Pathogens最新文献

{"title":"Highly Pathogenic H5N1 Influenza A Virus (IAV) in Blue-Winged Teal in the Mississippi Flyway Is Following the Historic Seasonal Pattern of Low-Pathogenicity IAV in Ducks.","authors":"David E Stallknecht, Deborah L Carter, Lyndon Sullivan-Brügger, Paul Link, Emily Ferraro, Ciara McCarty, Bruce Davis, Lynda Knutsen, James Graham, Rebecca L Poulson","doi":"10.3390/pathogens13111017","DOIUrl":"10.3390/pathogens13111017","url":null,"abstract":"<p><p>Highly pathogenic H5N1 (HP H5N1) influenza A virus (IAV) has been detected annually in North American ducks since its introduction during 2021, but it is unknown if this virus will follow the same seasonal and geographic patterns that have been observed with low-pathogenicity (LP) IAV in this reservoir. We monitored blue-winged teal in the Mississippi flyway prior to the detection of HP H5N1 and during two post-introduction migration cycles from spring 2022 to spring 2024, testing birds for infection and antibodies to IAV nucleoprotein (NP), hemagglutinin subtype H5, and neuraminidase subtype N1. Antigens representing clade 2.3.4.4b HP H5 and LP North American H5 were used for hemagglutination inhibition (HI) and virus neutralization (VN) tests for H5 antibodies. Virologic results were consistent with historic seasonal and geographic patterns reported for LP IAV with peak infections occurring in pre-migration staging areas in Minnesota during fall 2022. However, the high prevalence of the H5 subtype was exceptional compared to historic prevalence estimates at this same site and for the Mississippi flyway. HP H5N1 was detected on wintering areas in Louisiana and Texas during the fall of that same year and this was followed by an increase in estimated antibody prevalence to NP, H5, and N1 with no HP H5N1 detections during the wintering or spring migration periods of 2022/2023. HP H5N1 was not detected in Minnesota during fall 2023 but was detected from a single bird in Louisiana. However, a similar increase in antibody prevalence was observed during the winter and spring period of 2023 and 2024. Over the two migration cycles, there was a temporal shift in observed prevalence and relative titers against the H5 antigens with a higher proportion of ducks testing positive to the 2.3.4.4b H5 antigen and higher relative titer to that antigen compared to the representative LP North American H5 antigen. The seasonal and geographic patterns observed appear to be driven by population immunity during the migration cycle. Results support an initial high infection rate of HP H5N1 in blue-winged teal in the Mississippi flyway followed by a high prevalence of antibodies to NP, H5, and N1. Although prevalence was much reduced in the second migration cycle following introduction, it is not known if this pattern will persist in the longer term or affect historic patterns of subtype diversity in this reservoir.</p>","PeriodicalId":19758,"journal":{"name":"Pathogens","volume":"13 11","pages":""},"PeriodicalIF":3.3,"publicationDate":"2024-11-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11597780/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142731473","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Tick-Borne Encephalitis Virus Infection in a Two-Year-Old Child Returning from Switzerland (July-August 2023): Is It Time for TBE Immunization of Serbian Travelers?","authors":"Verica Simin, Ognjen Ležakov, Ivana Bogdan, Dragana Mijatović, Dragana Gazibara, Lidija Popović-Dragonjić, Gordana Vijatov Đurić, Borko Milanović, Jelena Kesić, Andrea Đuretić, Ágota Ábrahám, Zsófia Lanszki, Gábor Kemenesi, Brigitta Zana, Krisztián Bányai, Zbigniew Zając, Dejan Jakimovski, Alejandro Cabezas-Cruz, Pavle Banović","doi":"10.3390/pathogens13111013","DOIUrl":"10.3390/pathogens13111013","url":null,"abstract":"<p><p>Tick-borne encephalitis (TBE) is a vaccine-preventable viral infection that poses significant public health challenges, particularly in regions where tick-borne diseases are endemic. This case report describes a 2-year-old boy with confirmed abortive TBEV infection following a tick bite during travel to Switzerland. The patient developed fever and mild symptoms but did not exhibit central nervous system involvement. The case underscores the importance of raising awareness among healthcare providers and travelers from non-endemic areas, such as Serbia, about TBE risk and the potential benefits of preventive vaccination. Strategic immunization campaigns could mitigate the public health impact of travel-related TBE.</p>","PeriodicalId":19758,"journal":{"name":"Pathogens","volume":"13 11","pages":""},"PeriodicalIF":3.3,"publicationDate":"2024-11-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11597485/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142731610","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Molecular Analysis of <i>Aggregatibacter actinomycetemcomitans</i> ApiA, a Multi-Functional Protein.","authors":"Sera Jacob, Luciana Gusmao, Dipti Godboley, Senthil Kumar Velusamy, Nisha George, Helen Schreiner, Carla Cugini, Daniel H Fine","doi":"10.3390/pathogens13111011","DOIUrl":"10.3390/pathogens13111011","url":null,"abstract":"<p><p><i>Aggregatibacter actinomycetemcomitans</i> ApiA is a trimeric autotransporter outer membrane protein (Omp) that participates in multiple functions, enabling <i>A. actinomycetemcomitans</i> to adapt to a variety of environments. The goal of this study is to identify regions in the <i>apiA</i> gene responsible for three of these functions: auto-aggregation, buccal epithelial cell binding, and complement resistance. Initially, <i>apiA</i> was expressed in <i>Escherichia coli</i>. Finally, wild-type <i>A. actinomycetemcomitans</i> and an <i>apiA-</i>deleted version were tested for their expression in the presence and absence of serum and genes related to stress adaptation, such as oxygen regulation, catalase activity, and Omp proteins. Sequential deletions in specific regions in the <i>apiA</i> gene as expressed in <i>E. coli</i> were examined for membrane proteins, which were confirmed by microscopy. The functional activity of epithelial cell binding, auto-aggregation, and complement resistance were then assessed, and regions in the <i>apiA</i> gene responsible for these functions were identified. A region spanning amino acids 186-217, when deleted, abrogated complement resistance and Factor H (FH) binding, while a region spanning amino acids 28-33 was related to epithelial cell binding. A 13-amino-acid peptide responsible for FH binding was shown to promote serum resistance. An <i>apiA</i> deletion in a clinical isolate (IDH781) was created and tested in the presence and/or absence of active and inactive serum and genes deemed responsible for prominent functional activity related to <i>A. actinomycetemcomitans</i> survival using qRT-PCR. These experiments suggested that <i>apiA</i> expression in IDH781 is involved in global regulatory mechanisms that are serum-dependent and show complement resistance. This is the first study to identify specific <i>apiA</i> regions in <i>A. actinomycetemcomitans</i> responsible for FH binding, complement resistance, and other stress-related functions. Moreover, the role of <i>apiA</i> in overall gene regulation was observed.</p>","PeriodicalId":19758,"journal":{"name":"Pathogens","volume":"13 11","pages":""},"PeriodicalIF":3.3,"publicationDate":"2024-11-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11597641/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142731381","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"The Mechanisms of Developing Fungicide Resistance in <i>Fusarium graminearum</i> Causing Fusarium Head Blight and Fungicide Resistance Management.","authors":"Malini Anudya Jayawardana, Wannakuwattewaduge Gerard Dilantha Fernando","doi":"10.3390/pathogens13111012","DOIUrl":"10.3390/pathogens13111012","url":null,"abstract":"<p><p>Fusarium head blight (FHB), primarily caused by <i>Fusarium graminearum</i>, is one of the economically significant diseases in small grains. FHB causes severe damage to wheat production and grain quality. Several management strategies have been developed to control FHB, and chemical control through fungicides plays a significant role. Although fungicides have effectively controlled <i>F. graminearum</i> in the field, the continuous exposure causes a selection pressure in the pathogen population towards fungicide resistance. Several studies have identified fungicide-resistant <i>F. graminearum</i> isolates and fungicide-resistance mechanisms. Although new fungicides with a new mode of action can be introduced into the market, developing a new fungicide is time-consuming, and extra efforts are needed for testing, approvals, and registrations. Therefore, it is essential to strategize the methods to delay the fungicide resistance. This review focuses on the impact of several fungicide applications currently used on FHB, focusing on <i>Fusarium graminearum</i>, the status of the fungicide sensitivity for fungicide classes, the resistance mechanisms against fungicides, and the mitigation strategies to delay the development of fungicide resistance in the pathogen population. Studying the fungicide resistance mechanisms and the mitigation strategies will be helpful in the future to use the available fungicides against <i>F. graminearum</i> without losing its effectiveness.</p>","PeriodicalId":19758,"journal":{"name":"Pathogens","volume":"13 11","pages":""},"PeriodicalIF":3.3,"publicationDate":"2024-11-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11597361/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142731607","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Kinetics of RPR Decline in Pregnant Persons Treated for Syphilis in Pregnancy and Their Infants.","authors":"Danielle Schwartz, Alena Tse-Chang, Joan Robinson, Jennifer Gratrix, Petra Smyczek, Michael T Hawkes","doi":"10.3390/pathogens13111010","DOIUrl":"10.3390/pathogens13111010","url":null,"abstract":"<p><p>Congenital syphilis is a re-emerging infectious threat in areas of North America. The purpose of this study was to quantitatively describe the rate of decline of nontreponemal (rapid plasma reagin, RPR) titers in pregnant persons with syphilis and their infants. In a retrospective review, we included 120 pregnant persons with 563 reactive RPR measurements (median 5, range 2 to 11 per person) and 35 infants with 81 RPR measurements (median 2, range 2 to 6 per infant). First-order decay, second-order decay, and a mathematical model representing functional FcRn-mediated antibody recycling were fitted to individual patient RPR trajectories. The RPR titers decreased with a median half-life of 39 days (IQR 28-59) and 27 days (IQR 17-41) in birthing parents and infants, respectively. The half-life varied with the initial RPR titer, suggesting that the kinetics of RPR decline was not first-order. A mathematical model accounting for saturable antibody recycling explained the longevity of RPR reactivity, predicted the observed non-linear kinetics, and fit the empiric data well. In summary, RPR titers decline with a half-life of roughly one month; however, the elimination does not follow first-order kinetics. Saturable antibody recycling may explain the prolonged and non-linear elimination of nontreponemal antibodies.</p>","PeriodicalId":19758,"journal":{"name":"Pathogens","volume":"13 11","pages":""},"PeriodicalIF":3.3,"publicationDate":"2024-11-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11597520/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142731407","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"In Silico Genomic Analysis of Avian Influenza Viruses Isolated From Marine Seal Colonies.","authors":"Klaudia Chrzastek, Darrell R Kapczynski","doi":"10.3390/pathogens13111009","DOIUrl":"10.3390/pathogens13111009","url":null,"abstract":"<p><p>Genetically diverse avian influenza viruses (AIVs) are maintained in wild aquatic birds with increasingly frequent spillover into mammals, yet these represent a small proportion of the overall detections. The isolation of AIVs in marine mammals, including seals, has been reported sporadically over the last 45 years. Prior to 2016, all reports of AIVs detected in seals were of low-pathogenicity AIVs. In spite of this, the majority of reported AIV outbreaks caused fatal respiratory diseases, with harbor seals particularly susceptible to infection. The H5 clade 2.3.4.4b highly pathogenic AIV (HPAIV) was detected in seals for the first time in 2016. Recently, many cases of mass seal die-offs have occurred because of 2.3.4.4b HPAIV and are attributed to spillover from wild bird species. The potential for seal-to-seal transmission has been considered after the mass mortality of southern elephant seals off the coast of Argentina. Close contact between seals and wild birds, the rapid evolution of H5N1 AIVs, and the possibility of efficient mammal-to-mammal transmission are increasing concerns due to the potential for the establishment of a marine mammal reservoir and public health risks associated with the pandemic potential of the virus. This manuscript details the detection of AIVs in the seal population, comparing interesting features of various subtypes with an emphasis on avian-to-mammal-to-mammal transmission. Phylogenetic characterizations of the representative seal isolates were performed to demonstrate the relationships within the different virus isolates. Furthermore, we demonstrate that the reassortment events between different LPAIVs occurred before and after the viruses reached the seal population. The reassortment of viral segments plays an important role in the evolution of influenza viruses. Taken together, these data report on the 45 year history between seals and AIVs.</p>","PeriodicalId":19758,"journal":{"name":"Pathogens","volume":"13 11","pages":""},"PeriodicalIF":3.3,"publicationDate":"2024-11-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11597452/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142731491","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Quantifying the Molecular Properties of the Elk Chronic Wasting Disease Agent with Mass Spectrometry.","authors":"Christopher J Silva, Melissa L Erickson-Beltran, Eric D Cassmann, Justin J Greenlee","doi":"10.3390/pathogens13111008","DOIUrl":"10.3390/pathogens13111008","url":null,"abstract":"<p><p>Chronic wasting disease (CWD) is a prion disease afflicting wild and farmed elk. CWD prions (PrP^Sc) are infectious protein conformations that replicate by inducing a natively expressed prion protein (PrP^C) to refold into the prion conformation. Mass spectrometry was used to study the prions resulting from a previously described experimental inoculation of MM132, ML132, and LL132 elk with a common CWD inoculum. Chymotryptic digestion times and instrument parameters were optimized to yield a set of six peptides, TNMK, MLGSAMSRPL, LLGSAMSRPL, ENMYR, MMER, and VVEQMCITQYQR. These peptides were used to quantify the amount, the M132 and L132 polymorphic composition, and the extent of methionine oxidation of elk PrP^Sc. The amount (ng/g brain tissue) of PrP^Sc present in each sample was determined to be: MM132 (5.4 × 10² ± 7 × 10¹), ML132 (3.3 × 10² ± 6 × 10¹ and 3.6 × 10² ± 3 × 10¹) and LL132 (0.7 × 10² ± 1 × 10¹, 0.2 × 10² ± 0.2 × 10¹, and 0.2 × 10² ± 0.5 × 10¹). The proportion of L132 polymorphism in ML132 (heterozygous) PrP^Sc from CWD-infected elk was determined to be 43% ± 2% or 36% ± 3%. Methionine oxidation was detected and quantified for the M132 and L132 polymorphisms in the samples. In this way, mass spectrometry can be used to characterize prion strains at a molecular level.</p>","PeriodicalId":19758,"journal":{"name":"Pathogens","volume":"13 11","pages":""},"PeriodicalIF":3.3,"publicationDate":"2024-11-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11597226/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142731515","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"The Evolution of Wisteria Vein Mosaic Virus: A Case Study Approach to Track the Emergence of New Potyvirus Threats.","authors":"Massimiliano Morelli, Giusy D'Attoma, Pasquale Saldarelli, Angelantonio Minafra","doi":"10.3390/pathogens13111001","DOIUrl":"10.3390/pathogens13111001","url":null,"abstract":"<p><p>Wisteria vein mosaic virus (WVMV, <i>Potyvirus wisteriae</i>), a virus belonging to the genus <i>Potyvirus</i>, is responsible for Wisteria vein mosaic disease (WMD), a severe disease that affects <i>Wisteria</i>, a genus of garden plants acclaimed worldwide. Although probably originating in the Far East, WVMV infection was first reported in the US, and subsequently in numerous countries. Following the first molecular detection of an Italian isolate, WVMV Bari, its full-length genome was achieved using NGS barcoding technology. A PhyML phylogenetic analysis, supported by clustering algorithm validation, identified a clear separation between two phylogroups. One major clade comprised WVMV strains isolated from <i>Wisteria</i> spp. A second clade grouped three highly divergent strains, at the borderline species threshold, all found in non-wisteria hosts. Relying on a Relative Time Dated Tips (RTDT) molecular clock, the first emergence of WVMV clades has been traced back to around the 17th century. A network inference analysis confirmed the sharp separation between the two host-related phylogroups, also highlighting the presence of potential intermediate variants. Inter-population genetic parameters revealed a very high genetic differentiation in both populations, which was made reliable by statistically significant permutation tests. The migrant number (<i>N</i>m) and fixation index (<i>F</i>_ST) evidenced a restricted gene flow and strong population structures. According to the d<i>N</i>/d<i>S</i> ratio and negative neutrality tests, it was derived that purifying selection at the expense of non-silent variants is underway within WVMV populations. Targeting WVMV evolutionary traits, the present effort raised interesting questions about the underestimated potential of this culpably neglected species to spread in economically relevant crops. The main intention of our study is, therefore, to propose an evolution-based analysis approach that serves as a case study to investigate how other potyviruses or newly emerging viruses may spread.</p>","PeriodicalId":19758,"journal":{"name":"Pathogens","volume":"13 11","pages":""},"PeriodicalIF":3.3,"publicationDate":"2024-11-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11597161/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142731593","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Intestinal Carriage of Two Distinct <i>stx_2f</i>-Carrying <i>Escherichia coli</i> Strains by a Child with Uncomplicated Diarrhea.","authors":"Florence Crombé, Angela H A M van Hoek, Heleen Nailis, Frédéric Auvray, Toon Janssen, Denis Piérard","doi":"10.3390/pathogens13111002","DOIUrl":"10.3390/pathogens13111002","url":null,"abstract":"<p><p>Two distinct <i>stx_2f</i>-carrying <i>Escherichia coli</i> (<i>E. coli</i>) strains, isolated from a child with uncomplicated diarrhea fifteen weeks apart, were characterized by combining short- and long-read sequencing to compare their genetic relatedness. One strain was characterized as Shiga toxin-producing <i>E. coli</i> (STEC)/typical enteropathogenic <i>E. coli</i> (tEPEC) O63:H6 with a repertoire of virulence genes including <i>stx_2f</i>, <i>eae</i> (α2-subtype), <i>cdt</i>, and <i>bfpA</i>. The other STEC with serotype O157:H16, reported for the first time as <i>stx_2f</i>-carrying <i>Escherichia coli</i> in this study, possessed, in addition, <i>eae</i> (ε-subtype) and <i>cdt</i>, amongst other virulence-related genes. BLAST comparison showed that the <i>stx_2f</i>-harboring prophage sequences of both strains were highly homologous (99.6% identity and 96.1% coverage). These results were corroborated by core Stx2f phage Multilocus Sequence Typing (cpMLST) as the <i>stx_2f</i>-harboring prophages of both isolates clustered together when compared to those of 167 other human <i>stx_2f</i>-carrying <i>Escherichia coli</i>. Overall, the <i>stx_2f</i>-harboring prophages of the two distinct <i>E. coli</i> strains isolated from the present case were highly similar, suggesting that the <i>stx_2f</i>-harboring phage might have been transferred from the STEC/tEPEC O63:H6 strain to the atypical EPEC (aEPEC) O157:H16 strain in the gut of the child.</p>","PeriodicalId":19758,"journal":{"name":"Pathogens","volume":"13 11","pages":""},"PeriodicalIF":3.3,"publicationDate":"2024-11-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11597368/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142731554","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"<i>Mycoplasma bovis</i> Invades Non-Phagocytic Cells by Clathrin-Dependent Endocytic Pathways and Escapes from Phagocytic Vesicles.","authors":"Bin Li, Yabin Lu, Yaru Feng, Xiaolong Jiao, Qiuyu Zhang, Mengting Zhou, Yuyu Zhang, Jian Xu, Yuefeng Chu, Duoliang Ran","doi":"10.3390/pathogens13111003","DOIUrl":"10.3390/pathogens13111003","url":null,"abstract":"<p><p><i>Mycoplasma bovis</i> (<i>M. bovis</i>) is capable of causing pneumonia, arthritis, mastitis, and various other ailments in cattle of all age groups, posing a significant threat to the healthy progression of the worldwide cattle industry. The invasion of non-phagocytic host cells serves as a pivotal mechanism enabling <i>M. bovis</i> to evade the immune system and penetrate mucosal barriers, thereby promoting its spread. To investigate the differences in <i>M. bovis</i> invasion into four types of non-phagocytic cells (Madin-Darby bovine kidney (MDBK) cells, embryonic bovine lung (EBL) cells, bovine embryo tracheal (EBTr) cells and bovine turbinate (BT) cells) and further elucidate its invasion mechanism, this study first optimized the experimental methods for <i>M. bovis</i> invasion into cells. Utilizing laser scanning confocal microscopy, transmission electron microscopy, and high-content live-cell imaging systems, the invasion process of <i>M. bovis</i> into four types of non-phagocytic cells was observed. The invasion rates of three different strains of <i>M. bovis</i> (PG45, 07801, 08M) were quantified through the plate counting method. In order to clarify the specific pathway of <i>M. bovis</i> invasion into cells, chlorpromazine (CPZ), amiloride (AMI), and methyl-β-cyclodextrin (M-β-CD) were used to inhibit CLR-mediated clathrin-dependent endocytosis (CDE) pathway, macropinocytosis, and lipid raft pathway, respectively. Subsequently, the invasion rates of PG45 into these four types of cells were measured. Using siRNA technology, the expression of clathrin (CLR) in EBL cells was knocked down to further verify the role of CLR in the invasion process of <i>M. bovis</i>. The results showed that the optimal conditions for <i>M. bovis</i> to invade non-phagocytic cells were a multiplicity of infection (MOI) of 1000 and an optimal invasion time of 4 h. All three strains of <i>M. bovis</i> have the ability to invade the four types of non-phagocytic cells, yet their invasion abilities vary significantly. Observations from transmission electron microscopy further confirmed that at 120 min post-infection, PG45 had successfully invaded EBL cells and was present within endocytic vesicles. It is noteworthy that almost all PG45 successfully escaped from the endocytic vesicles after 240 min of infection had passed. Through chemical inhibition experiments and CLR protein knockdown experiments, it was found that when the CDE and lipid raft pathways were blocked or CLR protein expression was reduced, the invasion rates of PG45, 07801, and 08M in MDBK, EBL, EBTr, and BT cells were significantly decreased (<i>p</i> < 0.05). The above results indicate that <i>M. bovis</i> can invade all types of non-phagocytic cells through endocytic pathways involving CDE (clathrin-dependent endocytosis) or lipid raft-mediated endocytosis, and possesses the ability to escape from phagosomes.</p>","PeriodicalId":19758,"journal":{"name":"Pathogens","volume":"13 11","pages":""},"PeriodicalIF":3.3,"publicationDate":"2024-11-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11597682/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142731420","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}