cometadata/datacite-titles-descriptions-related-identifiers huggingface dataset:
DOI: 10.6084/m9.figshare.30485597.v1
title: Novel amino acid residues in listerolysin O drive phagosome escape and pathogenicity in <i>Listeria monocytogenes</i>

description: <i>Listeria monocytogenes</i>, a significant zoonotic foodborne intracellular pathogen, relies on its principal virulence factor, listeriolysin O (LLO), to escape from host cell phagosomes-a critical step for establishing intracellular infection and replication. LLO, composed of four structural domains with amino acids essential for its pore-forming abilities, still harbors many sites with undefined roles. We identified a critical domain within LLO (amino acids 251–255) crucial for bacterial pathogenesis and systematically investigated the individual roles of five amino acid sites within this domain. Our findings demonstrate that LLO K252 is dispensable for LLO functionality, residues F251, Q253, and Y255 are critical for LLO’s hemolytic activity and cytotoxicity. Mutation of I254 to alanine alters hemolytic activity, with effects dependent on environmental pH. Notably, alanine mutations at F251, Q253, and Y255A significantly impair bacterial proliferation in mouse organs and disruption of late phagosome membranes. In particular, mutations F251A and Y255A abolish late phagosome escape, underscoring their importance in host cell infection. This study has expanded our understanding of LLO’s role in <i>Listeria</i> pathogenicity and phagosome escape, offering new insights into the molecular mechanisms driving this pathogens’ virulence.
DOI: 10.6084/m9.figshare.30325563
title: Interaction with amoeba drives virulence-associated phenotypes in the <i>Candida haemulonii</i> complex

description: Predation by amoebae can induce adaptive traits in microorganisms, enhancing their resilience and pathogenic potential. This study investigated whether interaction with <i>Acanthamoeba castellanii</i> modulates the virulence of environmental fungi <i>Candida haemulonii</i> complex species. Our findings reveal that adaptation to an amoebic host induces significant phenotypic changes in <i>C. duobushaemulonii</i> isolated from a beetle’s gut, leading to four distinct phenotypes: smooth-pink, rough-pink, intense-pink, and hyphae-white. These variants exhibited increased aggregation and filamentation. Amoeba-adapted isolates produced over 15-fold more biofilm biomass and more than 2.2-fold greater extracellular matrix than non-adapted isolates. Transcriptional analysis revealed upregulation of virulence-associated genes, including dimorphism regulators <i>WOR1</i> ( &gt; 2.4-fold), <i>CPH1</i> (4.8-fold), and <i>HGC1</i> ( &gt; 1.9-fold) as well as biofilm-related genes <i>ALS1</i> ( &gt; 3.6-fold), <i>BCR1</i> ( &gt; 14.5-fold), and <i>EGF1</i> ( &gt; 1.6-fold), demonstrating amoeba-induced enhancement of pathogenic traits. Furthermore, adaptation enhanced fungal tolerance to environmental stressors and resulted in more than a 1.3-fold increase in the secretion of hydrolytic enzymes, including esterase, phospholipase, and secreted aspartyl proteinase. Pathogenicity assays using <i>Galleria mellonella</i> revealed significantly increased virulence of amoeba-adapted isolates, with a reduction in the lethal dose (LD<sub>50</sub>) by 71.5–97.1% compared to non-adapted controls (NA: 2.52 ± 0.22 × 10<sup>6</sup> CFU/mL; P10:HW: 7.17 ± 2.14 × 10<sup>4</sup> CFU/mL; P10:RP: 1.25 ± 0.15 × 10<sup>5</sup> CFU/mL). These findings suggest that environmental interaction with amoebae can promote the emergence of virulence-associated phenotypes in <i>C. duobushaemulonii</i>, providing a model for understanding how environmental pressures may facilitate fungal pathogenesis.
DOI: 10.6084/m9.figshare.30325563.v1
title: Interaction with amoeba drives virulence-associated phenotypes in the <i>Candida haemulonii</i> complex

description: Predation by amoebae can induce adaptive traits in microorganisms, enhancing their resilience and pathogenic potential. This study investigated whether interaction with <i>Acanthamoeba castellanii</i> modulates the virulence of environmental fungi <i>Candida haemulonii</i> complex species. Our findings reveal that adaptation to an amoebic host induces significant phenotypic changes in <i>C. duobushaemulonii</i> isolated from a beetle’s gut, leading to four distinct phenotypes: smooth-pink, rough-pink, intense-pink, and hyphae-white. These variants exhibited increased aggregation and filamentation. Amoeba-adapted isolates produced over 15-fold more biofilm biomass and more than 2.2-fold greater extracellular matrix than non-adapted isolates. Transcriptional analysis revealed upregulation of virulence-associated genes, including dimorphism regulators <i>WOR1</i> ( &gt; 2.4-fold), <i>CPH1</i> (4.8-fold), and <i>HGC1</i> ( &gt; 1.9-fold) as well as biofilm-related genes <i>ALS1</i> ( &gt; 3.6-fold), <i>BCR1</i> ( &gt; 14.5-fold), and <i>EGF1</i> ( &gt; 1.6-fold), demonstrating amoeba-induced enhancement of pathogenic traits. Furthermore, adaptation enhanced fungal tolerance to environmental stressors and resulted in more than a 1.3-fold increase in the secretion of hydrolytic enzymes, including esterase, phospholipase, and secreted aspartyl proteinase. Pathogenicity assays using <i>Galleria mellonella</i> revealed significantly increased virulence of amoeba-adapted isolates, with a reduction in the lethal dose (LD<sub>50</sub>) by 71.5–97.1% compared to non-adapted controls (NA: 2.52 ± 0.22 × 10<sup>6</sup> CFU/mL; P10:HW: 7.17 ± 2.14 × 10<sup>4</sup> CFU/mL; P10:RP: 1.25 ± 0.15 × 10<sup>5</sup> CFU/mL). These findings suggest that environmental interaction with amoebae can promote the emergence of virulence-associated phenotypes in <i>C. duobushaemulonii</i>, providing a model for understanding how environmental pressures may facilitate fungal pathogenesis.
DOI: 10.6084/m9.figshare.29465040
title: Emergence of porcine-derived hybrid <i>Escherichia coli</i> with dual-disease manifestations in intestinal and extraintestinal niches

description: <i>Escherichia coli</i>, a significant zoonotic pathogen, continuously evolves to produce hybrid strains with diverse pathogenic characteristics, threatening public health. These hybrid strains are capable of causing multiple diseases across various tissues within a single host or among genetically diverse hosts. They have been reported to exhibit heightened virulence, enhanced infectiousness, and increased resistance to environmental clearance. Here, we identified a hybrid aEPEC/ExPEC isolate, PE164, from the bloody stool of a piglet with severe diarrhoea, which exhibited both intestinal and extraintestinal pathogenic traits. Monitoring the emergence of PE164-like hybrids in porcine-derived <i>E. coli</i> is crucial for animal production and public health safety. Phylogenetic and virulence gene analyses of nearly 300 clinical isolates from diseased pigs revealed eight hybrid strains categorized into four types, with significant variations in pathogenicity confirmed through <i>in vitro</i> and <i>in vivo</i> models. Notably, two O108 isolates, PE150 and PE164, were classified as highly virulent strains. A fused gene cluster involved in colanic acid and O108 antigen biosynthesis (CAO108) was identified as a key determinant of their extraintestinal pathogenicity, raising concerns about O-antigens linked to other exopolysaccharide components to generate new high virulence factors. Furthermore, the type III secretion system (T3SS), a classic pathway for pathogenic <i>E. coli</i> to optimally colonize the host gut, enhanced bacterial immune evasion and contributed to the extraintestinal pathogenicity of PE164. These findings underscore the prevalence of hybrid pathogenic <i>E. coli</i> strains in pigs, presenting new challenges for disease control.
DOI: 10.6084/m9.figshare.29465040.v1
title: Emergence of porcine-derived hybrid <i>Escherichia coli</i> with dual-disease manifestations in intestinal and extraintestinal niches

description: <i>Escherichia coli</i>, a significant zoonotic pathogen, continuously evolves to produce hybrid strains with diverse pathogenic characteristics, threatening public health. These hybrid strains are capable of causing multiple diseases across various tissues within a single host or among genetically diverse hosts. They have been reported to exhibit heightened virulence, enhanced infectiousness, and increased resistance to environmental clearance. Here, we identified a hybrid aEPEC/ExPEC isolate, PE164, from the bloody stool of a piglet with severe diarrhoea, which exhibited both intestinal and extraintestinal pathogenic traits. Monitoring the emergence of PE164-like hybrids in porcine-derived <i>E. coli</i> is crucial for animal production and public health safety. Phylogenetic and virulence gene analyses of nearly 300 clinical isolates from diseased pigs revealed eight hybrid strains categorized into four types, with significant variations in pathogenicity confirmed through <i>in vitro</i> and <i>in vivo</i> models. Notably, two O108 isolates, PE150 and PE164, were classified as highly virulent strains. A fused gene cluster involved in colanic acid and O108 antigen biosynthesis (CAO108) was identified as a key determinant of their extraintestinal pathogenicity, raising concerns about O-antigens linked to other exopolysaccharide components to generate new high virulence factors. Furthermore, the type III secretion system (T3SS), a classic pathway for pathogenic <i>E. coli</i> to optimally colonize the host gut, enhanced bacterial immune evasion and contributed to the extraintestinal pathogenicity of PE164. These findings underscore the prevalence of hybrid pathogenic <i>E. coli</i> strains in pigs, presenting new challenges for disease control.
DOI: 10.6084/m9.figshare.28457552
title: Human SLAM-adapted canine distemper virus can enter human peripheral blood mononuclear cells and replicate in mice expressing human SLAM and defective for STAT1 expression

description: Canine distemper virus (CDV) is a member of the genus <i>Morbillivirus</i> with a worldwide distribution that causes fatal diseases in canids and marine mammals. In recent years, CDV has demonstrated the remarkable ability of pathogens to cross species barriers. The natural host range of CDV has expanded from Canidae to Primates, presumably attributed to ecological shifts and the emergence of viral variants. Therefore, it is important to investigate whether CDV can infect humans by adapting to the human signalling lymphocyte activation molecule (hSLAM) receptor to cross the species barrier. Through successive passaging and plaque cloning of a CDV wild-type strain (5804PeH) in Vero cells expressing hSLAM (Vero-hSLAM), we obtained an hSLAM adaptive strain, 5804PeH-VhS. The adapted CDV strain exhibited a D540G mutation within the receptor-binding domain (RBD) of the haemagglutinin (H) protein. The H<sup>D540G</sup> mutation has enhanced cell-cell fusion activity in Vero-hSLAM cells. This adaptation allowed the CDV strain to infect human peripheral blood mononuclear cells (PBMCs), particularly T lymphocytes and inhibited lymphocyte proliferation. Additionally, this strain could replicate in the lymphoid tissues of transgenic mice that express the hSLAM receptor, causing viraemia. However, the adapted strain did not spread to the epithelial cells or the central nervous system of the mice. While this adaptation indicates a potential risk, there is no definitive evidence that the virus can spread among humans.
DOI: 10.6084/m9.figshare.28457552.v1
title: Human SLAM-adapted canine distemper virus can enter human peripheral blood mononuclear cells and replicate in mice expressing human SLAM and defective for STAT1 expression

description: Canine distemper virus (CDV) is a member of the genus <i>Morbillivirus</i> with a worldwide distribution that causes fatal diseases in canids and marine mammals. In recent years, CDV has demonstrated the remarkable ability of pathogens to cross species barriers. The natural host range of CDV has expanded from Canidae to Primates, presumably attributed to ecological shifts and the emergence of viral variants. Therefore, it is important to investigate whether CDV can infect humans by adapting to the human signalling lymphocyte activation molecule (hSLAM) receptor to cross the species barrier. Through successive passaging and plaque cloning of a CDV wild-type strain (5804PeH) in Vero cells expressing hSLAM (Vero-hSLAM), we obtained an hSLAM adaptive strain, 5804PeH-VhS. The adapted CDV strain exhibited a D540G mutation within the receptor-binding domain (RBD) of the haemagglutinin (H) protein. The H<sup>D540G</sup> mutation has enhanced cell-cell fusion activity in Vero-hSLAM cells. This adaptation allowed the CDV strain to infect human peripheral blood mononuclear cells (PBMCs), particularly T lymphocytes and inhibited lymphocyte proliferation. Additionally, this strain could replicate in the lymphoid tissues of transgenic mice that express the hSLAM receptor, causing viraemia. However, the adapted strain did not spread to the epithelial cells or the central nervous system of the mice. While this adaptation indicates a potential risk, there is no definitive evidence that the virus can spread among humans.
DOI: 10.6084/m9.figshare.29979556
title: Investigating the differential microRNAs expression in young and aged <i>Drosophila melanogaster</i> following Flock House Virus infection

description: MicroRNAs (miRNAs) are small non-coding RNAs ~ 19–22 nt long that post-transcriptionally regulate their mRNA targets. In <i>Drosophila melanogaster</i>, the role of miRNAs has mostly been studied in regard to bacterial infection, yielding insights about their regulatory function in innate immunity. However, fewer studies have focused on viral infections, and importantly, how miRNAs modulate aging immune responses is not fully understood. Here, we performed small RNA-sequencing demonstrating that systemic Flock House Virus (FHV) infection of <i>Oregon-R</i> flies leads to differential microRNA expression and that this response differs with aging. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analyses identified cellular pathways and biological processes which may be regulated by dynamic expression of microRNAs during infection. For 17 candidate miRNAs, we tested <i>Drosophila</i> lines with <i>in vivo</i> miRNA knockdown for their survival of systemic FHV challenge. In response to infection, among miRNA knockdown lines, females consistently outlived males, and young flies generally outlived their aged counterparts. Furthermore, miRNA knockdown lines generally displayed increased susceptibility to viral infection in comparison to controls, particularly prominent among males. For one miRNA chosen for further study, <i>miR-311</i>, its dysregulation resulted in decreased survival independent of changes in viral load, suggesting a role in disease tolerance rather than resistance. Lastly, knockdown of the <i>immune deficiency</i> (<i>imd</i>) gene – a predicted target of <i>miR-311</i> - was associated with improved survival of FHV. This work identifies changes in miRNA expression in the aging antiviral response and highlights one miRNA with a role in disease tolerance to FHV in <i>Drosophila</i>.
DOI: 10.6084/m9.figshare.29979556.v1
title: Investigating the differential microRNAs expression in young and aged <i>Drosophila melanogaster</i> following Flock House Virus infection

description: MicroRNAs (miRNAs) are small non-coding RNAs ~ 19–22 nt long that post-transcriptionally regulate their mRNA targets. In <i>Drosophila melanogaster</i>, the role of miRNAs has mostly been studied in regard to bacterial infection, yielding insights about their regulatory function in innate immunity. However, fewer studies have focused on viral infections, and importantly, how miRNAs modulate aging immune responses is not fully understood. Here, we performed small RNA-sequencing demonstrating that systemic Flock House Virus (FHV) infection of <i>Oregon-R</i> flies leads to differential microRNA expression and that this response differs with aging. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analyses identified cellular pathways and biological processes which may be regulated by dynamic expression of microRNAs during infection. For 17 candidate miRNAs, we tested <i>Drosophila</i> lines with <i>in vivo</i> miRNA knockdown for their survival of systemic FHV challenge. In response to infection, among miRNA knockdown lines, females consistently outlived males, and young flies generally outlived their aged counterparts. Furthermore, miRNA knockdown lines generally displayed increased susceptibility to viral infection in comparison to controls, particularly prominent among males. For one miRNA chosen for further study, <i>miR-311</i>, its dysregulation resulted in decreased survival independent of changes in viral load, suggesting a role in disease tolerance rather than resistance. Lastly, knockdown of the <i>immune deficiency</i> (<i>imd</i>) gene – a predicted target of <i>miR-311</i> - was associated with improved survival of FHV. This work identifies changes in miRNA expression in the aging antiviral response and highlights one miRNA with a role in disease tolerance to FHV in <i>Drosophila</i>.
DOI: 10.6084/m9.figshare.28099610
title: Optimizing encephalomyocarditis virus VP1 protein assembly on pseudorabies virus envelope via US9 protein anchoring

description: Live herpesvirus-vectored vaccines are critical in veterinary medicine, but they can sometimes offer insufficient protection due to suboptimal antigen expression or localization. Encephalomyocarditis virus (EMCV) is a significant zoonotic threat, with VP1 protein as a key immunogen on its capsid. To enhance immunogenicity, we explored the use of recombinant pseudorabies virus (rPRV) as a vaccine vector against EMCV. In silico analysis indicated that fusing VP1 with US9 enhances the formation of a type II transmembrane heterodimer. We constructed six rPRV groups expressing different VP1 variants and found that VP1 fused with US9’s C-terminal (US9-VP1) enhances VP1’s membrane localization and its incorporation into the PRV envelope, unlike wild-type VP1. Immunogold electron microscopy illustrated that rPRV with deleted US8 and US9, supplemented with US8 regulatory sequence (rΔ89-U9VP1), improved VP1 incorporation into the viral envelope. Post-immunization, only rΔ89-U9VP1 provided 100% protection against EMCV in mice and induced high levels of virus-neutralizing antibodies in piglets. Additionally, rPRV expressing VP1 stimulated robust T-cell responses, as demonstrated by flow cytometry and ELISpot assays. This study introduces rPRV as a potential EMCV vaccine, demonstrating that the selection of the US9 C-terminal domain and US8 regulatory sequence significantly enhances the presentation of heterologous antigens, improving vaccine efficacy.
DOI: 10.6084/m9.figshare.28099610.v1
title: Optimizing encephalomyocarditis virus VP1 protein assembly on pseudorabies virus envelope via US9 protein anchoring

description: Live herpesvirus-vectored vaccines are critical in veterinary medicine, but they can sometimes offer insufficient protection due to suboptimal antigen expression or localization. Encephalomyocarditis virus (EMCV) is a significant zoonotic threat, with VP1 protein as a key immunogen on its capsid. To enhance immunogenicity, we explored the use of recombinant pseudorabies virus (rPRV) as a vaccine vector against EMCV. In silico analysis indicated that fusing VP1 with US9 enhances the formation of a type II transmembrane heterodimer. We constructed six rPRV groups expressing different VP1 variants and found that VP1 fused with US9’s C-terminal (US9-VP1) enhances VP1’s membrane localization and its incorporation into the PRV envelope, unlike wild-type VP1. Immunogold electron microscopy illustrated that rPRV with deleted US8 and US9, supplemented with US8 regulatory sequence (rΔ89-U9VP1), improved VP1 incorporation into the viral envelope. Post-immunization, only rΔ89-U9VP1 provided 100% protection against EMCV in mice and induced high levels of virus-neutralizing antibodies in piglets. Additionally, rPRV expressing VP1 stimulated robust T-cell responses, as demonstrated by flow cytometry and ELISpot assays. This study introduces rPRV as a potential EMCV vaccine, demonstrating that the selection of the US9 C-terminal domain and US8 regulatory sequence significantly enhances the presentation of heterologous antigens, improving vaccine efficacy.
DOI: 10.6084/m9.figshare.29817969
title: Riboflavin inhibits growth and reduces virulence of <i>Cryptococcus neoformans in vitro</i> by membrane disruption and excessive accumulation of reactive oxygen species and exhibits efficacy against pulmonary cryptococcosis and meningitis

description: The incidences of pulmonary cryptococcosis and meningitis cause significant morbidity and mortality. Effective and affordable drugs for treatment of cryptococcal meningitis are urgently needed. Drug reuse is an effective strategy for the development of new antifungals against <i>Cryptococcus neoformans</i> infection. In this study, riboflavin (RF) significantly inhibited growth of <i>C. neoformans</i> as determined by the broth microdilution and spot dilution methods. Moreover, RF significantly inhibited biofilm formation and reduced virulence (capsule, melanin, and urease). In addition, RF caused cell membrane damage, compromised cell wall integrity, and promoted accumulation of intracellular reactive oxygen species (ROS). RT-qPCR analysis confirmed that RF treatment up-regulated expression genes related to cell wall biosynthesis (<i>CHS3</i>, <i>CDA1</i>, and <i>FKS1</i>), the cell wall damage repair pathway (<i>Pkc1</i> and <i>Mpk1</i>), and virulence (<i>CAP59</i>, <i>Lac1</i>, <i>Lac2</i>), however, <i>Ure1</i> were down-regulated after RF treatment. Finally, in mouse models of intranasal and intravenous infection, RF treatment significantly reduced the fungal burden in multiple organs, reduced lung and brain damage, and decreased the levels of plasma interferon (IFN)-γ, tumor necrosis factor (TNF)-α, and interleukin (IL)-4 in the early stage of infection. These results showed that RF exerted significant antifungal effects for treatment of <i>C. neoformans</i> infection.
DOI: 10.6084/m9.figshare.29817969.v1
title: Riboflavin inhibits growth and reduces virulence of <i>Cryptococcus neoformans in vitro</i> by membrane disruption and excessive accumulation of reactive oxygen species and exhibits efficacy against pulmonary cryptococcosis and meningitis

description: The incidences of pulmonary cryptococcosis and meningitis cause significant morbidity and mortality. Effective and affordable drugs for treatment of cryptococcal meningitis are urgently needed. Drug reuse is an effective strategy for the development of new antifungals against <i>Cryptococcus neoformans</i> infection. In this study, riboflavin (RF) significantly inhibited growth of <i>C. neoformans</i> as determined by the broth microdilution and spot dilution methods. Moreover, RF significantly inhibited biofilm formation and reduced virulence (capsule, melanin, and urease). In addition, RF caused cell membrane damage, compromised cell wall integrity, and promoted accumulation of intracellular reactive oxygen species (ROS). RT-qPCR analysis confirmed that RF treatment up-regulated expression genes related to cell wall biosynthesis (<i>CHS3</i>, <i>CDA1</i>, and <i>FKS1</i>), the cell wall damage repair pathway (<i>Pkc1</i> and <i>Mpk1</i>), and virulence (<i>CAP59</i>, <i>Lac1</i>, <i>Lac2</i>), however, <i>Ure1</i> were down-regulated after RF treatment. Finally, in mouse models of intranasal and intravenous infection, RF treatment significantly reduced the fungal burden in multiple organs, reduced lung and brain damage, and decreased the levels of plasma interferon (IFN)-γ, tumor necrosis factor (TNF)-α, and interleukin (IL)-4 in the early stage of infection. These results showed that RF exerted significant antifungal effects for treatment of <i>C. neoformans</i> infection.