mSpherePub Date : 2024-11-12DOI: 10.1128/msphere.00770-24
Jie Gao, Ziming Wang, Wenjie Deng, Boxuan Sa, Xiaoxia Chen, Ruanhong Cai, Yi Yan, Nianzhi Jiao, Elaine Lai-Han Leung, Di Liu, Wei Yan
{"title":"Improved resolution of microbial diversity in deep-sea surface sediments using PacBio long-read 16S rRNA gene sequencing.","authors":"Jie Gao, Ziming Wang, Wenjie Deng, Boxuan Sa, Xiaoxia Chen, Ruanhong Cai, Yi Yan, Nianzhi Jiao, Elaine Lai-Han Leung, Di Liu, Wei Yan","doi":"10.1128/msphere.00770-24","DOIUrl":"https://doi.org/10.1128/msphere.00770-24","url":null,"abstract":"<p><p>16S rRNA gene sequencing is the gold standard for identifying microbial diversity in environmental communities. The Illumina short-read platform is widely used in marine environment studies due to its cost-effectiveness and high accuracy, but its limited read length restricts taxonomic identification mainly to genus or family levels. Recently, the PacBio long-read sequencing platform was developed. This method has exceptional base-level resolution exceeding 99%, thereby effectively mitigating the challenges associated with high error rates commonly observed in long-read sequencing technologies. However, few studies have compared the PacBio long-read and Illumina short-read platforms in marine deep-sea sediments. Here, the PacBio long-read and Illumina short-read platforms were compared with samples collected from the deep-sea surface sediments from the cold seep in the Shenhu area of the South China Sea offshore Pearl River Estuary. Comparisons revealed a more comprehensive taxonomic identification, α-diversity, and β-diversity by PacBio long-reads. The PacBio long-read platform exhibited higher classified rates and classified taxonomy at all levels, particularly at the species level. The PacBio long-read platform was also more accurate at capturing fine spatial-scale variations in microbial communities in sediments. Our studies will facilitate the selection of 16S rRNA sequencing platforms for investigating fine spatial-scale patterns in microbial communities in deep-sea surface sediments and serve as a crucial methodological reference for future studies on microbial diversity.</p><p><strong>Importance: </strong>The PacBio long-read platform, with its exceptional base-level resolution exceeding 99%, has advanced our comprehension of deep-sea microbial diversity. By comparing microbial community analyses conducted using the Illumina short-read and PacBio long-read sequencing platforms, we have provided an enhanced understanding of fine spatial-scale patterns in microbial community diversity with depth across a deep-sea sediment core, as well as methodological insights that will be valuable for future research in this field.</p>","PeriodicalId":19052,"journal":{"name":"mSphere","volume":" ","pages":"e0077024"},"PeriodicalIF":3.7,"publicationDate":"2024-11-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142624669","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
mSpherePub Date : 2024-11-12DOI: 10.1128/msphere.00821-24
Ui-Ju Lee, Joo-Han Gwak, Seungyeon Choi, Man-Young Jung, Tae Kwon Lee, Hojin Ryu, Samuel Imisi Awala, Wolfgang Wanek, Michael Wagner, Zhe-Xue Quan, Sung-Keun Rhee
{"title":"\"<i>Ca</i>. Nitrosocosmicus\" members are the dominant archaea associated with plant rhizospheres.","authors":"Ui-Ju Lee, Joo-Han Gwak, Seungyeon Choi, Man-Young Jung, Tae Kwon Lee, Hojin Ryu, Samuel Imisi Awala, Wolfgang Wanek, Michael Wagner, Zhe-Xue Quan, Sung-Keun Rhee","doi":"10.1128/msphere.00821-24","DOIUrl":"https://doi.org/10.1128/msphere.00821-24","url":null,"abstract":"<p><p>Archaea catalyzing the first step of nitrification in the rhizosphere possibly have an influence on plant growth and development. In this study, we found a distinct archaeal community, dominated by ammonia-oxidizing archaea (AOA), associated with the root system of pepper (<i>Capsicum anuum</i> L.) and ginseng plants (<i>Panax ginseng</i> C.A. Mey.) compared to bulk soil not penetrated by roots. While the abundance of total AOA decreased in the rhizosphere soils, AOA related to \"<i>Candidatus</i> Nitrosocosmicus,\" which harbor gene encoding manganese catalase (MnKat) in contrast to most other AOA, dominated the AOA community in the rhizosphere soils. For both plant species, the ratio of copy numbers of the AOA MnKat gene to the <i>amoA</i> gene (encoding the ammonia monooxygenase subunit A) was significantly higher in the rhizospheres than in bulk soils. In contrast to MnKat-negative strains from other AOA clades, the catalase activity of a representative isolate of \"<i>Ca.</i> Nitrosocosmicus\" was demonstrated. Members of this clade were enriched in H<sub>2</sub>O<sub>2</sub>-amended bulk soils, and constitutive expression of their MnKat gene was observed in both bulk and rhizosphere soils. Due to their abundance, <i>\"Ca.</i> Nitrosocosmicus<i>\"</i> members can be considered important players mediating the nitrification process in rhizospheres. The dominance of this MnKat-containing AOA in rhizospheres of agriculturally important plants hints at a previously overlooked AOA-plant interaction.</p><p><strong>Importance: </strong>Ammonia-oxidizing archaea (AOA) are widespread in terrestrial environments and outnumber other ammonia oxidizers in the rhizosphere, possibly exerting an influence on plant growth and development. However, little is known about the selection forces that shape their composition, functions, survival, and proliferation strategies in the rhizosphere. Here, we observed a distinct AOA community on root systems of two different plant species compared to bulk soil. Our results show that the <i>\"Ca.</i> Nitrosocosmicus<i>\"</i> clade, which possesses functional MnKat genes unlike most other AOA, dominated the rhizosphere soils. Moreover, members of this clade were enriched in H2O2-amended bulk soil, which mimics the ROS stress in root systems. While research on AOA-plant interactions in the rhizosphere is still in its infancy, these findings suggest that <i>\"Ca.</i> Nitrosocosmicus<i>\"</i> may be an important clade of AOA with potential AOA-plant interaction.</p>","PeriodicalId":19052,"journal":{"name":"mSphere","volume":" ","pages":"e0082124"},"PeriodicalIF":3.7,"publicationDate":"2024-11-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142624644","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
mSpherePub Date : 2024-11-11DOI: 10.1128/msphere.00750-24
James Blauwkamp, Sushma V Ambekar, Tahir Hussain, Gunnar R Mair, Josh R Beck, Sabrina Absalon
{"title":"Nuclear pore complexes undergo Nup221 exchange during blood-stage asexual replication of <i>Plasmodium</i> parasites.","authors":"James Blauwkamp, Sushma V Ambekar, Tahir Hussain, Gunnar R Mair, Josh R Beck, Sabrina Absalon","doi":"10.1128/msphere.00750-24","DOIUrl":"10.1128/msphere.00750-24","url":null,"abstract":"<p><p><i>Plasmodium</i> parasites, the causative agents of malaria, undergo closed mitosis without breakdown of the nuclear envelope. Unlike closed mitosis in yeast, <i>Plasmodium berghei</i> parasites undergo multiple rounds of asynchronous nuclear divisions in a shared cytoplasm. This results in a multinucleated organism prior to the formation of daughter cells within an infected red blood cell. During this replication process, intact nuclear pore complexes (NPCs) and their component nucleoporins play critical roles in parasite growth, facilitating selective bi-directional nucleocytoplasmic transport and genome organization. Here, we utilize ultrastructure expansion microscopy to investigate <i>P. berghei</i> nucleoporins at the single nucleus level throughout the 24-hour blood-stage replication cycle. Our findings reveal that these nucleoporins are distributed around the nuclei and organized in a rosette structure previously undescribed around the centriolar plaque, responsible for intranuclear microtubule nucleation during mitosis. By adapting the recombination-induced tag exchange system to <i>P. berghei</i> through a single plasmid tagging system, which includes the tagging plasmid as well as the Cre recombinase, we provide evidence of NPC formation dynamics, demonstrating Nup221 turnover during parasite asexual replication. Our data shed light on the distribution of NPCs and their homeostasis during the blood-stage replication of <i>P. berghei</i> parasites.</p><p><strong>Importance: </strong>Malaria, caused by <i>Plasmodium</i> species, remains a critical global health challenge, with an estimated 249 million cases and over 600,000 deaths in 2022, primarily affecting children under five. Understanding the nuclear dynamics of <i>Plasmodium</i> parasites, particularly during their unique mitotic processes, is crucial for developing novel therapeutic strategies. Our study leverages advanced microscopy techniques, such as ultrastructure expansion microscopy, to reveal the organization and turnover of nuclear pore complexes (NPCs) during the parasite's asexual replication. By elucidating these previously unknown aspects of NPC distribution and homeostasis, we provide valuable insights into the molecular mechanisms governing parasite mitosis. These findings deepen our understanding of parasite biology and may inform future research aimed at identifying new targets for anti-malarial drug development.</p>","PeriodicalId":19052,"journal":{"name":"mSphere","volume":" ","pages":"e0075024"},"PeriodicalIF":3.7,"publicationDate":"2024-11-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142624745","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
mSpherePub Date : 2024-11-11DOI: 10.1128/msphere.00376-24
Michele F S Dutra, Priscila A Silva, Jianchi Chen, Nelson A Wulff
{"title":"The complete genome sequence of \"<i>Candidatus</i> Liberibacter asiaticus\" strain 9PA and the characterization of field strains in the Brazilian citriculture.","authors":"Michele F S Dutra, Priscila A Silva, Jianchi Chen, Nelson A Wulff","doi":"10.1128/msphere.00376-24","DOIUrl":"https://doi.org/10.1128/msphere.00376-24","url":null,"abstract":"<p><p>\"<i>Candidatus</i> Liberibacter asiaticus\" (CLas) is associated with citrus huanglongbing, a severe disease with global importance that affects citrus production in Brazil. This study reports the first complete genome of a Brazilian strain of CLas. The genomic structure comparison of strain 9PA with those of 13 complete CLas genomes revealed 9,091 mismatches and 992 gaps/insertions, highlighting eight locally colinear blocks, among which six are in the prophage region. Phylogenetic analysis categorized 13 CLas genomes into two clusters with 9PA clustered with strains from China and the United States. Whole-genomic comparison identified diverse hypervariable genomic regions (HGRs). Three HGRs in the chromosomal region and three in the prophage region were selected and investigated by polymerase chain reaction. HGRs assessed from 68 samples, from medium- to high-huanglongbing incidence areas in Sao Paulo state, were grouped into haplotypes A to P. Haplotype A, which includes strain 9PA, is the second most prevalent, representing 19.1% of the samples. Haplotype B, the most common, accounts for 42.6%. Together with haplotype C, these make up 72% of the evaluated samples. The 9PA strain has prophage P-9PA-1, both integrated and circularized, and P-9PA-3, only found in a circularized form. Prophages show high identity with SC1 (83%) and P-JXGC-3 (98%). Co-occurrence of both type 1 and 3 prophages was observed in field samples. The approach employed provides insights into the Brazilian CLas population, providing markers for population studies and highlighting the prevalence of type 1 and 3 prophages in the population.</p><p><strong>Importance: </strong>CLas is a destructive pathogen responsible for causing the severe citrus disease known as huanglongbing. Our study presents the first fully sequenced Brazilian strain of CLas, designated as 9PA, and includes an analysis of two prophages occurring in this strain. The main objective of our research was to compare the genome features of this Brazilian strain with other fully sequenced genomes and to identify its hypervariable genetic regions. These regions were subsequently used to assess genomic variability within both the chromosomal and prophage regions in Brazilian isolates of CLas. Our findings offer valuable insights into the diversified adaptation of CLas.</p>","PeriodicalId":19052,"journal":{"name":"mSphere","volume":" ","pages":"e0037624"},"PeriodicalIF":3.7,"publicationDate":"2024-11-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142624747","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
mSpherePub Date : 2024-11-06DOI: 10.1128/msphere.00868-24
Allen Choi, Kevin Dong, Emily Williams, Lindsey Pia, Jordan Batagower, Paige Bending, Iris Shin, Daniel I Peters, Justin R Kaspar
{"title":"Erratum for Choi et al., \"Human saliva modifies growth, biofilm architecture, and competitive behaviors of oral streptococci\".","authors":"Allen Choi, Kevin Dong, Emily Williams, Lindsey Pia, Jordan Batagower, Paige Bending, Iris Shin, Daniel I Peters, Justin R Kaspar","doi":"10.1128/msphere.00868-24","DOIUrl":"https://doi.org/10.1128/msphere.00868-24","url":null,"abstract":"","PeriodicalId":19052,"journal":{"name":"mSphere","volume":" ","pages":"e0086824"},"PeriodicalIF":3.7,"publicationDate":"2024-11-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142583895","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
mSpherePub Date : 2024-11-06DOI: 10.1128/msphere.00594-24
Victoria Prieto-Echagüe
{"title":"mSphere of Influence: Venturing outside the biology canon with sex and gender.","authors":"Victoria Prieto-Echagüe","doi":"10.1128/msphere.00594-24","DOIUrl":"https://doi.org/10.1128/msphere.00594-24","url":null,"abstract":"<p><p>Victoria Prieto-Echagüe works in the field of signaling by primary cilia, adipogenesis, and obesity. In this mShpere of Influence article, she reflects on how gender studies, feminism, and societal movements such as #metoo may inform all areas of biomedical and health research. She describes how they inspired her to incorporate sex as a biological variable (SABV) principle to her research exploring sex-specific mechanisms in obesity and metabolic diseases and argues that incorporating SABV is crucial for advancing precision medicine and addressing healthcare inequities.</p>","PeriodicalId":19052,"journal":{"name":"mSphere","volume":" ","pages":"e0059424"},"PeriodicalIF":3.7,"publicationDate":"2024-11-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142583901","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
mSpherePub Date : 2024-11-06DOI: 10.1128/msphere.00727-24
Sarah Werner Lass, Bailey E Smith, Shaw Camphire, Rory A Eutsey, Jojo A Prentice, Saigopalakrishna S Yerneni, Ashni Arun, Andrew A Bridges, Jason W Rosch, James F Conway, Phil Campbell, N Luisa Hiller
{"title":"Pneumococcal extracellular vesicles mediate horizontal gene transfer via the transformation machinery.","authors":"Sarah Werner Lass, Bailey E Smith, Shaw Camphire, Rory A Eutsey, Jojo A Prentice, Saigopalakrishna S Yerneni, Ashni Arun, Andrew A Bridges, Jason W Rosch, James F Conway, Phil Campbell, N Luisa Hiller","doi":"10.1128/msphere.00727-24","DOIUrl":"10.1128/msphere.00727-24","url":null,"abstract":"<p><p>Bacterial cells secrete extracellular vesicles (EVs), the function of which is a matter of intense investigation. Here, we show that the EVs secreted by the human pathogen <i>Streptococcus pneumoniae</i> (pneumococcus) are associated with bacterial DNA on their surface and can deliver this DNA to the transformation machinery of competent cells. These findings suggest that EVs contribute to gene transfer in Gram-positive bacteria and, in doing so, may promote the spread of drug resistance genes in the population.IMPORTANCEThis work extends our understanding of horizontal gene transfer and the roles of extracellular vesicles in pneumococcus. This bacterium serves as the model for transformation, a process by which bacteria can take up naked DNA from the environment. Here, we show that extracellular vesicles secreted by the pneumococcus have DNA on their surface and that this DNA can be imported by the transformation machinery, facilitating gene transfer. Understanding EV-mediated gene transfer may provide new avenues to manage the spread of antibiotic drug resistance.</p>","PeriodicalId":19052,"journal":{"name":"mSphere","volume":" ","pages":"e0072724"},"PeriodicalIF":3.7,"publicationDate":"2024-11-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142583918","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
mSpherePub Date : 2024-10-29Epub Date: 2024-09-24DOI: 10.1128/msphere.00617-24
Shane Denecke, Madeline F Malfara, Kelly R Hodges, Nikki A Holmes, Andre R Williams, Julia H Gallagher-Teske, Julia M Pascarella, Abigail M Daniels, Geert Jan Sterk, Rob Leurs, Gordon Ruthel, Rachel Hoang, Megan L Povelones, Michael Povelones
{"title":"Adhesion of <i>Crithidia fasciculata</i> promotes a rapid change in developmental fate driven by cAMP signaling.","authors":"Shane Denecke, Madeline F Malfara, Kelly R Hodges, Nikki A Holmes, Andre R Williams, Julia H Gallagher-Teske, Julia M Pascarella, Abigail M Daniels, Geert Jan Sterk, Rob Leurs, Gordon Ruthel, Rachel Hoang, Megan L Povelones, Michael Povelones","doi":"10.1128/msphere.00617-24","DOIUrl":"10.1128/msphere.00617-24","url":null,"abstract":"<p><p>Trypanosomatids are single-celled parasites responsible for human and animal disease. Typically, colonization of an insect host is required for transmission. Stable attachment of parasites to insect tissues <i>via</i> their single flagellum coincides with differentiation and morphological changes. Although attachment is a conserved stage in trypanosomatid life cycles, the molecular mechanisms are not well understood. To study this process, we elaborate upon an <i>in vitro</i> model in which the swimming form of the trypanosomatid <i>Crithidia fasciculata</i> rapidly differentiates following adhesion to artificial substrates. Live imaging of cells transitioning from swimming to attached shows parasites undergoing a defined sequence of events, including an initial adhesion near the base of the flagellum immediately followed by flagellar shortening, cell rounding, and the formation of a hemidesmosome-like attachment plaque between the tip of the shortened flagellum and the substrate. Quantitative proteomics of swimming versus attached parasites suggests differential regulation of cyclic adenosine monophosphate (cAMP)-based signaling proteins. We have localized two of these proteins to the flagellum of swimming <i>C. fasciculata</i>; however, both are absent from the shortened flagellum of attached cells. Pharmacological inhibition of cAMP phosphodiesterases increased cAMP levels in the cell and prevented attachment. Further, treatment with inhibitor did not affect the growth rate of either swimming or established attached cells, indicating that its effect is limited to a critical window during the early stages of adhesion. These data suggest that cAMP signaling is required for attachment of <i>C. fasciculata</i> and that flagellar signaling domains may be reorganized during differentiation and attachment.IMPORTANCETrypanosomatid parasites cause significant disease burden worldwide and require insect vectors for transmission. In the insect, parasites attach to tissues, sometimes dividing as attached cells or producing motile, infectious forms. The significance and cellular mechanisms of attachment are relatively unexplored. Here, we exploit a model trypanosomatid that attaches robustly to artificial surfaces to better understand this process. This attachment recapitulates that observed <i>in vivo</i> and can be used to define the stages and morphological features of attachment as well as conditions that impact attachment efficiency. We have identified proteins that are enriched in either swimming or attached parasites, supporting a role for the cyclic AMP signaling pathway in the transition from swimming to attached. As this pathway has already been implicated in environmental sensing and developmental transitions in trypanosomatids, our data provide new insights into activities required for parasite survival in their insect hosts.</p>","PeriodicalId":19052,"journal":{"name":"mSphere","volume":" ","pages":"e0061724"},"PeriodicalIF":3.7,"publicationDate":"2024-10-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11520290/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142308182","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 the tolerable region for HiBiT tag insertion in the hepatitis B virus genome.","authors":"Asako Murayama, Hitomi Igarashi, Norie Yamada, Hussein Hassan Aly, Masaaki Toyama, Masanori Isogawa, Tetsuro Shimakami, Takanobu Kato","doi":"10.1128/msphere.00518-24","DOIUrl":"10.1128/msphere.00518-24","url":null,"abstract":"<p><p>A cell culture system that allows the reproduction of the hepatitis B virus (HBV) life cycle is indispensable to exploring novel anti-HBV agents. To establish the screening system for anti-HBV agents, we exploited the high affinity and bright luminescence (HiBiT) tag and comprehensively explored the regions in the HBV genome where the HiBiT tag could be inserted. The plasmids for the HiBiT-tagged HBV molecular clones with a 1.38-fold HBV genome length were prepared. The HiBiT tag was inserted into five regions: preS1, preS2, hepatitis B e (HBe), hepatitis B X (HBx), and hepatitis B polymerase (HB pol). HiBiT-tagged HBVs were obtained by transfecting the prepared plasmids into sodium taurocholate cotransporting polypeptide-transduced HepG2 (HepG2/NTCP) cells, and their infectivity was evaluated in human primary hepatocytes and HepG2/NTCP cells. Among the evaluated viruses, the infection of HiBiT-tagged HBVs in the preS1 or the HB pol regions exhibited a time-dependent increase of the hepatitis B surface antigen (HBsAg) level after infection to HepG2/NTCP cells as well as human primary hepatocytes. Immunostaining of the hepatitis B core (HBc) antigen in infected cells confirmed these viruses are infectious to those cells. However, the time-dependent increase of the HiBiT signal was only detected after infection with the HiBiT-tagged HBV in the preS1 region. The inhibition of this HiBiT-tagged HBV infection in human primary hepatocytes and HepG2/NTCP cells by the preS1 peptide could be detected by measuring the HiBiT signal. The infection system with the HiBiT-tagged HBV in HepG2/NTCP cells facilitates easy, sensitive, and high-throughput screening of anti-HBV agents and will be a useful tool for assessing the viral life cycle and exploring antiviral agents.</p><p><strong>Importance: </strong>Hepatitis B virus (HBV) is the principal causative agent of chronic hepatitis. Despite the availability of vaccines in many countries, HBV infection has spread worldwide and caused chronic infection. In chronic hepatitis B patients, liver inflammation leads to cirrhosis, and the accumulation of viral genome integration into host chromosomes leads to the development of hepatocellular carcinoma. The currently available treatment strategy cannot expect the eradication of HBV. To explore novel anti-HBV agents, a cell culture system that can detect HBV infection easily is indispensable. In this study, we examined the regions in the HBV genome where the high affinity and bright luminescence (HiBiT) tag could be inserted and established an HBV infection system to monitor infection by measuring the HiBiT signal by infecting the HiBiT-tagged HBV in sodium taurocholate cotransporting polypeptide-transduced HepG2 (HepG2/NTCP) cells. This system can contribute to screening for novel anti-HBV agents.</p>","PeriodicalId":19052,"journal":{"name":"mSphere","volume":" ","pages":"e0051824"},"PeriodicalIF":3.7,"publicationDate":"2024-10-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11520284/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142350727","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}
mSpherePub Date : 2024-10-29Epub Date: 2024-09-25DOI: 10.1128/msphere.00629-24
Kira A Griswold, Iaroslavna Vasylieva, Megan C Smith, Kay L Fiske, Olivia L Welsh, Alexa N Roth, Alan M Watson, Simon C Watkins, Danica M Sutherland, Terence S Dermody
{"title":"Sialic acid and PirB are not required for targeting of neural circuits by neurotropic mammalian orthoreovirus.","authors":"Kira A Griswold, Iaroslavna Vasylieva, Megan C Smith, Kay L Fiske, Olivia L Welsh, Alexa N Roth, Alan M Watson, Simon C Watkins, Danica M Sutherland, Terence S Dermody","doi":"10.1128/msphere.00629-24","DOIUrl":"10.1128/msphere.00629-24","url":null,"abstract":"<p><p>Serotype 3 (T3) strains of mammalian orthoreovirus (reovirus) spread to the central nervous system to infect the brain and cause lethal encephalitis in newborn mice. Although reovirus targets several regions in the brain, susceptibility to infection is not uniformly distributed. The neuronal subtypes and anatomic sites targeted throughout the brain are not precisely known. Reovirus binds several attachment factors and entry receptors, including sialic acid (SA)-containing glycans and paired immunoglobulin-like receptor B (PirB). While these receptors are not required for infection of some types of neurons, reovirus engagement of these receptors can influence neuronal infection in certain contexts. To identify patterns of T3 neurotropism, we used microbial identification after passive tissue clearance and hybridization chain reaction to stain reovirus-infected cells throughout intact, optically transparent brains of newborn mice. Three-dimensional reconstructions revealed in detail the sites targeted by reovirus throughout the brain volume, including dense infection of the midbrain and hindbrain. Using reovirus mutants incapable of binding SA and mice lacking PirB expression, we found that neither SA nor PirB is required for the infection of various brain regions. However, SA may confer minor differences in infection that vary by region. Collectively, these studies indicate that many regions in the brain of newborn mice are susceptible to reovirus and that patterns of reovirus infection are not dependent on reovirus receptors SA and PirB.IMPORTANCENeurotropic viruses invade the central nervous system (CNS) and target various cell types to cause disease manifestations, such as meningitis, myelitis, or encephalitis. Infections of the CNS are often difficult to treat and can lead to lasting sequelae or death. Mammalian orthoreovirus (reovirus) causes age-dependent lethal encephalitis in many young mammals. Reovirus infects neurons in several different regions of the brain. However, the complete pattern of CNS infection is not understood. We found that reovirus targets almost all regions of the brain and that patterns of tropism are not dependent on receptors sialic acid and paired immunoglobulin-like receptor B. These studies confirm that two known reovirus receptors do not completely explain the cell types infected in brain tissue and establish strategies that can be used to understand complete patterns of viral tropism in an intact brain.</p>","PeriodicalId":19052,"journal":{"name":"mSphere","volume":" ","pages":"e0062924"},"PeriodicalIF":3.7,"publicationDate":"2024-10-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11540169/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142350730","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}