation ability was compared using primary adherence assay, confocal laser scanning microscopy (CLSM) and scanning electron microscopy (SEM). The transcriptional level of biofilm formation–related genes (ace, gelE, efa, esp and fsrB) were detected. ResultsGenomic resequencing showed that 3RC and 58RC (Class B) were similar, while 5RC, 25RC, 31RC, 33RC and 37RC (Class A) were similar. Based on primary adherence assay, CLSM and SEM results, biofilm formation ability of Class B strains was lower, while Class A strains were higher when compared with control group (0.25 % glucose). Furthermore, compared with control group (0.25 % glucose), the transcriptional levels of ace, efa and fsrB genes were upregulated in all strains; the transcriptional levels of gelE were downregulated in Class B strains, upregulated in Class A strains; the transcriptional levels of esp of Class B strains were downregulated, while upregulated in 25RC, 31RC and 37RC (Class A), and not observed in 5RC and 33RC. ConclusionThe genotypes of wild type E. faecalis of different persistent periapical periodontitis teeth are different. The genotype differences and the transcription levels of related virulence genes (ace, gelE, efa, esp and fsrB) are related to the biological phenotype.

EFFECTS OF THE bap AND eno GENES ON THE EFFECTIVENESS OF DISINFECTANTS AGAINST COAGULASE-NEGATIVE STAPHYLOCOCCI

This study sought to determine the minimum bactericidal concentrations (MBCs) of didecyldimethylammonium chloride (DDAC), povidone iodine (PI), and chlorhexidine and the differences in these values among coagulase-negative staphylococci (CNSs) that harbor or do not harbor biofilm-related genes (eno, bap, and ica). Using the limiting dilution technique, the MBCs against three different disinfectants were estimated and compared using statistical methods. The results showed that the biofilm-gene-positive CNS isolates exhibited higher tolerance ([Formula: see text], upper-tailed test) to 2[Formula: see text]mg[Formula: see text]L[Formula: see text] DDAC and 10[Formula: see text]mg[Formula: see text]L[Formula: see text] povidone iodine ([Formula: see text], upper-tailed test). Different resistance capacities were found among these three biofilm formation-related genes. The isolates that contained the bap gene exhibited a higher tolerance level. This observation was confirmed by logistic regression models, which revealed that the bap (odds ratio [Formula: see text]) and eno (odds ratio [Formula: see text]) genes significantly affected the survival capacity. This study demonstrated that biofilm-gene-positive CNS isolates exhibited increased survival in the presence of higher concentrations of all three disinfectants and that both the bap and eno genes could affect the survival capacity. Among the three genes, the bap gene had the strongest effect on the survival capacity of CNSs in the presence of all three disinfectants. The appropriate use of disinfectants can control these biofilm-gene embedded CNSs more efficiently.

The response regulator ArcA enhances biofilm formation in the vpsT manner under the anaerobic condition in Vibrio cholerae

Vibrio cholerae, the agent of severe diarrheal disease cholera, is known to form biofilm to persist in the environmental and the host,s intestines. The bacteria execute a complex regulatory pathway producing virulence factors that allow colonization and cause disease in response to environmental signals in the intestine, including low oxygen-limited condition. VpsR and VpsT are primary regulators of the biofilm formation-regulatory network. In this study, we determined that anaerobic induction enhanced biofilm formation via the two component system, ArcB/A, which functions as a positive regulator of toxT expression. The biofilm formation has reduced approximately 2.4-fold in the ΔarcA mutant compared to the wild type in anaerobic condition. Chip-qPCR and EMSA assays confirmed that ArcA can bind directly to the vpsT promoter and then activates the expression of biofilm formation related genes, vpsA-K and vpsL-Q. Meanwhile, the ΔarcA mutant decreased the ability of colonization in intestine with CI (competition index) of 0.27 compared to wild type strain. These results suggest that ArcA links the expression of virulence and biofilm synthesis genes during anaerobic condition, and contributes to understand the complex relationship between biofilm formation and the intestinal signals during infection.

Molecular Mechanisms Underpinning Aggregation in Acidiphilium sp. C61 Isolated from Iron-Rich Pelagic Aggregates.

Iron-rich pelagic aggregates (iron snow) are hot spots for microbial interactions. Using iron snow isolates, we previously demonstrated that the iron-oxidizer Acidithrix sp. C25 triggers Acidiphilium sp. C61 aggregation by producing the infochemical 2-phenethylamine (PEA). Here, we showed slightly enhanced aggregate formation in the presence of PEA on different Acidiphilium spp. but not other iron-snow microorganisms, including Acidocella sp. C78 and Ferrovum sp. PN-J47. Next, we sequenced the Acidiphilium sp. C61 genome to reconstruct its metabolic potential. Pangenome analyses of Acidiphilium spp. genomes revealed the core genome contained 65 gene clusters associated with aggregation, including autoaggregation, motility, and biofilm formation. Screening the Acidiphilium sp. C61 genome revealed the presence of autotransporter, flagellar, and extracellular polymeric substances (EPS) production genes. RNA-seq analyses of Acidiphilium sp. C61 incubations (+/− 10 µM PEA) indicated genes involved in energy production, respiration, and genetic processing were the most upregulated differentially expressed genes in the presence of PEA. Additionally, genes involved in flagellar basal body synthesis were highly upregulated, whereas the expression pattern of biofilm formation-related genes was inconclusive. Our data shows aggregation is a common trait among Acidiphilium spp. and PEA stimulates the central cellular metabolism, potentially advantageous in aggregates rapidly falling through the water column.

Towards a better understanding of Pseudomonas putida biofilm formation in the presence of ZnO nanoparticles (NPs): Role of NP concentration

Elucidating the effects of nanoparticles (NPs) on key bacterial functions not only deepens our understanding of nano-toxicity mechanisms, but also guides us in the design criteria for manufacturing safe nanomaterials. In this study, bacterial growth, biofilm development and the expression of biofilm-related genes were monitored in Pseudomonas putida KT2440, a plant-beneficial bacterium, following exposure to ZnO NPs. Low concentrations of NPs (0.5–30 mg L−1) significantly promoted bacterial growth and biofilm formation, while higher concentrations (>30 mg L−1) significantly inhibited biofilm formation. Confocal laser scanning microscopy revealed that microscope slides coated with 0.5 mg L−1 of ZnO NPs showed enhanced bacterial colonization and biomass production, but at higher concentrations (250 mg L−1), biomass production was about 11 times lower than that of the substrate without NPs. Increased protein and sugar contents of the biofilm matrix corroborated the stimulating effects of low concentrations of ZnO NPs. Physiological data were supported by changes in the expression of genes associated with oxidative stress and biofilm development. ZnO NPs at 0.5 mg L−1 stimulated the expression of quorum sensing, lipopolysaccharide biosynthesis, and antibiotic resistance genes; high concentrations of ZnO NPs (250 mg L−1) down-regulated biofilm formation-related genes and up-regulated antioxidant genes. Our results indicate that long-term release of low concentrations of ZnO NPs to the environment would promote undesired biofilm formation and increased resistance to antibiotics.