{"title":"MtvS Regulates the Francisella Type V-A CRISPR-Cas System","authors":"Maj Brodmann, Luciano A Marraffini","doi":"10.1101/2024.09.12.612765","DOIUrl":"https://doi.org/10.1101/2024.09.12.612765","url":null,"abstract":"CRISPR-Cas systems endow bacteria and archaea with adaptive immune systems against mobile genetic elements, playing a fundamental role in shaping microbial communities. Many organisms harbor more than one CRISPR-Cas system, however whether and how these are differentially regulated is not known, in many instances due to the impossibility of studying CRISPR immunity in native hosts. Here we studied the regulation of endogenous type II-B and type V-A CRISPR-Cas systems in opportunistic human pathogen Francisella novicida U112. Fluorescence microscopy and transcriptomics experiments revealed that while the type II-B system is constitutively expressed, the type V-A CRISPR-Cas system is differentially expressed at stationary phase and high cell density. Using mass spectrometry and genetics we identified MtvS as a factor required for the differential expression of the type V-A CRISPR-Cas locus. Surprisingly, MtvS-dependent expression of the type V-A CRISPR-Cas system at high cell density is linked quorum sensing like behavior, even though F. novicida U112 lacks canonical quorum sensing genes. Our findings provide an example of how bacteria harboring multiple CRISPR systems regulate their expression.","PeriodicalId":501357,"journal":{"name":"bioRxiv - Microbiology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2024-09-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142214009","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Mallory L Myers, Michael T Conlon, John R Gallagher, DeMarcus D Woolfork, Noah D Khorrami, William B Park, Regan K Stradtman-Carvalho, Audray K Harris
{"title":"Analysis of polyclonal and monoclonal antibody to the influenza virus nucleoprotein in different oligomeric states","authors":"Mallory L Myers, Michael T Conlon, John R Gallagher, DeMarcus D Woolfork, Noah D Khorrami, William B Park, Regan K Stradtman-Carvalho, Audray K Harris","doi":"10.1101/2024.09.12.612748","DOIUrl":"https://doi.org/10.1101/2024.09.12.612748","url":null,"abstract":"Influenza virus nucleoprotein (NP) is one of the most conserved influenza proteins. Both NP antigen and anti-NP antibodies are used as reagents in influenza diagnostic kits, with applications in both clinical practice, and influenza zoonotic surveillance programs. Despite this, studies on the biochemical basis of NP diagnostic serology and NP epitopes are not as developed as for hemagglutinin (HA), the fast-evolving antigen which has been the critical component of current influenza vaccines. Here, we characterized the NP serology of mice, ferret, and human sera and the immunogenic effects of NP antigen presented as different structural complexes. Furthermore, we show that a classical anti-NP mouse mAb HB65 could detect NP in some commercial influenza vaccines. MAb HB65 bound a linear epitope with nanomolar affinity. Our analysis suggests that linear NP epitopes paired with their corresponding characterized detection antibodies could aid in designing and improving diagnostic technologies for influenza virus.","PeriodicalId":501357,"journal":{"name":"bioRxiv - Microbiology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2024-09-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142214012","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Jiexi Chen, Felix Goerdeler, Thapakorn Jaroentomeechai, Francisco X. Silva Hernandez, Xiaozhong Wang, Henrik Clausen, Yoshiki Narimatsu, Karla J Satchell
{"title":"Biantennary N-glycans As Receptors for MARTX Toxins in Vibrio Pathogenesis","authors":"Jiexi Chen, Felix Goerdeler, Thapakorn Jaroentomeechai, Francisco X. Silva Hernandez, Xiaozhong Wang, Henrik Clausen, Yoshiki Narimatsu, Karla J Satchell","doi":"10.1101/2024.09.12.611726","DOIUrl":"https://doi.org/10.1101/2024.09.12.611726","url":null,"abstract":"Multifunctional Autoprocessing Repeats-in-Toxin (MARTX) toxins are a diverse effector delivery platform of many Gram-negative bacteria that infect mammals, insects, and aquatic animal hosts. The mechanisms by which these toxins recognize host cell receptors for translocation of toxic effectors into the cell have remained elusive. Here, we map the first surface receptor-binding domain of a MARTX toxin from the highly lethal foodborne pathogen Vibrio vulnificus. This domain corresponds to a 273-amino acid sequence with predicted symmetrical immunoglobulin-like folds. We demonstrate that this domain binds internal N-acetylglucosamine on complex biantennary N-glycans with select preference for L1CAM and other N-glycoproteins with multiple N-glycans on host cell surfaces. This receptor binding domain is essential for V. vulnificus pathogenesis during intestinal infection. The identification of a highly conserved motif universally present as part of all N-glycans correlates with the V. vulnificus MARTX toxin boasting broad specificity and targeting nearly all cell types.","PeriodicalId":501357,"journal":{"name":"bioRxiv - Microbiology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2024-09-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142213990","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Emi Suzuki, Masaki Serata, Tomoyuki Sako, Sumie Sato, Tohru Iino, Hiroaki Tateno, Jun Hirabayashi
{"title":"Glyco-phenotyping of mutants of Lacticaseibacillus paracasei by lectin microarray","authors":"Emi Suzuki, Masaki Serata, Tomoyuki Sako, Sumie Sato, Tohru Iino, Hiroaki Tateno, Jun Hirabayashi","doi":"10.1101/2024.09.11.612535","DOIUrl":"https://doi.org/10.1101/2024.09.11.612535","url":null,"abstract":"We previously identified a gene cluster of <em>Lacticaseibacillus paracasei</em> strain Shirota (YIT 9029) for cell surface long-chain polysaccharides (LCPS-1) biosynthesis, which modulates YIT 9029 activity to induce cytokine production in immune cells, and showed that a lectin microarray can be useful for distinguishing the profile of bacterial cell-surface polysaccharide (PS) structures. Therefore, we isolated disruptive mutant strains of 51 genes predicted to be involved in cell wall PS biosynthesis in YIT 9029. Their binding profiles to lectins in conjunction with their binding abilities to YIT 9029-specific monoclonal antibody (MAb) were compared. The mutants defective in binding to the MAb all had defects within the <em>cps1</em> gene cluster. Some mutants partially bound to MAb, indicating that these genes may influence the synthesis and maturation of LCPS-1. Advanced lectin microarray analyzed the cell surface glycosylation properties of YIT 9029 and its mutants. YIT 9029 bound to a rhamnose-specific lectin CSA, and three additional lectins including an O-glycan binder (rDiscoidin II) and two mannose binders (rOrysata and rBanana). Lectin binding specificity was confirmed by a gene complementation assay for the <em>cps1C</em> gene and a carbohydrate inhibition assay. When the binding profiles of individual <em>cps1A</em> through <em>cps1J</em> knockout mutants were compared, typical and specific binding profiles patterns were observed, in which some similarities in the functions of each gene could be predicted. In conclusion, the combined use of lectin microarray and a YIT 9029 mutant strain library is a powerful tool for identifying unknown bacterial gene functions related to cell surface glycome.","PeriodicalId":501357,"journal":{"name":"bioRxiv - Microbiology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2024-09-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142214015","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Nour Diab, Eva Lena Stange, Chiun Huei Yong, Joerg Deiwick, Mihael Vucur, Tom Luedde, Michael Hensel, Natalia Torow, Kaiyi Zhang, Mathias Walter Hornef
{"title":"Phosphatase-independent suppression of mucosal inflammation and disease progression by Salmonella SopB","authors":"Nour Diab, Eva Lena Stange, Chiun Huei Yong, Joerg Deiwick, Mihael Vucur, Tom Luedde, Michael Hensel, Natalia Torow, Kaiyi Zhang, Mathias Walter Hornef","doi":"10.1101/2024.09.02.610793","DOIUrl":"https://doi.org/10.1101/2024.09.02.610793","url":null,"abstract":"Salmonella enterica subsp. enterica serovar Typhimurium (S. Typhimurium) translocates effector molecules via its Salmonella pathogenicity island (SPI)1 encoded type 3 secretion system (T3SS) to induce internalization by intestinal epithelial cells and manipulate cellular responses. Among these effector molecules, the Salmonella outer protein B (SopB) was shown to possess phosphatidyl-inositol phosphatase activity and induce bacterial internalisation, promote cell survival, influence endosomal trafficking and alter host cell signalling. Using a neonatal S. Typhimurium infection model, we here show that SopB in vivo suppresses early epithelial chemokine expression, delays mucosal immune cell recruitment, reduces barrier impairment by enterocyte necroptosis, and prevents disease progression and premature death. Unexpectedly, this immunosuppressive effect was independent of the phosphatidyl-inositol phosphatase and phosphotransferase activity of SopB but required an intact N-terminal domain. Thus, SopB exerts a potent phosphatase-independent immunosuppressive effect to delay local tissue inflammation and disease progression likely to promote host transmission.","PeriodicalId":501357,"journal":{"name":"bioRxiv - Microbiology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2024-09-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142227021","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Inorganic polyphosphate and the stringent response coordinately control cell division and cell morphology in Escherichia coli","authors":"Christopher W Hamm, Michael Jeffrey Gray","doi":"10.1101/2024.09.11.612536","DOIUrl":"https://doi.org/10.1101/2024.09.11.612536","url":null,"abstract":"Bacteria encounter numerous stressors in their constantly changing environments and have evolved many methods to deal with stressors quickly and effectively. One well known and broadly conserved stress response in bacteria is the stringent response, mediated by the alarmone (p)ppGpp. (p)ppGpp is produced in response to amino acid starvation and other nutrient limitations and stresses and regulates both the activity of proteins and expression of genes. <em>Escherichia coli</em> also makes inorganic polyphosphate (polyP), an ancient molecule evolutionary conserved across most bacteria and other cells, in response to a variety of stress conditions, including amino acid starvation. PolyP can act as an energy and phosphate storage pool, metal chelator, regulatory signal, and chaperone, among other functions. Here we report that <em>E. coli</em> lacking both (p)ppGpp and polyP have a complex phenotype indicating previously unknown overlapping roles for (p)ppGpp and polyP in regulating cell division, cell morphology, and metabolism. Disruption of either (p)ppGpp or polyP synthesis led to formation of filamentous cells, but simultaneous disruption of both pathways resulted in cells with heterogenous cell morphologies, including highly branched cells, severely mislocalized Z-rings, and cells containing substantial void spaces. These mutants also failed to grow when nutrients were limited, even when amino acids were added. These results provide new insights into the relationship between polyP synthesis and the stringent response in bacteria and point towards their having a joint role in controlling metabolism, cell division, and cell growth.","PeriodicalId":501357,"journal":{"name":"bioRxiv - Microbiology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2024-09-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142213993","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"ves1α genes expression is the major determinant of Babesia bovis-infected erythrocytes cytoadhesion to endothelial cells","authors":"Hassan Hakimi, Junya Yamagishi, Miako Sakaguchi, Guilherme G. Verocai, Shin-ichiro Kawazu, Masahito Asada","doi":"10.1101/2024.09.12.612601","DOIUrl":"https://doi.org/10.1101/2024.09.12.612601","url":null,"abstract":"<em>Babesia bovis</em> causes the most pathogenic form of babesiosis in cattle, resulting in high mortality in naive adults. This parasite invades red blood cells (RBCs) within the bovine hosts where they multiply and produce clinical disease. <em>Babesia bovis</em> exports numerous proteins into invaded RBCs changing its properties. Thus, the infected RBCs (iRBCs) are capable to cytoadhere in the microvasculature of internal organs and brain, leading to respiratory distress, neurologic signs, and mortality. Variant Erythrocyte Surface Antigen 1 (VESA1) is one of those exported proteins by <em>B. bovis</em> which represents a major virulence factor due to its central role in immune evasion by antigenic variation and intravascular parasite sequestration. VESA1 is a heterodimer protein encoded by <em>ves1α</em> and <em>ves1β</em> multigene family and localized on the ridges, the focal point for cytoadhesion. To gain further insights into the molecular mechanisms of cytoadhesion of <em>B. bovis</em>, we panned the parasites with bovine brain microvasculature endothelial cells, which resulted in obtaining several clones with different cytoadherence abilities. The transcriptome analysis of 2 high and 2 low cytoadherent clones revealed that <em>ves1α</em> sequences were diversified, likely resulting from genomic recombination. On the other hand, <em>ves1β</em> sequences were almost identical among these 4 clones. Insertion and expression of <em>ves1α</em> of a clone with high binding into <em>ef-1α</em> locus of a low binging clone increased cytoadherence confirming the role of <em>ves1α</em> suggested by our transcriptome data. Whole genome sequencing of cytoadherent clones revealed active locus of ves1 on chromosome 2. These results suggest that VESA1a proteins encoded by <em>ves1α</em> genes determine the cytoadherence specificity and/or cytoadherence strength of <em>B. bovis</em> and they are in the active site for recombination.","PeriodicalId":501357,"journal":{"name":"bioRxiv - Microbiology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2024-09-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142214014","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Comprehensive intra-host infection kinetics reveals high arbo-flavivirus transmission potential by neglected vector species, Aedes scutellaris","authors":"Yudthana Samung, Jutharat Pengon, Chatpong Pethrak, Phonchanan Pakparnich, Saranya Thaiudomsup, Kittitat Suksirisawat, Anon Phayakkaphol, Songpol Eiamsam-ang, Thipruethai Phanitchat, Channarong Sartsanga, Tararat Jantra, Patchara Sriwichai, Natapong Jupatanakul","doi":"10.1101/2024.09.12.612593","DOIUrl":"https://doi.org/10.1101/2024.09.12.612593","url":null,"abstract":"Background: Dengue virus (DENV) and Zika virus (ZIKV) are primarily transmitted by Aedes mosquitoes. As most studies on vector competence have focused on Aedes aegypti and Aedes albopictus while neglecting other Aedes species, it is possible that the transmission risks might be underestimated. it is necessary to examine additional species that could potentially serve as competent vectors. This is particularly important considering the potential expansion of their geographical range due to climate change or species-specific vector reduction interventions. Methodology/Principal Findings: In this study, we examined the infection kinetics and transmission potential of Aedes scutellaris from Thailand, comparing to Ae. aegypti and Ae. albopictus. Our findings demonstrated that Ae. scutellaris and Ae. albopictus had lower rates of midgut infection compared to Ae. aegypti due to smaller blood meal sizes during feeding. However, once the infection has established Ae. scutellaris exhibited efficient replication of ZIKV and DENV1-4 in the midguts, secondary organs, and salivary glands. Notably, Ae. scutellaris had a low salivary gland escape barrier, with comparable transmissibility as Ae. aegypti when inoculated with the same viral load. Conclusion: This study highlights the potential of Ae. scutellaris as a vector for DENV and ZIKV and emphasizes the importance of considering neglected mosquito species in arbovirus transmission and surveillance efforts.","PeriodicalId":501357,"journal":{"name":"bioRxiv - Microbiology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2024-09-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142213989","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Martina Castellan, Gianpiero Zamperin, Greta Foiani, Maira Zorzan, Maria Francesca Priore, Petra Drzewnioková, Erica Melchiotti, Marta Vascellari, Isabella Monne, Sergio Crovella, Stefania Leopardi, Paola De Benedictis
{"title":"Immunological findings of West Caucasian bat virus in an accidental host","authors":"Martina Castellan, Gianpiero Zamperin, Greta Foiani, Maira Zorzan, Maria Francesca Priore, Petra Drzewnioková, Erica Melchiotti, Marta Vascellari, Isabella Monne, Sergio Crovella, Stefania Leopardi, Paola De Benedictis","doi":"10.1101/2024.09.12.612599","DOIUrl":"https://doi.org/10.1101/2024.09.12.612599","url":null,"abstract":"The Lyssavirus genus includes seventeen neurotropic viral species which are able to cause rabies, an acute and almost invariably fatal encephalomyelitis of mammals. Rabies virus (RABV), the genus prototype, is a multi-host pathogen that undergoes multiple events of host-switching, thus occupying several geographical and ecological niches. In contrast, non-RABV lyssaviruses are mainly confined within a single natural host with rare spillover events never followed by adaptation to new accidental host species. In this scenario, unveiling the mechanisms underlying the host immune response against a virus is crucial to understand the dynamics of infection but also to predict the probability of colonization and adaptation to a new target species. Presently, the host response to lyssaviruses has only been partially explored, with the majority of data inferred from RABV infection, under the assumption that all members of the genus exhibit a similar behavior. Through our study we have investigated the immune response determined by the West Caucasian bat virus (WCBV). Indeed, WCBV has been recently associated with a spillover event to a domestic cat, raising concern about the risks for public health due to the circulation of the virus in its natural host. We selected the Syrian hamster as an animal model representative for an accidental host, and chose the intramuscular route in order to mimic the natural route of infection. In hamsters, WCBV was highly pathogenic, determining 100% lethality and mild encephalitis. In comparison with Duvenhage virus (DUVV) and RABV, we found that WCBV displayed an intermediate ability to promote cellular antiviral response, produce pro-inflammatory cytokines, recruit and activate lymphocytes in the hamsters’ central nervous system.","PeriodicalId":501357,"journal":{"name":"bioRxiv - Microbiology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2024-09-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142214016","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Xue Peng, Sophie Elizabeth Smith, Wanqi Huang, Jinlong Ru, Mohammadali Khan Mirzaei, Li Deng
{"title":"Metagenomic analyses of single phages and phage cocktails show instances of contamination with temperate phages and bacterial DNA","authors":"Xue Peng, Sophie Elizabeth Smith, Wanqi Huang, Jinlong Ru, Mohammadali Khan Mirzaei, Li Deng","doi":"10.1101/2024.09.12.612727","DOIUrl":"https://doi.org/10.1101/2024.09.12.612727","url":null,"abstract":"Increasing antibiotic resistance has led to renewed attention being paid to bacteriophage therapy. Commercial phage cocktails are available but often their contents of the phages are not well defined. Some metagenomic studies have been done to retrospectively characterise these cocktails, but little is known about the replication cycle of the included phages, or about the possible bacterial DNA contamination. In this study, published metagenomic sequences were reanalysed using recent advances in viromics tools. Signs of temperate phage contigs were found in all cocktail metagenomes, as well as host DNA, which could poses a risk as it may lead to horizontal gene transfer of virulence factors to commensals and pathogens. This suggests the need to implement further quality measures before using phage cocktails therapeutically.","PeriodicalId":501357,"journal":{"name":"bioRxiv - Microbiology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2024-09-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142214018","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}