Biology subjects

Vincent, A. T.

Publications and source records attributed to Vincent, A. T..

15 recordsLinked to original sources

bioRxiv2026

Isw2-mediated chromatin remodeling governs antifungal tolerance and heteroresistance in Candida albicans

Antifungal tolerance, unlike resistance, allows cells to grow slowly at concentrations above the minimum inhibitory concentration. While resistance mechanisms are well characterized, the pathways underlying tolerance remain elusive. Here, we performed a genetic screen of a transcriptional factor mutant library to identify regulators of azole tolerance in Candida albicans. This screen uncovered Isw2, the catalytic subunit of the ISW2 ATP-dependent chromatin remodeling complex, as a negative regulator of azole tolerance. Integrating transcriptomics, Isw2 DNA-binding profiles, and nucleosome-occupancy analyses revealed that Isw2 maintains a repressive chromatin architecture at the CRZ1 promoter, limiting the nucleosome-depleted region and suppressing CRZ1 transcription. Isw2 also modulates fluconazole heteroresistance and amphotericin B sensitivity through Crz1. In addition, we identified the copper-responsive transcription factor Mac1 as a context-dependent regulator of azole tolerance, acting negatively under copper limitation but positively under copper-replete conditions. Together, these findings reveal unexpected roles for chromatin remodeling and copper homeostasis in antifungal tolerance.

microbiology↗

bioRxiv2025

Crosstalk between the Ino80 complex and TOR signaling drives fungal adaptation to hypoxia through chromatin remodeling

In the human host, the opportunistic yeast Candida albicans must adapts to niches that vary markedly in oxygen tension, ranging from the hypoxic gastrointestinal and vaginal tracts to more oxygen-rich niches such as the skin. Thriving across this spectrum requires the capacity to dynamically rewire core metabolic pathways in response to fluctuating oxygen levels. Yet, despite the central role of oxygen in shaping fungal metabolism and fitness, the genetic basis underlying hypoxic adaptation in C. albicans remains incompletely understood. Here, we performed a genetic screen to identify genes required for adaptation to hypoxia (5% O2). This comprehensive approach revealed numerous genes spanning diverse metabolic and stress-related functions that are essential for optimal growth under oxygen-limiting conditions. Notably, this study uncovers a previously unrecognized role for the TOR (Target of Rapamycin) signaling pathway and the chromatin-remodeling complex Ino80 in orchestrating the fungal adaptive response to hypoxia. Integration of ino80 mutant transcriptomic data with Ino80 DNA-binding and nucleosome-occupancy profiles under conditional INO80 depletion revealed that this chromatin remodeler regulates the cellular phosphate demand associated with hypoxic exposure. The similarity between chromatin accessibility changes following TOR1 and INO80 depletion further suggests that Ino80 acts in concert with the TOR pathway to support growth and maintain phosphate homeostasis under low-oxygen conditions. Collectively, these findings provide new insight into how nutrient and oxygen signaling converge on chromatin remodeling, offering a mechanistic framework for understanding the regulatory logic underlying hypoxic adaptation in pathogenic fungi. ImportanceCandida albicans is an opportunistic fungal pathogen that colonizes diverse host niches, many of which are oxygen-limited. Understanding how this organism adapts to hypoxia is critical for elucidating the mechanisms underpinning both commensalism and pathogenicity of this yeast. Here, we reveal that the TOR signaling pathway and the Ino80 chromatin-remodeling complex are key regulators of fungal growth under low-oxygen conditions. Our data show that Ino80 coordinates phosphate homeostasis during hypoxia and functions in concert with TOR to maintain chromatin accessibility and metabolic adaptation. Our results uncover how oxygen and nutrient cues converge on chromatin remodeling, providing a framework for understanding how C. albicans and other human-associated fungi modulate their physiology to thrive and evolve in diverse host environments.

microbiology↗

bioRxiv2025

PqsE is conserved and functionally relevant in Pseudomonas aeruginosa

The bacterium Pseudomonas aeruginosa is an opportunistic pathogen responsible for several acute and chronic infections. It produces a diverse array of virulence and survival determinants, many of which are tightly regulated by three interlinked quorum sensing (QS) systems named las, rhl and pqs. RhlR, the transcriptional regulator of the rhl system, activates multiple virulence genes upon binding to its cognate autoinducer signal. Meanwhile, the pqs system relies on 4-hydroxy-2-alkylquinolines (HAQs) as signaling molecules to induce the transcriptional regulator MvfR (PqsR). MvfR then activates the transcription of the pqsABCDE operon, encoding enzymes for HAQ biosynthesis. The final gene in this operon encodes PqsE, a multifunctional protein unique to P. aeruginosa. Beyond its thioesterase activity in HAQ biosynthesis, PqsE stabilizes RhlR, facilitating its regulation of target genes, some of which are implicated in virulence. Because of its role in pathogenicity, PqsE is regarded as a promising therapeutic target for combating P. aeruginosa infections. While the role of PqsE towards the RhlR regulon is increasingly understood in prototypical P. aeruginosa strains such as PA14 and PAO1, its broader relevance as an anti-virulence target remains underexplored. Here, we confirm that PqsE is functionally relevant across a panel of twelve genetically diverse P. aeruginosa strains using metabolite quantification and phenotypic assays. Significant strain-to-strain variations further highlight the importance of studying QS regulation among diverse isolates. Moreover, this study underscores PqsE as a key QS regulator with a conserved role in coordinating virulence determinant production and social behaviors across diverse P. aeruginosa populations. IMPORTANCEPseudomonas aeruginosa is a versatile opportunistic pathogen, naturally tolerant and readily acquiring resistance to multiple antibiotics. Consequently, the World Health Organization identified this bacterium as a high priority pathogen for researching and developing new antimicrobial strategies. P. aeruginosa utilizes quorum sensing, a cell-to-cell communication system, to regulate the expression of several of its virulence factors. Here, we confirm that the PqsE protein is conserved and that its function in quorum sensing, especially towards the RhlR regulator, is maintained across a panel of twelve genetically diverse P. aeruginosa strains. Since PqsE is conserved and unique to this bacterium, it could serve as an ideal target for anti-virulence therapies, offering new alternatives to combat antimicrobial resistance.

microbiology↗

bioRxiv2025

Met32 governs transcriptional control of sulfur metabolic flexibility and resistance to reactive sulfur species in the human fungal pathogen Candida albicans

Although considerable advances have been made in understanding the metabolic machinery that enables bacteria to utilize sulfur sources, many aspects of this process remain understudied in fungi. To explore the genetic circuit by which the highly prevalent human opportunistic yeast Candida albicans controls sulfur utilization, we characterized the transcriptional landscape associated with sulfur starvation in this fungus. We identified many desulfonation enzymes that were differentially modulated and showed that Jlp12, a sulfonate/-ketoglutarate dioxygenase, was critical for the utilization of different sulfur sources found in many niches of the human host. We also uncovered that the Zinc-finger transcription factor Met32 acts as a master regulator, that modulates genes of sulfur utilization including Jlp12. Importantly, we found that C. albicans Met32 exclusively regulates sulfur utilization genes, while in the Saccharomyces cerevisiae lineage, it controls methionine biosynthesis. This work also identified Seo13 as the first major facilitator superfamily transporter in fungi that transports the alternative sulfur source glutathione, under the direct control of Met32. Furthermore, we showed that Met32 modulates C. albicans tolerance to sulfite excess by tuning the basal transcriptional level of the superoxide dismutase Sod1. This underscores the dual role of Met32 in the breakdown of sulfur-containing metabolites and the neutralization of the resulting reactive sulfur species (RSS). Our study delineates a new mechanism by which fungal pathogens utilize sulfur sources and neutralize RSS and underscores its importance in fungal fitness in vivo. ImportanceCandida albicans is the most prevalent fungal colonizer of humans and it is also the first cause of disseminated fungal infections leading to a high mortality rate. The ability of this yeast to metabolize a plethora of carbon and nitrogen sources inside the host is a critical asset for both the commensal and the pathogenic lifestyles of this yeast. Thus, these pathways represent attractive targets for antifungal therapy. While sulfur is an essential nutritional element for all living organisms, its contribution to fungal virulence remains understudied. Here, we describe new players of sulfur utilization metabolism in C. albicans and underline their importance in supporting fungal virulence. This work emphasizes the significance of targeting sulfur metabolic flexibility to manage fungal infections.

microbiology↗

bioRxiv2025

Manganese homeostasis modulates glucan and chitin unmasking in the opportunistic yeast Candida albicans

Candida albicans is a commensal fungus and also the most prevalent human fungal pathogen. The ability of this opportunistic yeast to acquire and maintain homeostatic levels of manganese (Mn), particularly in the metal-limited host environment, is an important determinant of its fitness. Recent studies have underscored the importance of Mn acquisition through members of Smf transporters, in C. albicans virulence and its ability to withstand various stresses. In the present study, we undertook transcriptional profiling in the mutant of the Mn transporter Smf12 under restricted Mn availability to identify processes that are directly affected in defective Mn uptake. Our analysis revealed that smf12 displayed a transcriptional pattern suggestive of a cell wall defect, with many transcripts associated with cell wall biogenesis being differentially regulated. smf12 together with smf11, a mutant of the closest homolog of Smf12, exhibited hypersensitivity to cell wall stressors and an altered cell wall ultrastructure. The smf mutants also exhibited unmasking of both {beta}-glucan and chitin, which unexpectedly resulted in a decreased rate of phagocytosis by macrophages, suggesting impaired recognition or internalization--an observation that challenges the prevailing paradigm. Furthermore, we showed that Mn-mediated unmasking of {beta}-glucan required modulation of glucanase activity and was not mediated through the calcineurin pathway. This study uncovers a novel role for Mn in maintaining cell wall integrity and modulating the exposure of fungal antigenic determinants, further emphasizing the critical role of this metal in supporting the opportunistic nature of C. albicans.

microbiology↗

bioRxiv2024

The vprAB-ompV-virK operon of Vibrio cholerae senses antimicrobial peptides and activates the expression of multiple resistance systems

Antimicrobial peptides are small cationic molecules produced by eukaryotic cells to combat infection, as well as by bacteria for niche competition. Polymyxin B (PmB), a cationic cyclic antimicrobial peptide, is used prophylactically in livestock for infection prevention and as a last-resort treatment for multidrug-resistant bacterial infections in humans. In this study, a transcriptomic analysis in Vibrio cholerae showed that expression of the uncharacterized gene ompV is stimulated in response to PmB. We found that ompV is organized in a conserved four-gene operon with the two-component system vprAB (carRS) and virK in V. cholerae, and that these genes are also upregulated in response to PmB treatment. A virK deletion mutant was more sensitive to the human cathelicidin LL-37 than the wild-type strain, while an ompV mutant was more sensitive to PmB and LL-37, suggesting that both OmpV and VirK contribute to antimicrobial resistance in V. cholerae. This increased sensitivity to antimicrobial peptides was not due to membrane destabilization or reduced sequestration by membrane vesicles as a result of ompV deletion. Instead, our transcriptomic analysis showed that the efflux pump vexAB, a known effector of PmB resistance, was also upregulated in the presence of PmB in an ompV-dependent manner. Examination of the predicted structure of OmpV revealed a lateral opening in the {beta}-barrel wall with access to an electronegative pocket in the barrel lumen that can accommodate PmB. Such an interaction could facilitate intracellular signaling through a conformational change in OmpV. This is the first evidence of a specialized operon governing multiple systems for antimicrobial resistance in V. cholerae. Author SummaryIn this study, we identified the first specialized operon controlling multiple systems of antimicrobial resistance in V. cholerae. The operon encodes the two-component system vprAB, which activates the main mechanism of polymyxin B resistance in V. cholerae, and the uncharacterized genes ompV and virK. We provide evidence that OmpV and VirK are implicated in antimicrobial resistance and show that OmpV has a membrane-accessible lateral opening into a pocket that could accommodate the antimicrobial peptide polymyxin B. We propose that OmpV acts as an outer membrane sensor that signals the presence of antimicrobial peptides to activate the expression of the operon, leading to the activation of multiple mechanisms of resistance, including modifications of the outer membrane and the multi-drug efflux system vexAB.

microbiology↗

bioRxiv2023

Manganese homeostasis modulates fungal virulence and stress tolerance in Candida albicans

Due to the scarcity of transition metals within the human host, fungal pathogens have evolved sophisticated mechanisms to uptake and utilize these micronutrients at the infection interface. While considerable attention was turned to iron and copper acquisition mechanisms and their importance in fungal fitness, less was done regarding either the role of manganese (Mn) in infectious processes or the cellular mechanism by which fungal cells achieve their Mn- homeostasis. Here, we undertook transcriptional profiling in the pathogenic fungus Candida albicans experiencing both Mn starvation and excess to capture biological processes that are modulated by this metal. We uncovered that Mn scarcity influences diverse processes associated with fungal fitness including invasion of host cells and antifungal sensitivity. We show that Mn levels influence the abundance of iron and zinc emphasizing the complex crosstalk between metals. Deletion of SMF12, a member of Mn Nramp transporters confirmed its contribution to Mn uptake. smf12 was unable to form hyphae and damage host cells and exhibited sensitivity to azoles. We found that the unfolded protein response (UPR), likely activated by decreased glycosylation under Mn limitation, was required to restore growth when cells were shifted from a Mn-starved to a Mn-repleted medium. RNA-seq profiling of cells exposed to Mn excess revealed that UPR was also activated. Furthermore, the UPR signaling axis Ire1-Hac1 was required to bypass Mn toxicity. Collectively, this study underscores the importance of Mn homeostasis in fungal virulence, and comprehensively provides a portrait of biological functions that are modulated by Mn in a fungal pathogen.

microbiology↗

bioRxiv2023

The Adr1 transcription factor directs regulation of the ergosterol pathway and azole resistance in C. albicans

Transcription factors play key roles in cellular regulation and are critical in the control of drug resistance in the fungal pathogen Candida albicans. We found that activation of the transcription factor C4_02500C_A (Adr1) conferred significant resistance against fluconazole. In Saccharomyces cerevisiae, Adr1 is a carbon-source-responsive zinc-finger transcription factor required for transcription of the glucose-repressed gene ADH1 and of genes required for ethanol, glycerol, and fatty acid utilization. Motif scanning of promoter elements suggests that Adr1 may be rewired in fungi and governs the ergosterol synthesis pathway in C. albicans. Because previous studies have identified the zinc-cluster transcription factor Upc2 as a regulator of the ergosterol pathway in both fungi, we examined the relationship of Adr1 and Upc2 in sterol biosynthesis in C. albicans. Phenotypic profiles of either ADR1 and UPC2 modulation in C. albicans showed differential growth in the presence of fluconazole; either adr1 or upc2 homozygous deletion results in sensitivity to the drug while their activation generates a fluconazole resistant strain. The rewiring from ergosterol synthesis to fatty acid metabolism involved all members of the Adr1 regulon except the alcohol dehydrogenase Adh1, which remains under Adr1 control in both circuits and may have been driven by the lifestyle of S. cerevisiae, which requires the ability to both tolerate and process high concentrations of ethanol.

genetics↗

bioRxiv2022

Adaptive radioresistance of Enterohemorrhagic Escherichia coli O157:H7 results in genomic loss of shiga toxin-encoding prophages

Enterohemorragic Escherichia coli (EHEC) O157:H7 is a foodborne pathogen producing shiga toxins (Stx1 and Stx2), can cause hemorrhagic diarrhea, and life-threatening infections. O157:H7 strain EDL933 carries prophages CP-933V and BP-933W that encode shiga toxin genes (stx1 and stx2 respectively). The aim of this work was to investigate the mechanisms of adaptive resistance of EHEC strain EDL933 to a typically "lethal" dose of {gamma}-irradiation (1.5 kGy). Adaptive selection through six passages of exposure to 1.5 kGy resulted in the loss of CP-933V and BP-933W prophages from the genome and mutations within three genes: wrbA, rpoA, and Wt_02639 (molY). Three selected EHEC clones that became irradiation-adapted to the 1.5 kGy dose (C1, C2 and C3) demonstrated increased resistance to oxidative stress, sensitivity to acid pH, and decreased cytotoxicity to Vero cells. To confirm that loss of prophages plays a role in increased radioresistance, C1 and C2 clones were exposed to bacteriophage containing lysates. Although, phage BP-933W could lysogenize C1, C2, and E.coli K-12 strain MG1655, it was not found to have integrated into the bacterial chromosome in C1-{Phi} and C2-{Phi} lysogens. Interestingly, for the E. coli K-12 lysogen (K12-{Phi}), BP-933W DNA had integrated at the wrbA gene (K12-{Phi}). Both C1-{Phi} and C2-{Phi} lysogens regained sensitivity to oxidative stress, were more effectively killed by a 1.5 kGy {gamma}-irradiation dose and had regained cytotoxicity and acid resistance phenotypes. Further, the K12-{Phi} lysogen became cytotoxic, more sensitive to {gamma}-irradiation and oxidative stress and slightly more acid resistant. ImportanceGamma ({gamma})-irradiation of food products can provide an effective means of eliminating bacterial pathogens such as enterohemorrhagic Escherichia coli (EHEC) O157:H7, a significant foodborne pathogen that can cause severe disease due to the production of Shiga toxins. To decipher the mechanisms of adaptive resistance of the O157:H7 strain EDL933, we evolved clones of this bacterium resistant to a lethal dose of {gamma}-irradiation by repeatedly exposing bacterial cells to irradiation following a growth restoration over six successive passages. Our findings provide evidence that adaptive selection involved modifications in the bacterial genome including deletion of the CP-933V and BP-933W prophages. These mutations in EHAC O157:H7 resulted in loss of stx1, stx2, loss of cytotoxicity to epithelial cells and decreased resistance to acidity, critical virulence determinants of EHEC, concomitant with increased resistance to lethal irradiation and oxidative stress. These findings demonstrate that the potential adaptation of EHEC to high doses of radiation would involve elimination of the Stx encoding phages and likely lead to a substantial attenuation of virulence.

microbiology↗

bioRxiv2022

Pseudomonas aeruginosa strains from both clinical and environmental origins readily adopt a stable small colony variant (SCV) phenotype resulting from single mutations in c-di-GMP pathways

A subpopulation of Small Colony Variants (SCVs) is a frequently observed feature of Pseudomonas aeruginosa isolated from colonized cystic fibrosis lungs. Since most SCVs have until now been isolated from clinical samples, it remains unclear how widespread is the ability of P. aeruginosa to develop this phenotype and the genetic mechanism(s) behind SCVs emergence according to the origin of the isolate. In the present work, we investigated the ability of 22 P. aeruginosa isolates from various environmental origins to, under laboratory culture conditions, spontaneously adopt a SCV-like smaller alternative morphotype distinguishable from the ancestral parent strain. We found that all the P. aeruginosa strains tested could adopt a SCV phenotype, regardless of their origin. Whole genome sequencing of SCVs obtained from clinical and environmental sources revealed single mutations exclusively in two distinct c-di-GMP signaling pathways, Wsp and YfiBNR. We conclude that the ability to switch to a SCV phenotype is a conserved feature of P. aeruginosa and results from the acquisition of a stable genetic mutation, regardless of the origin of the strain. IMPORTANCEP. aeruginosa is an opportunistic pathogen that thrives in many environments. It poses a significant health concern, notably because this bacterium is the most prevalent pathogen found in the lungs of people with cystic fibrosis. In infected hosts, its persistence is considered related to the emergence of an alternative small colony variant (SCV) phenotype. By reporting the distribution of P. aeruginosa SCVs in various non-clinical environments and the involvement of c-di-GMP in SCV emergence from both clinical and environmental strains, this work contributes to understanding a conserved adaptation mechanism used by P. aeruginosa to adapt readily in all environments. Hindering this adaptation strategy could help control P. aeruginosa persistent infection.

microbiology↗

bioRxiv2021

Experimental evolution of Vibrio cholerae: Identification of genes involved in motility in presence of polymyxin B

Vibrio cholerae is the bacteria responsible for the cholera disease and a natural inhabitant of aquatic environments. Biofilm formation is important for human colonization and environmental survival. Motility is essential for adhesion and biofilm formation by V. cholerae. In a previous study, we showed that motility and biofilm formation are altered in the presence of sub-inhibitory concentrations of polymyxin B in V. cholerae. In this study, we performed an experimental evolution to identify genes rescuing the motility in the presence of polymyxin B. Mutations in 5 genes have been identified: ihfA, vacJ (mlaA), mlaF, dacB and ccmH. The details of these mutations, their potential impact on the function of the proteins they encode and on the motility in presence of polymyxin B are discussed.

microbiology↗

bioRxiv2021

Pseudomonas aeruginosa isolates defective in function of the LasR quorum sensing regulator are frequent in diverse environmental niches

The saprophyte Pseudomonas aeruginosa is a versatile opportunistic pathogen causing infections in immunocompromised individuals. To facilitate its adaptation to a large variety of niches, this bacterium exploits population density-dependant gene regulation systems called quorum sensing. In P. aeruginosa, three distinct but interrelated quorum sensing systems (las, rhl and pqs) regulate the production of many survival and virulence functions. In prototypical strains, the las system, through its transcriptional regulator LasR, is important for the full activation of the rhl and pqs systems. Still, LasR-deficient isolates have been reported, mostly sampled from the lungs of people with cystic fibrosis, where they are considered selected by the chronic infection environment. In this study, we show that a defect in LasR activity appears to be an actually widespread mechanism of adaptation in this bacterium. Indeed, we found abundant LasR-defective isolates sampled from hydrocarbon-contaminated soils, hospital sink drains, and meat/fish market environments, using an approach based on phenotypic profiling, supported by gene sequencing. Interestingly, several LasR-defective isolates maintain an active rhl system or are deficient in pqs system signaling. The high prevalence of a LasR-defective phenotype among environmental P. aeruginosa isolates questions the role of quorum sensing in niche adaptation.

microbiology↗

bioRxiv2020

The oxidative stress response and virulence of pathogenic Leptospira are controlled by the interplay of two peroxide stress regulators

Pathogenic Leptospira are the causative agents of leptospirosis, the most widespread zoonotic infectious disease. Leptospirosis is a potentially severe and life-threatening emerging disease with highest burden in sub-tropical areas and impoverished populations. Mechanisms allowing pathogenic Leptospira to survive inside a host and induce acute leptospirosis are not fully understood. The ability to resist deadly oxidants produced by the host during infection is pivotal for Leptospira virulence. We have previously shown that genes encoding defenses against oxidants in L. interrogans are repressed by PerRA (encoded by LIMLP_10155), a peroxide stress regulator of the Fur family. In this study, we describe the identification and characterization of another putative PerR-like regulator (LIMLP_05620) in L. interrogans. Protein sequence and phylogenetic analyses indicated that LIMLP_05620 displayed all the canonical PerR amino acid residues and is restricted to pathogenic Leptospira clades. We therefore named this PerR-like regulator PerRB. In L. interrogans, the PerRB regulon is distinct from that of PerRA. While a perRA mutant had a greater tolerance to peroxide, inactivating perRB led to a higher tolerance to superoxide, suggesting that these two regulators have a distinct function in the adaptation of L. interrogans to oxidative stress. The concomitant inactivation of perRA and perRB resulted in a higher tolerance to both peroxide and superoxide and, unlike the single mutants, a double perRAperRB mutant was avirulent. Interestingly, this correlated with major changes in gene and non-coding RNA expression. Notably, several virulence-associated genes (clpB, ligA/B, and lvrAB) were repressed. By obtaining a double mutant in a pathogenic Leptospira strain, our study has uncovered an interplay of two PerRs in the adaptation of Leptospira to oxidative stress with a putative role in virulence and pathogenicity, most likely through the transcriptional control of a complex regulatory network. Author summaryLeptospirosis is a widespread infectious disease responsible for over one million of severe cases and 60 000 fatalities annually worldwide. This neglected and emerging disease has a worldwide distribution, but it mostly affects populations from developing countries in sub-tropical areas. The causative agents of leptospirosis are pathogenic bacterial Leptospira spp. There is a considerable deficit in our knowledge of these atypical bacteria, including their virulence mechanisms. In addition to the Leptospira PerRA regulator that represses defenses against peroxide, we have identified and characterized a second PerR regulator in pathogenic Leptospira species (PerRB) that participates in Leptospira tolerance to superoxide. Phenotypic and transcriptomic analyses of single PerRA and PerRB mutants suggest that the two PerRs fulfill distinct functions in the adaptation to oxidative stress. Concomitant inactivation of PerRA and PerRB resulted in a higher tolerance to both peroxide and superoxide. Moreover, the perRAperRB mutant lost its virulence. Major changes in gene expression, including a decreased expression of several virulence factors, were observed in the double perRAperRB mutant. Our study suggests that PerRA and PerRB cooperate to orchestrate a complex regulatory network involved in Leptospira virulence.

microbiology↗

bioRxiv2019

Genome sequencing and functional characterization of a Dictyopanus pusillus fungal extract offers a promising alternative for lignocellulose pretreatment of oil palm residues

The pretreatment of biomass is a critical requirement of bio-renewable fuel production from lignocellulose. Although current processes primarily involve chemical and physical approaches, the biological breakdown of lignin using enzymes and microorganisms is quickly becoming an interesting eco-friendly alternative to classical processes. As a result, bioprospection of wild fungi from naturally occurring lignin-rich sources remains a suitable method to uncover and isolate new species exhibiting ligninolytic activity. In this study, wild species of white rot fungi were collected from Colombian forests based on their natural wood decay ability and high capacity to secrete oxidoreductases with high affinity for phenolic polymers such as lignin. Based on high activity obtained from solid-state fermentation using a lignocellulose source from oil palm as matrix, we describe the isolation and whole-genome sequencing of Dictyopanus pusillus, a wild basidiomycete fungus exhibiting ABTS oxidation as an indication of laccase activity. Functional characterization of a crude enzymatic extract identified laccase activity as the main enzymatic contributor to fungal extracts, an observation supported by the identification of 13 putative genes encoding for homologous laccases in the genome. To the best of our knowledge, this represents the first report of an enzymatic extract exhibiting laccase activity in the Dictyopanus genera, offering means to exploit this species and its enzymes for the delignification process of lignocellulosic by-products from oil palm.

bioengineering↗

bioRxiv2019

Escherichia coli O157:H7 responds to phosphate starvation by modifying LPS involved in biofilm formation

In open environments such as water, enterohemorrhagic Escherichia coli O157:H7 responds to inorganic phosphate (Pi) starvation by inducing the Pho regulon controlled by PhoB. The phosphate-specific transport (Pst) system is the high-affinity Pi transporter. In the {Delta}pst mutant, PhoB is constitutively activated and regulates the expression of genes from the Pho regulon. In E. coli O157:H7, the {Delta}pst mutant, biofilm, and autoagglutination were increased. In the double-deletion mutant {Delta}pst {Delta}phoB, biofilm and autoagglutination were similar to the wild-type strain, suggesting that PhoB is involved. We investigated the relationship between PhoB activation and enhanced biofilm formation by screening a transposon mutant library derived from {Delta}pst mutant for decreased autoagglutination and biofilms mutants. Lipopolysaccharide (LPS) genes involved in the synthesis of the LPS core were identified. Transcriptomic studies indicate the influence of Pi-starvation and pst mutation on LPS biosynthetic gene expression. LPS analysis indicated that the O-antigen was deficient in the {Delta}pst mutant. Interestingly, waaH, encoding a glycosyltransferase associated with LPS modifications in E. coli K-12, was highly expressed in the {Delta}pst mutant of E. coli O157:H7. Deletion of waaH from the {Delta}pst mutant and from the wild-type strain grown in Pi-starvation conditions decreased the biofilm formation but without affecting LPS. Our findings suggest that LPS core is involved in the autoagglutination and biofilm phenotypes of the {Delta}pst mutant and that WaaH plays a role in biofilm in response to Pi-starvation. This study highlights the importance of Pi-starvation in biofilm formation of E. coli O157:H7, which may affect its transmission and persistence. IMPORTANCEEnterohemorrhagic Escherichia coli O157:H7 is a human pathogen responsible for bloody diarrhea and renal failures. In the environment, O157:H7 can survive for prolonged periods of time under nutrient-deprived conditions. Biofilms are thought to participate in this environmental lifestyle. Previous reports have shown that the availability of extracellular inorganic phosphate (Pi) affected bacterial biofilm formation; however, nothing was known about O157:H7 biofilm formation. Our results show that O157:H7 membrane undergoes modifications upon PhoB activation leading to increased biofilm formation. A mutation in the Pst system results in reduced amount of the smooth type LPS and that this could influence the biofilm composition. This demonstrates how the E. coli O157:H7 adapts to Pi starvation increasing its ability to occupy different ecological niches.

microbiology↗