Mechanisms Of Biofilm Formation Research Articles

Improved mini-Tn7 Delivery Plasmids for Fluorescent Labeling of Stenotrophomonas maltophilia.

Fluorescently labeled bacterial cells have become indispensable for many aspects of microbiological research, including studies on biofilm formation as an important virulence factor of various opportunistic bacteria of environmental origin such as Stenotrophomonas maltophilia. Using a Tn7-based genomic integration system, we report the construction of improved mini-Tn7 delivery plasmids for labeling of S. maltophilia with sfGFP, mCherry, tdTomato and mKate2 by expressing their codon-optimized genes from a strong, constitutive promoter and an optimized ribosomal binding site. Transposition of the mini-Tn7 transposons into single neutral sites located on average 25 nucleotides downstream of the 3'-end of the conserved glmS gene of different S. maltophilia wild-type strains did not have any adverse effects on the fitness of their fluorescently labeled derivatives. This was demonstrated by comparative analyses of growth, resistance profiles against 18 antibiotics of different classes, the ability to form biofilms on abiotic and biotic surfaces, also independent of the fluorescent protein expressed, and virulence in Galleria mellonella. It is also shown that the mini-Tn7 elements remained stably integrated in the genome of S. maltophilia over a prolonged period of time in the absence of antibiotic selection pressure. Overall, we provide evidence that the new improved mini-Tn7 delivery plasmids are valuable tools for generating fluorescently labeled S. maltophilia strains that are indistinguishable in their properties from their parental wild-type strains. IMPORTANCE The bacterium S. maltophilia is an important opportunistic nosocomial pathogen that can cause bacteremia and pneumonia in immunocompromised patients with a high rate of mortality. It is now considered as a clinically relevant and notorious pathogen in cystic fibrosis patients but has also been isolated from lung specimen of healthy donors. The high intrinsic resistance to a wide range of antibiotics complicates treatment and most likely contributes to the increasing incidence of S. maltophilia infections worldwide. One important virulence-related trait of S. maltophilia is the ability to form biofilms on any surface, which may result in the development of increased transient phenotypic resistance to antimicrobials. The significance of our work is to provide a mini-Tn7-based labeling system for S. maltophilia to study the mechanisms of biofilm formation or host-pathogen interactions with live bacteria under non-destructive conditions.

Attachment characteristics and kinetics of biofilm formation by Staphylococcus aureus on ready-to-eat cooked beef contact surfaces.

Staphylococcus aureus is a food-borne pathogen that quickly forms biofilm on meat contact surfaces and thus poses a serious threat to the safety of the meat industry. This study evaluated the attachment, survival, and growth of S. aureus biofilm with exposure to environmental factors in the meat industry by simulated ready-to-eat (RTE) cooked beef product contamination scenarios. The results indicated that the meat-borne S. aureus biofilm formation dynamic could be divided into four different phases: initial adhesion (4-12h), exponential (12-24h), slow growth (1-3 days), and stationary (3-7 days). Meat-borne S. aureus has strong adhesion and biofilm formation ability, and its biofilm exhibits persistence, high-intensity metabolic activity, aerotaxis, and strain heterogeneity. This study has also demonstrated that in the long-term existence of meat-borne S. aureus biofilm on stainless steel and plexiglass surfaces(>7 days, 7.2-8.8 log CFU/cm2 ), expose to RTE cooked beef products, may cause it to become high-risk contaminated food. Meat-borne S. aureus that forms a dense and rough concave-convex in the shape of biofilm architecture was observed by scanning electron microscopy, consisting of complex components and adhesion of living and dead cells. This was further confirmed by the meat-borne S. aureus biofilm on the stainless steel surface by attenuated total reflectance Fourier transformed infrared spectroscopy, and the dominant peaks in biofilm spectra were mainly associated with proteins, polysaccharides, amino acid residues, and phospholipids(>50%). These findings may help in the identification of the main sources of contamination within the meat industry and the subsequent establishment of strategies for biofilm prevention and removal. PRACTICAL APPLICATION: This study revealed the meat-borne S. aureus biofilm formation mechanism and found that it exhibited strong colonization and biofilm-forming ability, which can persist on the contact surfaces of ready-to-eat beef products. These initial findings could provide information on the behavior of meat-borne S. aureus biofilm attached to meat contact surfaces under conditions commonly encountered in meat environments, which help to support the determination of the main sources of contamination within the meat industry and the subsequent establishment of strategies for biofilm prevention and removal. It was also helpful in controlling biofilm contamination and improving meat safety to minimize it.

Influence of interspecies interactions on biomass and extracellular polymeric substances of bacterial biofilms

BackgroundStudies with emphasis on pure and mixed-species biofilms are of significant importance in understanding biofilm formation mechanisms during microbial infections. This research aims to evaluate pure- and dual-species biofilms of Escherichia coli (code A), Staphylococcus aureus (code B), Klebsiella pneumoniae (code C) and Pseudomonas aeruginosa (code D) pathogenic bacterial species and their production of biofilm exopolysaccharides at laboratory scale. MethodsBiofilm biomass (A595) of pure- and dual-species cultures was determined by means of a microtiter plate assay in triplicate using a microplate photometer (Fisher Scientific, type-357). Extracellular polymeric substances (EPS) and soluble microbial products (SMP) were extracted from the biofilm cells (pure- and dual-species cultures) using the alkaline-heat extraction method. Dry weights (g/L) of EPS and SMP were determined by drying the samples at 105 °C for 8 hours. ResultsKlebsiella pneumoniae biofilm biomass accounted for a 28-72% greater biofilm biomass than the other bacteria. Experimental values of dual-species biofilm biomasses were in the range of 6% to 30% over theoretical values. The experimental value of one dual-species (bacteria B + D) biofilm biomass was 30% higher than its expected value. Decrease or increase in the dual-species biofilm biomass of either bacteria A+C or bacteria B+C combinations was totally dependent on the cell density of bacteria C. Conclusions Biofilm biomass of pure-species cultures was totally species-dependent, and the biofilm biomass of four species was in the following order: bacteria C > D > A > B. Relation between biofilm biomasses and SMP or EPS was inconsistent.

Effects of organic carbon sources on algal biofilm formation and insight into mechanism

Algal biofilm cultivation has great potential in producing high biomass yields and reducing harvesting costs. Organic carbon sources can promote algal growth in suspended cultivation, but their effects on algal biofilm formation and related mechanism have rarely been studied. In this study, we determined the most suitable organic carbon source and its concentration for the formation of algal biofilm, and structure/composition characteristics of extracellular polymeric substances (EPS) from algal biofilm. The results showed that the glycerol concentration of 1000 mg C L−1 in algal biofilm system was the most suitable condition to contribute to the biofilm formation and enhance attached biomass among the treatments. The algal biofilm at the glycerol concentration excreted the contents of relatively high EPS and its protein (EPS-PN), which were beneficial to forming the biofilm. The high EPS-PN content was mainly attributed to the increased hydrophobic proline and alanine contents but declined hydrophilic glutamic acid for improving algal biofilm formation. These findings enhanced our understanding of the formation mechanism of algal biofilm under the organic carbon source condition and provided the theoretical basis for algal biofilm formation.

The characteristic evolution and formation mechanism of hybrid microalgae biofilm and its application in mariculture wastewater treatment

Microalgae biofilm had important applications in both energy and environmental fields. However, the current research on the formation mechanism of microalgae biofilm was vague. And the treatment of mariculture wastewater by microalgae biofilm was rare. Herein, a kind of natural hybrid microalgae was applied in this study to investigate the mechanism of microalgae biofilm formation and the treatment of mariculture wastewater under different ammonia nitrogen loads (13 mg/L, 25 mg/L, 37 mg/L). The results indicated that filamentous algae were first to attach to the carrier and then formed a web-like structure. Later, more non-filamentous algae and bacteria involved in the formation of biofilm. Protein in extracellular polymeric substances (EPS) was the key substance for the initial colonization. The NN functional group, tryptophan, high molecular weight of EPS promoted microalgae biofilm formation. When the ammonia nitrogen concentration was 25 mg/L, the microalgae biofilm reached the maximum biomass (1.79 mg/cm2) and the highest nitrogen and phosphorus removal efficiency (98% of NH4+-N, 84% of TN, 68% of PO4--P). The study provides a theoretical basis for understanding the mechanism of natural hybrid microalgae biofilm formation and its application for mariculture wastewater treatment.

Transcriptome analysis of the biofilm formation mechanism of Vibrio parahaemolyticus under the sub-inhibitory concentrations of copper and carbenicillin

Biofilm formation of Vibrio parahaemolyticus enhanced its tolerance to the environment, but caused many serious problems to food safety and human health. In this paper, the effects of copper and carbenicillin (CARB) stress on the formation of the biofilms of V. parahaemolyticus organisms were studied, and RNA sequencing technology was used to compare the differences in transcriptome profiles of the biofilm-related genes of V. parahaemolyticus organisms under different sub-inhibitory stresses. The results proved that V. parahaemolyticus had a large growth difference under the two stresses, copper and CARB at 1/2 minimal inhibitory concentration (MIC), and it could form a stable biofilm under both stress conditions. The amount of biofilm formed under CARB stress was significantly higher than that of copper stress (p < 0.05). Based on the analysis of transcriptome sequencing results 323, 1,550, and 1,296 significantly differential expressed genes were identified in the three treatment groups namely 1/2 MIC CARB, Cu2+, and Cu2++CARB. Through COG annotation, KEGG metabolic pathway analysis and gene expression analysis related to biofilm formation, the functional pathways of transcriptome changes affecting V. parahaemolyticus were different in the three treatment groups, and the CARB treatment group was significantly different from the other two groups. These differences indicated that the ABC transport system, two-component system and quorum sensing were all involved in the biofilm formation of the V. parahaemolytic by regulating flagellar motility, extracellular polysaccharides and extracellular polymer synthesis. Exploring the effects of different stress conditions on the transcriptome of V. parahaemolyticus could provide a basis for future research on the complex network system that regulates the formation of bacterial biofilms.

Comparative genome identification of accessory genes associated with strong biofilm formation in Vibrio parahaemolyticus

Vibrio parahaemolyticus biofilms on the seafood processing plant surfaces are a potential source of seafood contamination and subsequent food poisoning. Strains differ in their ability to form biofilm, but little is known about the genetic characteristics responsible for biofilm development. In this study, pangenome and comparative genome analysis of V. parahaemolyticus strains reveals genetic attributes and gene repertoire that contribute to robust biofilm formation. The study identified 136 accessory genes that were exclusively present in strong biofilm forming strains and these were functionally assigned to the Gene Ontology (GO) pathways of cellulose biosynthesis, rhamnose metabolic and catabolic processes, UDP-glucose processes and O antigen biosynthesis (p < 0.05). Strategies of CRISPR-Cas defence and MSHA pilus-led attachment were implicated via Kyoto Encyclopedia of Genes and Genomes (KEGG) annotation. Higher levels of horizontal gene transfer (HGT) were inferred to confer more putatively novel properties on biofilm-forming V. parahaemolyticus. Furthermore, cellulose biosynthesis, a neglected potential virulence factor, was identified as being acquired from within the order Vibrionales. The cellulose synthase operons in V. parahaemolyticus were examined for their prevalence (22/138, 15.94 %) and were found to consist of the genes bcsG, bcsE, bcsQ, bcsA, bcsB, bcsZ, bcsC. This study provides insights into robust biofilm formation of V. parahaemolyticus at the genomic level and facilitates: identification of key attributes for robust biofilm formation, elucidation of biofilm formation mechanisms and development of potential targets for novel control strategies of persistent V. parahaemolyticus.

Molecular mechanisms of biofilm formation in Staphylococci bacteria: How antibiotic resistant bacteria is evolving to become more resilient to mechanical force loads.

Among the arsenal of virulence factors used by Staphylococcus aureus bacteria, adhesins play critical roles participating actively on the formation of biofilms. Combining single molecule force spectroscopy in vitro and in silico, we have discovered the extreme mechanostability of the interaction between pathogenic adhesins and proteins of the human extracellular matrix. Here we describe how Staphylococci adhesins use a catch-bond mechanism to hold tight to their human target peptides under high-force loads, while allowing for thermal dissociation in a low-force regime. Furthermore, we employed bioinformatic tools to retrieve and align nearly 200 proteins of the bacterial adhesin superfamily. Using AI-based protein structure prediction, we modelled, together with their human target, adhesins from methicillin susceptible strains of S. aureus (MSSA) and methicillin resistant strains of S. aureus (MRSA). Using GPU-accelerated NAMD 3, thousands of steered molecular dynamics (SMD) simulations were performed using a wide-sampling paradigm. Our protocol reveled that mechanostability is preserved among all strains with a pattern of higher forces for the MRSA strains. In summary, our work shows that the complex formed by Staphylococci adhesins with proteins of the human extracellular matrix is, by far, the most mechanically stable protein complex know to date, surpassing the mechanostability of the widely employed streptavidin:biotin complex by an order of magnitude. To keep such mechanostable complex, a catch-bond mechanism, where bonds live longer under mechanical load, is used by these adhesins. Additionally, we have recently discovered that the mechanostability of the complex is increasing with the evolution of these bacteria to become resistant to methicillin. With increasing prevalence of multidrug resistant bacterial infections, this new finding could be exploited for the development of antiadhesion strategies as an alternative to antibiotics.

Extensively drug-resistant Acinetobacter baumannii: role of conjugative plasmids in transferring resistance.

Acinetobacter baumannii is one of the most successful pathogens that can cause difficult-to-treat nosocomial infections. Outbreaks and infections caused by multi-drug resistant A. baumannii are prevalent worldwide, with only a few antibiotics are currently available for treatments. Plasmids represent an ideal vehicle for acquiring and transferring resistance genes in A. baumannii. Five extensively drug-resistant A. baumannii clinical isolates from three major Jordanian hospitals were fully sequenced. Whole-Genome Sequences (WGS) were used to study the antimicrobial resistance and virulence genes, sequence types, and phylogenetic relationship of the isolates. Plasmids were characterized In-silico, followed by conjugation, and plasmid curing experiments. Eight plasmids were recovered; resistance plasmids carrying either aminoglycosides or sulfonamide genes were detected. Chromosomal resistance genes included blaOXA-66, blaOXA-91, and blaOXA-23, and the detected virulence factors were involved in biofilm formation, adhesion, and many other mechanisms. Conjugation and plasmid curing experiments resulted in the transfer or loss of several resistance phenotypes. Plasmid profiling along with phylogenetic analyses revealed high similarities between two A. baumannii isolates recovered from two different intensive care units (ICU). The high similarities between the isolates of the study, especially the two ICU isolates, suggest that there is a common A. baumannii strain prevailing in different ICU wards in Jordanian hospitals. Three resistance genes were plasmid-borne, and the transfer of the resistance phenotype emphasizes the role and importance of conjugative plasmids in spreading resistance among A. baumannii clinical strains.

Treatment of nitrogen-rich wastewater by mixed aeration combined with bioaugmentation in a sequencing batch biofilm reactor: Biofilm formation and nitrogen-removal capacity analysis

The mixed aerated and bioaugmented sequencing batch biofilm reactor (SBBR) was developed to improve the efficiency of nitrogen-rich wastewater treatment, and the mechanism of biofilm formation and nitrogen removal was analyzed. The results showed that the addition of aerobic denitrifiers could shorten the start-up time and significantly improve the biofilm formation ability of SBBRs. The presence of nanobubbles could make the biofilm more compact and secrete more EPS, which promoted the activity and stability of the biofilm. During the stable operation stage, the bioaugmented SBBR under mixed aeration mode showed the best nitrogen removal performance and the removal rates of NH4+-N (95.65) and TN (94.88%） were increased by 19.86% and 14.36% compared with the control group, respectively. Microbial community structure and nitrogen metabolism pathway analysis revealed that strain ZS1 colonized the reactor. Hyphomonas, Defluviicoccus and Hydrogenophaga played important roles in N removal in the mixed aerated and bioaugmented SBBR. Meanwhile, the suitable oxidative environment created by mixed aeration made autotrophic nitrifiers（mainly Nitrospira and Nitrosomonas）and aerobic denitrifiers (mainly Hydrogenophaga, Hyphomonas and Pseudofulvimonas) coexist successfully, increasing the relative abundance of the key functional genes (amoABC, hao, napAB genes) in nitrification and denitrification processes to improve the N removal performance. Furthermore, mixed aeration combined with bioaugmented stimulated the nirA, gltBD and GLUD genes, thus promoting the process of nitrogen assimilation and providing a pathway of the direct removal of NH4+-N.

Non-deacetylated poly-N-acetylglucosamine-hyperproducing Staphylococcus aureus undergoes immediate autoaggregation upon vortexing.

Biofilms are microbial communities of cells embedded in a matrix of extracellular polymeric substances generated and adhering to each other or to a surface. Cell aggregates formed in the absence of a surface and floating pellicles that form biofilms at the air-liquid interface are also considered to be a type of biofilm. Staphylococcus aureus is a well-known cause of biofilm infections and high-molecular-weight polysaccharides, poly-N-acetylglucosamine (PNAG) is a main constituent of the biofilm. An icaADBC operon comprises major machinery to synthesize and extracellularly secrete PNAG. Extracellular PNAG is partially deacetylated by IcaB deacetylase, and the positively charged PNAG hence interacts with negatively charged cell surface to form the major component of biofilm. We previously reported a new regulator of biofilm (Rob) and demonstrated that Rob binds to a unique 5-bp motif, TATTT, present in intergenic region between icaADBC operon and its repressor gene icaR in Yu et al. The deletion of the 5-bp motif induces excessive adherent biofilm formation. The real function of the 5-bp motif is still unknown. In an attempt to isolate the 5-bp motif deletion mutant, we isolated several non-adherent mutants. They grew normally in turbid broth shaking culture but immediately auto-aggregated upon weak vortexing and sedimented as a lump resulting in a clear supernatant. Whole genome sequencing of the mutants identified they all carried mutations in icaB in addition to deletion of the 5-bp motif. Purification and molecular characterization of auto-aggregating factor in the culture supernatant of the mutant identified that the factor was a massively produced non-deacetylated PNAG. Therefore, we created a double deficient strain of biofilm inhibitory factors (5-bp motif, icaR, rob) and icaB to confirm the aggregation phenomenon. This peculiar phenomenon was only observed in Δ5bpΔicaB double mutant but not in ΔicaR ΔicaB or ΔrobΔicaB mutant. This study explains large amount of extracellularly produced non-deacetylated PNAG by Δ5bpΔicaB double mutation induced rapid auto-aggregation of S. aureus cells by vortexing. This phenomenon indicated that Staphylococcus aureus may form biofilms that do not adhere to solid surfaces and we propose this as a new mechanism of non-adherent biofilm formation of S. aureus.

Mixotrophy broadens the ecological niche range of the iron oxidizerSideroxydanssp. CL21 isolated from an iron-rich peatland

Sideroxydans sp. CL21 is a microaerobic, acid-tolerant Fe(II)-oxidizer, isolated from the Schlöppnerbrunnen fen. Since the genome size of Sideroxydans sp. CL21 is 21% larger than that of the neutrophilic Sideroxydans lithotrophicus ES-1, we hypothesized that strain CL21 contains additional metabolic traits to thrive in the fen. The common genomic content of both strains contains homologs of the putative Fe(II) oxidation genes, mtoAB and cyc2. A large part of the accessory genome in strain CL21 contains genes linked to utilization of alternative electron donors, including NiFe uptake hydrogenases, and genes encoding lactate uptake and utilization proteins, motility and biofilm formation, transposable elements, and pH homeostasis mechanisms. Next, we incubated the strain in different combinations of electron donors and characterized the fen microbial communities. Sideroxydans spp. comprised 3.33% and 3.94% of the total relative abundance in the peatland soil and peatland water, respectively. Incubation results indicate Sideroxydans sp. CL21 uses H2 and thiosulfate, while lactate only enhances growth when combined with Fe, H2, or thiosulfate. Rates of H2 utilization were highest in combination with other substrates. Thus, Sideroxydans sp. CL21 is a mixotroph, growing best by simultaneously using substrate combinations, which helps to thrive in dynamic and complex habitats.

Fungistatic effect of Moringa oleifera Lam. on the metabolism changes of Candida albicans

Context: Candida albicans is a pathological agent that triggers oral candidiasis because C. albicans can adapt to changes to increase growth and adhesion through biofilm formation mechanisms. Moringa oleifera Lam. has been reported to have fungistatic properties and increases the metabolism change of C. albicans. Aims: To evaluate the fungistatic effect of M. oleifera leaves ethanolic extract (MOLE) on the metabolism changes of C. albicans cells associated with growth and biofilm formation. Methods: The assessment of metabolism changes (stress response and metabolic alterations) of C. albicans by the action of MOLE was performed by mean of FTIR, growth assessment by spectrophotometry, biofilm formation with 1% crystal violet, also read by spectrophotometry, and observation of biofilm mass with a microscope. Results: MOLE showed substantial absorption values based on topological polar surface area (&lt;140 Å2). Concentrations of 25% and 6.25% of MOLE increased the stress response (metabolism changes) of C. albicans (66-75%), meanwhile 50% and nystatin (100.000 IU/mL) were similar in inducing metabolism changes of C. albicans. All concentrations of M. oleifera could inhibit the growth of C. albicans at all incubation times (24, 48, and 72 h) with an Optical Density (OD) of 0.02-0.05 (&lt;300 CFU/mL) and were able to degrade the biofilm formation of C. albicans on a scale substantial at 24 and 48 h (OD&gt;0.4), and moderate scale at 72 h (OD 0.2-0.39). Conclusions: The extract of M. oleifera has increased metabolism changes (stress response) of C. albicans cells, which correlate with the ability to inhibit growth and biofilm formation for 24, 48, and 72 h.

Abstract 2722: Mechanisms of host adhesion and biofilm formation mediated by the staphylococcal surface protein Aap

Abstract 2722: Mechanisms of host adhesion and biofilm formation mediated by the staphylococcal surface protein Aap

Mechanism of biofilm formation in ESKAPE organism and their treatment

One of the major global issues is the urgency of nosocomial infections, biofilm development, and antibiotic resistance. The ESKAPE pathogens are a significant factor in these problems (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter species). These pathogens, often known as hospital-acquired infections (HAIs), are common in hospital settings and present one of the most significant difficulties in treatment. The development of biofilms, in which the microbial cells adapt to a multicellular lifestyle by getting trapped inside the extracellular polymeric matrix, is a key biological concept in clinical contexts. For the creation of novel antimicrobial agents, as well as for the repurposing of currently accessible medications or pre-clinical substances and the broader use of combination therapies, it is essential to comprehend the process by which these bacteria build biofilms. The Pathomechanisms of these bacterial biofilm formations and alternate strategies to prevent biofilm formation in hospital management are highlighted in this review.

Microbial Biofilms for Environmental Bioremediation of Heavy Metals: a Review.

Heavy metal pollution caused due to various industrial and mining activities poses a serious threat to all forms of life in the environment because of the persistence and toxicity of metal ions. Microbial-mediated bioremediation including microbial biofilms has received significant attention as a sustainable tool for heavy metal removal as it is considered safe, effective, and feasible. The biofilm matrix is dynamic, having microbial cells as major components with constantly changing and evolving microenvironments. This review summarizes the bioremediation potential of bacterial biofilms for different metal ions. The composition and mechanism of biofilm formation along with interspecies communication among biofilm-forming bacteria have been discussed. The interaction of biofilm-associated microbes with heavy metals takes place through a variety of mechanisms. These include biosorption and bioaccumulation in which the microbes interact with the metal ions leading to their conversion from a highly toxic form to a less toxic form. Such interactions are facilitated via the negative charge of the extracellular polymeric substances on the surface of the biofilm with the positive charge of the metal ions and the high cell densities and high concentrations of cell-cell signaling molecules within the biofilm matrix. Furthermore, the impact of the anodic and cathodic redox potentials in a bioelectrochemical system (BES) for the reduction, removal, and recovery of numerous heavy metal species provides an interesting insight into the bacterial biofilm-mediated bioelectroremediation process. The review concludes that biofilm-linked bioremediation is a viable option for the mitigation of heavy metal pollution in water and ecosystem recovery.

A Review of Biofilm Formation of Staphylococcus aureus and Its Regulation Mechanism.

Bacteria can form biofilms in natural and clinical environments on both biotic and abiotic surfaces. The bacterial aggregates embedded in biofilms are formed by their own produced extracellular matrix. Staphylococcus aureus (S. aureus) is one of the most common pathogens of biofilm infections. The formation of biofilm can protect bacteria from being attacked by the host immune system and antibiotics and thus bacteria can be persistent against external challenges. Therefore, clinical treatments for biofilm infections are currently encountering difficulty. To address this critical challenge, a new and effective treatment method needs to be developed. A comprehensive understanding of bacterial biofilm formation and regulation mechanisms may provide meaningful insights against antibiotic resistance due to bacterial biofilms. In this review, we discuss an overview of S. aureus biofilms including the formation process, structural and functional properties of biofilm matrix, and the mechanism regulating biofilm formation.

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.

Phenylalanine Promotes Biofilm Formation of Meyerozyma caribbica to Improve Biocontrol Efficacy against Jujube Black Spot Rot.

During storage and transportation after harvest, the jujube fruit is susceptible to black spot rot, which is caused by Alternaria alternata. The present study aimed to evaluate the effectiveness of the yeast Meyerozyma caribbica in controlling A. alternata in postharvest jujube fruits, and to explore the biofilm formation mechanism. The results showed that M. caribbica treatment significantly reduced the A. alternata decay in jujube fruits. M. caribbica could rapidly colonize jujube fruit wounds, adhering tightly to hyphae of A. alternata, and accompanied by the production of extracellular secretions. In in vitro experiments, we identified that M. caribbica adhered to polystyrene plates, indicating a strong biofilm-forming ability. Furthermore, we demonstrated that M. caribbica can secrete phenylethanol, a quorum sensing molecule which can affect biofilm development. Phenylalanine (a precursor substance for phenylethanol synthesis) enhanced the secretion of phenylethanol and promoted the formation of M. caribbica biofilms. Meanwhile, phenylalanine enhanced the biological control performance of M. caribbica against jujube black spot rot. Our study provided new insights that enhance the biological control performance of antagonistic yeast.

Directions in the development of modern and promising antimicrobial agents

The use of antibiotics is becoming increasingly limited. This is mainly due to the development of resistance to pathogenic bacteria, and, over time, more and more bacteria will become resistant to antimicrobials. This problem inevitably leads to the conclusion that studies into alternative methods of combating pathogens, which are necessary to develop sufficiently reliable and effective therapies for bacterial infections, are indispensable. This review highlights some recent developments in conventional antibiotic and non-antibiotic treatment strategies. It has been shown that traditional antibacterial targets include derivatives of known antibiotic classes, new chemical classes with new targets, as well as unknown or undefined agents with unclear targets. Promising strategies for combating microbial pathogens have been identified, including new targets, namely, toxin secretion systems, biofilm formation, and adhesion mechanisms that affect quorum sensing of microbial populations. In addition, it is important to use new antimicrobial agents with other, non-antibiotic, mechanisms of action: phage and phage-derived peptides, microbiota-modulating therapies, and enhancing immune response.