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1.
PLoS Pathog ; 16(9): e1008867, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32925969

RESUMO

Surface attachment, an early step in the colonization of multiple host environments, activates the virulence of the human pathogen P. aeruginosa. However, the downstream toxins that mediate surface-dependent P. aeruginosa virulence remain unclear, as do the signaling pathways that lead to their activation. Here, we demonstrate that alkyl-quinolone (AQ) secondary metabolites are rapidly induced upon surface association and act directly on host cells to cause cytotoxicity. Surface-induced AQ cytotoxicity is independent of other AQ functions like quorum sensing or PQS-specific activities like iron sequestration. We further show that packaging of AQs in outer-membrane vesicles (OMVs) increases their cytotoxicity to host cells but not their ability to stimulate downstream quorum sensing pathways in bacteria. OMVs lacking AQs are significantly less cytotoxic, suggesting these molecules play a role in OMV cytotoxicity, in addition to their previously characterized role in OMV biogenesis. AQ reporters also enabled us to dissect the signal transduction pathways downstream of the two known regulators of surface-dependent virulence, the quorum sensing receptor, LasR, and the putative mechanosensor, PilY1. Specifically, we show that PilY1 regulates surface-induced AQ production by repressing the AlgR-AlgZ two-component system. AlgR then induces RhlR, which can induce the AQ biosynthesis operon under specific conditions. These findings collectively suggest that the induction of AQs upon surface association is both necessary and sufficient to explain surface-induced P. aeruginosa virulence.


Assuntos
Proteínas de Bactérias/metabolismo , Infecções por Pseudomonas/metabolismo , Pseudomonas aeruginosa , Quinolonas/farmacologia , Percepção de Quorum/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Fatores de Virulência/metabolismo , Células A549 , Animais , Humanos , Camundongos , Infecções por Pseudomonas/patologia , Pseudomonas aeruginosa/metabolismo , Pseudomonas aeruginosa/patogenicidade
2.
PLoS Pathog ; 16(8): e1008822, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32866204

RESUMO

Shigella flexneri invades host cells by entering within a bacteria-containing vacuole (BCV). In order to establish its niche in the host cytosol, the bacterium ruptures its BCV. Contacts between S. flexneri BCV and infection-associated macropinosomes (IAMs) formed in situ have been reported to enhance BCV disintegration. The mechanism underlying S. flexneri vacuolar escape remains however obscure. To decipher the molecular mechanism priming the communication between the IAMs and S. flexneri BCV, we performed mass spectrometry-based analysis of the magnetically purified IAMs from S. flexneri-infected cells. While proteins involved in host recycling and exocytic pathways were significantly enriched at the IAMs, we demonstrate more precisely that the S. flexneri type III effector protein IpgD mediates the recruitment of the exocyst to the IAMs through the Rab8/Rab11 pathway. This recruitment results in IAM clustering around S. flexneri BCV. More importantly, we reveal that IAM clustering subsequently facilitates an IAM-mediated unwrapping of the ruptured vacuole membranes from S. flexneri, enabling the naked bacterium to be ready for intercellular spread via actin-based motility. Taken together, our work untangles the molecular cascade of S. flexneri-driven host trafficking subversion at IAMs to develop its cytosolic lifestyle, a crucial step en route for infection progression at cellular and tissue level.


Assuntos
Disenteria Bacilar , Shigella flexneri , Transdução de Sinais , Vacúolos , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Disenteria Bacilar/genética , Disenteria Bacilar/metabolismo , Células HeLa , Humanos , Shigella flexneri/genética , Shigella flexneri/metabolismo , Shigella flexneri/patogenicidade , Vacúolos/genética , Vacúolos/metabolismo , Vacúolos/microbiologia , Fatores de Virulência/genética , Fatores de Virulência/metabolismo , Proteínas rab de Ligação ao GTP/genética , Proteínas rab de Ligação ao GTP/metabolismo
3.
Nat Commun ; 11(1): 4894, 2020 09 29.
Artigo em Inglês | MEDLINE | ID: mdl-32994400

RESUMO

Identification of the complete set of translated genes of viruses is important to understand viral replication and pathogenesis as well as for therapeutic approaches to control viral infection. Here, we use chemical proteomics, integrating bio-orthogonal non-canonical amino acid tagging and high-resolution mass spectrometry, to characterize the newly synthesized herpes simplex virus 1 (HSV-1) proteome in infected cells. In these infected cells, host cellular protein synthesis is shut-off, increasing the chance to preferentially detect viral proteomes. We identify nine previously cryptic orphan protein coding sequences whose translated products are expressed in HSV-1-infected cells. Functional characterization of one identified protein, designated piUL49, shows that it is critical for HSV-1 neurovirulence in vivo by regulating the activity of virally encoded dUTPase, a key enzyme that maintains accurate DNA replication. Our results demonstrate that cryptic orphan protein coding genes of HSV-1, and probably other large DNA viruses, remain to be identified.


Assuntos
Encefalite por Herpes Simples/virologia , Herpesvirus Humano 1/patogenicidade , Pirofosfatases/metabolismo , Proteínas Virais/metabolismo , Fatores de Virulência/metabolismo , Animais , Encéfalo/patologia , Encéfalo/virologia , Chlorocebus aethiops , Replicação do DNA , Modelos Animais de Doenças , Encefalite por Herpes Simples/patologia , Feminino , Genes Virais/genética , Células HEK293 , Células HeLa , Herpesvirus Humano 1/genética , Humanos , Camundongos , Biossíntese de Proteínas , Proteômica/métodos , Células Vero , Proteínas Virais/genética , Fatores de Virulência/genética , Replicação Viral
4.
Nat Commun ; 11(1): 4382, 2020 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-32873802

RESUMO

Fusarium graminearum is a causal agent of Fusarium head blight (FHB) and a deoxynivalenol (DON) producer. In this study, OSP24 is identified as an important virulence factor in systematic characterization of the 50 orphan secreted protein (OSP) genes of F. graminearum. Although dispensable for growth and initial penetration, OSP24 is important for infectious growth in wheat rachis tissues. OSP24 is specifically expressed during pathogenesis and its transient expression suppresses BAX- or INF1-induced cell death. Osp24 is translocated into plant cells and two of its 8 cysteine-residues are required for its function. Wheat SNF1-related kinase TaSnRK1α is identified as an Osp24-interacting protein and shows to be important for FHB resistance in TaSnRK1α-overexpressing or silencing transgenic plants. Osp24 accelerates the degradation of TaSnRK1α by facilitating its association with the ubiquitin-26S proteasome. Interestingly, TaSnRK1α also interacts with TaFROG, an orphan wheat protein induced by DON. TaFROG competes against Osp24 for binding with the same region of TaSnRKα and protects it from degradation. Overexpression of TaFROG stabilizes TaSnRK1α and increases FHB resistance. Taken together, Osp24 functions as a cytoplasmic effector by competing against TaFROG for binding with TaSnRK1α, demonstrating the counteracting roles of orphan proteins of both host and fungal pathogens during their interactions.


Assuntos
Proteínas Fúngicas/metabolismo , Fusarium/patogenicidade , Doenças das Plantas/imunologia , Proteínas de Plantas/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Triticum/microbiologia , Fatores de Virulência/metabolismo , Resistência à Doença , Fusarium/imunologia , Fusarium/metabolismo , Interações Hospedeiro-Patógeno/imunologia , Doenças das Plantas/microbiologia , Proteínas de Plantas/genética , Proteínas de Plantas/imunologia , Plantas Geneticamente Modificadas , Complexo de Endopeptidases do Proteassoma/metabolismo , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/imunologia , Proteólise , Tricotecenos/metabolismo , Triticum/imunologia
5.
PLoS Pathog ; 16(8): e1008781, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32810179

RESUMO

Chagas disease is caused by Trypanosoma cruzi, a protozoan parasite that has a heterogeneous population composed of a pool of strains with distinct characteristics, including variable levels of virulence. In previous work, transcriptome analyses of parasite genes after infection of human foreskin fibroblasts (HFF) with virulent (CL Brener) and non-virulent (CL-14) clones derived from the CL strain, revealed a reduced expression of genes encoding parasite surface proteins in CL-14 compared to CL Brener during the final steps of the intracellular differentiation from amastigotes to trypomastigotes. Here we analyzed changes in the expression of host genes during in vitro infection of HFF cells with the CL Brener and CL-14 strains by analyzing total RNA extracted from cells at 60 and 96 hours post-infection (hpi) with each strain, as well as from uninfected cells. Similar transcriptome profiles were observed at 60 hpi with both strains compared to uninfected samples. However, at 96 hpi, significant differences in the number and expression levels of several genes, particularly those involved with immune response and cytoskeleton organization, were observed. Further analyses confirmed the difference in the chemokine/cytokine signaling involved with the recruitment and activation of immune cells such as neutrophils upon T. cruzi infection. These findings suggest that infection with the virulent CL Brener strain induces a more robust inflammatory response when compared with the non-virulent CL-14 strain. Importantly, the RNA-Seq data also exposed an unexplored role of fibroblasts as sentinel cells that may act by recruiting neutrophils to the initial site of infection. This role for fibroblasts in the regulation of the inflammatory response during infection by T. cruzi was corroborated by measurements of levels of different chemokines/cytokines during in vitro infection and in plasma from Chagas disease patients as well as by neutrophil activation and migration assays.


Assuntos
Doença de Chagas/metabolismo , Fibroblastos , Regulação da Expressão Gênica , Redes Reguladoras de Genes , Ativação de Neutrófilo , Neutrófilos , Trypanosoma cruzi/metabolismo , Doença de Chagas/genética , Doença de Chagas/patologia , Fibroblastos/metabolismo , Fibroblastos/parasitologia , Fibroblastos/patologia , Humanos , Neutrófilos/metabolismo , Neutrófilos/parasitologia , Neutrófilos/patologia , Trypanosoma cruzi/genética , Trypanosoma cruzi/patogenicidade , Fatores de Virulência/genética , Fatores de Virulência/metabolismo
6.
Virology ; 550: 1-7, 2020 11.
Artigo em Inglês | MEDLINE | ID: mdl-32853833

RESUMO

Avian coronavirus infectious bronchitis virus (IBV) is an important pathogen threatening poultry production worldwide. Here, two recombinant IBVs (rYN-1a-aYN and rYN-1b-aYN) were generated in which ORF1a or ORF1b of the virulent YN genome were replaced by the corresponding regions from the attenuated strain aYN. The pathogenicity and virulence of rIBVs were evaluated in ovo and in vivo. The results revealed that mutations in the ORF1a gene during passage in embryonated eggs caused the decreased pathogenicity of virulent IBV YN strain, proven by determination of virus replication in ECEs and CEK cells, the observation of clinical signs, gross lesions, microscopic lesions, tracheal ciliary activity and virus distribution in chickens following exposure to rIBVs. However, mutations in ORF1b had no obvious effect on virus replication in both ECEs and CEK cells, or pathogenicity in chickens. Our findings demonstrate that the replicase 1a gene of avian coronavirus IBV is a determinant of pathogenicity.


Assuntos
Infecções por Coronavirus/veterinária , Vírus da Bronquite Infecciosa/patogenicidade , Doenças das Aves Domésticas/patologia , RNA Replicase/genética , Vírus Reordenados/patogenicidade , Proteínas Virais/genética , Fatores de Virulência/genética , Animais , Linhagem Celular , Embrião de Galinha , Galinhas , Infecções por Coronavirus/patologia , Infecções por Coronavirus/transmissão , Infecções por Coronavirus/virologia , Cricetulus , Células Epiteliais/patologia , Células Epiteliais/virologia , Expressão Gênica , Vida Livre de Germes , Vírus da Bronquite Infecciosa/genética , Vírus da Bronquite Infecciosa/crescimento & desenvolvimento , Mutação , Doenças das Aves Domésticas/transmissão , Doenças das Aves Domésticas/virologia , RNA Replicase/metabolismo , Vírus Reordenados/genética , Vírus Reordenados/crescimento & desenvolvimento , Proteínas Virais/metabolismo , Virulência , Fatores de Virulência/metabolismo , Replicação Viral
7.
PLoS Pathog ; 16(8): e1008835, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32785253

RESUMO

Hyaloperonospora arabidopsidis (Hpa) is an oomycete pathogen causing Arabidopsis downy mildew. Effector proteins secreted from the pathogen into the plant play key roles in promoting infection by suppressing plant immunity and manipulating the host to the pathogen's advantage. One class of oomycete effectors share a conserved 'RxLR' motif critical for their translocation into the host cell. Here we characterize the interaction between an RxLR effector, HaRxL21 (RxL21), and the Arabidopsis transcriptional co-repressor Topless (TPL). We establish that RxL21 and TPL interact via an EAR motif at the C-terminus of the effector, mimicking the host plant mechanism for recruiting TPL to sites of transcriptional repression. We show that this motif, and hence interaction with TPL, is necessary for the virulence function of the effector. Furthermore, we provide evidence that RxL21 uses the interaction with TPL, and its close relative TPL-related 1, to repress plant immunity and enhance host susceptibility to both biotrophic and necrotrophic pathogens.


Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/imunologia , Interações Hospedeiro-Patógeno/imunologia , Oomicetos/fisiologia , Doenças das Plantas/imunologia , Imunidade Vegetal/imunologia , Fatores de Virulência/metabolismo , Arabidopsis/microbiologia , Proteínas de Arabidopsis/genética , Regulação da Expressão Gênica de Plantas , Doenças das Plantas/microbiologia , Virulência , Fatores de Virulência/genética
8.
PLoS One ; 15(8): e0236713, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32750089

RESUMO

Coagulase-negative staphylococci (CoNS) are the most common isolates from blood culture in neonates resulting in high mortality and morbidity. This study investigated CoNS obtained from blood cultures of neonates for antibiotic resistance and virulence factors, and possible association with inflammatory response (C-reactive protein). A total of 93 CoNS isolates were collected from 76 blood cultures of neonates at the Maternity hospital in Kuwait in a six-month period and investigated for susceptibility to antibiotics, carriage of staphylococcal cassette chromosome mec (SCCmec), and virulence-associated genes. The 93 CoNS isolates consisted of S. epidermidis (76; 81.7%), S. capitis (12; 12.9%), S. hominis (2; 2.1%), S. warneri (2; 2.1%) and S. haemolyticus (1; 1.0%). Eighty-six (92.4%) of the isolates were resistant to cefoxitin (MR-CoNS) while 49 (52.7%) expressed multi-antibiotic resistance. The methicillin-resistant isolates (MR-CoNS) carried SCCmec III, SCCmec IVa and four combinations of SCCmec types including SCCmec types I+IVa (one S. warneri and 25 S. epidermidis isolates), types I+III (one S. epidermidis isolate), types III+IVa (six S. epidermidis isolates) and types I+III+IVa (one S. epidermidis isolate). The most common virulence-related genes were icaC, seb, arc detected in 69.7%, 60.5%, 40.8% of the isolates respectively. Two isolates were positive for tst1. No association between C-reactive protein and antibiotic resistance or virulence factors was established. This study revealed that S. epidermidis carrying different SCCmec genetic elements, was the dominant CoNS species isolated from neonatal blood cultures with 90.3% and 36.6% of the isolates positive for genes for biofilm and ACME production respectively.


Assuntos
Antibacterianos/farmacologia , Farmacorresistência Bacteriana , Recém-Nascido Prematuro , Infecções Estafilocócicas/microbiologia , Staphylococcus/efeitos dos fármacos , Toxinas Bacterianas/genética , Biofilmes , Coagulase/metabolismo , Feminino , Genes Bacterianos , Humanos , Recém-Nascido , Kuweit , Masculino , Testes de Sensibilidade Microbiana , Estudos Retrospectivos , Staphylococcus/enzimologia , Staphylococcus/isolamento & purificação , Staphylococcus/metabolismo , Fatores de Virulência/genética , Fatores de Virulência/metabolismo
9.
PLoS One ; 15(7): e0222747, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32639982

RESUMO

Physiology-based differentiation of SH genes and Hemileia vastatrix races is the principal method employed for the characterization of coffee leaf rust resistance. Based on the gene-for-gene theory, nine major rust resistance genes (SH1-9) have been proposed. However, these genes have not been characterized at the molecular level. Consequently, the lack of molecular data regarding rust resistance genes or candidates is a major bottleneck in coffee breeding. To address this issue, we screened a BAC library with resistance gene analogs (RGAs), identified RGAs, characterized and explored for any SH related candidate genes. Herein, we report the identification and characterization of a gene (gene 11), which shares conserved sequences with other SH genes and displays a characteristic polymorphic allele conferring different resistance phenotypes. Furthermore, comparative analysis of the two RGAs belonging to CC-NBS-LRR revealed more intense diversifying selection in tomato and grape genomes than in coffee. For the first time, the present study has unveiled novel insights into the molecular nature of the SH genes, thereby opening new avenues for coffee rust resistance molecular breeding. The characterized candidate RGA is of particular importance for further biological function analysis in coffee.


Assuntos
Café/genética , Resistência à Doença/genética , Genoma de Planta , Sequência de Aminoácidos , Basidiomycota/fisiologia , Sítios de Ligação , Café/classificação , Biblioteca Gênica , Lycopersicon esculentum/classificação , Lycopersicon esculentum/genética , Fases de Leitura Aberta/genética , Filogenia , Doenças das Plantas/microbiologia , Proteínas de Plantas/química , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Polimorfismo Genético , Alinhamento de Sequência , Fatores de Virulência/genética , Fatores de Virulência/metabolismo , Vitis/classificação , Vitis/genética
10.
Nat Commun ; 11(1): 3571, 2020 07 16.
Artigo em Inglês | MEDLINE | ID: mdl-32678094

RESUMO

Pathogenic bacteria of the genus Bartonella can induce vasoproliferative lesions during infection. The underlying mechanisms are unclear, but involve secretion of an unidentified mitogenic factor. Here, we use functional transposon-mutant screening in Bartonella henselae to identify such factor as a pro-angiogenic autotransporter, called BafA. The passenger domain of BafA induces cell proliferation, tube formation and sprouting of microvessels, and drives angiogenesis in mice. BafA interacts with vascular endothelial growth factor (VEGF) receptor-2 and activates the downstream signaling pathway, suggesting that BafA functions as a VEGF analog. A BafA homolog from a related pathogen, Bartonella quintana, is also functional. Our work unveils the mechanistic basis of vasoproliferative lesions observed in bartonellosis, and we propose BafA as a key pathogenic factor contributing to bacterial spread and host adaptation.


Assuntos
Bartonella/patogenicidade , Neovascularização Patológica/metabolismo , Transdução de Sinais , Sistemas de Secreção Tipo V/metabolismo , Fatores de Crescimento do Endotélio Vascular/metabolismo , Fatores de Virulência/metabolismo , Animais , Bartonella/classificação , Bartonella/genética , Proliferação de Células , Perfilação da Expressão Gênica , Células Endoteliais da Veia Umbilical Humana/citologia , Células Endoteliais da Veia Umbilical Humana/metabolismo , Células Endoteliais da Veia Umbilical Humana/microbiologia , Humanos , Camundongos , Neovascularização Patológica/genética , Neovascularização Patológica/microbiologia , Domínios Proteicos , Sistemas de Secreção Tipo V/química , Sistemas de Secreção Tipo V/genética , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Fatores de Virulência/química , Fatores de Virulência/genética
11.
PLoS Pathog ; 16(7): e1008650, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32628723

RESUMO

Toxoplasma gondii is an obligate intracellular parasite that can invade any nucleated cell of any warm-blooded animal. In a previous screen to identify virulence determinants, disruption of gene TgME49_305140 generated a T. gondii mutant that could not establish a chronic infection in mice. The protein product of TgME49_305140, here named TgPL3, is a 277 kDa protein with a patatin-like phospholipase (PLP) domain and a microtubule binding domain. Antibodies generated against TgPL3 show that it is localized to the apical cap. Using a rapid selection FACS-based CRISPR/Cas-9 method, a TgPL3 deletion strain (ΔTgPL3) was generated. ΔTgPL3 parasites have defects in host cell invasion, which may be caused by reduced rhoptry secretion. We generated complementation clones with either wild type TgPL3 or an active site mutation in the PLP domain by converting the catalytic serine to an alanine, ΔTgPL3::TgPL3S1409A (S1409A). Complementation of ΔTgPL3 with wild type TgPL3 restored all phenotypes, while S1409A did not, suggesting that phospholipase activity is necessary for these phenotypes. ΔTgPL3 and S1409A parasites are also virtually avirulent in vivo but induce a robust antibody response. Vaccination with ΔTgPL3 and S1409A parasites protected mice against subsequent challenge with a lethal dose of Type I T. gondii parasites, making ΔTgPL3 a compelling vaccine candidate. These results demonstrate that TgPL3 has a role in rhoptry secretion, host cell invasion and survival of T. gondii during acute mouse infection.


Assuntos
Proteínas de Protozoários/metabolismo , Toxoplasma/patogenicidade , Toxoplasmose/metabolismo , Fatores de Virulência/metabolismo , Animais , Camundongos , Fosfolipases/genética , Fosfolipases/metabolismo , Proteínas de Protozoários/genética , Toxoplasma/genética , Toxoplasmose/enzimologia , Virulência
12.
Eur J Clin Microbiol Infect Dis ; 39(10): 1821-1830, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32557327

RESUMO

Helicobacter pylori (H. pylori) infection is associated with some gastric diseases, such as gastritis, peptic ulcer, and gastric cancer. CagA and VacA are known virulence factors of H. pylori, which play a vital role in severe clinical outcomes. Additionally, the expression of outer membrane proteins (OMPs) helps H. pylori attach to gastric epithelial cells at the primary stage and increases the virulence of H. pylori. In this review, we have summarized the paralogs of H. pylori OMPs, their genomic loci, and the different receptors of OMPs identified so far. We focused on five OMPs, BabA (HopS), SabA (HopP), OipA (HopH), HopQ, and HopZ, and one family of OMPs: Hom. We highlight the coexpression of OMPs with other virulence factors and their relationship with clinical outcomes. In conclusion, OMPs are closely related to the pathogenic processes of adhesion, colonization, persistent infection, and severe clinical consequences. They are potential targets for the prevention and treatment of H. pylori-related diseases.


Assuntos
Proteínas da Membrana Bacteriana Externa/genética , Proteínas da Membrana Bacteriana Externa/metabolismo , Infecções por Helicobacter/microbiologia , Helicobacter pylori/genética , Helicobacter pylori/patogenicidade , Fatores de Virulência/metabolismo , Células Epiteliais/microbiologia , Expressão Gênica , Humanos , Fatores de Virulência/genética
14.
PLoS One ; 15(6): e0235020, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32584855

RESUMO

The pathogenesis of Salmonella Typhimurium depends on the bacterium's ability to survive and replicate within host cells. The formation and maintenance of a unique membrane-bound compartment, termed the Salmonella-containing vacuole (SCV), is essential for S. Typhimurium pathogenesis. SCV-bound S. Typhimurium induces formation of filamentous tubules that radiate outwards from the SCV, termed Salmonella-induced filaments (SIFs). SIF formation is concomitant with the onset of replication within host epithelial cells. SIF biogenesis, formation and maintenance of the SCV, and the intracellular positioning of the SCV within the host cell requires translocation of bacterial proteins (effectors) into the host cell. Effectors secreted by the type III secretion system encoded on Salmonella pathogenicity island 2 (T3SS2) function to interfere with host cellular processes and promote both intracellular survival and replication of S. Typhimurium. Seven T3SS2-secreted effectors, SifA, SopD2, PipB2, SteA, SseJ, SseF, and SseG have previously been implicated to play complementary, redundant, and/or antagonistic roles with respect to SIF biogenesis, intracellular positioning of the SCV, and SCV membrane dynamics modulation during infection. We undertook a systematic study to delineate the contribution of each effector to these processes by (i) deleting all seven of these effectors in a single S. Typhimurium strain; and (ii) deleting combinations of multiple effectors based on putative effector function. Using this deletion mutant library, we show that each of SIF biogenesis, intracellular SCV localization, intramacrophage replication, colonization, and virulence depends on the activities of multiple effectors. Together, our data demonstrates the complex interplay between these seven effectors and highlights the necessity to study T3SS2-secreted effectors as groups, rather than studies of individual effectors.


Assuntos
Proteínas de Bactérias , Translocação Bacteriana/genética , Ilhas Genômicas , Mucosa Intestinal , Infecções por Salmonella , Salmonella typhimurium , Fatores de Virulência , Animais , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Feminino , Células HeLa , Humanos , Mucosa Intestinal/metabolismo , Mucosa Intestinal/microbiologia , Mucosa Intestinal/patologia , Camundongos , Células RAW 264.7 , Infecções por Salmonella/genética , Infecções por Salmonella/metabolismo , Infecções por Salmonella/patologia , Salmonella typhimurium/genética , Salmonella typhimurium/metabolismo , Salmonella typhimurium/patogenicidade , Células THP-1 , Fatores de Virulência/genética , Fatores de Virulência/metabolismo
15.
Int J Nanomedicine ; 15: 2957-2969, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32425527

RESUMO

Experimental and epidemiological evidence shows that parasites, particularly helminths, play a central role in balancing the host immunity. It was demonstrated that parasites can modulate immune responses via their excretory/secretory (ES) and some specific proteins. Extracellular vesicles (EVs) are nano-scale particles that are released from eukaryotic and prokaryotic cells. EVs in parasitological studies have been mostly employed for immunotherapy of autoimmune diseases, vaccination, and diagnosis. EVs can carry virulence factors and play a central role in the development of parasites in host cells. These molecules can manipulate the immune responses through transcriptional changes. Moreover, EVs derived from helminths modulate the immune system via provoking anti-inflammatory cytokines. On the other hand, EVs from parasite protozoa can induce efficient immunity, that makes them useful for probable next-generation vaccines. In addition, it seems that EVs from parasites may provide new diagnostic approaches for parasitic infections. In the current study, we reviewed isolation methods, functions, and applications of parasite's EVs in immunotherapy, vaccination, and diagnosis.


Assuntos
Vesículas Extracelulares/metabolismo , Interações Hospedeiro-Parasita/imunologia , Imunoterapia/métodos , Parasitos/citologia , Animais , Doenças Autoimunes/terapia , Citocinas/metabolismo , Vesículas Extracelulares/química , Vesículas Extracelulares/classificação , Humanos , Imunidade , Parasitos/patogenicidade , Doenças Parasitárias/diagnóstico , Vacinação , Vacinas/imunologia , Fatores de Virulência/metabolismo
16.
Proc Natl Acad Sci U S A ; 117(22): 12387-12393, 2020 06 02.
Artigo em Inglês | MEDLINE | ID: mdl-32409599

RESUMO

Microbiota, host and dietary metabolites/signals compose the rich gut chemical environment, which profoundly impacts virulence of enteric pathogens. Enterohemorrhagic Escherichia coli (EHEC) engages a syringe-like machinery named type-III secretion system (T3SS) to inject effectors within host cells that lead to intestinal colonization and disease. We previously conducted a high-throughput screen to identify metabolic pathways that affect T3SS expression. Here we show that in the presence of arginine, the arginine sensor ArgR, identified through this screen, directly activates expression of the genes encoding the T3SS. Exogenously added arginine induces EHEC virulence gene expression in vitro. Congruently, a mutant deficient in arginine transport (ΔartP) had decreased virulence gene expression. ArgR also augments murine disease caused by Citrobacter rodentium, which is a murine pathogen extensively employed as a surrogate animal model for EHEC. The source of arginine sensed by C. rodentium is not dietary. At the peak of C. rodentium infection, increased arginine concentration in the colon correlated with down-regulation of the host SLC7A2 transporter. This increase in the concentration of colonic arginine promotes virulence gene expression in C. rodentium Arginine is an important modulator of the host immune response to pathogens. Here we add that arginine also directly impacts bacterial virulence. These findings suggest that a delicate balance between host and pathogen responses to arginine occur during disease progression.


Assuntos
Citrobacter rodentium/metabolismo , Infecções por Enterobacteriaceae/microbiologia , Escherichia coli Êntero-Hemorrágica/metabolismo , Infecções por Escherichia coli/microbiologia , Regulação Bacteriana da Expressão Gênica , Animais , Arginina/metabolismo , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Citrobacter rodentium/genética , Citrobacter rodentium/patogenicidade , Escherichia coli Êntero-Hemorrágica/genética , Escherichia coli Êntero-Hemorrágica/patogenicidade , Humanos , Camundongos , Camundongos Endogâmicos C3H , Virulência , Fatores de Virulência/genética , Fatores de Virulência/metabolismo
17.
Proc Natl Acad Sci U S A ; 117(23): 12763-12771, 2020 06 09.
Artigo em Inglês | MEDLINE | ID: mdl-32461369

RESUMO

Aphids are sap-feeding insects that colonize a broad range of plant species and often cause feeding damage and transmit plant pathogens, including bacteria, viruses, and viroids. These insects feed from the plant vascular tissue, predominantly the phloem. However, it remains largely unknown how aphids, and other sap-feeding insects, establish intimate long-term interactions with plants. To identify aphid virulence factors, we took advantage of the ability of the green peach aphid Myzus persicae to colonize divergent plant species. We found that a M. persicae clone of near-identical females established stable colonies on nine plant species of five representative plant eudicot and monocot families that span the angiosperm phylogeny. Members of the novel aphid gene family Ya are differentially expressed in aphids on the nine plant species and are coregulated and organized as tandem repeats in aphid genomes. Aphids translocate Ya transcripts into plants, and some transcripts migrate to distal leaves within several plant species. RNAi-mediated knockdown of Ya genes reduces M. persicae fecundity, and M. persicae produces more progeny on transgenic plants that heterologously produce one of the systemically migrating Ya transcripts as a long noncoding (lnc) RNA. Taken together, our findings show that beyond a range of pathogens, M. persicae aphids translocate their own transcripts into plants, including a Ya lncRNA that migrates to distal locations within plants, promotes aphid fecundity, and is a member of a previously undescribed host-responsive aphid gene family that operate as virulence factors.


Assuntos
Afídeos/patogenicidade , Magnoliopsida/parasitologia , Transporte de RNA , RNA Longo não Codificante/metabolismo , Fatores de Virulência/metabolismo , Animais , Afídeos/genética , Proteínas de Insetos/genética , RNA Longo não Codificante/genética , Fatores de Virulência/genética
18.
PLoS Pathog ; 16(5): e1008342, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32365117

RESUMO

Chitinases are important enzymes that contribute to the generation of carbon and nitrogen from chitin, a long chain polymer of N-acetylglucosamine that is abundant in insects, fungi, invertebrates and fish. Although mammals do not produce chitin, chitinases have been identified in bacteria that are key virulence factors in severe respiratory, gastrointestinal and urinary diseases. However, it is unclear how these enzymes are able to carry out this dual function. Legionella pneumophila is the causative agent of Legionnaires' disease, an often-fatal pneumonia and its chitinase ChiA is essential for the survival of L. pneumophila in the lung. Here we report the first atomic resolution insight into the pathogenic mechanism of a bacterial chitinase. We derive an experimental model of intact ChiA and show how its N-terminal region targets ChiA to the bacterial surface after its secretion. We provide the first evidence that L. pneumophila can bind mucins on its surface, but this is not dependent on ChiA. This demonstrates that additional peripheral mucin binding proteins are also expressed in L. pneumophila. We also show that the ChiA C-terminal chitinase domain has novel Zn2+-dependent peptidase activity against mammalian mucin-like proteins, namely MUC5AC and the C1-esterase inhibitor, and that ChiA promotes bacterial penetration of mucin gels. Our findings suggest that ChiA can facilitate passage of L. pneumophila through the alveolar mucosa, can modulate the host complement system and that ChiA may be a promising target for vaccine development.


Assuntos
Quitinases/metabolismo , Legionella pneumophila/metabolismo , Acetilglucosamina/metabolismo , Proteínas de Bactérias/metabolismo , Quitina/metabolismo , Quitinases/fisiologia , Regulação Bacteriana da Expressão Gênica/genética , Doença dos Legionários/metabolismo , Metais , Mucina-1/metabolismo , Mucinas/metabolismo , Proteólise , Relação Estrutura-Atividade , Fatores de Virulência/metabolismo
19.
Biofouling ; 36(3): 351-367, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32401555

RESUMO

Pseudomonas aeruginosa and Serratia marcescens are prominent members belonging to the group of ESKAPE pathogens responsible for Urinary Tract Infections (UTI) and nosocomial infections. Both the pathogens regulate several virulence factors, including biofilm formation through quorum sensing (QS), an intercellular communication mechanism. The present study describes the anti-biofilm and QS quenching effect of thiazolinyl-picolinamide based palladium(II) complexes against P. aeruginosa and S. marcescens. Palladium(II) complexes showed quorum sensing inhibitory potential in inhibiting swarming motility behaviour, pyocyanin production and other QS mediated virulence factors in both P. aeruginosa and S. marcescens. In addition, the establishment of biofilms was prevented on palladium (II) coated catheters. Overall, the present study demonstrates that thiazolinyl-picolinamide based palladium (II) complexes will be a promising strategy to combat device-mediated UTI infections.


Assuntos
Antibacterianos/farmacologia , Biofilmes/efeitos dos fármacos , Complexos de Coordenação/farmacologia , Paládio/farmacologia , Ácidos Picolínicos/química , Tiazóis/química , Cateteres Urinários/microbiologia , Antibacterianos/química , Antibacterianos/toxicidade , Biofilmes/crescimento & desenvolvimento , Sobrevivência Celular/efeitos dos fármacos , Complexos de Coordenação/química , Complexos de Coordenação/toxicidade , Infecção Hospitalar/prevenção & controle , Humanos , Células MCF-7 , Paládio/química , Paládio/toxicidade , Pseudomonas aeruginosa/efeitos dos fármacos , Pseudomonas aeruginosa/metabolismo , Piocianina/metabolismo , Percepção de Quorum/efeitos dos fármacos , Serratia marcescens/efeitos dos fármacos , Serratia marcescens/metabolismo , Infecções Urinárias/microbiologia , Infecções Urinárias/prevenção & controle , Virulência , Fatores de Virulência/metabolismo
20.
PLoS Pathog ; 16(4): e1007979, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32298382

RESUMO

Brucella species are facultative intracellular Gram-negative bacteria relevant to animal and human health. Their ability to establish an intracellular niche and subvert host cell pathways to their advantage depends on the delivery of bacterial effector proteins through a type IV secretion system. Brucella Toll/Interleukin-1 Receptor (TIR)-domain-containing proteins BtpA (also known as TcpB) and BtpB are among such effectors. Although divergent in primary sequence, they interfere with Toll-like receptor (TLR) signaling to inhibit the innate immune responses. However, the molecular mechanisms implicated still remain unclear. To gain insight into the functions of BtpA and BtpB, we expressed them in the budding yeast Saccharomyces cerevisiae as a eukaryotic cell model. We found that both effectors were cytotoxic and that their respective TIR domains were necessary and sufficient for yeast growth inhibition. Growth arrest was concomitant with actin depolymerization, endocytic block and a general decrease in kinase activity in the cell, suggesting a failure in energetic metabolism. Indeed, levels of ATP and NAD+ were low in yeast cells expressing BtpA and BtpB TIR domains, consistent with the recently described enzymatic activity of some TIR domains as NAD+ hydrolases. In human epithelial cells, both BtpA and BtpB expression reduced intracellular total NAD levels. In infected cells, both BtpA and BtpB contributed to reduction of total NAD, indicating that their NAD+ hydrolase functions are active intracellularly during infection. Overall, combining the yeast model together with mammalian cells and infection studies our results show that BtpA and BtpB modulate energy metabolism in host cells through NAD+ hydrolysis, assigning a novel role for these TIR domain-containing effectors in Brucella pathogenesis.


Assuntos
Proteínas de Bactérias/metabolismo , Brucella abortus/crescimento & desenvolvimento , Brucelose/metabolismo , Hidrolases/metabolismo , NAD/metabolismo , Saccharomyces cerevisiae/crescimento & desenvolvimento , Fatores de Virulência/metabolismo , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Brucella abortus/metabolismo , Brucelose/microbiologia , Células HeLa , Humanos , Conformação Proteica , Domínios e Motivos de Interação entre Proteínas , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Fatores de Virulência/genética
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