Biofilm Formation Gene Research Articles

Identification of the antibacterial action mechanism of curcumin on Streptococcus mutans through transcriptome profiling

ObjectiveThe purpose of this study was to explore the effect and mechanism responsible for how curcumin affects the biofilm formation by Streptococcus mutans (S. mutans). DesignThe antibacterial activity of curcumin was evaluated by measuring the minimum inhibitory concentration (MIC) and the minimum bactericidal concentration (MBC). The mass of the biofilm was measured by crystal violet staining. Transcriptome sequencing was used to obtain all the transcript information associated with the biological activity of curcumin-treated S. mutans. Real-time quantitative PCR (qRT-PCR) was performed to examine the expression levels of related biofilm formation genes. ResultsThe MIC value for curcumin was 64 μM. Curcumin inhibited the formation of a biofilm by S. mutans and degraded mature biofilms. A gene ontology enrichment analysis showed that the DEGs were significantly relevant to biofilm formation. In addition, 17 significantly enriched Kyoto Encyclopedia of Genes and Genomes pathways (p ≤ 0.01) were identified and were potentially associated with the biochemical metabolic processes of S. mutans. DEGs associated with the biofilm formation of S. mutants, including gtfB, gtfC, rgpG, spaP, spxA1, spxA2, bacA, lrgB, and gshAB. The qRT-PCR results were consistent with transcriptome sequencing that the expression levels of gtfB, gtfC, rgpG, and spaP significantly decreased in the curcumin-treated group, whereas the expression levels of spx1, spx2, bacA, lrgB, and gshAB were up-regulated. ConclusionsCurcumin showed marked inhibitory effects against the formation of biofilms by S. mutans and degradation of formed biofilms.

Genomic Insights and Functional Analysis Reveal Plant Growth Promotion Traits of Paenibacillus mucilaginosus G78.

Paenibacillus mucilaginosus has widely been reported as a plant growth-promoting rhizobacteria (PGPR). However, the important genomic insights into plant growth promotion in this species remain undescribed. In this study, the genome of P. mucilaginosus G78 was sequenced using Illumina NovaSeq PE150. It contains 8,576,872 bp with a GC content of 58.5%, and was taxonomically characterized. Additionally, a total of 7337 genes with 143 tRNAs, 41 rRNAs, and 5 ncRNAs were identified. This strain can prohibit the growth of the plant pathogen, but also has the capability to form biofilm, solubilize phosphate, and produce IAA. Twenty-six gene clusters encoding secondary metabolites were identified, and the genotypic characterization indirectly proved its resistant ability to ampicillin, bacitracin, polymyxin and chloramphenicol. The putative exopolysaccharide biosynthesis and biofilm formation gene clusters were explored. According to the genetic features, the potential monosaccharides of its exopolysaccharides for P. mucilaginosus G78 may include glucose, mannose, galactose, fucose, that can probably be acetylated and pyruvated. Conservation of the pelADEFG compared with other 40 Paenibacillus species suggests that Pel may be specific biofilm matrix component in P. mucilaginosus. Several genes relevant to plant growth-promoting traits, i.e., IAA production and phosphate solubilization are well conserved compared with other 40 other Paenibacillus strains. The current study can benefit for understanding the plant growth-promoting traits of P. mucilaginosus as well as its potential application in agriculture as PGPR.

The Antibacterial Effect of Boron Compounds and Evaluation of the Effects on Biofilm Formation in the Infection Model of Klebsiella pneumoniae on the HepG2 Cell Line

Aim: Klebsiella pneumoniae causes hospital-acquired pneumonia, urinary tract infection and bacteremia in immunocompromised patients. Klebsiella pneumoniae, which has become more common recently, causes antibiotic resistance as well as pyogenic liver abscesses and hematogenous metastatic spread in humans. Developing antibiotic resistance complicates the treatment of liver infections. In our study, we aimed to evaluate the effect of boron compounds in an infection model created by Klebsiella pneumoniae 700603. Materials and Methods: Minimum inhibitory concentration and fractional inhibitory concentration studies, resistance gene levels, and HepG2 cell analyses were performed and evaluated. Results: We determined the low and high minimum inhibitory concentration values of boron components, sodium perborate monohydrate and etidote, respectively. In addition, sodium perborate monohydrate is also effective on biofilm formation. Our findings have shown that boron compounds are more effective when used in a combination. In the toxicity model created in the cellular study, the boron compounds cytotoxic effect decreased due to their antibacterial effects. Conclusion: İt seems that boron compounds are effective, and the positive effect increases when used together.

Reinforcement amid genetic diversity in the Candida albicans biofilm regulatory network.

Biofilms of the fungal pathogen Candida albicans include abundant long filaments called hyphae. These cells express hypha-associated genes, which specify diverse virulence functions including surface adhesins that ensure biofilm integrity. Biofilm formation, virulence, and hypha-associated gene expression all depend upon the transcription factor Efg1. This transcription factor has been characterized extensively in the C. albicans type strain SC5314 and derivatives, but only recently has its function been explored in other clinical isolates. Here we define a principal set of Efg1-responsive genes whose expression is significantly altered by an efg1Δ/Δ mutation across 17 clinical isolates. This principal gene set includes 68 direct Efg1 targets, whose 5' regions are bound by Efg1 in five clinical isolates, and 42 indirect Efg1 targets, whose 5' regions are not detectably bound by Efg1. Three direct Efg1 target genes encode transcription factors-BRG1, UME6, and WOR3 -whose increased expression in an efg1Δ/Δ mutant restores expression of multiple indirect and direct principal targets, as well as biofilm formation ability. Although BRG1 and UME6 are well known positive regulators of hypha-associated genes and biofilm formation, WOR3 is best known as an antagonist of Efg1 in the sexual mating pathway. We confirm the positive role of WOR3 in biofilm formation with the finding that a wor3Δ/Δ mutation impairs biofilm formation in vitro and in an in vivo biofilm model. Positive control of Efg1 direct target genes by other Efg1 direct target genes-BRG1, UME6, and WOR3 -may buffer principal Efg1-responsive gene expression against the impact of genetic variation in the C. albicans species.

A Coculture of Enterobacter and Comamonas Species Reduces Cadmium Accumulation in Rice.

The accumulation of cadmium (Cd) in plants is strongly impacted by soil microbes, but its mechanism remains poorly understood. Here, we report the mechanism of reduced Cd accumulation in rice by coculture of Enterobacter and Comamonas species. In pot experiments, inoculation with the coculture decreased Cd content in rice grain and increased the amount of nonbioavailable Cd in Cd-spiked soils. Fluorescence in situ hybridization and scanning electron microscopy detection showed that the coculture colonized in the rhizosphere and rice root vascular tissue and intercellular space. Soil metagenomics data showed that the coculture increased the abundance of sulfate reduction and biofilm formation genes and related bacterial species. Moreover, the coculture increased the content of organic matter, available nitrogen, and potassium and increased the activities of arylsulfatase, β-galactosidase, phenoloxidase, arylamidase, urease, dehydrogenase, and peroxidase in soils. In subsequent rice transcriptomics assays, we found that the inoculation with coculture activated a hypersensitive response, defense-related induction, and mitogen-activated protein kinase signaling pathway in rice. Heterologous protein expression in yeast confirmed the function of four Cd-binding proteins (HIP28-1, HIP28-4, BCP2, and CID8), a Cd efflux protein (BCP1), and three Cd uptake proteins (COPT4, NRAM5, and HKT6) in rice. Succinic acid and phenylalanine were subsequently proved to inhibit rice divalent Cd [Cd(II)] uptake and activate Cd(II) efflux in rice roots. Thus, we propose a model that the coculture protects rice against Cd stress via Cd immobilization in soils and reducing Cd uptake in rice. [Formula: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Curcumin and 10-undecenoic acid as natural quorum sensing inhibitors of LuxS/AI-2 of Bacillus subtilis and LasI/LasR of Pseudomonas aeruginosa

The quorum sensing (QS) system is related to cell-to-cell communication as a function of population density, which regulates several physiological functions including biofilm formation and virulence gene expression. QS inhibitors have emerged as a promising strategy to tackle virulence and biofilm development. Among a wide variety of phytochemicals, many of them have been described as QS inhibitors. Driven by their promising clues, this study aimed to identify active phytochemicals against LuxS/autoinducer-2 (AI-2) (as the universal QS system) from Bacillus subtilis and LasI/LasR (as a specific QS system) of Pseudomonas aeruginosa, through in silico analysis followed by in vitro validation. The optimized virtual screening protocols were applied to screen a phytochemical database containing 3479 drug-like compounds. The most promising phytochemicals were curcumin, pioglitazone hydrochloride, and 10-undecenoic acid. In vitro analysis corroborated the QS inhibitory activity of curcumin and 10-undecenoic acid, however, pioglitazone hydrochloride showed no relevant effect. Inhibitory effects on LuxS/AI-2 QS system triggered reduction of 33–77% by curcumin (at 1.25–5 µg/mL) and 36–64% by 10-undecenoic acid (at 12.5–50 µg/mL). Inhibition of LasI/LasR QS system was 21% by curcumin (at 200 µg/mL) and 10–54% by 10-undecenoic acid (at 15.625–250 µg/mL). In conclusion, in silico analysis allowed the identification of curcumin and, for the first time, 10-undecenoic acid (showing low cost, high availability, and low toxicity) as alternatives to counteract bacterial pathogenicity and virulence, avoiding the imposition of selective pressure usually related to classic industrial disinfection and antibiotics therapy.

Antibacterial and anti-biofilm activities of Disaspidin BB against Staphylococcus epidermidis.

Staphylococcus epidermidis infections are an important concern in worldwide, especially when associated with biofilms, and resistance of this agent to many drugs makes the situation even worse. We investigated the inhibitory effect of Disaspidin BB obtained from plant extracts and purifications on clinical S. epidermidis strains and their biofilms, and preliminarily investigated its mechanism of of its anti-biofilm activity. The broth dilution method was used to determine the minimum inhibitory concentrations (MIC) of Disaspidin BB on 11 clinical S. epidermidis strains (MIC value of 0.63 ~ 2.5 μg/ml). SEP-05 was found to be erythromycin-resistant (MIC value>8 μg/ml) and Disaspidin BB sensitive with an MIC value of 0.63 μg/ml. The time-kill curve assay indicated that the antibacterial activity of Disaspidin BB against SEP-05 with concentration dependence. The metabolic activity and total biomass of the drug-treated SEP-05 biofilm in each stage were significantly inhibited by the crystalline violet and XTT assay, and the scavenging effect of Disaspidin BB on SEP-05 biofilm was also confirmed by SEM observation. The results of real-time quantitative PCR showed that subinhibitory concentrations Disaspidin BB can inhibit biofilm formation by affecting the expression level of key genes (aap, atlE, icaA, luxS, recA) in SEP-05 biofilm formation. In addition, the content of polysaccharides, proteins and extracellular DNA in biofilm matrix after the intervention of Disaspidin BB was significantly reduced, and it was tentatively determined that the ability of SEP-05 biofilm formation and its stability were thus disturbed. The results show that Disaspidin BB has promising antibacterial effect on erythromycin-resistant S. epidermidis and significant scavenging effect on its biofilm, which provides a theoretical basis for the further development of BB as a new drug for the treatment of skin infections caused by S. epidermidis.

Characterization of high fluoride resistant Pseudomonas aeruginosa species isolated from water samples

Groundwater fluoride contamination is one of the most serious toxicological environmental issues in India and around the world. Water samples were taken from Natham taluk and screened for fluoride resistant bacteria. Initially, twenty-four fluoride resistant colonies were selected from 50 mM NaF supplemented LB agar plates. On blood agar plates, all isolates showed β-haemolysis, confirming their status as pathogens. Virulence factors (algD, plcH, toxA, gyrB, rhlC, lasB) and biofilm-forming genes (ppyR, pelA, pslA) were identified in these isolates by PCR analysis. The fluoride ion transporter ‘crcB’ was successfully amplified from these isolates by gene-specific PCR. Genus Pseudomonas and P. aeruginosa species-specific PCR analysis identified that all strains as belonging to the P. aeruginosa species. Besides, three high fluoride resistance strains were selected based on high fluoride resistance and confirmed as P. aeruginosa species by 16S rRNA sequencing and NCBI blast analysis.

Hemolysin function of Listeria is related to biofilm formation: transcriptomics analysis

Listeriolysin O (LLO) is the main virulence protein of Listeria monocytogenes (LM), that helps LM escape lysosomes. We previously found that the cellular immune response elicited by L.ivanovii (LI) is weaker than that elicited by LM. We speculated that this may be related to the function of ivanolysin O (ILO). Here, we constructed hemolysin gene deletion strain, LIΔilo, and a modified strain, LIΔilo::hly, in which ilo was replaced by hly. Prokaryotic transcriptome sequencing was performed on LI, LIΔilo, and LIΔilo::hly. Transcriptome differences between the three strains were compared, and genes and pathways with significant differences between the three strains were analyzed. Prokaryotic transcriptome sequencing results revealed the relationship of ilo to the ribosome, quorum sensing, and phosphotransferase system (PTS) pathways, etc. LIΔilo exhibited attenuated biofilm formation ability compared to LI. Biofilm formation was significantly recovered or even increased after replenishing hly. After knocking out ilo, the relative expression levels of some virulence genes, including sigB, prfA, actA, smcL, and virR, were up-regulated compared to LI. After replenishing hly, these genes were down-regulated compared to LIΔilo. The trend and degree of such variation were not completely consistent when cultured in media containing only monosaccharides or disaccharides. The results confirmed that hemolysin is related to some important biological properties of Listeria, including biofilm formation and virulence gene expression levels. This is the first comprehensive study on ILO function at the transcriptomic level and the first evidence of a relationship between Listeria hemolysin and biofilm formation.

The Landscape of Gene Expression during Hyperfilamentous Biofilm Development in Oral Candida albicans Isolated from a Lung Cancer Patient

The filamentation ability of Candida albicans represents one of the main virulence factors allowing for host tissue penetration and biofilm formation. The aim of this paper was to study the genetic background of the hyperfilamentous biofilm development in vitro in C. albicans isolated from the oral cavity of a lung cancer patient. Analyzed C. albicans isolates (CA1, CA2, CA3) were chosen based on their different structures of mature biofilm. The CA3 isolate possessing hyperfilamentation properties and forming high biofilm was compared with CA1 and CA2 isolates exhibiting low or average biofilm-forming ability, respectively. The detailed biofilm organization was studied with the use of confocal scanning laser microscopy. The whole transcriptome analysis was conducted during three stages of biofilm development (24 h, 48 h, 72 h). In contrast to CA1 and/or CA2 isolate, the CA3 isolate was characterized by a significant upregulation of genes encoding for cell wall proteins (HWP1, PGA13, PGA44, ALS3) and candidalysin (ECE1), as well as being involved in iron metabolism (FRE1, ALS3), sulfur metabolism (HAL21), the degradation of aromatic compounds (HQD2), and membrane transport (DIP5, PHO89, TNA1). In contrast, some genes (SCW11, FGR41, RBE1) in the CA3 were downregulated. We also observed the overexpression of a few genes over time-mainly FRE1, ATX1, CSA2 involved in iron metabolism. This is the first insight into the potential function of multiple genes in the hyperfilamentous biofilm formation in C. albicans, primarily isolated from host tissue, which may have an important clinical impact on cancer patients. Moreover, the presented data can lay the foundation for further research on novel pathogen-specific targets for antifungal drugs.

Identification of novel genes involved in the biofilm formation process of Avian Pathogenic Escherichia coli (APEC).

Avian pathogenic Escherichia coli (APEC) is the etiological agent of avian colibacillosis, a leading cause of economic loss to the poultry industry worldwide. APEC causes disease using a diverse repertoire of virulence factors and has the ability to form biofilms, which contributes to the survival and persistence of APEC in harsh environments. The objective of this study was to identify genes most widespread and important in APEC that contribute to APEC biofilm formation. Using the characterized APEC O18 as the template strain, a total of 15,660 mutants were randomly generated using signature tagged mutagenesis and evaluated for decreased biofilm formation ability using the crystal violet assay. Biofilm deficient mutants were sequenced, and a total of 547 putative biofilm formation genes were identified. Thirty of these genes were analyzed by PCR for prevalence among 109 APEC isolates and 104 avian fecal E. coli (AFEC) isolates, resulting in nine genes with significantly greater prevalence in APEC than AFEC. The expression of these genes was evaluated in the wild-type APEC O18 strain using quantitative real-time PCR (qPCR) in both the exponential growth phase and the mature biofilm phase. To investigate the role of these genes in biofilm formation, isogenic mutants were constructed and evaluated for their biofilm production and planktonic growth abilities. Four of the mutants (rfaY, rfaI, and two uncharacterized genes) displayed significantly decreased biofilm formation, and of those four, one (rfaI) displayed significantly decreased growth compared to the wild type. Overall, this study identified novel genes that may be important in APEC and its biofilm formation. The data generated from this study will benefit further investigation into the mechanisms of APEC biofilm formation.

Plasmid parB contributes to uropathogenic Escherichia coli colonization in vivo by acting on biofilm formation and global gene regulation.

The endogenous plasmid pUTI89 harbored by the uropathogenic Escherichia coli (UPEC) strain UTI89 plays an important role in the acute stage of infection. The partitioning gene parB is important for stable inheritance of pUTI89. However, the function of partitioning genes located on the plasmid in pathogenesis of UPEC still needs to be further investigated. In the present study, we observed that disruption of the parB gene leads to a deficiency in biofilm formation in vitro. Moreover, in a mixed infection with the wild type strain and the parB mutant, in an ascending UTI mouse model, the mutant displayed a lower bacterial burden in the bladder and kidneys, not only at the acute infection stage but also extending to 72 hours post infection. However, in the single infection test, the reduced colonization ability of the parB mutant was only observed at six hpi in the bladder, but not in the kidneys. The colonization capacity in vivo of the parB-complemented strain was recovered. qRT-PCR assay suggested that ParB could be a global regulator, influencing the expression of genes located on both the endogenous plasmid and chromosome, while the gene parA or the operon parAB could not. Our study demonstrates that parB contributes to the virulence of UPEC by influencing biofilm formation and proposes that the parB gene of the endogenous plasmid could regulate gene expression globally.

Effects of chlorogenic acid on antimicrobial, antivirulence, and anti-quorum sensing of carbapenem-resistant Klebsiella pneumoniae.

The rise in infections caused by the hypervirulent carbapenem-resistant Klebsiella pneumoniae (hv-CRKP) is an emergent threat to public health. We assessed the effects of chlorogenic acid (CA), a natural phenolic compound, on antibacterial, antivirulence, and anti-quorum sensing (QS) of hv-CRKP. Five hv-CRKP were selected for antimicrobial susceptibility test and confirmed to carry virulence genes and carbapenem-resistant genes by polymerase chain reaction (PCR). Subsequently, a series of time-kill assay, determinations of protease activity and capsule content, biofilm-related experiment, scanning electron microscopy (SEM) and transmission electron microscope (TEM) observation, G. mellonella infection model, quantitative real-time PCR (qRT-PCR) of QS-related genes and biofilm formation genes, as well as AI-2 binding test were conduct to verify the effect of CA on hv-CRKP. Five CRKP strains showed varying degrees of resistance to antibacterial agents. All strains carried the bla KPC-2 gene, primarily carrying rmpA2, iucA, and peg-344. CA showed no effect on CRKP growth at the 1/2 minimum inhibitory concentration (MIC), 1/4 MIC, and 1/8 MIC, CA could reduce the production of extracellular protease and capsular polysaccharides, and improve the survival rate of larvae in Galleria mellonella (G. mellonella) infection model. By means of crystal violet staining and scanning electron microscopy experiments, we observed that CA can inhibit the formation of CRKP biofilm. On the quantitative real-time PCR analysis, the expression of the luxS, mrkA and wbbm genes in most CRKP strains appeared downregulated because of the CA treatment. Besides, CA significantly inhibited the effect of AI-2 activity of BB170. Our study suggests that CA can be an effective antimicrobial, antivirulent compound that can target QS in hv-CRKP infections, thus providing a new therapeutic direction for treating bacterial infections.

Electron cryo-microscopy reveals the structure of the archaeal thread filament

Pili are filamentous surface extensions that play roles in bacterial and archaeal cellular processes such as adhesion, biofilm formation, motility, cell-cell communication, DNA uptake and horizontal gene transfer. The model archaeaon Sulfolobus acidocaldarius assembles three filaments of the type-IV pilus superfamily (archaella, archaeal adhesion pili and UV-inducible pili), as well as a so-far uncharacterised fourth filament, named “thread”. Here, we report on the cryo-EM structure of the archaeal thread. The filament is highly glycosylated and consists of subunits of the protein Saci_0406, arranged in a head-to-tail manner. Saci_0406 displays structural similarity, but low sequence homology, to bacterial type-I pilins. Thread subunits are interconnected via donor strand complementation, a feature reminiscent of bacterial chaperone-usher pili. However, despite these similarities in overall architecture, archaeal threads appear to have evolved independently and are likely assembled by a distinct mechanism.

Doxycycline combined manure microbes to enhances biofilm formation of the soil plastisphere and increases the surface bio-risk of microplastics vehicle

Microbial biofilms can protect bacteria against environmental stresses, such as antibiotics. Microplastics (MPs) provide attachment sites for microbes to form biofilms in aquatic systems; however, in terrestrial systems, research is still lacking, especially in croplands under fertilization and antibiotic exposure. The present study revealed the effects of different doxycycline (DOX) concentrations on the soil plastisphere by examining biofilm formation regularity and the risk posed by MP-attached strains during manuring. The results showed that soil MP biofilm formation could be enhanced by DOX exposure, especially in low-DOX-concentration group over 60 days, and the biofilm formation gene (BFG) expression level on MPs was stronger in low-DOX-exposure group. DOX exposure increased the abundance of most tet genes on the MPs, and the intl1 abundance was highly increased in the DOX exposure groups (enhanced to 1.07 × 108–8.37 × 107 copies/g). DOX exposure increased the diversity of bacteria that were both potential antibiotic resistance gene (ARG) hosts and biofilm-producing strains on MPs, and low DOX exposure enhanced the abundance of potential pathogens on MPs. The correlations among biofilms, ARGs and DOX on soil MPs were promoted by DOX exposure. Further experiments also proved that both MPs and DOX existing enhanced the biofilm production ability of the strains, and the biofilm production ability of the strains attached to the MPs was stronger than that of planktonic cells. This study revealed that DOX exposure, typically low concentrations of DOX, could increase ARG transfer and the abundance of residual pathogens on MPs through biofilm production and the associated protection during manuring.

Association between antibiotic resistance, biofilm formation and lasB gene in Pseudomonas aeruginosa isolated from different clinical specimens.

Association between antibiotic resistance, biofilm formation and lasB gene in Pseudomonas aeruginosa isolated from different clinical specimens.

Analysis of the Antimicrobial and Anti-Biofilm Activity of Natural Compounds and Their Analogues against Staphylococcus aureus Isolates.

(1) Background: Staphylococcus aureus (S. aureus) is one of the most frequent causes of biofilm-associated infections. With the emergence of antibiotic-resistant, especially methicillin-resistant S. aureus (MRSA), there is an urgent need to discover novel inhibitory compounds against this clinically important pathogen. In this study, we evaluated the antimicrobial and anti-biofilm activity of 11 compounds, including phenyl propenes and phenolic aldehydes, eugenol, ferulic acid, sinapic acid, salicylaldehyde, vanillin, cinnamoyl acid, and aldehydes, against drug-resistant S. aureus isolates. (2) Methods: Thirty-two clinical S. aureus isolates were obtained from Alkhidmat Diagnostic Center and Blood Bank, Karachi, Pakistan, and screened for biofilm-forming potential, and susceptibility/resistance against ciprofloxacin, chloramphenicol, ampicillin, amikacin, cephalothin, clindamycin, streptomycin, and gentamicin using the Kirby-Bauer disk diffusion method. Subsequently, 5 representative clinical isolates were selected and used to test the antimicrobial and anti-biofilm potential of 11 compounds using both qualitative and quantitative assays, followed by qPCR analysis to examine the differences in the expression levels of biofilm-forming genes (ica-A, fnb-B, clf-A and cna) in treated (with natural compounds and their derivatives) and untreated isolates. (3) Results: All isolates were found to be multi-drug resistant and dominant biofilm formers. The individual Minimum Inhibitory Concentration (MIC) of natural compounds and their analogues ranged from 0.75–160 mg/mL. Furthermore, the compounds, Salicylaldehyde (SALI), Vanillin (VAN), α-methyl-trans-cinnamaldehyde (A-MT), and trans-4-nitrocinnamic acid (T4N) exhibited significant (15–92%) biofilm inhibition/reduction percentage capacity at the concentration of 1–10 mg/mL. Gene expression analysis showed that salicylaldehyde, α-methyl-trans-cinnamaldehyde, and α-bromo-trans-cinnamaldehyde resulted in a significant (p < 0.05) downregulation of the expression of ica-A, clf-A, and fnb-A genes compared to the untreated resistant isolate. (4) Conclusions: The natural compounds and their analogues used in this study exhibited significant antimicrobial and anti-biofilm activity against S. aureus. Biofilms persist as the main concern in clinical settings. These compounds may serve as potential candidate drug molecules against biofilm forming S. aureus.

Culture Supernatant of Enterococcus faecalis Promotes the Hyphal Morphogenesis and Biofilm Formation of Candida albicans.

(1) Background: Enterococcus faecalis and Candida albicans are often isolated from infected root canals. The interaction between these two species is not clear enough. Therefore, the objective of this study was to investigate the effect of E. faecalis on the biofilm formation, hyphal morphogenesis and virulence gene expression of C. albicans. (2) Methods: We used the culture supernatant of E. faecalis (CSE) to treat the biofilms of C. albicans. Then, crystal violet staining and colony-forming unit (CFU) counting were performed to evaluate biofilm biomass. Scanning electron microscopy (SEM) and confocal laser scanning microscope (CLSM) were applied to observe fungal morphology. Subsequently, exopolymeric substances (EPS) production, cellular surface hydrophobicity (CSH) and adhesion force of biofilms were investigated by CLSM, water–hydrocarbon two-phase assay and atomic force microscopy (AFM), respectively. Finally, the expression of C. albicans virulence genes (ALS1, ALS3, HWP1 and EFG1) were measured by RT-qPCR assay. (3) Results: The exposure of CSE promoted the biofilm formation and hyphal morphogenesis of C. albicans, increased the EPS production, CSH and adhesion force of C. albicans biofilms, and increased the expression level of EFG1. (4) Conclusions: Our data indicated that CSE promoted the hyphal morphogenesis and biofilm formation of C. albicans.

Inhibition of growth, biofilm formation, virulence, and surface attachment of Agrobacterium tumefaciens by cinnamaldehyde derivatives.

Agrobacterium tumefaciens, a soil-borne, saprophytic plant pathogen that colonizes plant surfaces and induces tumors in a wide range of dicotyledonous plants by transferring and expressing its T-DNA genes. The limited availabilities and efficacies of current treatments necessitate the exploration of new anti-Agrobacterium agents. We examined the effects of trans-cinnamaldehyde (t-CNMA) and its derivatives on the cell surface hydrophobicity, exopolysaccharide and exo-protease production, swimming motility on agar, and biofilm forming ability of A. tumefaciens. Based on initial biofilm inhibition results and minimum inhibitory concentration (MIC) data, 4-nitro, 4-chloro, and 4-fluoro CNMAs were further tested. 4-Nitro, 4-chloro, and 4-fluoro CNMA at ≥150 μg/ml significantly inhibited biofilm formation by 94–99%. Similarly, biofilm formation on polystyrene or nylon was substantially reduced by 4-nitro and 4-chloro CNMAs as determined by optical microscopy and scanning electron microscopy (SEM) and 3-D spectrum plots. 4-Nitro and 4-chloro CNMAs induced cell shortening and concentration- and time-dependently reduced cell growth. Virulence factors were significantly and dose-dependently suppressed by 4-nitro and 4-chloro CNMAs (P ≤ 0.05). Gene expressional changes were greater after 4-nitro CNMA than t-CNMA treatment, as determined by qRT-PCR. Furthermore, some genes essential for biofilm formation, motility, and virulence genes significantly downregulated by 4-nitro CNMA. Seed germination of Raphanus sativus was not hindered by 4-nitro or 4-fluoro CNMA at concentrations ≤200 μg/ml, but root surface biofilm formation was severely inhibited. This study is the first to report the anti-Agrobacterium biofilm and anti-virulence effects of 4-nitro, 4-chloro, and 4-fluoro CNMAs and t-CNMA and indicates that they should be considered starting points for the development of anti-Agrobacterium agents.

Biofilms as a microbial hazard in the food industry: A scoping review.

Biofilms pose a serious public health hazard with a significant economic impact on the food industry. The present scoping review is designed to analyse the literature published during 2001-2020 on biofilm formation of microbes, their detection methods, and association with antimicrobial resistance (if any). The peer-reviewed articles retrieved from 04 electronic databases were assessed using PRISMA-ScR guidelines. From the 978 preliminary search results, a total of 88 publications were included in the study. On analysis, the commonly isolated pathogens were Listeria monocytogenes, Staphylococcus aureus, Salmonella spp., Escherichia coli, Bacillus spp., Vibrio spp., Campylobacter jejuni and Clostridium perfringens. The biofilm-forming ability of microbes was found to be influenced by various factors such as attachment surfaces, temperature, presence of other species, nutrient availability etc. A total of 18 studies characterized the biofilm-forming genes, particularly for S. aureus, Salmonella spp., and E. coli. In most studies, polystyrene plate and/or stainless-steel coupons were used for biofilm formation, and the detection was carried out by crystal violet assays and/or by plate counting method. The strain-specific significant differences in biofilm formation were observed in many studies, and few studies carried out analysis of multi-species biofilms. The association between biofilm formation and antimicrobial resistance was not clearly defined. Further, viable but non-culturable form of the foodborne pathogens is posing an unseen (by conventional cultivation techniques) but potent threat to the food safety. The present review recommends the need for carrying out systematic surveys and risk analysis of biofilms in food chain to highlight the evidence-based public health concerns, especially in regions where microbiological food hazards are quite prevalent.