Cronobacter sakazakii, an opportunity foodborne pathogen, could contaminate a broad range of food materials and cause life-threatening symptoms in infants. The bacterial envelope structure contribute to bacterial environment tolerance, biofilm formation and virulence in various in Gram-negative bacteria. DsbA and PepP are two important genes related to the biogenesis and stability of bacterial envelope. In this study, the DsbA and PepP were deleted in C. sakazakii to evaluate their contribution to stress tolerance and virulence of the pathogen. The bacterial environment resistance assays showed DsbA and PepP are essential in controlling C. sakazakii resistance to heat and desiccation in different mediums, as well as acid, osmotic, oxidation and bile salt stresses. DsbA and PepP also played an important role in regulating biofilm formation and motility. Furthermore, DsbA and PepP deletion weaken C. sakazakii adhesion and invasion in Caco-2, intracellular survival and replication in RAW 264.7. qRT-PCR results showed that DsbA and PepP of C. sakazakii played roles in regulating the expression of several genes associated with environment stress tolerance, biofilm formation, bacterial motility and cellular invasion. These findings indicate that DsbA and PepP played an important regulatory role in the environment resisitance, biofilm formation and virulence of C. sakazakii, which enrich understanding of genetic determinants of adaptability and virulence of the pathogen.

Antibiotic resistance in soil introduced by organic fertilizer application pose a globally recognized threat to human health. Insect organic fertilizer may be a promising alternative due to its low antibiotic resistance. However, it is not yet clear how to regulate soil microbes to reduce antibiotic resistance in organic fertilizer agricultural application. In this study, we investigated soil microbes and antibiotic resistome under black soldier fly organic fertilizer (BOF) application in pot and field systems. Our study shows that BOF could stimulate ARB (antibiotic resistant - bacteria) - suppressive Bacillaceae in the soil microbiome and reduce antibiotic resistome. The carbohydrate transport and metabolism pathway of soil Bacillaceae was strengthened, which accelerated the synthesis and transport of polysaccharides to form biofilm to antagonistic soil ARB, and thus reduced the antibiotic resistance. We further tested the ARB - suppressive Bacillus spp. in a microcosm assay, which resulted in a significant decrease in the presence of ARGs and ARB together with higher abundance in key biofilm formation gene (epsA). This knowledge might help to the development of more efficient bio-fertilizers aimed at mitigating soil antibiotic resistance and enhancing soil health, in particular, under the requirements of global \"One Health\".

Bacterial ribonuclease E (RNase E) is vital for posttranscriptional regulation by degrading and processing RNA. The RraA protein inhibits RNase E activity through protein-protein interactions, exerting a global regulatory effect on gene expression. However, the specific role of RraA remains unclear. In this study, we investigated rraA expression in Vibrio alginolyticus ZJ-T and identified three promoters responsible for its expression, resulting in transcripts with varying 5\'-UTR lengths. During the stationary phase, rraA was significantly posttranscriptionally inhibited. Deletion of rraA had no impact on bacterial growth in rich medium Luria-Bertani broth with salt (LBS) but resulted in decreased biofilm formation and increased resistance to polymyxin B. Transcriptome analysis revealed 350 differentially expressed genes (DEGs) between the wild type and the rraA mutant, while proteome analysis identified 267 differentially expressed proteins (DEPs). Integrative analysis identified 55 genes common to both DEGs and DEPs, suggesting that RraA primarily affects gene expression at the posttranscriptional level. KEGG (Kyoto Encyclopedia of Genes and Genomes) analysis demonstrated that RraA facilitates the conversion of fatty acids, propionic acid, and branched-chain amino acids to acetyl-CoA while enhancing amino acid and peptide uptake. Notably, RraA positively regulates the expression of virulence-associated genes, including those involved in biofilm formation and the type VI secretion system. This study expands the understanding of the regulatory network of RraA through transcriptome analysis, emphasizing the importance of proteomic analysis in investigating posttranscriptional regulation.IMPORTANCERraA is an inhibitor protein of ribonuclease E that interacts with and suppresses its endonucleolytic activity, thereby playing a widespread regulatory role in the degradation and maturation of diverse mRNAs and noncoding small RNAs. However, the physiological functions and associated regulon of RraA in Vibrio alginolyticus have not been fully elucidated. Here, we report that RraA impacts virulence-associated physiological processes, namely, antibiotic resistance and biofilm formation, in V. alginolyticus. By conducting an integrative analysis of both the transcriptome and proteome, we revealed the involvement of RraA in carbon metabolism, amino acid catabolism, and transport, as well as in the type VI secretion system. Collectively, these findings elucidate the regulatory influence of RraA on multiple pathways associated with metabolism and pathogenesis in V. alginolyticus.

UNASSIGNED: Persistent infections caused by Helicobacter pylori (H. pylori), which are resistant to antibiotic treatment, pose a growing global public health concern. Biofilm formation is known to be associated with persistent infections due to its role in enhancing antimicrobial resistance and the tolerance of many pathogenic bacteria.
UNASSIGNED: This study aims to evaluate the biofilm formation of clinical isolates of H. pylori and its impact on antibiotic eradication.
UNASSIGNED: The thickness, morphology, and structure of biofilms derived from nine H. pylori strains were examined using confocal laser scanning microscopy, scanning electron microscopy, and transmission electron microscopy. Subsequently, the susceptibility of both planktonic and biofilm bacteria was assessed through the determination of minimum inhibitory concentration and minimum biofilm eradication concentration for amoxicillin, clarithromycin, levofloxacin, and tetracycline.
UNASSIGNED: The results revealed varying biofilm thicknesses and densities among the strains, characterised by the presence of numerous filaments intertwining and connecting bacterial cells. Additionally, several cases exhibited susceptibility based on MIC measurements but resistance according to MBEC measurements, with MBEC indicating a higher resistance rate. Pearson Correlation analysis demonstrated a positive correlation between biofilm thickness and MBEC results (0 < r < 1), notably significant for amoxicillin (r = 0.801, P = 0.009) and tetracycline (r = 0.696, P = 0.037).
UNASSIGNED: Different strains of H. pylori exhibit variations in their capacity to release outer membrane vesicles (OMVs) and form biofilms. Biofilm formation can influence the effectiveness of amoxicillin and tetracycline in eradicating susceptible bacterial strains.

Microbial community adaptability to pH stress plays a crucial role in biofilm formation. This study aims to investigate the regulatory mechanisms of exogenous putrescine on pH stress, as well as enhance understanding and application for the technical measures and molecular mechanisms of biofilm regulation. Findings demonstrated that exogenous putrescine acted as a switch-like distributor affecting microorganism pH stress, thus promoting biofilm formation under acid conditions while inhibiting it under alkaline conditions. As pH decreases, the protonation degree of putrescine increases, making putrescine more readily adsorbed. Protonated exogenous putrescine could increase cell membrane permeability, facilitating its entry into the cell. Subsequently, putrescine consumed intracellular H+ by enhancing the glutamate-based acid resistance strategy and the γ-aminobutyric acid metabolic pathway to reduce acid stress on cells. Furthermore, putrescine stimulated ATPase expression, allowing for better utilization of energy in H+ transmembrane transport and enhancing oxidative phosphorylation activity. However, putrescine protonation was limited under alkaline conditions, and the intracellular H+ consumption further exacerbated alkali stress and inhibits cellular metabolic activity. Exogenous putrescine promoted the proportion of fungi and acidophilic bacteria under acidic stress and alkaliphilic bacteria under alkali stress while having a limited impact on fungi in alkaline biofilms. Increasing Bdellovibrio under alkali conditions with putrescine further aggravated the biofilm decomposition. This research shed light on the unclear relationship between exogenous putrescine, environmental pH, and pH stress adaptability of biofilm. By judiciously employing putrescine, biofilm formation could be controlled to meet the needs of engineering applications with different characteristics.IMPORTANCEThe objective of this study is to unravel the regulatory mechanism by which exogenous putrescine influences biofilm pH stress adaptability and understand the role of environmental pH in this intricate process. Our findings revealed that exogenous putrescine functioned as a switch-like distributor affecting the pH stress adaptability of biofilm-based activated sludge, which promoted energy utilization for growth and reproduction processes under acidic conditions while limiting biofilm development to conserve energy under alkaline conditions. This study not only clarified the previously ambiguous relationship between exogenous putrescine, environmental pH, and biofilm pH stress adaptability but also offered fresh insights into enhancing biofilm stability within extreme environments. Through the modulation of energy utilization, exerting control over biofilm growth and achieving more effective engineering goals could be possible.

Vibrio parahaemolyticus possesses two distinct type VI secretion systems (T6SS), namely T6SS1 and T6SS2. T6SS1 is predominantly responsible for adhesion to Caco-2 and HeLa cells and for the antibacterial activity of V. parahaemolyticus, while T6SS2 mainly contributes to HeLa cell adhesion. However, it remains unclear whether the T6SS systems have other physiological roles in V. parahaemolyticus. In this study, we demonstrated that the deletion of icmF2, a structural gene of T6SS2, reduced the biofilm formation capacity of V. parahaemolyticus under low salt conditions, which was also influenced by the incubation time. Nonetheless, the deletion of icmF2 did not affect the biofilm formation capacity in marine-like growth conditions, nor did it impact the flagella-driven swimming and swarming motility of V. parahaemolyticus. IcmF2 was found to promote the production of the main components of the biofilm matrix, including extracellular DNA (eDNA) and extracellular proteins, and cyclic di-GMP (c-di-GMP) in V. parahaemolyticus. Additionally, IcmF2 positively influenced the transcription of cpsA, mfpA, and several genes involved in c-di-GMP metabolism, including scrJ, scrL, vopY, tpdA, gefA, and scrG. Conversely, the transcription of scrA was negatively impacted by IcmF2. Therefore, IcmF2-dependent biofilm formation was mediated through its effects on the production of eDNA, extracellular proteins, and c-di-GMP, as well as its impact on the transcription of cpsA, mfpA, and genes associated with c-di-GMP metabolism. This study confirmed new physiological roles for IcmF2 in promoting biofilm formation and c-di-GMP production in V. parahaemolyticus.

UNASSIGNED: Microorganisms in biofilms are particularly difficult to control because of their increased survival and antibiotic resistance. Allicin and domiphen were employed to inhibit the microbial growth and biofilm formation of Staphylococcus aureus, Escherichia coli, and Candida albicans strains.
UNASSIGNED: Broth microdilution method and checkerboard assay were conducted to determine the efficacy of allicin combined with domiphen against S. aureus, E. coli, and C. albicans. Microbial biofilm formation was measured using the crystal violet staining method and fluorescence microscopy. And the total viable count of the biofilm cells on material surface after the treatment with antimicrobial reagents was calculated with the plate count technique.
UNASSIGNED: The two drugs showed synergistic effects against the pathogens with a fractional bactericidal concentration of less than 0.38. The combination of 64 μg/mL allicin with 1 μg/mL domiphen dispersed over 50% of the biofilm mass of S. aureus, E. coli, and C. albicans. In addition, the drug combination reduced the total viable counts of E. coli and C. albicans biofilm cells on stainless steel and polyethylene surfaces by more than 102 CFU/mL.
UNASSIGNED: The combination of allicin and domiphen is an effective strategy for efficiently decreasing biofilms formation on various industrial materials surfaces.

Bacillus licheniformis is one of the major spore-forming bacteria with great genotypic diversity in raw milk, dairy ingredients, final dairy products, and is found throughout the dairy processing continuum. Though being widely used as a probiotic strain, this species also serves as a potential risk in the dairy industry based on its roles in foodborne illness and dairy spoilage. Biofilm formation of B. licheniformis in combined with the heat resistance of its spores, make it impossible to prevent the presence of B. licheniformis in final dairy products by traditional cleaning and disinfection procedures. Despite the extensive efforts on the identification of B. licheniformis from various dairy samples, no reviews have been reported on both hazard and benefits of this spore-former. This review discusses the prevalence of B. licheniformis from raw milk to commercial dairy products, biofilm formation and spoilage potential of B. licheniformis, and its potential prevention methods. In addition, the potential benefits of B. licheniformis in the dairy industry were also summarized.

Understanding bacterial adhesion at the nanoscale is crucial for elucidating biofilm formation, enhancing biosensor performance, and designing advanced biomaterials. However, the dynamics of the critical transition from reversible to irreversible adhesion has remained elusive due to analytical constraints. Here, we probed this adhesion transition, unveiling nanoscale, step-like bacterial approaches to substrates using a plasmonic imaging technique. This method reveals the discontinuous nature of adhesion, emphasizing the complex interplay between bacterial extracellular polymeric substances (EPS) and substrates. Our findings not only deepen our understanding of bacterial adhesion but also have significant implications for the development of theoretical models for biofilm management. By elucidating these nanoscale step-like adhesion processes, our work provides avenues for the application of nanotechnology in biosensing, biofilm control, and the creation of biomimetic materials.

The presence and accumulation of both, plastics and antibiotics in soils may lead to the colonization, selection, and propagation of soil bacteria with certain metabolic traits, e.g., antibiotic resistance, in the plastisphere. However, the impact of plastic-antibiotic tandem on the soil ecosystem functioning, particularly on microbial function and metabolism remains currently unexplored. Herein, we investigated the competence of soil bacteria to colonize plastics and degrade 13C-labeled sulfamethoxazole (SMX). Using single-cell imaging, isotope tracers, soil respiration and SMX mineralization bulk measurements we show that microbial colonization of polyethylene (PE) and polystyrene (PS) surfaces takes place within the first 30 days of incubation. Morphologically diverse microorganisms were colonizing both plastic types, with a slight preference for PE substrate. CARD-FISH bacterial cell counts on PE and PS surfaces formed under SMX amendment ranged from 5.36 × 103 to 2.06 × 104, and 2.06 × 103 to 3.43 × 103 hybridized cells mm-2, respectively. Nano-scale Secondary Ion Mass Spectrometry measurements show that 13C enrichment was highest at 130 days with values up to 1.29 atom%, similar to those of the 13CO2 pool (up to 1.26 atom%, or 22.55 ‰). Independent Mann-Whitney U test showed a significant difference between the control plastisphere samples incubated without SMX and those in 13C-SMX incubations (P < 0.001). Our results provide direct evidence demonstrating, at single-cell level, the capacity of bacterial colonizers of plastics to assimilate 13C-SMX from contaminated soils. These findings expand our knowledge on the role of soil-seeded plastisphere microbiota in the ecological functioning of soils impacted by anthropogenic stressors.