Expression Of Adhesins Research Articles

Epinephrine and norepinephrine regulate the expression of virulence factors in Gallibacterium anatis

Gallibacterium anatis is a member of the Pasteurellaceae family and is an opportunistic pathogen that causes gallibacteriosis in chickens. Stress plays a relevant role in promoting the development of pathogenicity in G. anatis. Epinephrine (E) and norepinephrine (NE) are relevant to stress; however, their effects on G. anatis have not been elucidated. In this work, we evaluated the effects of E and NE on the growth, biofilm formation, expression of adhesins, and proteases of two G. anatis strains, namely, the hemolytic 12656-12 and the nonhemolytic F149T biovars. E (10 μM/mL) and NE (30 and 50 μM/mL) increased the growth of G. anatis 12656-12 by 20 % and 25 %, respectively. E did not affect the growth of F149T, whereas 40 μM/mL NE decreased bacterial growth by 25 %. E and NE at a dose of 30–50 μM/mL upregulated five fibrinogen adhesins in the 12565-12 strain, whereas no effect was observed in the F149T strain. NE increased proteolytic activity in both strains, whereas E diminished proteolytic activity in the 12656-12 strain. E and NE reduced biofilm formation (30 %) and increased Congo red binding (15 %) in both strains. QseBC is the E and NE two-component detection system most common in bacteria. The qseC gene, which is the E and NE receptor in bacteria, was identified in the genomic DNA of the 12565-12 and F149TG. anatis strains via PCR amplification. Our results suggest that QseC can detect host changes in E and NE concentrations and that catecholamines can modulate the expression of several virulence factors in G. anatis.

Conditional expression of flagellar motility, curli fimbriae, and biofilms in Shiga toxin- producing Escherichia albertii.

Escherichia albertii is an emerging foodborne pathogen. We previously reported that some avian Shiga toxin-producing E. albertii strains exhibited higher or comparable cytotoxicity in Vero-d2EGFP cells with several enterohemorrhagic E. coli (EHEC) outbreak strains. To better understand the environmental persistence of this pathogen, comparative genomics and phenotypic assays were applied to assess adhesion capability, motility, and biofilm formation in E. albertii. Among the 108 adherence-related genes, those involved in biogenesis of curli fimbriae, hemorrhagic E. coli pilus, type 1 fimbriae, and Sfm fimbriae were conserved in E. albertii. All 20 E. albertii strains carried a complete set of primary flagellar genes that were organized into four gene clusters, while five strains possessed genes related to the secondary flagella, also known as lateral flagella. Compared to EHEC strain EDL933, the eight chemotaxis genes located within the primary flagellar gene clusters were deleted in E. albertii. Additional deletion of motility genes flhABCD and motBC was identified in several E. albertii strains. Swimming motility was detected in three strains when grown in LB medium, however, when grown in 5% TSB or in the pond water-supplemented with 10% pigeon droppings, an additional four strains became motile. Although all E. albertii strains carried curli genes, curli fimbriae were detected only in four, eight, and nine strains following 24, 48, and 120 h incubation, respectively. Type 1 fimbriae were undetectable in any of the strains grown at 37°C or 28°C. Strong biofilms were detected in strains that produced curli fimbriae and in a chicken isolate that was curli fimbriae negative but carried genes encoding adhesive fimbriae K88, a signature of enterotoxigenic E. coli strains causing neonatal diarrhea in piglets. In all phenotypic traits examined, no correlation was revealed between the strains isolated from different sources, or between the strains with and without Shiga toxin genes. The phenotypic variations could not be explained solely by the genetic diversity or the difference in adherence genes repertoire, implying complex regulation in expression of various adhesins. Strains that exhibited a high level of cytotoxicity and were also proficient in biofilm production, may have potential to emerge into high-risk pathogens.

The Hog1 MAPK substrate governs Candida glabrata-epithelial cell adhesion via the histone H2A variant.

CgHog1, terminal kinase of the high-osmolarity glycerol signalling pathway, orchestrates cellular response to multiple external stimuli including surplus-environmental iron in the human fungal pathogen Candida glabrata (Cg). However, CgHog1 substrates remain unidentified. Here, we show that CgHog1 adversely affects Cg adherence to host stomach and kidney epithelial cells in vitro, but promotes Cg survival in the iron-rich gastrointestinal tract niche. Further, CgHog1 interactome and in vitro phosphorylation analysis revealed CgSub2 (putative RNA helicase) to be a CgHog1 substrate, with CgSub2 also governing iron homeostasis and host adhesion. CgSub2 positively regulated EPA1 (encodes a major adhesin) expression and host adherence via its interactor CgHtz1 (histone H2A variant). Notably, both CgHog1 and surplus environmental iron had a negative impact on CgSub2-CgHtz1 interaction, with CgHTZ1 or CgSUB2 deletion reversing the elevated adherence of Cghog1Δ to epithelial cells. Finally, the surplus-extracellular iron led to CgHog1 activation, increased CgSub2 phosphorylation, elevated CgSub2-CgHta (canonical histone H2A) interaction, and EPA1 transcriptional activation, thereby underscoring the iron-responsive, CgHog1-induced exchange of histone partners of CgSub2. Altogether, our work mechanistically defines how CgHog1 couples Epa1 adhesin expression with iron abundance, and point towards specific chromatin composition modification programs that probably aid fungal pathogens align their adherence to iron-rich (gut) and iron-poor (blood) host niches.

Host-derived protease promotes aggregation of Staphylococcus aureus by cleaving the surface protein SasG.

Staphylococcus aureus is one of the leading causes of hospital-acquired infections, many of which begin following attachment and accumulation on indwelling medical devices or diseased tissue. These infections are often linked to the establishment of biofilms, but another often overlooked key characteristic allowing S. aureus to establish persistent infection is the formation of planktonic aggregates. Such aggregates are physiologically similar to biofilms and protect pathogens from innate immune clearance and increase antibiotic tolerance. The cell-wall-associated protein SasG has been implicated in biofilm formation via mechanisms of intercellular aggregation but the mechanism in the context of disease is largely unknown. We have previously shown that the expression of cell-wall-anchored proteins involved in biofilm formation is controlled by the ArlRS-MgrA regulatory cascade. In this work, we demonstrate that the ArlRS two-component system controls aggregation, by repressing the expression of sasG by activation of the global regulator MgrA. We also demonstrate that SasG must be proteolytically processed by a non-staphylococcal protease to induce aggregation and that strains expressing functional full-length sasG aggregate significantly upon proteolysis by a mucosal-derived host protease found in human saliva. We used fractionation and N-terminal sequencing to demonstrate that human trypsin within saliva cleaves within the A domain of SasG to expose the B domain and induce aggregation. Finally, we demonstrated that SasG is involved in virulence during mouse lung infection. Together, our data point to SasG, its processing by host proteases, and SasG-driven aggregation as important elements of S. aureus adaptation to the host environment.IMPORTANCEHere, we demonstrate that the Staphylococcus aureus surface protein SasG is important for cell-cell aggregation in the presence of host proteases. We show that the ArlRS two-component regulatory system controls SasG levels through the cytoplasmic regulator MgrA. We identified human trypsin as the dominant protease triggering SasG-dependent aggregation and demonstrated that SasG is important for S. aureus lung infection. The discovery that host proteases can induce S. aureus aggregation contributes to our understanding of how this pathogen establishes persistent infections. The observations in this study demonstrate the need to strengthen our knowledge of S. aureus surface adhesin function and processing, regulation of adhesin expression, and the mechanisms that promote biofilm formation to develop strategies for preventing chronic infections.

Human PRH1, PRH2 susceptibility and resistance and Streptococcus mutans virulence phenotypes specify different microbial profiles in caries

Lifestyle- and sucrose-dependent polymicrobial ecological shifts are a primary cause of caries in populations with high caries prevalence. In populations with low prevalence, PRH1, PRH2 susceptibility and resistance phenotypes may interact with the Streptococcus mutans adhesin cariogenicity phenotype to affect caries progression, but studies are lacking on how these factors affect the microbial profile of caries. We analysed how the residency and infection profiles of S.mutans adhesin (SpaP A/B/C and Cnm/Cbm) phenotypes and commensal streptococci and lactobacilli influenced caries progression in a prospective case-referent sample of 452 Swedish adolescents with high (P4a), moderate (P6), and low (P1) caries PRH1, PRH2 phenotypes. Isolates of S.mutans from participants were analysed for adhesin expression and glycosylation and invitro and in situ mechanisms related to caries activity. Among adolescents with the resistant (P1) phenotype, infection with S.mutans high-virulence phenotypes was required for caries progression. In contrast, with highly (P4a) or moderately (P6) susceptible phenotypes, caries developed from a broader polymicrobial flora that included moderately cariogenic oral commensal streptococci and lactobacilli and S.mutans phenotypes. High virulence involved unstable residency and fluctuating SpaP ABC, B-1, or Cnm expression/glycosylation phenotypes, whereas low/moderate virulence involved SpaP A phenotypes with stable residency. Adhesin phenotypes did not display changes in individual host residency but were paired within individuals and geographic regions. These results suggest that receptor PRH1, PRH2 susceptibility and resistance and S.mutans adhesin virulence phenotypes specify different microbial profiles in caries. Swedish Research Council and funding bodies listed in the acknowledgement section.

Methanogenic partner influences cell aggregation and signalling of Syntrophobacterium fumaroxidans

For several decades, the formation of microbial self-aggregates, known as granules, has been extensively documented in the context of anaerobic digestion. However, current understanding of the underlying microbial-associated mechanisms responsible for this phenomenon remains limited. This study examined morphological and biochemical changes associated with cell aggregation in model co-cultures of the syntrophic propionate oxidizing bacterium Syntrophobacterium fumaroxidans and hydrogenotrophic methanogens, Methanospirillum hungatei or Methanobacterium formicicum. Formerly, we observed that when syntrophs grow for long periods with methanogens, cultures tend to form aggregates visible to the eye. In this study, we maintained syntrophic co-cultures of S. fumaroxidans with either M. hungatei or M. formicicum for a year in a fed-batch growth mode to stimulate aggregation. Millimeter-scale aggregates were observed in both co-cultures within the first 5 months of cultivation. In addition, we detected quorum sensing molecules, specifically N-acyl homoserine lactones, in co-culture supernatants preceding the formation of macro-aggregates (with diameter of more than 20 μm). Comparative transcriptomics revealed higher expression of genes related to signal transduction, polysaccharide secretion and metal transporters in the late-aggregation state co-cultures, compared to the initial ones. This is the first study to report in detail both biochemical and physiological changes associated with the aggregate formation in syntrophic methanogenic co-cultures.Keypoints• Syntrophic co-cultures formed mm-scale aggregates within 5 months of fed-batch cultivation.• N-acyl homoserine lactones were detected during the formation of aggregates.• Aggregated co-cultures exhibited upregulated expression of adhesins- and polysaccharide-associated genes.Graphical abstract

Spermidine constitutes a key determinant of motility and attachment of Salmonella Typhimurium through a novel regulatory mechanism

Spermidine is a poly-cationic molecule belonging to the family of polyamines and is ubiquitously present in all organisms. Salmonella synthesizes, and harbours specialized transporters to import spermidine. A group of polyamines have been shown to assist in Salmonella Typhimurium's virulence and regulation of Salmonella pathogenicity Inslad 1 (SPI-1) genes and stress resistance; however, the mechanism remains elusive. The virulence trait of Salmonella depends on its ability to employ multiple surface structures to attach and adhere to the surface of the target cells before invasion and colonization of the host niche. Our study discovers the mechanism by which spermidine assists in the early stages of Salmonella pathogenesis. For the first time, we report that Salmonella Typhimurium regulates spermidine transport and biosynthesis processes in a mutually inclusive manner. Using a mouse model, we show that spermidine is critical for invasion into the murine Peyer’s patches, which further validated our in vitro cell line observation. We show that spermidine controls the mRNA expression of fimbrial (fimA) and non-fimbrial adhesins (siiE, pagN) in Salmonella and thereby assists in attachment to host cell surfaces. Spermidine also regulated the motility through the expression of flagellin genes by enhancing the translation of sigma-28, which features an unusual start codon and a poor Shine-Dalgarno sequence. Besides regulating the formation of the adhesive structures, spermidine tunes the expression of the two-component system BarA/SirA to regulate SPI-1 encoded genes. Thus, our study unravels a novel regulatory mechanism by which spermidine exerts critical functions during Salmonella Typhimurium pathogenesis.

Discovering genetic determinants for cell-to-cell adhesion in two prevalent conjugative lactococcal plasmids

Plasmids pNP40 and pUC11B encode two prevalent yet divergent conjugation systems, which have been characterized in detail recently. Here, we report the elucidation of the putative adhesins of the pNP40 and pUC11B conjugation systems, encoded by traAd and trsAd, respectively. Despite their significant sequence divergence, TraAd and TrsAd represent the most conserved component between the pNP40- and the pUC11B-encoded conjugation systems and share similar peptidoglycan-hydrolase domains. Protein structure prediction using AlphaFold2 highlighted the structural similarities between their predicted domains, as well as the potential homo-dimeric state of both proteins. Expression of the putative surface adhesins resulted in a cell clumping phenotype not only among cells expressing these surface adhesins but also between adhesin-expressing and non-producing cells. Furthermore, mutant derivatives of plasmids pNP40 or pUC11B carrying a mutation in traAd or trsAd, respectively, were shown to act as efficient donors provided the corresponding recipient expresses either traAd or trsAd, thus demonstrating in trans reciprocal complementarity of these proteins in conjugation systems.

Multilocus-sequence typing reveals clonality of Staphylococcus aureus in atopic dermatitis.

Atopic dermatitis is exacerbated by Staphylococcus aureus which is capable of displacing not only the physiological microbiota, but also other strains of its own species. Analyzes of molecular characteristics and relationship of S. aureus strains present in different microniches are lacking. To determine the relationship of S. aureus isolates from the lesional skin, nonlesional skin and anterior nares of patients with atopic dermatitis using multilocus-sequence typing and review the characteristics of the dominant clones. Sixty-three individuals with active atopic dermatitis were enrolled. Ten moderate-to-severe patients (SCORAD≥25) colonized by S. aureus within all analyzed locations were included in the multilocus-sequence typing analysis. The most prevalent sequence types were 7 (10/30 strains, 33.3%), 15, and 97 (both 5/30 strains, 16.7%) all of which were associated with the expression of adhesins and toxins promoting chronic microbial dysbiosis, skin barrier damage and inflammation. Six patients (60%) were carriers of clonal S. aureus strains within all analyzed locations, three patients (30%) - within the lesional skin and nonlesional skin, and one patient (10%) - within nonlesional skin and the anterior nares. The results imply that the identified S. aureus lineages are better adapted to dominate the microbiota in atopic dermatitis. Decontaminating the identified reservoirs of S. aureus, i.e. the anterior nares and nonlesional skin, could reduce atopic dermatitis severity.

Expression of hes, iha, and tpsA codified in locus of adhesion and autoaggregation and their involvement in the capability of shiga toxin-producing Escherichia coli strains to adhere to epithelial cells

ObjectivesShiga toxin-producing Escherichia coli strains LAA-positive are important cause of human infection. The capability to adhere to epithelial cells is a key virulence trait, and genes codified in LAA pathogenicity island could be involved in the adhesion during the pathogenesis of LAA-positive STEC strains. Thus, our objectives were to compare hes-negative and hes-positive STEC strains in their adherence capability to epithelial cells (HEp-2) and to evaluate the expression levels of the hes, iha, and tpsA in the bacteria adhered and non-adhered to HEp-2 cells. These genes are encoded in LAA, and are virulence factors that participate in adhesion and autoaggregation.ResultsWe could not observe differences between the adhesion of strains but also in the expression level of of hes, iha, and tpsA. Genes encoded in LAA contribute to the adhesion phenotype though the expression of STEC adhesins is a coordinated event that depends not only the strain but also on the environment as well as its genetic background. Therefore, the results of this study suggest that LAA ,the most prevalent PAI among LEE-negative STEC strains, plays a role in pathogenesis.

A mucin-regulated adhesin determines the spatial organization and inflammatory character of a bacterial symbiont in the vertebrate gut

In a healthy gut, microbes are often aggregated with host mucus, yet the molecular basis for this organization and its impact on intestinal health are unclear. Mucus is a viscous physical barrier separating resident microbes from epithelia, but it also provides glycan cues that regulate microbial behaviors. Here, we describe a mucin-sensing pathway in an Aeromonas symbiont of zebrafish, Aer01. In response to the mucin-associated glycan N-acetylglucosamine, a sensor kinase regulates the expression of an aggregation-promoting adhesin we named MbpA. Upon MbpA disruption, Aer01 colonizes to normal levels but is largely planktonic and more pro-inflammatory. Increasing cell surface MbpA rescues these traits. MbpA-like adhesins are common in human-associated bacteria, and the expression of an Akkermansia muciniphila MbpA-like adhesin in MbpA-deficient Aer01 restores lumenal aggregation and reverses its pro-inflammatory character. Our work demonstrates how resident bacteria use mucin glycans to modulate behaviors congruent with host health.

Lactiplantibacillus plantarum ZJ316 Reduces Helicobacter pylori Adhesion and Inflammation by Inhibiting the Expression of Adhesin and Urease Genes.

The present study aims to investigate the anti-Helicobacter pylori (H. pylori) effects of Lactiplantibacillus plantarum ZJ316 (L. plantarum ZJ316) both in vitro and in vivo. This study finds that L. plantarum ZJ316 effectively suppresses H. pylori adhesion in inhibition (Pre-ZJ316), competition (Co-ZJ316), and displacement (Post-ZJ316) assays, and Pre-ZJ316 displaying the most potent inhibitory effect with an impressive inhibition ratio of 70.14%. Upon anti-adhesion, L. plantarum ZJ316 significantly downregulates the expression of H. pylori virulence genes, including ureA, ureB, flaA, and sabA, with inhibition ratios of 46.83%, 24.02%, 21.42%, and 62.38% at 2 h, respectively. In addition, L. plantarum ZJ316 is observed to reduce the level of interleukin 8 (IL-8) and improve cell viability in infected AGS cells. Furthermore, in vivo studies show that supplementation with L. plantarum ZJ316 effectively hinders H. pylori colonization and significantly suppresses the infiltration of immune cells and IL-8 production with H. pylori infection, protecting host from inflammatory damage. L. plantarum ZJ316 exhibits excellent adhesion inhibition on H. pylori, and may be used as a probiotic candidate in the prevention or adjuvant therapy of gastric disease caused by H. pylori.

Pathogen-induced dormancy in liquid limits gastrointestinal colonization of Caenorhabditis elegans

ABSTRACT Colonization is generally considered a prerequisite for infection, but this event is context-dependent, as evidenced by the differing ability of the human pathogen Pseudomonas aeruginosa to efficiently colonize Caenorhabditis elegans on agar but not in liquid . In this study, we examined the impact of the environment, pathogen, host, and their interactions on host colonization. We found that the transition to a liquid environment reduces food uptake by about two-fold. Also expression of specific adhesins was significantly altered in liquid-based assays for P. aeruginosa, suggesting that it may be one factor driving diminished colonization. Unexpectedly, host immune pathways did not appear to play a significant role in decreased colonization in liquid. Although knocking down key immune pathways (e.g. daf-16 or zip-2), either alone or in combination, significantly reduced survival, the changes in colonization were very small. In spite of the limited bacterial accumulation in the liquid setting, pathogenic colonization was still required for the virulence of Enterococcus faecalis. In addition, we found that a pathogen-induced dormancy was displayed by C. elegans in liquid medium after pathogen exposure, resulting in cessation of pharyngeal pumping and a decrease in bacterial intake. We conclude that poor colonization in liquid is likely due to a combination of environmental factors and host-pathogen interactions. These results provide new insights into mechanisms for colonization in different models, enabling pathogenesis models to be fine-tuned to more accurately represent the conditions seen in human infections so that new tools for curbing bacterial and fungal infections can be developed.

Abstract 2202: Role of FadA secreting Fusobacterium nucleatum in initiation and progression of colorectal cancer

Abstract Colorectal cancer (CRC) is a global disease with poor prognosis. There is an alarming increase in incidence of CRC in younger patients along with well recognized racial disparities. Recent data implicate a role of microbiome in initiation and progression of CRC. Fusobacterium nucleatum (Fn), an oral opportunistic pathogen, has emerged as one of the common pathogens associated with CRC. Fn transformation from commensal to pathogen requires expression of amyloid-like FadA adhesin that plays a key role in its attachment to the colonic mucosal cells resulting in inflammatory, oncogenic, and metastatic changes. CRC initiates as polyps that go on to invade the mucosal layers and metastasize. This study analyzed the significance of Fn as a biomarker in CRC as well as to understand the etiology and metastasis of CRC. Deidentified and delinked formalin fixed paraffin embedded tissue sections from cancerous, precancerous and non-cancerous colorectal biopsies were analyzed for Fn presence and markers in epithelial to mesenchymal transition (EMT) in CRC through immunofluorescence. EMT was studied through biomarkers such as E-cadherin, which is an epithelial cell surface protein, vimentin as an intracellular filament protein normally expressed in mesenchymal cells, and N-cadherin, a cell adhesion molecule that is shown to express aberrantly in some solid tumor cells undergoing EMT. As expected, E-cadherin was abundantly expressed on the surface of epithelial cells while vimentin was found in the stromal cells in the non-cancerous tissue sections. Interestingly, the E-cadherin expression was lower in the cancerous tissue and some cancerous specimens also exhibited the vimentin positive foci within epithelial cells signifying partial EMT. N-cadherin expression was not observed in any of the specimens. The presence of Fn as analyzed through the FadA staining correlated with decline in the E-cadherin expression in some cancer specimens. To further analyze the significance of Fn, a meta-analytic study was conducted from 20 studies that indicated a significant presence of Fn not only in CRC tissues but also in fecal specimens from patients with CRC. A whole genome sequencing analysis corroborated with the abundance of Fn in CRC tissue specimens. These data suggest that analysis for Fn presence offers an attractive opportunity for intervention and targeted treatment for chemo resistant tumors. EMT analysis provides an additional corroboration in early detection of cancer progression which is pivotal in successful treatment of CRC. Citation Format: Angela Huang, Anjana Saxena, Mintoo Patel. Role of FadA secreting Fusobacterium nucleatum in initiation and progression of colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2202.

Cell-cell adhesion control in multicellular assemblies under the influence of external fluid flow.

Understanding the dynamics and viscoelastic properties of growing multicellular bacterial assemblies under external fluid flow is important for many applications. What behaviors are possible, and how they can be controlled with the strength and selectivity of cell-cell adhesins is largely unknown. Here we take a synthetic biology approach by controlling the expression of synthetic cell-cell adhesins in bacterial aggregates attached to a surface while subjecting these aggregates to external fluid flows. We experimentally characterize the phase space of possible behaviors of these aggregates based on flow conditions and adhesion strength. We identify a rich set of behaviors including cluster growth, cluster deformation, and intermittent cell shedding. We present a biophysical model explaining the observed behaviors. These results provide fundamental insights for active matter research, and will inform the behavior of clinical biofilms, the evolutionary constraints of multicellular life, and have applications like active biomaterials, among others.

Testosterone and estradiol modify the expression of adhesins and biofilm formation in Actinobacillus seminis.

Actinobacillus seminis is the causal agent of epididymitis and has other effects on the reproductive tracts of small ruminants and bovines. This bacterium causes infection when luteinizing (LH) or follicle-stimulating hormones increase, and hosts reach sexual maturity. LH induces female ovulation and male testosterone production, suggesting that these hormones affect A. seminis pathogenicity. In the present study, we evaluated the effect of testosterone (1-5ng/ml) or estradiol (5-25 pg/ml) added to culture medium on the in vitro growth, biofilm production, and adhesin expression of A. seminis. Estradiol does not promote the growth of this bacterium, whereas testosterone increased A. seminis planktonic growth 2-fold. Both hormones induced the expression of the elongation factor thermo unstable (EF-Tu) and phosphoglycerate mutase (PGM), proteins that A. seminis uses as adhesins. Estradiol (5 or 10 pg/ml) decreased biofilm formation by 32%, whereas testosterone, even at 5ng/ml, showed no effect. Both hormones modified the concentrations of carbohydrates and eDNA in biofilms by 50%. Amyloid proteins are characterized by their capacity to bind Congo red (CR) dye. Actinobacillus seminis binds CR dye, and this binding increases in the presence of 5-20 pg/ml estradiol or 4ng/ml testosterone. The A. seminis EF-Tu protein was identified as amyloid-like protein (ALP). The effect of sexual hormones on the growth and expression of virulence factors of A. seminis seems to be relevant for its colonization and permanence in the host.

Regulation in the Expression of Virulence Factors of Helicobacter pylori by the Interaction between AGS Cells and H. pylori

Helicobacter pylori (H. pylori) was defined as a Class 1 pathogenic carcinogen by WHO causing chronic inflammation in the stomach, thereby increasing the risk of gastric cancer. Various virulence factors are involved in the mechanism of gastric cancer caused by H. pylori infection. These virulence factors usually show different expression levels depending on the environment of H. pylori, which can affect the risk of gastric cancer. In this study, the differences in the expression levels of major virulence factors of H. pylori depending on the environment were investigated by comparing expression levels of H. pylori cultured with AGS or alone. As a result, there was no difference in the expression of adhesins of alpA, sabA, and babA even after co-culture with AGS cells. In addition, the co-culture environment did not induce a difference in the expression levels of flaA and ureB. On the other hand, H. pylori co-cultured with AGS cells showed low expression levels of cagA, groEL, and oipA and high expression of vacA compared to H. pylori cultured alone. Our results suggest that not only the presence or absence of virulence factor genes but also differences in expression levels should be considered when evaluating the risk of gastric cancer after H. pylori infection based on the virulence factors.

Efficacy of Qingre Huashi decoction on infection of : inhibiting adhesion, antioxidant, and anti-inflammation.

To investigate the phytochemical profile of Qingre Huashi decoction (, QHD) and evaluate the mechanisms rationalizing the use of QHD against ()-associated gastritis. QHD is composed of 11 herbs, which was prepared by a fixed Pharmacy and concentrated into clear paste. High-performance liquid chromatography coupled with quadrupole time-of-flight tandem mass spectrometry (HPLC-QTOF/MS) was used to detect the phytochemical profile of QHD. The toxicity of QHD against and human gastric epithelial cells was evaluated by the toxicology test and cell counting kit-8 assay. The adhesion model was constructed by incubating and gastric mucosal epithelial cells for 2 h. The urease assay was used to examine the anti-adhesion effects of QHD, and gene expression of adhesins was evaluated by quantitative polymerase chain reaction. The antioxidant activity was assessed by DCFH-DA labeling. To evaluate the anti-inflammatory effect, the levels of pro-inflammatory cytokines in the culture supernatant were detected by enzyme-linked immunosorbent assay. HPLC-QTOF/MS profiling indicated the presence of primary compounds 1-20 in QHD. Drug concentration was determined as 1, 2, and 5 mg/mL by the toxic concentration of QHD against and human gastric epithelial cells. QHD prevented adhesion to the human gastric epithelial cells and reduced levels of reactive oxygen species. QHD also reduced the level of interleukin-8 and other pro-inflammatory cytokines that were upregulated by infection. QHD could inhibit adhesion, and exert antioxidant and anti-inflammatory effects .

Prevalence of the fimbrial operon mrkABCD, mrkA expression, biofilm formation and effect of biocides on biofilm formation in carbapenemase-producing Klebsiella pneumoniae isolates belonging or not belonging to high-risk clones

The role of mrkA adhesin expression, biofilm production, biofilm viability and biocides in the biofilm of carbapenemase-producing Klebsiella pneumoniae isolates was investigated. Seventeen isolates representing different sequence types and carbapenemases were investigated. mrkA expression was determined by real-time reverse transcription polymerase chain reaction. Biofilm production (25°C and 37°C, with and without humidity) was determined by the crystal violet assay. The effect of isopropanol, povidone-iodine, sodium hypochlorite, chlorhexidine digluconate, benzalkonium chloride, ethanol and triclosan on biofilm was determined. The effect of povidone-iodine on biofilm biomass and thickness was also determined by confocal laser scanning microscopy. mrkA expression ranged from 28.2 to 1.3 [high or intermediate level; 64% of high-risk (HR) clones] and from 21.5 to 1.3 (50% of non-HR clones). At 25°C, biofilm formation was observed in 41% of isolates (absence of humidity) and 35% of isolates (presence of humidity), whereas at 37°C, biofilm formation was observed in 76% of isolates with and without humidity. At 25°C, biofilm producers were more frequently observed in HR clones (45% with humidity and 55% without humidity) than non-HR clones (17% with and without humidity). Biofilm viability from day 21 was higher at 25°C than 37°C. The greatest decrease in biofilm formation was observed with povidone-iodine (29% decrease), which also decreased biofilm thickness. Biofilm formation in carbapenemase-producing K. pneumoniae is related to mrkA expression. Biofilm formation is affected by temperature (37°C>25°C), whereas humidity has little effect. Biofilm viability is affected by temperature (25°C>37°C). At 25°C, HR clones are more frequently biofilm producers than non-HR clones. Povidone-iodine can decrease biofilm production and biofilm thickness.

Competitiveness during Dual-Species Biofilm Formation of Fusarium oxysporum and Candida albicans and a Novel Treatment Strategy.

During an infection, a single or multispecies biofilm can develop. Infections caused by non-dermatophyte molds, such as Fusarium spp. and yeasts, such as Candida spp., are particularly difficult to treat due to the formation of a mixed biofilm of the two species. Fusarium oxysporum is responsible for approximately 20% of human fusariosis, while Candida albicans is responsible for superficial mucosal and dermal infections and for disseminated bloodstream infections with a mortality rate above 40%. This study aims to investigate the interactions between C. albicans and F. oxysporum dual-species biofilm, considering variable formation conditions. Further, the ability of the WMR peptide, a modified version of myxinidin, to eradicate the mixed biofilm when used alone or in combination with fluconazole (FLC) was tested, and the efficacy of the combination of WMR and FLC at low doses was assessed, as well as its effect on the expression of some biofilm-related adhesin and hyphal regulatory genes. Finally, in order to confirm our findings in vivo and explore the synergistic effect of the two drugs, we utilized the Galleria mellonella infection model. We concluded that C. albicans negatively affects F. oxysporum growth in mixed biofilms. Combinatorial treatment by WMR and FLC significantly reduced the biomass and viability of both species in mature mixed biofilms, and these effects coincided with the reduced expression of biofilm-related genes in both fungi. Our results were confirmed in vivo since the synergistic antifungal activity of WMR and FLC increased the survival of infected larvae and reduced tissue invasion. These findings highlight the importance of drug combinations as an alternative treatment for C. albicans and F. oxysporum mixed biofilms.