Biofilm Formation Ability Research Articles

Transcriptomic Analyses to Unravel Cronobacter sakazakii Resistance Pathways.

The proliferation of antibiotic usage has precipitated the emergence of drug-resistant variants of bacteria, thereby augmenting their capacity to withstand pharmaceutical interventions. Among these variants, Cronobacter sakazakii (C. sakazakii), prevalent in powdered infant formula (PIF), poses a grave threat to the well-being of infants. Presently, global contamination by C. sakazakii is being observed. Consequently, research endeavors have been initiated to explore the strain's drug resistance capabilities, alterations in virulence levels, and resistance mechanisms. The primary objective of this study is to investigate the resistance mechanisms and virulence levels of C. sakazakii induced by five distinct antibiotics, while concurrently conducting transcriptomic analyses. Compared to the susceptible strains prior to induction, the drug-resistant strains exhibited differential gene expression, resulting in modifications in the activity of relevant enzymes and biofilm secretion. Transcriptomic studies have shown that the expression of glutathione S-transferase and other genes were significantly upregulated after induction, leading to a notable enhancement in biofilm formation ability, alongside the existence of antibiotic resistance mechanisms associated with efflux pumps, cationic antimicrobial peptides, and biofilm formation pathways. These alterations significantly influence the strain's resistance profile.

Klebsiella Pneumoniae – Taxonomy, Occurrence, Identification, Virulence Factors and Pathogenicity

Abstract Gram-negative bacilli Klebsiella pneumoniae are among the most important pathogens responsible for healthcare-associated infections (HAIs). These bacteria often have high pathogenic and epidemic potential, contributing to infection outbreaks worldwide. K. pneumoniae is part of the natural microbiota of humans. At the same time, as an opportunistic microorganism, when the host organism is weakened, it can cause serious infections such as pneumonia, urinary tract infections, septic infections and intra-organ abscesses. Widespread distribution in nature and exceptional adaptability provide K. pneumoniae with the opportunity to master new niches in the hospital environment, which poses a threat to hospitalized patients. Also, the bacteria are increasingly causing life-threatening infections in the non-hospital environment. The pathogenicity of K. pneumoniae is determined by the presence of many virulence factors such as capsular polysaccharide (CPS, K antigen), lipopolysaccharid (LPS, O antigen), fimbrial and non-fimbrial adhesins, siderophores (aero-bactin, enterobactin, salmochelin and yersiniabactin), heat-stabile and heat-labile enterotoxins, cytotoxins and biofilm-forming ability. Currently, hypervirulent strains of K. pneumoniae (hvKp) equipped with new virulence traits constitute a significant danger. The paper presents these bacteria concerning the global threat arising from the dynamic spread of hvKp strains in hospitals in Poland and worldwide.

Virulence Factors and Susceptibility to Ciprofloxacin, Vancomycin, Triclosan, and Chlorhexidine among Enterococci from Clinical Specimens, Food, and Wastewater.

Enterococcus faecalis and E. faecium are opportunistic pathogens commonly found in the microbiota of humans and other animals as well as in the environment. This article presents the results of antimicrobial susceptibility testing using phenotypic methods (broth microdilution and standardized disk diffusion) on selected clinical, food, and wastewater isolates of E. faecalis and E. faecium. The isolates were divided into subgroups based on their sensitivity to the following antibiotics: vancomycin (VAN) and ciprofloxacin (CIP), and biocides triclosan (TCL) and chlorhexidine (CHX). The study also investigated in vitro virulence factors, including biofilm formation ability, cell surface hydrophobicity (CSH) and β-hemolysis, to explore aspects of pathogenesis. In our study, regardless of the isolation source, VAN-resistant (VAN-R) and CIP-resistant (CIP-R) E. faecalis and E. faecium were detected. The highest proportion of CIP-R strains was found among clinical isolates of E. faecalis and E. faecium, with clinical E. faecium also showing the highest proportion of VAN-R strains. But the highest proportion of VAN-R E. faecalis strains was found in wastewater samples. The highest TCL MIC90 values for E. faecalis were found in wastewater isolates, while for E. faecium, the highest TCL MIC90 values were observed in food isolates. The highest CHX MIC90 values for both E. faecalis and E. faecium were identified in clinical specimens. The results obtained for E. faecalis did not indicate differences in TCL MIC and CHX MIC values with respect to sensitivity to VAN and CIP. Higher CHX MIC50 and CHX MIC90 values were obtained for CIP-R and VAN-R E. faecium. Among the tested isolates, 97.75% of the E. faecalis isolates produced biofilm, while 72.22% of the E. faecium isolates did so as well. In biofilm-forming strength categories III and IV, statistically significantly higher proportions of CIP-susceptible (CIP-S) and VAN-susceptible (VAN-S) E. faecalis were determined. In category III, there is no statistically significant difference in E. faecium CIP sensitivity. In category IV, we had a significantly higher proportion of CIP-R strains. On the other hand, the association between the moderate or strong category of biofilm formation and E. faecium VAN susceptibility was not significant. E. faecalis isolated from wastewater had a CSH index (HI) ≥ 50%, categorizing them as "moderate", while all the other strains were categorized as "low" based on the CSH index. Among the E. faecalis isolates, cell surface hydrophobicity indices differed significantly across isolation sources. In contrast, E. faecium isolates showed similar hydrophobicity indices across isolation sources, with no significant difference found. Moreover, no correlation was found between the enterococcal cell surface hydrophobicity and biofilm formation in vitro. After anaerobic incubation, β-hemolytic activity was confirmed in 19.10% of the E. faecalis and 3.33% of the E. faecium strains.

Trajectory analysis of Bacillus subtilis in micro-droplets.

In order to study Bacillus subtilis biofilm formation in microdroplets, we use microfluidics technology to make the droplets and confocal microscopy to capture bacterial movement and biofilm formation in the droplets. We develop a multi-target tracking methodology, using a YOLOv5 detector to identify cells and a DeepSORT algorithm to track cell movements. We find that Bacillus subtilis bacteria with autonomous migration and biofilm-forming ability prefer to cluster and swarm near the microdroplet surface, rather than in the droplet interior. Bacterial mobility depends on phenotype and spatial location within the droplet. The motile cells move about 3.5 times faster than the matrix-producing cells. When the cells are near the wall of the droplet, the direction of the motion of motile cells is along that wall. When the cells are inside the droplet, the direction of the motion of motile cells is disordered, i.e., there is no clear directional or goal-oriented movement. This contrast increases the cell contact probability and facilitates the formation of a Bacillus subtilis biofilm in the droplet. Furthermore, we develop a mathematical model to describe the motion behavior of Bacillus subtilis in microdroplets, which is useful for exploring the influence of motility on biofilm formation.

The role of aroA and ppk1 in Aeromonas veronii pathogenicity and the efficacy evaluation of mutant strain AV-ΔaroA/ppk1 as a live attenuated vaccine

Aeromonas veronii is an opportunistic pathogen that poses great threat to aquaculture and human health, so there is an urgent need for green and efficient methods to deal with its infection. In this study, single and double gene deletion strains (AV-ΔaroA, AV-Δppk1 and AV-ΔaroA/ppk1) that can be stably inherited were constructed. Pathogenicity test showed that the toxicity of AV-ΔaroA and AV-ΔaroA/ppk1 was significantly lower compared to wild-type A. veronii. Biological characterization analysis revealed that the decrease in pathogenicity might be due to the declined growth, motility, biofilm formation abilities and the expression of virulence-related genes in mutants. Subsequently, we evaluated the efficacy of AV-ΔaroA/ppk1 as a live attenuated vaccine (LAV). Safety assessment experiments showed that AV-ΔaroA/ppk1 injected at a concentration of 3 × 107 CFU/mL was safe for C. carassius. The relative percentage survival of AV-ΔaroA/ppk1 was 67.85 %, significantly higher than that of the inactivated A. veronii, which had an RPS of 54.84 %. This improved protective effect was mainly attributed to the increased levels of A. veronii specific IgM antibody, enhanced alkaline phosphatase, lysozyme and superoxide dismutase activities, as well as higher expression levels of several immune related genes. Together, these findings deepen our understanding of the functional roles of aroA and ppk1 in A. veronii pathogenicity, provide a good candidate of LAV for A. veronii.

Examining the Frequency of Selected Genes Involved in Biofilm Formation in Clinical Isolates of Acinetobacter Baumannii

Introduction: Acinetobacter baumannii, a non-fermenting Gram-negative coccobacillus, exhibits high resistance to antimicrobial compounds. Biofilm formation is a crucial feature in many Acinetobacter species, contributing to their antibiotic resistance. The present study aimed to investigate the prevalence of ompA, csuA, bap and pgaB genes in clinical isolates of Acinetobacter baumannii with biofilm formation ability. Methods: In this cross-sectional study, 49 isolates of Acinetobacter baumannii were collected from patients hospitalized in the health centers of Borujerd City. Acinetobacter baumannii isolates were confirmed by biochemical tests. In these isolates, the biofilm production ability was investigated by microtitreplate method. Then, using PCR method and specific primers, ompA, csuA, bap and pgaB genes were identified.The collected data were analyzed descriptively and analytically using SPSS version 16 software. Data analysis was done with Chi-square test, Fisher's exact test, and P<0.05 was considered as the basis of significance. Results: The presence of ompA, csuA, bap, and pgaB genes was detected in 87%, 92%, 98%, and 100% of the isolates, respectively. Additionally, the microtitreplate method revealed that biofilm formation was strong in 3 isolates (6%), moderate in 17 isolates (35%), and weak in 29 isolates (59%). Conclusion: The prevalence of genes associated with biofilm formation in Acinetobacter baumannii isolates was high. This suggests that the studied isolates possess a significant ability to form biofilm structures.

VPA0198, a GGDEF domain-containing protein, affects the motility and biofilm formation of Vibrio parahaemolyticus and is regulated by quorum sensing associated regulators

Cyclic di-GMP (c-di-GMP), a ubiquitous secondary messenger in bacteria, affects multiple bacterial behaviors including motility and biofilm formation. c-di-GMP is synthesized by diguanylate cyclase harboring a GGDEF domain and degraded by phosphodiesterase harboring an either EAL or HD-GYP domain. Vibrio parahaemolyticus, the leading cause of seafood-associated gastroenteritis, harbors more than 60 genes involved in c-di-GMP metabolism. However, roles of most of these genes including vpa0198, which encodes a GGDEF-domain containing protein, are still completely unknown. AphA and OpaR are the master quorum sensing (QS) regulators operating at low (LCD) and high cell density (HCD), respectively. QsvR integrates into QS to control gene expression via direct regulation of AphA and OpaR. In this study, we showed that deletion of vpa0198 remarkably reduced c-di-GMP production and biofilm formation, whereas promoted the swimming motility of V. parahaemolyticus. Overexpression of VPA0198 in the vpa0198 mutant strain significantly reduced the swimming and swarming motility and enhanced the biofilm formation ability of V. parahaemolyticus. In addition, transcription of vpa0198 was under the collective regulation of AphA, OpaR and QsvR. AphA activated the transcription of vpa0198 at LCD, whereas QsvR and OpaR coordinately and directly repressed vpa0198 transcription at HCD, thereby leading to a cell density-dependent expression of vpa0198. Therefore, this work expanded the knowledge of synthetic regulatory mechanism of c-di-GMP in V. parahaemolyticus.

Cultivation in long-term simulated microgravity is detrimental to pyocyanin production and subsequent biofilm formation ability of Pseudomonas aeruginosa.

Given plans for humans to engage in prolonged space travel, we investigated biofilm and pigment/virulence factor formation in Pseudomonas aeruginosa when cultivated in microgravity. These bacteria are opportunistic pathogens in immunocompromised individuals. Previous studies of space travelers have shown some immune system diminutions. Hence, our studies shed some light on how prolonged cultivation of bacteria in simulated microgravity conditions affect their growth characteristics.

Insights into macrolide resistance in Arcobacter butzleri: potential resistance mechanisms and impact on bacterial fitness and virulence.

Macrolides are recommended for treating the emerging enteropathogen Arcobacter butzleri; nonetheless, this bacterium often exhibits highly variable resistance rates, and the mechanisms behind this resistance phenotype remain largely unexplored. To understand the phenotypic and genotypic consequences associated with the acquisition of erythromycin resistance in A. butzleri, as well as the effects on the fitness of this species. Resistant strains resulting from spontaneous mutations and adaptive laboratory evolution under increasing erythromycin concentrations were examined regarding their cross-resistance and collateral susceptibility profiles. Genetic causes of phenotypic antibiotic resistance were analysed by sequencing and bioinformatics, with functional correlation through ethidium bromide accumulation assays. Growth profiles in the presence and absence of erythromycin, motility and biofilm formation abilities were assessed to detect potential changes in fitness and virulence. Clones from spontaneous mutation rate evolution demonstrated decreased susceptibility to erythromycin and other classes of antibiotics, associated with mutations in the transcriptional repressor areR, causing overexpression of the AreABC efflux pump. In turn, WGS analysis of the evolved strain showed additional mutations in the ribosomal proteins L4 and L22 and in the areR gene. Furthermore, the acquisition of macrolide resistance altered A. butzleri virulence and entailed a high biological cost. The findings of this study have proved that efflux activity contributes synergistically with mutations in the ribosomal proteins L4 and L22 to A. butzleri's high-level macrolide resistance. The results further suggest an impact on the bacterial physiology and virulence, with the increased fitness cost justifying the low worldwide prevalence of high-level resistant circulating strains.

Genetic Characterization, Antibiotic Resistance, and Virulence Genes Profiling of Bacillus cereus Strains from Various Foods in Japan.

Bacillus cereus sensu stricto is a foodborne pathogen that causes food poisoning. Their spore and biofilm-forming abilities persist in various environments and foods. This study investigated the prevalence, virulence, antibiotic resistance, and genetic diversity of B. cereus s. s. strains isolated from various food samples. Of 179 samples, 22.34% were positive for B. cereus s. s., with significantly high detection rates in milk products and raw chicken meat. Forty strains were isolated from positive samples. Matrix-assisted laser desorption ionization/time of flight mass spectrometry analysis revealed nine distinct clusters and multi-locus sequence typing revealed 34 sequence types including 23 novel sequences, demonstrating high genetic diversity among the isolates. PCR analysis revealed that all the strains contained at least one toxin gene, but none contained the cytK gene. Antibiotic resistance tests revealed that all isolates were classified as multidrug-resistant, with high resistance levels, particularly to β-lactam antibiotics and vancomycin, but were susceptible to gentamicin. All isolates showed variations in biofilm formation. This study highlights the significant public health risk due to B. cereus s. s. and underscores the need for stringent monitoring and control measures in food production to manage antimicrobial resistance and ensure food safety.

Study the relationship between antibiotic resistance and the biofilm-forming ability of bacteria isolated from cases of osteomyelitis after surgery

Study the relationship between antibiotic resistance and the biofilm-forming ability of bacteria isolated from cases of osteomyelitis after surgery

Diverse polysaccharide production and biofilm formation abilities of clinical Klebsiella pneumoniae.

Klebsiella pneumoniae infections have become a growing threat for human health. The lack of understanding of the relationship between antibiotic resistance, mucoviscosity, and biofilm formation in clinical isolates impedes our abilities to effectively predict K. pneumoniae infection outcomes. These traits are also associated with fitness in natural populations and more specifically within a host. The Multidrug-Resistant Organism Repository and Surveillance Network offers a unique opportunity into the genetic and phenotypic variabilities in the K. pneumoniae isolates encountered in the clinics today. To this end, we compared the genetic profiles of these isolates with the phenotypic biofilm formation abilities, percent mucoviscosity, and growth rates. We found most isolates formed limited biofilm, although a select group of isolates could form extremely robust biofilms. Variation in biofilm formation could not be explained by difference in growth rate, suggesting specific genetic and physical determinants. Interestingly, the most mucoid strains in the populations were lacking the genetic element regulating the mucoid phenotype and three of these isolates were able to form robust biofilms. There was a significant phenotype-genotype correlation with decreased biofilm formation and an insertion sequence in the transcriptional activator of the type III fimbrial system. Finally, confocal microscopy highlighted the structural and spatial heterogeneity of biofilm among the most robust biofilm formers not detected by traditional methods. The combination of phenotypic, genomic and image analyses allowed us to reveal an unexpected phenotypic diversity and an intricate relation between growth, mucoviscosity and specific virulence-associated genetic determinants.

Molecular and phenotypic characterization of biofilm formation and antimicrobial resistance patterns of uropathogenic staphylococcus haemolyticus isolates in Casablanca, Morocco

This study aimed to establish the correlation between antibiotic resistance and biofilm formation by Staphylococcus haemolyticus and to examine the impact of sub-inhibitory concentrations of antibiotics (sub-MICs) on biofilm formation.Antibiotic susceptibility testing was conducted using the disk diffusion method, and biofilm formation was determined using Congo red agar and microtiter plate methods. Antibiotic resistance and biofilm-associated genes were detected using polymerase chain reaction.The majority of the twenty-one S. haemolyticus isolates were multidrug-resistant, methicillin-resistant (MRSH) and biofilm producers, including 43 % of moderate biofilm producers. A significant correlation was observed between MRSH and MSSH isolates in terms of biofilm production. Vancomycin, gentamicin, and ciprofloxacin at their sub-MICs tended to promote biofilm formation. The eno gene was present in 76.2 % of strains, followed by aap, and atlE.This study revealed a strong correlation between the biofilm-forming ability and antibiotic resistance in S. haemolyticus, which underlines a crucial public health issue.

Characterization of biofilm-forming ability and antibiotic resistance profiles of clinical Pseudomonas aeruginosa isolates from patients with cystic fibrosis

Background: Pseudomonas aeruginosa is a key pathogen in cystic fibrosis (CF) associated infections, known for its biofilm-forming ability and resistance to multiple antibiotics. Understanding the relationship between biofilm production and antibiotic resistance profiles in clinical isolates can guide treatment strategies. Aims and Objectives: The aims of this study were to investigate the biofilm-forming abilities and antibiotic resistance profiles of clinical P. aeruginosa isolates from patients with CF and to determine the relationship between these two factors and their impact on treatment efficacy. Materials and Methods: A cross-sectional study was conducted on 100 clinical isolates of P. aeruginosa from CF patients. Biofilm-forming capacity was categorized as strong, moderate, or weak based on quantitative assays. Antibiotic resistance was assessed for ciprofloxacin, tobramycin, ceftazidime, meropenem, and piperacillin/tazobactam. Multi-drug resistance was defined as resistance to three or more antibiotic classes. Statistical correlations between biofilm-forming capacity and resistance levels were evaluated using the Chi-square tests. Results: Fifty-six percentages of isolates were strong biofilm producers, while 34% and 10% were moderate and weak producers, respectively. The highest antibiotic resistance was observed against ciprofloxacin (60%), followed by tobramycin (50%) and ceftazidime (45%). Forty percentages of the isolates were classified as multi-drug resistant. Strong biofilm producers demonstrated a significantly high correlation with antibiotic resistance (P<0.05). Two predominant clonal groups were identified among the isolates, suggesting a possible clonal spread of resistance traits. Conclusion: The study confirms a strong association between robust biofilm production and heightened antibiotic resistance in P. aeruginosa isolates from CF patients. These findings highlight the need for targeted therapeutic strategies to disrupt biofilm formation and curb resistance spread.

Multi-functional properties of lactic acid bacteria strains derived from canine feces.

Probiotics, especially Lactic Acid Bacteria (LAB), can promote the health of host animals in a variety of ways, such as regulating intestinal flora and stimulating the host's immune system. In this study, 206 LAB strains were isolated from 48 canine fecal samples. Eleven LAB strains were selected based on growth performance, acid and bile salt resistance. The 11 candidates underwent comprehensive evaluation for probiotic properties, including antipathogenic activity, adhesion, safety, antioxidant capacity, and metabolites. The results of the antipathogenic activity tests showed that 11 LAB strains exhibited strong inhibitory effect and co-aggregation ability against four target pathogens (E. coli, Staphylococcus aureus, Salmonella braenderup, and Pseudomonas aeruginosa). The results of the adhesion test showed that the 11 LAB strains had high cell surface hydrophobicity, self-aggregation ability, biofilm-forming ability and adhesion ability to the Caco-2 cells. Among them, Lactobacillus acidophilus (L177) showed strong activity in various adhesion experiments. Safety tests showed that 11 LAB strains are sensitive to most antibiotics, with L102, L171, and L177 having the highest sensitivity rate at 85.71%, and no hemolysis occurred in all strains. Antioxidant test results showed that all strains showed good H2O2 tolerance, high scavenging capacity for 1, 1-diphenyl-2-trinitrophenylhydrazine (DPPH) and hydroxyl (OH-). In addition, 11 LAB strains can produce high levels of metabolites including exopolysaccharide (EPS), γ-aminobutyric acid (GABA), and bile salt hydrolase (BSH). This study provides a thorough characterization of canine-derived LAB strains, highlighting their multifunctional potential as probiotics. The diverse capabilities of the strains make them promising candidates for canine dietary supplements, offering a holistic approach to canine health. Further research should validate their efficacy in vivo to ensure their practical application.

Anti-virulence potential of iclaprim, a novel folic acid synthesis inhibitor, against Staphylococcus aureus

Infections caused by Staphylococcus aureus pose a significant global public problem. Therefore, new antibiotics and therapeutic strategies are needed to combat this pathogen. This investigation delves into the effects of iclaprim, a newly discovered inhibitor of folic acid synthesis, on S. aureus virulence. The phenotypic and genotypic effects of iclaprim were thoroughly examined in relation to virulence factors, biofilm formation, and dispersal, as well as partial virulence-encoding genes associated with exoproteins, adherence, and regulation in S. aureus MW2, N315, and ATCC 25923. Then, the in vivo effectiveness of iclaprim on S. aureus pathogenicity was explored by a Galleria mellonella larvae infection model. The use of iclaprim at sub-inhibitory concentrations (sub-MICs) resulted in a reduction of α-hemolysin (Hla) production and a differential effect on the activity of coagulase in S. aureus strains. The results of biofilm formation and eradication assay showed that iclaprim was highly effective in depolymerizing the mature biofilm of S. aureus strains at concentrations of 1 MIC or greater, however, inhibited the biofilm-forming ability of only strains N315 and ATCC 25923 at sub-MICs. Interestingly, treatment of strains with sub-MICs of iclaprim resulted in significant stimulation or suppression of most virulence-encoding genes expression. Iclaprim did not affect the production of δ-hemolysin or staphylococcal protein A (SpA), nor did it impact the total activity of proteases, nucleases, and lipases. In vivo testing showed that sub-MICs of iclaprim significantly improves infected larvae survival. The present study offered valuable insights towards a better understating of the influence of iclaprim on different strains of S. aureus. The findings suggest that iclaprim may have potential as an anti-virulence and antibiofilm agent, thus potentially mitigating the pathogenicity of S. aureus and improving clinical outcomes associated with infections caused by this pathogen.Key points• Iclaprim effectively inhibits α-hemolysin production and biofilm formation in a strain-dependent manner and was an excellent depolymerizing agent of mature biofilm• Iclaprim affected the mRNA expression of virulence-encoding genes associated with exoproteins, adherence, and regulation• In vivo study in G. mellonella larvae challenged with S. aureus exhibited that iclaprim improves larvae survival

Interactive effects of osmotic, acid, heat, cold, and freezing stresses on the biofilm formation ability of Salmonella serotypes

AbstractFood‐related stresses such as heating and freezing may influence the biofilm formation ability of bacteria. This study aimed to investigate the main and interactive effects of food‐related stresses on the biofilm formation potential of Salmonella strains isolated from meat. Salmonella enteritidis, Salmonella typhi, and Salmonella typhimurium were subjected to osmotic, acid, heat, cold, and freezing stresses. The colorimetric microtiter plate method was used to measure the biofilm formation ability as a response to the stresses. Among the main effects, freezing time had the most significant effect on the biofilm formation responses of three Salmonella serovars. Freezing reduced the biofilm formation ability of Salmonella typhimurium and Salmonella enteritidis but increased that of Salmonella typhi (p < 0.05). Among the interaction effects, the most significant effect on the biofilm formation response of Salmonella typhi was the interaction between pH and heat, which had a negative effect on the biofilm formation response. In contrast, the interaction between osmotic pressure and cold stresses was the most significant interactive effect on the biofilm formation responses of Salmonella enteritidis and Salmonella typhimurium, which had an increasing effect. This study concluded that the food‐related stresses could change the biofilm formation capacity of Salmonella serotypes, and each serotype might show different biofilm formation abilities in response to different stresses.

The multifaceted role of TolA protein in promoting survival, biofilm formation and virulence of avian pathogenic Escherichia coli

Avian pathogenic Escherichia coli (APEC) can spread beyond the intestines and cause systemic infections, leading to various clinical manifestations, including airsacculitis, pericarditis, perihepatitis and colisepticemia. The mechanisms facilitating this extraintestinal infections are not fully understood. In this study, we investigate how the tolA gene affects APEC virulence by encoding a protein involved in maintaining outer membrane integrity. We constructed a tolA deletion mutant of APEC strain E058 and evaluated its growth and survival in various environments, including in vitro cultures and in vivo infection models in chickens. We found that the motility-defective ΔtolA mutant exhibits reduced biofilm formation ability and weakened resistance to the environmental stresses, suggesting an important role for TolA in APEC's survival. The lack of tolA gene affects the bacterial ability to resist the host's immune system, such as complement-mediated serum killing or phagocytosis, as shown by the serum killing and macrophage phagocytosis assays. Additionally, in vivo infection studies using chickens demonstrated that the ΔtolA mutant displayed attenuated virulence, evidenced by reduced mortality and lower tissue bacterial burden. Reverse transcription quantitative real-time PCR (RT-qPCR) analysis revealed that inactivation of tolA led to downregulation of virulence genes associated with serum resistance (traT) and flagellar biosynthesis (fliR). Taken together, our findings demonstrate the multifaceted role of TolA protein in promoting the survival, immune evasion, biofilm formation, and virulence of APEC E058. This suggests that targeting TolA could potentially offer new strategies for combating APEC infections.

Carbon Source and Substrate Surface Affect Biofilm Formation by the Plant-Associated Bacterium Pseudomonas donghuensis P482.

The ability of bacteria to colonize diverse environmental niches is often linked to their competence in biofilm formation. It depends on the individual characteristics of a strain, the nature of the colonized surface (abiotic or biotic), or the availability of certain nutrients. Pseudomonas donghuensis P482 efficiently colonizes the rhizosphere of various plant hosts, but a connection between plant tissue colonization and the biofilm formation ability of this strain has not yet been established. We demonstrate here that the potential of P482 to form biofilms on abiotic surfaces and the structural characteristics of the biofilm are influenced by the carbon source available to the bacterium, with glycerol promoting the process. Also, the type of substratum, polystyrene or glass, impacts the ability of P482 to attach to the surface. Moreover, P482 mutants in genes associated with motility or chemotaxis, the synthesis of polysaccharides, and encoding proteases or regulatory factors, which affect biofilm formation on glass, were fully capable of colonizing the root tissue of both tomato and maize hosts. Investigating the role of cellular factors in biofilm formation using these plant-associated bacteria shows that the ability of bacteria to form biofilm on abiotic surfaces does not necessarily mirror its ability to colonize plant tissues. Our research provides a broader perspective on the adaptation of these bacteria to various environments.

Evaluation of In Vitro Biofilm Formation of Helicobacter pylori in Different Culture Media

Objective: Biofilms are surface-attached cell communities that play a role in the survival of bacteria. Helicobacter pylori, a gram-negative pathogen that colonizes the human gastric mucosa, forms biofilms, causing treatment failure and the risk of developing peptic ulcers, gastritis and gastric cancer in infected individuals. The aim of the study is to evaluate the biofilm formation abilities of H. pylori ATCC 26695 and three clinical strains in different culture media. Material and Method: Biofilm formation characteristics of H. pylori strains using different culture media were evaluated, and the crystal violet (CV) staining method (measured at OD 595 ) was used. Various media were used for incubating H. pylori strains: Brucella broth (BB), TSB with 10% FBS, BHI with 10% FBS, BB with 10% FBS, BB with 10% FBS + 0.25% glucose, and BB with 10% FBS + 1% glucose (incubated for 3 days). Additionally, BB with 1% FBS, BB with 10% FBS, and BB with 5% inactivated human serum were incubated for 2 and 4 days at 37°C under microaerophilic conditions. Results: It was observed that 5% inactived human serum was more effective in biofilm formation of H. pylori ATCC 26695 and three clinical strains. However, there was no biofilm production in the strains cultured with Brucella broth alone and that the strains cultured with TSB + 10% FBS could not form a strong biofilm compared to other media. Conclusion: Different culture media used for H. pylori ATCC 26695 and three clinical strains affect biofilm formation. It is thought that in vitro experiments to prevent biofilm formation may provide a solution to the prevention of H. pylori infection.