mSphere最新文献

{"title":"<i>Plasmodium falciparum</i> leucine-rich repeat 5 disruption alters the transcription progression during asexual and sexual stage development.","authors":"Camilla V Pires, Chengqi C Q Wang, Min Zhang, Chiara Micchelli, Prem Prakash, Jenna Oberstabler, Julian C Rayner, Thomas D Otto, John H Adams","doi":"10.1128/msphere.00186-26","DOIUrl":"10.1128/msphere.00186-26","url":null,"abstract":"<p><p>How <i>Plasmodium falciparum</i> responds to stresses such as antimalarial drug pressure and uses them to trigger developmental decisions such as the production of sexual gametocytes remains poorly understood. Here, we investigated the function of the leucine-rich repeat protein LRR5 (PF3D7_1432400). Previous genome-wide <i>piggyBac</i> mutagenesis established that disruptions of LRR5 increase sensitivity to artemisinin and heat-shock and enhance gametocyte production. Using an optimized single-cell RNA sequencing approach applied to unsynchronized parasite cultures, we profiled transcriptional heterogeneity across the intraerythrocytic developmental cycle in wild-type NF54 parasites and an isogenic LRR5 <i>piggyBac</i> mutant (LRR5pB). Disruption of LRR5 reshaped the parasite transcriptome across multiple stages, with marked downregulation of pathways involved in protein folding, protein export, and host-cell remodeling, including Maurer's cleft components, the PTEX export machinery, and the exported chaperone HSP70-x. In contrast, cytoplasmic heat-shock proteins and translation-associated genes were upregulated, suggesting a compensatory stress response. Notably, LRR5pB parasites displayed premature activation of gametocyte-associated transcriptional programs, including elevated expression of the early gametocyte marker Pfs16, which was confirmed at the protein level. Pseudotime analysis further revealed accelerated transcriptional progression with mistimed expression of trophozoite-, merozoite-, and gametocyte-associated gene clusters. Together, these findings suggest LRR5 as a link between proteostasis, host-cell remodeling, and developmental timing in <i>P. falciparum</i>.IMPORTANCEMalaria parasites must survive environmental stress while deciding whether to continue asexual replication in the blood stream or instead commit to sexual gametocytes for transmission. Understanding how these processes are coordinated is critical for developing new strategies to block both disease and transmission. In this study, we used single-cell transcriptomics to examine how disruption of the <i>P. falciparum</i> protein LRR5 alters parasite development. By analyzing individual parasites from unsynchronized cultures, we uncovered hidden changes in stress-response pathways, host-cell remodeling, and early activation of genes linked to sexual development. Our findings show that loss of LRR5 weakens the parasite's ability to manage stress while prematurely priming parasites for gametocyte formation. These results suggest LRR5 as a link to stress adaptation to transmission potential.</p>","PeriodicalId":19052,"journal":{"name":"mSphere","volume":" ","pages":"e0018626"},"PeriodicalIF":4.0,"publicationDate":"2026-07-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13410765/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148252534","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Tools for genetic manipulation of the endemic fungal pathogen <i>Emergomyces africanus</i> and application of a fluorescent reporter strain in infection models.","authors":"Lucian Duvenage, Alisha Chetty, Darren D Thomson, Elizabeth R Ballou, Nelesh P Govender, Chad A Rappleye, J Claire Hoving","doi":"10.1128/msphere.00180-26","DOIUrl":"10.1128/msphere.00180-26","url":null,"abstract":"<p><p><i>Emergomyces africanus</i> is a thermally dimorphic fungal pathogen endemic to Southern Africa, which can cause fatal systemic infections in persons with advanced HIV disease. Its mechanisms of pathogenesis are not well understood. Characterization of virulence traits in this pathogen requires appropriate molecular tools for genetic manipulation. Molecular technologies developed for the transformation of <i>Histoplasma capsulatum</i> were adapted for use in <i>E. africanus. Agrobacterium</i>-mediated transformation was used to generate a reporter strain expressing green fluorescent protein (GFP). The <i>E. africanus</i> GFP reporter strain facilitated the study of yeast interaction with macrophages <i>in vitro</i> and allowed the identification of infected phagocyte cell types in the mouse lung by flow cytometry. <i>E. africanus</i> could also maintain episomal plasmids with telomere-like sequences to introduce expression constructs without genome modification. Using this plasmid system, RNA interference constructs were used to knock down the expression of cell wall α(1,3)-glucan by targeting the transcripts of the α-glucan synthase (<i>AGS1</i>). An episomal CRISPR/Cas9 system was evaluated for <i>E. africanus</i>, which effectively disrupted <i>GFP</i> in a reporter strain and enabled the generation of a <i>URA5</i> uracil auxotroph. These tools and strains will facilitate future studies to elucidate the mechanisms of pathogenesis of <i>E. africanus</i>.</p><p><strong>Importance: </strong><i>Emergomyces africanus</i> is an opportunistic fungal pathogen affecting persons with advanced HIV disease in South Africa. The biology and pathogenesis of <i>E. africanus</i> are not well understood, as the importance of the disease caused by this fungus (emergomycosis) has only been recognized in recent years, and molecular studies have been impaired by the lack of genetic technologies. In this work, we describe tools and methods for the genetic modification of this pathogen, which will accelerate future studies investigating how the fungus causes disease in the human host. These essential tools include (i) the ability to create fluorescent reporter strains, such as the green fluorescent protein <i>E. africanus</i> strain described here, which facilitates tracking the spread of the fungus during infection and enhances microscopy studies, (ii) methods for knocking down gene expression in <i>E. africanus</i>, and (iii) the permanent disruption of genes through CRISPR/Cas9 gene editing.</p>","PeriodicalId":19052,"journal":{"name":"mSphere","volume":" ","pages":"e0018026"},"PeriodicalIF":4.0,"publicationDate":"2026-07-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13410962/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148369149","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Enteric populations of <i>Escherichia coli</i> are likely to be resistant to phages due to O antigen expression.","authors":"Brandon A Berryhill, Teresa Gil-Gil, Kylie B Burke, Jake Fontaine, Catherine E Brink, Mason G Harvill, David A Goldberg, Jareth N Navas, Kerrie L May, Marcin Grabowicz, Konstantinos T Konstantinidis, Bruce R Levin, Michael H Woodworth","doi":"10.1128/msphere.00386-26","DOIUrl":"10.1128/msphere.00386-26","url":null,"abstract":"<p><p>Metagenomic data provide evidence that bacteriophage (phage) abound in the enteric microbiomes of humans. However, the contribution of these viruses in shaping the bacterial composition of the gut microbiome and how these phages are maintained remain unclear. We performed experiments with 756 combinations of 54 <i>Escherichia coli</i> and nine phage isolates from four fecal microbiota transplantation (FMT) doses and five laboratory phages as samples of non-dysbiotic human enteric microbiota. We also developed a mathematical model of the population and evolutionary dynamics of bacteria and phage. Our experiments predict that as a consequence of the production of the O antigen, most of the <i>E. coli</i> in the human enteric microbiome will be resistant to infections with the array of co-occurring phages. Our modeling suggests that phages are maintained in these enteric communities due to the high rates of transition between the O antigen-resistant and -sensitive states. Based on our observations and predictions from this theory, we postulate that the phage found in the human gut are likely to play a little role in shaping the strain composition of <i>E. coli</i> of healthy individuals. Although we only investigated <i>E. coli</i>, the mechanism of resistance described here is shared among most of the gram-negative bacteria. Evidence is provided that, as a consequence of O antigen-mediated resistance, the genetically diverse array of bacteriophage in the gut microbiome of humans plays little or no role in determining the densities and distribution of the genetically diverse strain <i>E. coli</i> in this habitat. Our mathematical model predicts and our experiments support the hypothesis that the phage present in the gut microbiome are maintained by replication on the minority of sensitive bacteria generated by the leakiness of O antigen-mediated resistance.IMPORTANCEBacteriophages (phages) are abundant in the human gut, yet whether these viruses shape the bacterial communities living there remains unresolved. Using <i>Escherichia coli</i> and phages isolated from the stool of healthy fecal microbiota transplantation (FMT) donors, together with a mathematical model, we show that the vast majority of gut <i>E. coli</i> are resistant to co-occurring phages because they express the O antigen, a surface structure that masks the receptors phages use to attach. Despite this widespread resistance, phages persist by replicating on a small, continually regenerated subpopulation of sensitive cells, a phenomenon we term leaky resistance. These findings suggest that phages play a little role in determining which <i>E. coli</i> strains dominate the healthy human gut. Because the O antigen is broadly expressed across gram-negative bacteria, this mechanism likely extends well beyond <i>E. coli</i> and helps explain why isolating therapeutic phages against many pathogens is difficult.</p>","PeriodicalId":19052,"journal":{"name":"mSphere","volume":" ","pages":"e0038626"},"PeriodicalIF":4.0,"publicationDate":"2026-07-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13410980/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148391371","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"From gene detection to resistome ecology: targeted hybrid capture in AMR surveillance.","authors":"Giulia Santarelli, Barbara Fiori, Cataldo Maria Mannavola, Roberto Rosato, Giordana Cafaro, Francesca Romana Monzo, Maurizio Sanguinetti, Flavio De Maio","doi":"10.1128/msphere.00221-26","DOIUrl":"10.1128/msphere.00221-26","url":null,"abstract":"&lt;p&gt;&lt;p&gt;Antimicrobial resistance (AMR) surveillance requires approaches capable of capturing the full resistome beyond culture-based and targeted molecular diagnostics. Here, we compared a commercial real-time PCR assay for carbapenemase detection with a targeted hybrid capture next-generation sequencing (xHYB) approach for comprehensive antimicrobial resistance gene (ARG) profiling in rectal swab samples. The xHYB panel showed high concordance with PCR for &lt;i&gt;bla&lt;sub&gt;KPC&lt;/sub&gt;&lt;/i&gt; and &lt;i&gt;bla&lt;sub&gt;NDM&lt;/sub&gt;&lt;/i&gt;, with overall agreement varying by gene target. Sequencing demonstrated higher analytical sensitivity, detecting additional ARGs not identified by PCR, particularly among &lt;i&gt;OXA&lt;/i&gt;-like variants. Sequencing read counts were inversely correlated with PCR cycle threshold values, supporting a semi-quantitative relationship between methods, while no association was observed with total DNA concentration. Beyond single-gene detection, resistome analysis revealed significant compositional differences between carbapenemase-positive and -negative samples. Positive samples displayed a broader multidrug resistance profile, including β-lactamases, aminoglycoside-modifying enzymes, tetracycline, sulfonamide, macrolide, trimethoprim resistance genes, and multidrug efflux systems. Comparative analyses across resistance databases showed variability at gene and peptide-marker levels, whereas gene-family-level profiles were more consistent. Overall, targeted xHYB sequencing enables sensitive detection of clinically relevant ARGs and reveals structured resistome patterns associated with carbapenemase carriage, supporting its integration into AMR surveillance strategies. The xHYB panel showed high concordance with PCR for &lt;i&gt;bla&lt;sub&gt;KPC&lt;/sub&gt;&lt;/i&gt; and &lt;i&gt;bla&lt;sub&gt;NDM&lt;/sub&gt;&lt;/i&gt;, with overall agreement varying by gene target. Sequencing demonstrated higher analytical sensitivity, detecting additional ARGs not identified by PCR, particularly among &lt;i&gt;OXA&lt;/i&gt;-like variants. Sequencing read counts were inversely correlated with PCR cycle threshold values, supporting a semi-quantitative relationship between methods, while no association was observed with total DNA concentration.IMPORTANCEAntimicrobial resistance is a growing global health threat driven by the spread of resistance genes among bacteria, particularly in the human gut, which acts as a major reservoir. This study focuses on intestinal antimicrobial resistance genes from human rectal swabs, highlighting that the presence of carbapenemases was associated not only with single resistance genes but with a broader and more complex multidrug resistance profile. This indicates that conventional diagnostic methods may underestimate the true resistance potential of microbial communities. By providing a more comprehensive view of resistance gene diversity, this approach can improve surveillance and help identify high-risk resistance patterns earlier. These findings support the use of sequencing-based methods to comple","PeriodicalId":19052,"journal":{"name":"mSphere","volume":" ","pages":"e0022126"},"PeriodicalIF":4.0,"publicationDate":"2026-07-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13410987/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148251858","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Exploring life's hidden majority: microbial dark matter symposium highlights.","authors":"Zachary N Flamholz, Sayali A Mulay, Victor Leshyk, J Gregory Caporaso, Jonathan A Eisen, Libusha Kelly, Karen G Lloyd, Magdalena R Osburn, Mircea Podar, Simon Roux, S Aaron B Regberg, S Emil Ruff, Braden Tierney, Scott Tighe, Elizabeth Trembath-Reichert, Kasthuri Venkateswaran, Tanja Woyke, Kristopher M Locken, Haley M Sapers, Katrine Whiteson","doi":"10.1128/msphere.00587-25","DOIUrl":"10.1128/msphere.00587-25","url":null,"abstract":"<p><p>The Microbial Dark Matter Symposium held on August 28-29, 2025, in Laguna Beach, Orange County, CA, convened a multidisciplinary group of scientists to address the vast unknowns in microbial life-from uncultured taxa and uncharacterized proteins to elusive viruses and spacefaring microbes. Set against a scenic coastal backdrop, the symposium highlighted advances in single-cell genomics, proximity ligation sequencing, and artificial intelligence-ready bioinformatics, while also probing the limits of microbial persistence, metabolism, and ecological distribution. Sessions explored microbial dark matter from multiple dimensions: cultivability, where new strategies are enabling recovery of elusive microbes; functional ambiguity, where metagenomic dark zones are illuminated by computational annotation; and genomic representation, where single-cell methods bridge gaps left by shotgun community sequencing. Researchers shared breakthroughs in identifying atmospheric microbiomes, \"dark oxygen\" production in groundwater ecosystems, and microbial survival on the International Space Station. The symposium emphasized integration of methods, disciplines, and ecosystems, advancing a collective push to illuminate the microbial dark matter on Earth and beyond. By highlighting emerging tools, pressing questions, and cross-domain insights, the symposium underscored the need for collaborative, open, and adaptive approaches to study the microbial unknown. The meeting marks a pivotal moment in microbiology, where cultivating knowledge of the uncultivated promises transformative understanding of life, everywhere.</p>","PeriodicalId":19052,"journal":{"name":"mSphere","volume":" ","pages":"e0058725"},"PeriodicalIF":4.0,"publicationDate":"2026-07-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13410755/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148422705","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Optimizing an avian influenza vaccine using a novel Bacterial Enzymatic Combinatorial Chemistry (BECC) TLR4 adjuvant.","authors":"Devon V Riley, Lauren Baracco, Sayan Das, Brandon M Tenaglia, Sydney Speed, Carly Dillen, Juliahna Hayes, Samanta Del Veliz, Haye Nijhuis, Valerie Le Sage, Hannah W Despres, Lynda Coughlan, Weina Sun, Matthew B Frieman, Robert K Ernst","doi":"10.1128/msphere.00171-26","DOIUrl":"10.1128/msphere.00171-26","url":null,"abstract":"<p><p>The development of broadly protective and dose-sparing influenza vaccines remains a critical challenge, particularly for zoonotic H5N1 strains with pandemic potential. This study evaluates BECC470s, a synthetic TLR4 adjuvant, for its ability to enhance the immunogenicity and protective efficacy of recombinant H5 hemagglutinin (rHA) vaccination in murine models. BECC470s-adjuvanted rHA elicited robust IgG1/IgG2a antibody responses and complete survival following homologous 2004 H5N1 challenge in a prime-boost model. Although BECC470s broadened antibody binding to both variable HA head and conserved stalk domains by ELISA, functional neutralizing antibody responses were restricted to the matched 2004 H5N1 isolate, with no detectable neutralization of H5N1 viruses isolated in 2022 or 2024. These data indicate that BECC470s enhances the magnitude and apparent cross-reactivity of binding antibody responses while maintaining strain-specific neutralizing activity, supporting its potential as an adjuvant for next-generation influenza vaccines while underscoring the need for further optimization to achieve true cross-neutralizing protection.IMPORTANCEBECC470s is a synthetic TLR4 agonist adjuvant that enhances the breadth and potency of recombinant H5 hemagglutinin vaccination in mice. By driving balanced antibody responses, complete protection against homologous challenge, and neutralizing activity targeting both HA head and stalk, BECC470s outperforms the comparator TLR4 adjuvant. These preliminary findings indicate that BECC470s enables antigen sparing while increasing antigen immunogenicity, supporting its potential translational application to current H5N1 vaccines.</p>","PeriodicalId":19052,"journal":{"name":"mSphere","volume":" ","pages":"e0017126"},"PeriodicalIF":4.0,"publicationDate":"2026-07-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13410986/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148422876","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Inheritance of four-membrane-bound structures in the \"apicoplast-minus\" <i>Plasmodium falciparum</i>.","authors":"Wei Xu, Ikechukwu Nwankwo, Sean T Prigge, Hangjun Ke","doi":"10.1128/msphere.00024-26","DOIUrl":"10.1128/msphere.00024-26","url":null,"abstract":"<p><p>Most apicomplexan parasites contain a plastid-derived organelle called the apicoplast, which originated through secondary endosymbiosis. As a result of this evolutionary trajectory, the non-photosynthetic apicoplast is surrounded by four membranes and contains many bacterial-like, druggable targets. It is widely accepted that asexual malaria parasites (<i>Plasmodium falciparum</i>) can thrive under antibiotic treatment if supplemented with high concentrations of isopentenyl pyrophosphate (IPP, 200 µM), and these IPP-rescued parasites are thought to lack the apicoplast and its 35 kb genome but possess many vesicles. However, our findings suggest that this \"apicoplast-minus\" concept may warrant reconsideration. Using live-cell microscopy, we observed that apicoplast-derived vesicles closely colocalize with mitochondria in late schizonts and are faithfully segregated into merozoites during schizogony, suggesting that these vesicles are inherited from the previous cycle rather than newly synthesized. Furthermore, immuno-electron microscopy (immuno-EM) revealed that the apicoplast-minus parasites possess structures surrounded by four membranes, in addition to single-membrane-surrounded entities. The presence of four-membrane-bound structures suggests that the apicoplast has not truly disappeared in the apicoplast-minus <i>P. falciparum</i> but has remained in a distinct, diminished form. We propose that these distinct four-membrane-bound structures still retain essential biochemical and/or structural functions, which act as barriers to the complete loss of apicoplast when the parasites face antibiotic stress and IPP rescue.</p><p><strong>Importance: </strong>The plant-like organelle named apicoplast is essential for malaria parasites and is a major antimalarial drug target. For more than a decade, scientists have believed that malaria parasites in the blood stages could dispense with the apicoplast if they were supplied with a critical metabolite made by the organelle, leading to the idea of \"apicoplast-minus\" parasites. Our results challenge this long-standing view. We find that even when the apicoplast is disrupted, the organelle remains in a highly reduced form. This apicoplast-derived organelle is inherited as parasites continue their life cycles, suggesting that it contains essential functions even when the organelle is disrupted. Our data reveal an unexpected level of complexity in apicoplast biology and open new doors for future identification of essential apicoplast-derived pathways that cannot be easily bypassed.</p>","PeriodicalId":19052,"journal":{"name":"mSphere","volume":" ","pages":"e0002426"},"PeriodicalIF":4.0,"publicationDate":"2026-07-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13410965/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148345981","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Cysteine-S-nitrosylation inhibits ROP5-mediated immune evasion in <i>Toxoplasma gondii</i>.","authors":"S L Lempke, P DiMare, B Nichols, L F Fitzsimmons, A Chaudhry, V C Grebe, M L Reese, S E Ewald","doi":"10.1128/msphere.00309-26","DOIUrl":"10.1128/msphere.00309-26","url":null,"abstract":"<p><p>Reactive nitrogen species (RNS) are a mechanism to control microbial infections conserved across the host species of the obligate intracellular parasite <i>Toxoplasma gondii</i>. Cysteine S-nitrosylation (SNO) is a reversible post-translational modification that controls complex cell behaviors by regulating protein interactions and signal transduction events. Here, we identified a cluster of <i>T. gondii</i> secreted effector proteins that are SNO-modified in a host inducible nitric oxide synthase (iNOS)-dependent manner. Among these were the rhoptry protein 5 (ROP5) paralogs, major virulence determinants in <i>T. gondii</i> and an immunodominant antigen in B6 mice. ROP5 was necessary for Type I and Type II parasites to evade IFN-γ-mediated immune clearance in iNOS-deficient macrophages. RNS led to the loss of ROP5 association with the parasitophorous vacuole membrane (PVM), which is necessary for the known functions of ROP5. Infection with ROP5 knockout parasites rescued the susceptibility of iNOS-deficient mice to infection with Type II <i>T. gondii</i>. Together, these data indicate that RNS can promote cell-autonomous parasite clearance by inhibiting the function of ROP5 alleles at the PVM.</p><p><strong>Importance: </strong>RNS are necessary for cell-autonomous immunity to <i>T. gondii</i> infection; however, the molecular mechanisms by which RNS regulate parasite control remain poorly understood. Our findings support a model in which post-translational modification of ROP5 by RNS is a conserved mechanism of inhibiting the functions of divergent ROP5 paralogs. These data provide a specific example of how host RNS are used to counter <i>T. gondii</i> immune evasion effectors that can be applied to understand how nitrosylation regulates the function of other parasite effectors and the role of RNS in the control of other intracellular pathogens.</p>","PeriodicalId":19052,"journal":{"name":"mSphere","volume":" ","pages":"e0030926"},"PeriodicalIF":4.0,"publicationDate":"2026-07-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13410961/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148352579","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A novel method for depletion of dengue virus non-structural protein 1 antibodies to determine serotype exposure history.","authors":"Ying Yi Zheng, M Jane Morwitzer, Michael K McCracken, Heather Friberg, Gregory D Gromowski, Jeffrey R Currier","doi":"10.1128/msphere.00043-26","DOIUrl":"10.1128/msphere.00043-26","url":null,"abstract":"<p><p>Nearly 4 billion people worldwide reside in dengue virus (DENV) endemic areas. Primary dengue infection generates both type-specific and cross-reactive immunity. While serotype-specific immunity provides long-term protection against homologous virus challenge, cross-reactive immunity is short-lived and, upon subsequent heterotypic exposure, can increase the risk of severe clinical outcomes. Therefore, it is critical to measure type-specific immunity. The current resource-intensive and technically demanding approach of removing antibodies using purified whole virions and subsequently assessing type-specific immunity through neutralization assays is practical only for small-scale studies. We developed a novel, straightforward method that uses transfected suspension cells stably expressing DENV non-structural protein 1 (NS1) <i>de novo</i> to selectively deplete targeted serotype-specific and cross-reactive NS1 antibodies from plasma and serum samples. The same cell lines are then used to establish an antibody profile of the depleted plasma to uncover type-specific serotype exposure history. This approach to antibody depletion offers several advantages over column- and bead-based depletion methods, such as (i) ease of antigenic substrate production; (ii) simplicity of the depletion procedure; and (iii) interpretability of the data.</p><p><strong>Importance: </strong>Accurately determining dengue serotype exposure remains challenging because current serotyping methods are labor-intensive, costly, and often rely on engineered viral antigens or antibodies to capture native immune components. We developed a streamlined approach that removes NS1-specific antibodies from dengue convalescent plasma using transfected cells that express NS1 <i>de novo</i>. This procedure enables the characterization of the depleted sample for the antibodies left behind to understand the relative levels of different antibody specificities and uncover an individual's dengue exposure history. The knowledge of viral antigen-antibody profiles refines the interpretation of downstream functional assays and establishes a practical framework for dissecting dengue immunity with greater precision.</p>","PeriodicalId":19052,"journal":{"name":"mSphere","volume":" ","pages":"e0004326"},"PeriodicalIF":4.0,"publicationDate":"2026-07-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13410990/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148422721","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Adaptation of <i>Anaplasma phagocytophilum</i> to the tick vector is controlled by the transcriptional regulator Tr1.","authors":"EricaRose Warwick, Rachel Burt, Jeffrey T Badigian, Daniel Howell, Kyle T Swallow, Chloe Leach, Azeza M Falghoush, Dana K Shaw, Ian T Cadby, Jason M Park","doi":"10.1128/msphere.00872-25","DOIUrl":"10.1128/msphere.00872-25","url":null,"abstract":"<p><p>Rickettsial pathogens are strictly dependent on the cellular biology of their hosts for survival and replication. Predominantly transmitted by blood-feeding arthropods, these vector-borne pathogens are forced to adapt between the disparate environments of their mammalian host and arthropod vector. To achieve this, the Rickettsial bacteria <i>Anaplasma phagocytophilum</i> undergo extensive transcriptional reprogramming, with over 41% of its genes differentially transcribed between mammals and <i>Ixodes scapularis</i> ticks. How the bacterium orchestrates this dramatic transcriptional reprogramming is not understood. The gene <i>tr1</i> encodes a Helix-Turn-Helix DNA-binding protein that is exclusively expressed during tick infection. Herein, we show that <i>tr1</i> is essential for <i>A. phagocytophilum</i> survival in ticks and regulates the transcription of other genes necessary to adapt to the arthropod vector. We demonstrate that Tr1 is a DNA-binding protein that recognizes promoters of tick-specific genes in <i>A. phagocytophilum</i>, including secreted effector <i>ateA</i>, alternate components of the type IV secretion system (T4SS), and membrane proteins. Our findings demonstrate that Tr1 is a master regulator of genes critical for <i>A. phagocytophilum</i> adaptation to the tick.IMPORTANCETick-borne pathogens are a persistent threat to human and animal health worldwide. These pathogens must be capable of surviving in both the arthropod vector and the mammalian hosts to successfully complete their lifecycle. To achieve this, these pathogens reciprocally regulate genes that are specific for either mammalian or tick infection. The mechanism orchestrating this switch remains undefined. In this study, we identify a transcriptional regulator controlling how the tick-borne agent for granulocytic anaplasmosis, <i>Anaplasma phagocytophilum</i>, adapts to life in the tick. Disabling this transcriptional switch and the genes it controls renders the bacteria unable to survive in the arthropod vector. Understanding how this central regulator and the genes under its control impact tick infection could lead to interventions that disrupt the cycle of transmission, thereby preventing disease.</p>","PeriodicalId":19052,"journal":{"name":"mSphere","volume":" ","pages":"e0087225"},"PeriodicalIF":4.0,"publicationDate":"2026-07-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13410764/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148251910","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}