mBio最新文献

{"title":"Noncanonical amino acids as prophage inducers for protein regulation in bacteria-based delivery systems.","authors":"Hongfang Liu, Sijia Shen, Qi Xu, Yuyang Wang, Kejing Qi, Bowen Lu, Bing Tang, Min Wu, Fei Gan","doi":"10.1128/mbio.03988-24","DOIUrl":"10.1128/mbio.03988-24","url":null,"abstract":"<p><p>Genetically engineered bacteria represent a promising drug delivery tool for disease treatment. The development of new strategies for specific and independent protein regulation is necessary, especially for combination protein drug therapy. Using the well-studied <i>Escherichia coli</i> phage λ as a model system, we applied <u>n</u>on<u>c</u>anonical <u>a</u>mino <u>a</u>cid<u>s</u> (ncAAs) as novel inducers for protein regulation in a bacteria-based delivery system. Screening the permissive sites of the Cro protein revealed that incorporation of AlocK at the K8 site with the <i>Mb</i>PylRS-349F/tRNA^Pyl system produced a functional Cro-K8AlocK variant. Using an engineered λ lysogen expressing the <i>Mb</i>PylRS-349F/tRNA^Pyl pair, Cro-8X, and the reporter mNeonGreen, <i>in vitro</i> and <i>in vivo</i> experiments showed that AlocK led to bacterial lysis through prophage activation and the release of mNeonGreen. If <i>mNeonGreen</i> was integrated into the λ prophage genome, λ phages released due to AlocK induction delivered the reporter gene into the recipient <i>E. coli</i> strain, enabling mNeonGreen expression. Furthermore, insertion of pIF at the F14 site with the <i>Af</i>pIFRS/tRNA^Tyr pair produced a functional Cro-F14pIF variant. Importantly, <i>Af</i>pIFRS/tRNA^Tyr and <i>Mb</i>PylRS-349F/tRNA^Pyl pairs were confirmed to be mutually orthogonal. In a mixture of two engineered λ lysogens expressing different aaRS/tRNAs, Cro-ncAAs, and reporter proteins, AlocK and pIF independently induced bacterial lysis and activated the expression of mNeonGreen and mCherry in the recipient <i>E. coli</i> strain. Collectively, the proposed bacteria-based delivery system provides two options for protein delivery and enables independent regulation of multiple proteins with ncAAs, offering a novel approach for <i>in situ</i> protein regulation and combination therapy.</p><p><strong>Importance: </strong>The use of genetically engineered bacteria as drug delivery vectors has attracted more and more attention in recent years. A key issue with bacteria-based delivery systems is how to regulate multiple protein drugs. Based on genetic code expansion technology, we developed a new strategy of using ncAAs as small molecular inducers for <i>in situ</i> protein regulation and engineered λ phage lysogen into a bacteria-based delivery system that can function in two delivery modes. Furthermore, this strategy enables independent regulation of multiple proteins by different ncAAs, offering important implications for combination therapy. This approach requires minimal genetic engineering efforts, and similar strategies can be applied to engineer other prophage-bacteria systems or study phage biology. This work expands the therapeutic applications of ncAAs and lysogenic phages.</p>","PeriodicalId":18315,"journal":{"name":"mBio","volume":" ","pages":"e0398824"},"PeriodicalIF":5.1,"publicationDate":"2025-04-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143625331","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Clade 2.3.4.4b H5N1 neuraminidase has a long stalk, which is in contrast to most highly pathogenic H5N1 viruses circulating between 2002 and 2020.","authors":"Enikő Hermann, Florian Krammer","doi":"10.1128/mbio.03989-24","DOIUrl":"10.1128/mbio.03989-24","url":null,"abstract":"<p><p>Since 2020, H5N1 highly pathogenic avian influenza (HPAI) viruses of clade 2.3.4.4b have been rapidly spreading in wild birds but have also caused a large number of mammalian infections and more than 70 known human cases. Importantly, this H5N1 clade has also crossed the species barrier into dairy cattle in the US in late 2023/early 2024. The neuraminidase (NA) protein of the N1 subtype can feature truncations in its stalk domain, which have been identified as putative virulence factors in poultry but seem to have a negative impact on transmission in mammals. Since its emergence, the vast majority of HPAI H5N1 A/goose/Guangdong/1/1996-lineage isolates have featured this truncated version of the NA stalk domain. Here, we report that this changed with the 2020 expansion of clade 2.3.4.4b H5N1 and that the majority of isolates-including the strains circulating in dairy cattle-feature a long NA stalk domain.IMPORTANCEWhile the truncated version of the N1 neuraminidase stalk domain may be associated with increased virulence in poultry, the long version of the stalk domain has been associated with increased transmissibility in mammals. The vast majority of highly pathogenic H5N1 of clade 2.3.4.4b that is currently circulating globally features the long stalk version of the neuraminidase, which may increase the risk for these viruses to become human-to-human transmissible.</p>","PeriodicalId":18315,"journal":{"name":"mBio","volume":" ","pages":"e0398924"},"PeriodicalIF":5.1,"publicationDate":"2025-04-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143502254","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Amplicon sequencing and culture-dependent approaches reveal core bacterial endophytes aiding freezing stress tolerance in alpine Rosaceae plants.","authors":"Malek Marian, Livio Antonielli, Ilaria Pertot, Michele Perazzolli","doi":"10.1128/mbio.01418-24","DOIUrl":"10.1128/mbio.01418-24","url":null,"abstract":"<p><p>Wild plants growing in alpine regions are associated with endophytic microbial communities that may support plant growth and survival under cold conditions. The structure and function of endophytic bacterial communities were characterized in flowers, leaves, and roots of three alpine Rosaceae plants in Alpine areas using a combined amplicon sequencing and culture-dependent approaches to determine the role of core taxa on plant freezing stress tolerance. Amplicon sequencing analysis revealed that plant tissue, collection site, and host plant are the main factors affecting the richness, diversity, and taxonomic structure of endophytic bacterial communities in alpine Rosaceae plants. Core endophytic bacterial taxa were identified as 31 amplicon sequence variants highly prevalent across all plant tissues. Psychrotolerant bacterial endophytes belonging to the core taxa of <i>Duganella</i>, <i>Erwinia</i>, <i>Pseudomonas</i>, and <i>Rhizobium</i> genera mitigated freezing stress in strawberry plants, demonstrating the beneficial role of endophytic bacterial communities and their potential use for cold stress mitigation in agriculture.IMPORTANCEFreezing stress is one of the major abiotic stresses affecting fruit production in Rosaceae crops. Current strategies to reduce freezing damage include physical and chemical methods, which have several limitations in terms of costs, efficacy, feasibility, and environmental impacts. The use or manipulation of plant-associated microbial communities was proposed as a promising sustainable approach to alleviate cold stress in crops, but no information is available on the possible mitigation of freezing stress in Rosaceae plants. A combination of amplicon sequencing, culture-dependent, and plant bioassay approaches revealed the beneficial role of the endophytic bacterial communities in alpine Rosaceae plants. In particular, we showed that culturable psychrotolerant bacterial endophytes belonging to the core taxa of <i>Duganella</i>, <i>Erwinia</i>, <i>Pseudomonas</i>, and <i>Rhizobium</i> genera can mitigate freezing stress on strawberry seedlings. Overall, this study demonstrates the potential use of psychrotolerant bacterial endophytes for the development of biostimulants for cold stress mitigation in agriculture.</p>","PeriodicalId":18315,"journal":{"name":"mBio","volume":" ","pages":"e0141824"},"PeriodicalIF":5.1,"publicationDate":"2025-04-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143492441","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Classical swine fever virus recruits ALIX and ESCRT-III to facilitate viral budding.","authors":"Jinxia Chen, Hanfei Yang, Mingyue Wan, Yan Cheng, Jishan Bai, Yuhang Li, Jing Chen, Bingqian Zhao, Fei Gao, Bin Zhou","doi":"10.1128/mbio.02618-24","DOIUrl":"10.1128/mbio.02618-24","url":null,"abstract":"<p><p>Classical swine fever virus (CSFV) incurs substantial economic losses in the global swine industry due to its persistent emergence and re-emergence across various countries. However, the precise mechanisms governing CSFV budding remain inadequately understood. Our study elucidates that the endosomal sorting complex required for transport (ESCRT)-associated protein ALIX, in conjunction with ESCRT-III, plays a pivotal role in orchestrating CSFV budding. Genomic sequence analysis identified a critical interaction between the YPXnL late domain on the E2 protein and ALIX. Through immunoprecipitation and structural domain deletion assays, we demonstrated that the ALIX Bro1 domain specifically recognized viral particles by binding to the YPXnL motif. Immunoelectron and transmission electron microscopy further confirmed that, upon infection, ALIX accumulated at the periphery of subcellular organelles, including COPII vesicles, endosomes, and the Golgi apparatus, thereby facilitating CSFV budding. Our findings also revealed that ESCRT-III subunits CHMP2B, CHMP4B, CHMP7, and VPS4A interacted with ALIX to expedite CSFV budding. Notably, Rab8 activated by Kif4A contributed to the release of CSFV particles by interacting with ALIX and directing ALIX-containing vesicles along microtubules toward the cytosol. Our study demonstrates that ALIX specifically recognizes E2 and orchestrates the recruitment of ESCRT-III and Rab8 to facilitate the vesicular budding of CSFV particles from the Golgi apparatus to the cytosol. Ultimately, virus-laden vesicles propelled by Kif4A are transported along microtubules to the plasma membrane for release. Our findings offer the first comprehensive elucidation of the CSFV budding process and contribute to the identification of antiviral targets, thereby advancing the development of antiviral therapeutics.IMPORTANCEThe endosomal sorting complex required for transport (ESCRT) machinery plays a pivotal role in the sorting of membrane proteins in eukaryotic cells and regulating various stages of infection for numerous viruses. Previous studies have underscored the indispensable role of ESCRT in the cellular entry and replication of classical swine fever virus (CSFV). However, the precise mechanisms by which ESCRT recognizes CSFV particles and initiates viral vesicle budding have remained elusive. This study reveals that the Bro1 domain of ALIX initiates viral budding proximal to the Golgi apparatus by specifically recognizing the YPXnL late domain on the CSFV E2 protein. Mechanistically, ALIX and ESCRT-III facilitate Rab8-regulated endosomal transport of CSFV particles from the Golgi apparatus to the plasma membrane. Subsequently, virions are propelled by the kinesin Kif4A along microtubules for egress into the extracellular space. In summary, these findings significantly advance our understanding of CSFV pathogenesis and offer valuable insights into the vesicular transport and budding mechanisms of CSFV particles.</p>","PeriodicalId":18315,"journal":{"name":"mBio","volume":" ","pages":"e0261824"},"PeriodicalIF":5.1,"publicationDate":"2025-04-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143492444","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Quorum sensing employs a dual regulatory mechanism to repress T3SS gene expression.","authors":"Payel Paul, Ram Podicheti, Logan J Geyman, Elizabeth N Baker, Kai Papenfort, Douglas B Rusch, Julia C van Kessel","doi":"10.1128/mbio.00106-25","DOIUrl":"10.1128/mbio.00106-25","url":null,"abstract":"<p><p>The type III secretion system (T3SS) is a needle-like complex used by numerous bacterial pathogens in host infection to inject exotoxins into the host cell cytoplasm. The T3SS is a known virulence factor in the shrimp pathogen <i>Vibrio campbellii</i>. The ~40 genes comprising the <i>V. campbellii</i> T3SS are regulated by a network of transcription factors in response to changes in the cell's environment: cell density (quorum sensing; QS), temperature, calcium, and host cell contact. Under positive environmental stimuli, the master T3SS transcription factor ExsA activates the expression of the four structural T3SS operons required for needle formation. Previous studies identified a key role of the master QS transcription factor LuxR: repression of <i>exsA</i> transcription via DNA binding at the <i>exsBA</i> promoter. Here, we uncovered a new regulatory role of LuxR: post-translational repression of ExsA activity via transcriptional repression of the gene encoding the anti-anti-activator ExsC. In <i>V. campbellii</i>, ExsC is a positive regulator of T3SS transcription; deletion of <i>exsC</i> decreases ExsA-dependent transcription activation of the T3SS structural promoters. Through genetic epistasis and <i>in vitro</i> biochemical assays, we show that LuxR directly binds the <i>exsC</i> promoter upstream of ExsA and represses transcription of <i>exsC</i>. Our findings collectively show that <i>V. campbellii</i> responds to high cell density signals to shut down ExsA-dependent expression of the T3SS via two mechanisms. We postulate that this dual regulatory mechanism by LuxR enables both the rapid inactivation of existing ExsA protein and blocks its further synthesis, leading to a rapid shutdown of T3SS activity at high cell density.</p><p><strong>Importance: </strong><i>Vibrio campbellii</i> utilizes the type III secretion system (T3SS) as a mechanism of pathogenesis, which is a highly studied \"injectisome\" complex that delivers exotoxins into host cells during infection. The T3SS pathogenicity island in <i>V. campbellii</i> comprises ~40 genes that are organized into four structural operons. In this study, we determined that quorum sensing-a method of bacterial communication-regulates T3SS genes both at the transcriptional and post-translational levels to shut down T3SS gene expression at high population densities.</p>","PeriodicalId":18315,"journal":{"name":"mBio","volume":" ","pages":"e0010625"},"PeriodicalIF":5.1,"publicationDate":"2025-04-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143492686","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Blanking on blanks: few insect microbiota studies control for contaminants.","authors":"Elisabeth M Williamson, Tobin J Hammer, Katja Hogendoorn, Raphael Eisenhofer","doi":"10.1128/mbio.02658-24","DOIUrl":"10.1128/mbio.02658-24","url":null,"abstract":"<p><p>Research on insect-microbe relationships is booming, with DNA sequencing being the most commonly used method to describe insect microbiota. However, sequencing is vulnerable to contamination, especially when the sample has low microbial biomass. Such low-biomass samples are common across insect taxa, developmental stages, and tissue types. Identifying putative contaminants is essential to distinguish between true microbiota and introduced contaminant DNA. It is therefore important that studies control for contamination, but how often this is done is unknown. To investigate the status quo of contamination control, we undertook a systematic literature review to quantify the prevalence of negative control usage and contamination control across the literature on insect microbiota (specifically bacterial communities) over a 10 year period. Two-thirds of the 243 insect microbiota studies evaluated had not included blanks (negative controls), and only 13.6% of the studies sequenced these blanks and controlled for contamination in their samples. Our findings highlight a major lack of contamination control in the field of insect microbiota research. This result suggests that a number of microbes reported in the literature may be contaminants as opposed to insect-associated microbiota and that more rigorous contamination control is needed to improve research reliability, validity, and reproducibility. Based on our findings, we recommend the previously developed guidelines outlined in the RIDE checklist, with the addition of one more guideline. We refer to this as the RIDES checklist, which stands for Report methodology, Include negative controls, Determine the level of contamination, Explore contamination downstream, and State the amount of off-target amplification.IMPORTANCEOur systematic review reveals a major lack of methodological rigor within the field of research on insect-associated microbiota. The small percentage of studies that control for contamination suggests that an unknown but potentially considerable number of bacteria reported in the literature could be contaminants. The implication of this finding is that true microbiota may be masked or misrepresented, especially in insects with low microbial biomass.</p>","PeriodicalId":18315,"journal":{"name":"mBio","volume":" ","pages":"e0265824"},"PeriodicalIF":5.1,"publicationDate":"2025-04-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143492442","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Passage-attenuated Powassan virus LI9P protects mice from lethal LI9 challenge and links envelope residue D308 to neurovirulence.","authors":"Grace E Himmler, Megan C Mladinich, Jonas N Conde, Elena E Gorbunova, Marissa R Lindner, Hwan Keun Kim, Erich R Mackow","doi":"10.1128/mbio.00065-25","DOIUrl":"10.1128/mbio.00065-25","url":null,"abstract":"<p><p>Powassan virus (POWV) is an emergent tick-borne flavivirus that causes lethal encephalitic disease and chronic neurologic deficits in surviving patients. POWV-LI9 is a tick-derived isolate that causes neurovirulent disease and age-dependent lethality in mice. Serial passage of VeroE6 cells infected with LI9 resulted in eight amino acid changes in a POWV strain LI9P. LI9P fails to cause neurological sequelae, or lethality in C57BL/6 mice yet elicits neutralizing POWV antibody responses and protects mice from lethal LI9 challenge. Analysis revealed that LI9, but not LI9P, is present at high levels in the CNS, suggesting that LI9P is restricted from neuroinvasion or CNS replication. LI9 and LI9P are distinguished by a D308N envelope change within a domain associated with cell attachment. We evaluated the roles of Env-Domain III residue changes in LI9 virulence and LI9P attenuation using recombinant POWVs (recPOWVs) generated by reverse genetics. Remarkably, mutating D308N in LI9 completely abolished viral lethality and neuroinvasion in 50-week-old mice, reflecting the avirulent phenotype of LI9P. Analysis of the reciprocal N308D change in LI9P only partially restored neuroinvasion and lethality to the LI9P-N308D mutant, indicating that further LI9P residue changes contribute to LI9P attenuation. Consistent with differences in neuroinvasion, we found that rapid LI9P RNA synthesis and corresponding early IFN induction may contribute to LI9P clearance. Collectively, these findings define D308 as a determinant of POWV neuroinvasion and lethality, suggest potential mechanisms for restricted LI9P CNS entry, and reveal passage-attenuated LI9P as a candidate POWV vaccine platform.</p><p><strong>Importance: </strong>Powassan virus (POWV) infection causes a 10% lethal encephalitis, resulting in chronic neurological symptoms in half of survivors. POWV is transmitted in as short as 15 min following tick attachment, demonstrating the need for the development of POWV vaccines and therapeutics. Mechanisms of POWV neurovirulence remain to be defined to inform vaccine and therapeutic design. Cell culture passage has successfully been used to generate live-attenuated flavivirus vaccines. Accordingly, we serially passaged POWV LI9-infected VeroE6 cells and isolated an attenuated POWV strain, LI9P, that fails to cause neurologic sequelae or murine lethality. LI9P elicits neutralizing antibody responses, protects mice from a lethal WT POWV challenge, and is a potential POWV vaccine. Analysis of attenuating mutations in LI9P revealed that changing envelope residue D308N alone in LI9 prevents POWV neurovirulence and lethality in immunocompetent mice. Altogether, this study defines viral determinants of POWV pathogenesis and attenuating mutations that inform the development of live-attenuated POWV vaccines.</p>","PeriodicalId":18315,"journal":{"name":"mBio","volume":" ","pages":"e0006525"},"PeriodicalIF":5.1,"publicationDate":"2025-04-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143492684","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Spontaneous lung colonization in the cystic fibrosis rat model is linked to gastrointestinal obstruction.","authors":"Mikayla Murphree-Terry, Johnathan D Keith, Ashley M Oden, Susan E Birket","doi":"10.1128/mbio.03883-24","DOIUrl":"10.1128/mbio.03883-24","url":null,"abstract":"<p><p>Cystic fibrosis (CF) is a genetic disease caused by mutations in the cystic fibrosis transmembrane conductance regulator (<i>CFTR</i>) gene, resulting in CFTR protein dysfunction. CFTR dysfunction has multi-organ consequences, leading to dehydrated mucus that is adherent to epithelia. In the lungs, this leads to recalcitrant infections with bacteria such as <i>Pseudomonas aeruginosa</i>. In the gut, mucus-laden feces can adhere to the intestines, resulting in distal intestinal obstruction syndrome (DIOS). There is limited information on how lung colonization and DIOS are correlated in people with CF (pwCF). In this novel work, we describe the development of spontaneous lung colonization of CF pathogens in young (<3 months old) CF rats, preceding the development of DIOS. Once DIOS is established, the lung microbiome becomes predominated by taxa also observed in the feces. Induced infection with <i>P. aeruginosa</i> in the CF rats reflects data found in pwCF, as once CF rats are infected, they retain a higher relative abundance of <i>P. aeruginosa</i> than their healthy agemates. Finally, we found that ivacaftor treatment favors a healthier gut microbiome in CF rats, decreasing the relative abundance of <i>Escherichia coli</i>. These results indicate that the CF rat model is recapitulative of human CF disease with the spontaneous lung colonization of traditional CF pathogens and maintenance of <i>P. aeruginosa</i> after induced infection. Furthermore, these results indicate a possible role for the gut-lung axis in lung colonization and DIOS in CF.IMPORTANCEThese data describe for the first time the development of spontaneous lung colonization in the cystic fibrosis (CF) rat model, a hallmark aspect of human CF disease. We also find that CF rats infected with <i>Pseudomonas aeruginosa</i> maintain higher relative abundance following chronic infection as compared to healthy rats, similar to those is seen in people with CF. Additionally, we describe the possible contribution of the gut-lung axis linking lung health with distal intestinal obstruction syndrome, a relationship largely unexplored in the context of CF.</p>","PeriodicalId":18315,"journal":{"name":"mBio","volume":" ","pages":"e0388324"},"PeriodicalIF":5.1,"publicationDate":"2025-04-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143557318","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"<i>Aspergillus fumigatus</i> secondary metabolite pyripyropene is important for the dual biofilm formation with <i>Pseudomonas aeruginosa</i>.","authors":"Patricia Alves de Castro, Daniel Yuri Akiyama, Camila Figueiredo Pinzan, Thaila Fernanda Dos Reis, Endrews Delbaje, Peter Rocha, Mario Augusto Izidoro, Sérgio Schenkman, Shinya Sugimoto, Norio Takeshita, Karin Steffen, Jessica L Aycock, Stephen K Dolan, Antonis Rokas, Taícia Fill, Gustavo H Goldman","doi":"10.1128/mbio.00363-25","DOIUrl":"10.1128/mbio.00363-25","url":null,"abstract":"<p><p>The human pathogenic fungus <i>Aspergillus fumigatus</i> establishes dual biofilm interactions in the lungs with the pathogenic bacterium <i>Pseudomonas aeruginosa</i>. Screening of 21 <i>A</i>. <i>fumigatus</i> null mutants revealed seven mutants (two G protein-coupled receptors, three mitogen-activated protein kinase receptors, a Gα protein, and one histidine kinase receptor) with reduced biofilm formation, specifically in the presence of <i>P. aeruginosa</i>. Transcriptional profiling and metabolomics analysis of secondary metabolites produced by one of these mutants, Δ<i>gpaB</i> (<i>gpaB</i> encodes a Gα protein), showed GpaB controls the production of several important metabolites for the dual biofilm interaction, including pyripyropene A, a potent inhibitor of mammalian acyl-CoA cholesterol acyltransferase. Deletion of <i>pyr2</i>, encoding a non-reducing polyketide synthase essential for pyripyropene biosynthesis, showed reduced <i>A. fumigatus</i> Δ<i>pyr2-P. aeruginosa</i> biofilm growth, altered macrophage responses, and attenuated mouse virulence in a chemotherapeutic murine model. We identified pyripyropene as a novel player in the ecology and pathogenic interactions of this important human fungal pathogen.IMPORTANCE<i>Aspergillus fumigatus</i> and <i>Pseudomonas aeruginosa</i> are two important human pathogens. Both organisms establish biofilm interactions in patients affected with chronic lung pulmonary infections, such as cystic fibrosis (CF) and chronic obstructive pulmonary disease. Colonization with <i>A. fumigatus</i> is associated with an increased risk of <i>P. aeruginosa</i> colonization in CF patients, and disease prognosis is poor when both pathogens are present. Here, we identified <i>A. fumigatus</i> genetic determinants important for the establishment of <i>in vitro</i> dual <i>A. fumigatus-P. aeruginosa</i> biofilm interactions. Among them, an <i>A. fumigatus</i> Gα protein GpaB is important for this interaction controlling the production of the secondary metabolite pyripyropene. We demonstrate that the lack of pyripyropene production decreases the dual biofilm interaction between the two species as well as the virulence of <i>A. fumigatus</i> in a chemotherapeutic murine model of aspergillosis. These results reveal a complete novel role for this secondary metabolite in the ecology and pathogenic interactions of this important human fungal pathogen.</p>","PeriodicalId":18315,"journal":{"name":"mBio","volume":" ","pages":"e0036325"},"PeriodicalIF":5.1,"publicationDate":"2025-04-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143649752","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Cryo-EM structure of the <i>Pseudomonas aeruginosa</i> MexY multidrug efflux pump.","authors":"William D Gregor, Rakesh Maharjan, Zhemin Zhang, Lucius Chiaraviglio, Nithya Sastry, Meng Cui, James E Kirby, Edward W Yu","doi":"10.1128/mbio.03826-24","DOIUrl":"10.1128/mbio.03826-24","url":null,"abstract":"<p><p><i>Pseudomonas aeruginosa</i>, a Gram-negative pathogen, has emerged as one of the most highly antibiotic-resistant bacteria worldwide and subsequently has become a leading cause of healthcare-associated, life-threatening infections. <i>P. aeruginosa</i> multidrug efflux Y (MexY) is an efflux pump that belongs to the resistance-nodulation-cell division (RND) superfamily. It is a major determinant for resistance to aminoglycosides in this opportunistic pathogen. However, the detailed molecular mechanisms involved in aminoglycoside recognition and extrusion by MexY have not been elucidated. Here, we report the cryo-electron microscopy structure of MexY to a resolution of 3.63 Å. The structure directly indicates two plausible pathways for drug export. It also suggests that MexY is capable of picking up antibiotics via the ceiling of the central cavity formed by the MexY trimer. Molecular dynamics simulations depict that MexY is able to use a tunnel connecting the central cavity to the funnel of the trimer to export its substrates.</p><p><strong>Importance: </strong>Here, we report the cryo-electron microscopy structure of the MexY multidrug efflux pump, posing the possibility that this pump is capable of capturing antibiotics from both the central cavity and the periplasmic cleft of the pump. The results indicate that MexY may utilize charged residues to bind and export drugs, mediating resistance to these antibiotics.</p>","PeriodicalId":18315,"journal":{"name":"mBio","volume":" ","pages":"e0382624"},"PeriodicalIF":5.1,"publicationDate":"2025-04-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143557315","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}