RESUMEN

Gallibacterium anatis is a member of the Pasteurellaceae family and is an opportunistic pathogen that causes gallibacteriosis in chickens. Stress plays a relevant role in promoting the development of pathogenicity in G. anatis. Epinephrine (E) and norepinephrine (NE) are relevant to stress; however, their effects on G. anatis have not been elucidated. In this work, we evaluated the effects of E and NE on the growth, biofilm formation, expression of adhesins, and proteases of two G. anatis strains, namely, the hemolytic 12656-12 and the nonhemolytic F149T biovars. E (10 µM/mL) and NE (30 and 50 µM/mL) increased the growth of G. anatis 12656-12 by 20 % and 25 %, respectively. E did not affect the growth of F149T, whereas 40 µM/mL NE decreased bacterial growth by 25 %. E and NE at a dose of 30-50 µM/mL upregulated five fibrinogen adhesins in the 12565-12 strain, whereas no effect was observed in the F149T strain. NE increased proteolytic activity in both strains, whereas E diminished proteolytic activity in the 12656-12 strain. E and NE reduced biofilm formation (30 %) and increased Congo red binding (15 %) in both strains. QseBC is the E and NE two-component detection system most common in bacteria. The qseC gene, which is the E and NE receptor in bacteria, was identified in the genomic DNA of the 12565-12 and F149TG. anatis strains via PCR amplification. Our results suggest that QseC can detect host changes in E and NE concentrations and that catecholamines can modulate the expression of several virulence factors in G. anatis.

Asunto(s)

Biopelículas , Pollos , Epinefrina , Regulación Bacteriana de la Expresión Génica , Norepinefrina , Pasteurellaceae , Factores de Virulencia , Factores de Virulencia/genética , Factores de Virulencia/metabolismo , Norepinefrina/farmacología , Norepinefrina/metabolismo , Epinefrina/farmacología , Biopelículas/crecimiento & desarrollo , Biopelículas/efectos de los fármacos , Pasteurellaceae/genética , Pasteurellaceae/patogenicidad , Pasteurellaceae/efectos de los fármacos , Pasteurellaceae/metabolismo , Animales , Regulación Bacteriana de la Expresión Génica/efectos de los fármacos , Adhesinas Bacterianas/genética , Adhesinas Bacterianas/metabolismo , Péptido Hidrolasas/metabolismo , Péptido Hidrolasas/genética , Enfermedades de las Aves de Corral/microbiología , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Infecciones por Pasteurellaceae/microbiología , Infecciones por Pasteurellaceae/veterinaria

RESUMEN

BACKGROUND: Auranofin is an approved anti-rheumatic drug that has a broad-range inhibitory action against several microorganisms, including human pathogenic fungi. The auranofin activity against Histoplasma capsulatum, the dimorphic fungus that causes histoplasmosis, has not been properly addressed. Since there are few therapeutic options for this life-threatening systemic mycosis, this study evaluated the effects of auranofin on H. capsulatum growth and expression of virulence factors. METHODOLOGY/PRINCIPAL FINDINGS: Minimal inhibitory and fungicidal concentrations (MIC and MFC, respectively) of auranofin against 15 H. capsulatum strains with distinct genetic backgrounds were determined using the yeast form of the fungus and a microdilution protocol. Auranofin activity was also assessed on a macrophage model of infection and on a Tenebrio molitor invertebrate animal model. Expression of virulence-related genes was compared between auranofin treated and untreated H. capsulatum yeast cells using a quantitative PCR assay. Auranofin affected the growth of different strains of H. capsulatum, with MIC and MFC values ranging from 1.25 to 5.0 µM and from 2.5 to >10 µM, respectively. Auranofin was able to kill intracellular H. capsulatum yeast cells and conferred protection against the fungus in the experimental animal model of infection. Moreover, the expression of catalase A, HSP70, superoxide dismutase, thioredoxin reductase, serine proteinase, cytochrome C peroxidase, histone 2B, formamidase, metallopeptidase, Y20 and YPS3 proteins were reduced after six hours of auranofin treatment. CONCLUSIONS/SIGNIFICANCE: Auranofin is fungicidal against H. capsulatum and reduces the expression of several virulence-related genes, which makes this anti-rheumatic drug a good candidate for new medicines against histoplasmosis.

Asunto(s)

Antifúngicos , Auranofina , Histoplasma , Pruebas de Sensibilidad Microbiana , Histoplasma/efectos de los fármacos , Histoplasma/genética , Histoplasma/patogenicidad , Auranofina/farmacología , Animales , Antifúngicos/farmacología , Factores de Virulencia/genética , Histoplasmosis/microbiología , Histoplasmosis/tratamiento farmacológico , Macrófagos/microbiología , Macrófagos/efectos de los fármacos , Ratones , Tenebrio/microbiología , Virulencia/efectos de los fármacos , Modelos Animales de Enfermedad , Humanos

RESUMEN

In humans and Drosophila melanogaster, the functional convergence of the endosomal sorting complex required for transport (ESCRT) machinery that is in charge of selecting ubiquitinated proteins for sorting into multivesicular bodies, and the retromer, that is the complex responsible for protein recycling to the plasma membrane and Golgi apparatus. ESCRT and retromer complexes are codependent for protein sorting recycling, degradation, and secretion. In this article, we studied the EhVps35 C isoform (referred to as EhVps35), that is the central member of the Entamoeba histolytica retromer, and its relation with the ESCRT machinery during sorting and protein recycling events and their involvement virulence. Our findings revealed that EhVps35 interacts with at least 300 proteins that participate in multiple cellular processes. Laser confocal and transmission electronic microscopy images, as well as secretion assays, revealed that EhVps35 is secreted in vesicles together with EhVps23 and EhADH (both ESCRT machinery proteins). In addition, immunoprecipitation, immunofluorescence, and molecular docking assays revealed the relationship among EhVps35 and other ESCRT machinery proteins. Red blood cell stimulus increased EhVps35 secretion, and the knockdown of the Ehvps35 gene in trophozoites reduced their capacity to migrate and invade tissues. This also impacts the cellular localization of ubiquitin, EhVps23 (ESCRT-I), and EhVps32 (ESCRT-III) proteins, strongly suggesting their functional relationship. Our results, taken together, give evidence that EhVps35 is a key factor in E. histolytica virulence mechanisms and that it, together with the ESCRT machinery components and other regulatory proteins, is involved in vesicle trafficking, secretion, migration, and cell proliferation.

Asunto(s)

Complejos de Clasificación Endosomal Requeridos para el Transporte , Entamoeba histolytica , Transporte de Proteínas , Proteínas Protozoarias , Entamoeba histolytica/metabolismo , Entamoeba histolytica/patogenicidad , Entamoeba histolytica/genética , Complejos de Clasificación Endosomal Requeridos para el Transporte/metabolismo , Complejos de Clasificación Endosomal Requeridos para el Transporte/genética , Animales , Proteínas Protozoarias/metabolismo , Proteínas Protozoarias/genética , Humanos , Virulencia , Simulación del Acoplamiento Molecular , Eritrocitos/parasitología , Eritrocitos/metabolismo , Factores de Virulencia/metabolismo , Entamebiasis/parasitología

RESUMEN

The aim of this study was to identify, using proteomics, the molecular alterations caused by human serum exposure to Klebsiella pneumoniae ACH2. The analysis was performed under two different conditions, native serum from healthy donors and heat-inactivated serum (to inactivate the complement system), and at two different times, after 1 and 4 h of serum exposure. More than 1,000 bacterial proteins were identified at each time point. Enterobactin, a siderophore involved in iron uptake, and proteins involved in translation were upregulated at 1 h, while the chaperone ProQ and the glyoxylate cycle were identified after 4 h. Enzymes involved in the stress response were downregulated, and the SOD activity was validated using an enzymatic assay. In addition, an intricate metabolic adaptation was observed, with pyruvate and thiamine possibly involved in survival and virulence in the first hour of serum exposure. The addition of exogenous thiamine contributes to bacterial growth in human serum, corroborating this result. During 4 h of serum exposure, the glyoxylate cycle (GC) probably plays a central role, and the addition of exogenous succinate suppresses the GC, inducing a decrease in serum resistance. Therefore, serum exposure causes important changes in iron acquisition, the expression of virulence factors, and metabolic reprogramming, which could contribute to bacterial serum resistance.

Asunto(s)

Proteínas Bacterianas , Klebsiella pneumoniae , Humanos , Klebsiella pneumoniae/patogenicidad , Proteínas Bacterianas/metabolismo , Proteínas Bacterianas/genética , Evasión Inmune , Suero/metabolismo , Proteómica/métodos , Factores de Virulencia/metabolismo , Hierro/metabolismo , Tiamina/farmacología , Tiamina/metabolismo , Interacciones Huésped-Patógeno , Infecciones por Klebsiella/microbiología , Infecciones por Klebsiella/inmunología , Glioxilatos/metabolismo , Reprogramación Metabólica

RESUMEN

BACKGROUND: Klebsiella pneumoniae is the major cause of nosocomial infections worldwide and is related to a worsening increase in Multidrug-Resistant Bacteria (MDR) and virulence genes that seriously affect immunosuppressed patients, long-stay intensive care patients, elderly individuals, and children. Whole-Genome Sequencing (WGS) has resulted in a useful strategy for characterizing the genomic components of clinically important bacteria, such as K. pneumoniae, enabling them to monitor genetic changes and understand transmission, highlighting the risk of dissemination of resistance and virulence associated genes in hospitals. In this study, we report on WGS 14 clinical isolates of K. pneumoniae from a pediatric hospital biobank of Guayaquil, Ecuador. RESULTS: The main findings revealed pronounced genetic heterogeneity among the isolates. Multilocus sequencing type ST45 was the predominant lineage among non-KPC isolates, whereas ST629 was found more frequently among KPC isolates. Phylogenetic analysis suggested local transmission dynamics. Comparative genomic analysis revealed a core set of 3511 conserved genes and an open pangenome in neonatal isolates. The diversity of MLSTs and capsular types, and the high genetic diversity among these isolates indicate high intraspecific variability. In terms of virulence factors, we identified genes associated with adherence, biofilm formation, immune evasion, secretion systems, multidrug efflux pump transporters, and a notably high number of genes related to iron uptake. A large number of these genes were detected in the ST45 isolate, whereas iron uptake yersiniabactin genes were found exclusively in the non-KPC isolates. We observed high resistance to commonly used antibiotics and determined that these isolates exhibited multidrug resistance including ß-lactams, aminoglycosides, fluoroquinolones, quinolones, trimetropins, fosfomycin and macrolides; additionally, resistance-associated point mutations and cross-resistance genes were identified in all the isolates. We also report the first K. pneumoniae KPC-3 gene producers in Ecuador. CONCLUSIONS: Our WGS results for clinical isolates highlight the importance of MDR in neonatal K. pneumoniae infections and their genetic diversity. WGS will be an imperative strategy for the surveillance of K. pneumoniae in Ecuador, and will contribute to identifying effective treatment strategies for K. pneumoniae infections in critical units in patients at stratified risk.

Asunto(s)

Farmacorresistencia Bacteriana Múltiple , Genoma Bacteriano , Hospitales Pediátricos , Klebsiella pneumoniae , Filogenia , Secuenciación Completa del Genoma , Humanos , Ecuador , Klebsiella pneumoniae/genética , Klebsiella pneumoniae/aislamiento & purificación , Klebsiella pneumoniae/efectos de los fármacos , Farmacorresistencia Bacteriana Múltiple/genética , Niño , Infecciones por Klebsiella/microbiología , Infecciones por Klebsiella/epidemiología , Factores de Virulencia/genética , Tipificación de Secuencias Multilocus , Preescolar , Lactante , Variación Genética

RESUMEN

Introduction. Staphylococcus aureus is a leading agent in community-acquired bacteraemia (CAB) and has been linked to elevated mortality rates and methicillin resistance in Costa Rica.Gap statement and aim. To update and enhance previous data obtained in this country, we analysed the clinical manifestations of 54 S. aureus CAB cases in a tertiary hospital and delineated the sequence types (STs), virulome, and resistome of the implicated isolates.Methodology. Clinical information was retrieved from patient files. Antibiotic susceptibility profiles were obtained with disc diffusion and automated phenotypic tests. Genomic data were exploited to type the isolates and for detection of resistance and virulence genes.Results. Primary infections predominantly manifested as bone and joint infections, followed by skin and soft tissue infections. Alarmingly, 70% of patients continued to exhibit positive haemocultures beyond 48 h of treatment modification, with nearly a quarter requiring mechanical ventilation or developing septic shock. The 30-day mortality rate reached an alarming 40%. More than 60% of the patients were found to have received suboptimal or inappropriate antibiotic treatment, and there was an alarming tendency towards the overuse of third-generation cephalosporins as empirical treatment. Laboratory tests indicated elevated creatinine levels, leukocytosis, and bandaemia within the first 24 h of hospitalization. However, most showed improvement after 48 h. The isolates were categorized into 13 STs, with a predominance of representatives from the clonal complexes CC72 (ST72), CC8 (ST8), CC5 (ST5, ST6), and CC1 (ST188). Twenty-four isolates tested positive for mecA, with ST72 strains accounting for 20. In addition, we detected genes conferring acquired resistance to aminoglycosides, MLSB antibiotics, trimethoprim/sulfamethoxazole, and mutations for fluoroquinolone resistance in the isolate collection. Genes associated with biofilm formation, capsule synthesis, and exotoxin production were prevalent, in contrast to the infrequent detection of enterotoxins or exfoliative toxin genes.Conclusions. Our findings broaden our understanding of S. aureus infections in a largely understudied region and can enhance patient management and treatment strategies.

Asunto(s)

Antibacterianos , Bacteriemia , Infecciones Comunitarias Adquiridas , Pruebas de Sensibilidad Microbiana , Infecciones Estafilocócicas , Staphylococcus aureus , Centros de Atención Terciaria , Humanos , Costa Rica/epidemiología , Centros de Atención Terciaria/estadística & datos numéricos , Infecciones Comunitarias Adquiridas/microbiología , Infecciones Comunitarias Adquiridas/epidemiología , Infecciones Comunitarias Adquiridas/mortalidad , Infecciones Estafilocócicas/microbiología , Infecciones Estafilocócicas/epidemiología , Infecciones Estafilocócicas/tratamiento farmacológico , Infecciones Estafilocócicas/mortalidad , Bacteriemia/microbiología , Bacteriemia/epidemiología , Bacteriemia/mortalidad , Bacteriemia/tratamiento farmacológico , Masculino , Staphylococcus aureus/genética , Staphylococcus aureus/efectos de los fármacos , Staphylococcus aureus/aislamiento & purificación , Persona de Mediana Edad , Femenino , Anciano , Adulto , Antibacterianos/uso terapéutico , Antibacterianos/farmacología , Anciano de 80 o más Años , Adulto Joven , Adolescente , Factores de Virulencia/genética , Niño

RESUMEN

INTRODUCTION: Candida albicans is the most common opportunistic pathogen causing fungal infections worldwide, especially in high-risk patients. Its pathogenicity is related to virulence factors gene expression, such as hyphal growth (HWP1), cell adhesion (ALS3), and protease secretion (SAP1) during infection spreading mechanisms. In recent years, an increase in non-albicans Candida infections has been reported, which may present coinfection or competitive interactions with C. albicans, potentially aggravating the patient's condition. This study aims to evaluate the expression of genes related to virulence factors of C. albicans and non-albicans Candida during planktonic stage. METHODS: C. albicans (ATCC MYA-3573) as well as with three clinical strains (C. albicans DCA53, C. tropicalis DCT6, and C. parapsilosis DCP1) isolated from blood samples, were grown in 24-well plates at 37°C for 20 h, either in monocultures or mixed cultures. Quantitative real-time polymerase chain reaction was used to evaluate the expression levels of the genes HWP1, ALS3, and SAP1 in cells collected during the planktonic stage. In addition, hyphal filamentation was observed using a Scanning Electron Microscope. RESULTS: The overexpression of HWP1 and ASL3 genes in mixed growth conditions between C. albicans and non-albicans Candida species suggests a synergistic relationship as well as an increased capacity for hyphal growth and adhesion. In contrast, C. parapsilosis versus C. tropicalis interaction shows an antagonistic relationship during mixed culture, suggesting a decreased virulence profile of C. parapsilosis during initial coinfection with C. tropicalis. CONCLUSION: The expression of HWP1, ALS3, and SAP1 genes associated with virulence factors varies under competitive conditions among species of the genus Candida during planktonic stage.

Asunto(s)

Candida albicans , Proteínas Fúngicas , Factores de Virulencia , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Humanos , Factores de Virulencia/genética , Candida albicans/patogenicidad , Candida albicans/genética , Virulencia/genética , Hifa/genética , Regulación Fúngica de la Expresión Génica , Candidiasis/microbiología , Ácido Aspártico Endopeptidasas/genética , Ácido Aspártico Endopeptidasas/metabolismo , Plancton/genética , Candida/patogenicidad , Candida/genética , Glicoproteínas de Membrana

RESUMEN

The formamidase (FMD) enzyme plays an important role in fungal thriving by releasing a secondary nitrogen source as a product of its activity. In Paracoccidioides species, previous studies have demonstrated the upregulation of this enzyme in a wide range of starvation and infective-like conditions. However, Paracoccidioides lutzii formamidase has not yet been defined as a virulence factor. Here, by employing in vivo infections using an fmd-silenced strain in Galleria mellonella larvae model, we demonstrate the influence of formamidase in P. lutzii's immune stimulation and pathogenicity. The formamidase silencing resulted in improper arrangement of the nodules, poor melanogenesis and decreased fungal burden. Thus, we suggest that formamidase may be a piece composing the process of molecular recognition by Galleria immune cells. Furthermore, formamidase silencing doubled the observed survival rate of the larvae, demonstrating its importance in fungal virulence in vivo. Therefore, our findings indicate that formamidase contributes to Galleria's immune incitement and establishes the role of this enzyme as a P. lutzii virulence factor.

Asunto(s)

Larva , Mariposas Nocturnas , Paracoccidioides , Factores de Virulencia , Animales , Factores de Virulencia/genética , Factores de Virulencia/metabolismo , Larva/microbiología , Mariposas Nocturnas/microbiología , Paracoccidioides/patogenicidad , Paracoccidioides/enzimología , Paracoccidioides/genética , Virulencia , Paracoccidioidomicosis/microbiología , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Modelos Animales de Enfermedad , Silenciador del Gen

RESUMEN

Escherichia coli is a frequent pathogen isolated from bloodstream infections. This study aimed to characterize the genetic features of EC092, an E. coli strain isolated from bacteremia that harbors enteroaggregative E. coli (EAEC) genetic markers, indicating its hybrid pathogenic potential. Whole-genome sequencing showed that EC092 belongs to phylogroup B1, ST278, and serotype O165:H4. Genes encoding virulence factors such as fimbriae, toxins, iron-uptake systems, autotransporter proteins (Pet, Pic, Sat, and SepA), and secretion systems were detected, as well as EAEC virulence genes (aggR, aatA, aaiC, and aap). EC092 was found to be closely related to the other EAEC prototype strains and highly similar in terms of virulence to three EAEC strains isolated from diarrhea. The genomic neighborhood of pet, pic, sat, sepA, and the EAEC virulence genes of EC092 and its three genetically related fecal EAEC strains showed an identical genomic organization and nucleotide sequences. Also, EC092 produced and secreted Pet, Pic, Sat, and SepA in the culture supernatant and resisted the bactericidal activity of normal human serum. Our results demonstrate that the strain EC092, isolated from bacteremia, is a hybrid pathogenic extraintestinal E. coli (ExPEC)/EAEC with virulence features that could mediate both extraintestinal and intestinal infections.

Asunto(s)

Bacteriemia , Infecciones por Escherichia coli , Escherichia coli , Genoma Bacteriano , Factores de Virulencia , Humanos , Bacteriemia/microbiología , Escherichia coli/genética , Escherichia coli/patogenicidad , Factores de Virulencia/genética , Infecciones por Escherichia coli/microbiología , Secuenciación Completa del Genoma , Virulencia/genética , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Filogenia , Genómica/métodos

RESUMEN

Cases of diphtheria, even in immunized individuals, are still reported in several parts of the world, including in Brazil. New outbreaks occur in Europe and other continents. In this context, studies on Corynebacterium diphtheriae infections are highly relevant, both for a better understanding of the pathogenesis of the disease and for controlling the circulation of clones and antimicrobial resistance genes. Here we present a case of cutaneous infection by multidrug-resistant Corynebacterium diphtheriae and provide its whole-genome sequencing. Genomic analysis revealed resistance genes, including tet(W), sul1, cmx, rpoB2, rbpA and mutation in rpoB. We performed phylogenetic analyzes and used the BRIG to compare the predicted resistance genes with those found in genomes from other significant isolates, including those associated with some outbreaks. Virulence factors such as spaD, srtBC, spaH, srtDE, surface-anchored pilus proteins (sapD), nonfimbrial adhesins (DIP0733, DIP1281, and DIP1621), embC and mptC (putatively involved in CdiLAM), sigA, dtxR and MdbA (putatively involved) in post-translational modification, were detected. We identified the CRISPR-Cas system in our isolate, which was classified as Type II-U based on the database and contains 15 spacers. This system functions as an adaptive immune mechanism. The strain was attributed to a new sequence type ST-928, and phylogenetic analysis confirmed that it was related to ST-634 of C. diphtheriae strains isolated in French Guiana and Brazil. In addition, since infections are not always reported, studies with the sequence data might be a way to complement and inform C. diphtheriae surveillance.

Asunto(s)

Sistemas CRISPR-Cas , Corynebacterium diphtheriae , Rifampin , Factores de Virulencia , Corynebacterium diphtheriae/genética , Corynebacterium diphtheriae/patogenicidad , Corynebacterium diphtheriae/efectos de los fármacos , Humanos , Factores de Virulencia/genética , Rifampin/farmacología , Mutación , Filogenia , Difteria/microbiología , Genoma Bacteriano , Proteínas Bacterianas/genética , Farmacorresistencia Bacteriana Múltiple/genética

RESUMEN

Atypical enteropathogenic Escherichia coli (aEPEC) is a significant cause of diarrhea in low- and middle-income countries. Certain aEPEC strains, including the Brazilian representative strain of serotype O51:H40 called aEPEC 1711-4, can use flagella to attach to, invade, and persist in T84 and Caco-2 intestinal cells. It can also translocate from the gut to extraintestinal sites in a rat model. Although various aspects of the virulence of this strain were studied and the requirement of a type III secretion system for the efficiency of the invasion process was demonstrated, the expression of the locus of enterocyte effacement (LEE) genes during the invasion and intracellular persistence remains unclear. To address this question, the expression of flagella and the different LEE operons was evaluated during kinetic experiments of the interaction of aEPEC 1711-4 with enterocytes in vitro. The genome of the strain was also sequenced. The results showed that flagella expression remained unchanged, but the expression of eae and escJ increased during the early interaction and invasion of aEPEC 1711-4 into Caco-2 cells, and there was no change 24 h post-infection during the persistence period. The number of actin accumulation foci formed on HeLa cells also increased during the 6-h analysis. No known gene related to the invasion process was identified in the genome of aEPEC 1711-4, which was shown to belong to the global EPEC lineage 10. These findings suggest that the LEE components and the intimate adherence promoted by intimin are necessary for the invasion and persistence of aEPEC 1711-4, but the detailed mechanism needs further study.IMPORTANCEAtypical enteropathogenic Escherichia coli (aEPEC) is a major cause of diarrhea, especially in low- and middle-income countries, like Brazil. However, due to the genome heterogeneity of each clonal group, it is difficult to comprehend the pathogenicity of this strain fully. Among aEPEC strains, 1711-4 can invade eukaryotic cells in vitro, cross the gut barrier, and reach extraintestinal sites in animal models. By studying how different known aEPEC virulence factors are expressed during the invasion process, we can gain insight into the commonalities of this phenotype among other aEPEC strains. This will help in developing preventive measures to control infections caused by invasive strains. No known virulence-encoding genes linked to the invasion process were found. Nevertheless, additional studies are still necessary to evaluate the role of other factors in this phenotype.

Asunto(s)

Enterocitos , Escherichia coli Enteropatógena , Infecciones por Escherichia coli , Proteínas de Escherichia coli , Flagelos , Serogrupo , Escherichia coli Enteropatógena/genética , Escherichia coli Enteropatógena/patogenicidad , Escherichia coli Enteropatógena/metabolismo , Humanos , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Enterocitos/microbiología , Células CACO-2 , Infecciones por Escherichia coli/microbiología , Flagelos/genética , Flagelos/metabolismo , Virulencia/genética , Fosfoproteínas/genética , Fosfoproteínas/metabolismo , Regulación Bacteriana de la Expresión Génica , Adhesión Bacteriana/genética , Animales , Brasil , Factores de Virulencia/genética , Factores de Virulencia/metabolismo , Operón/genética , Ratas

RESUMEN

The current study aimed to detect virulence, hetero-pathogenicity, and hybridization genes in Escherichia coli strains, previously isolated from cloacal swabs in commercial breeding psittacines and zoological collections, via multiplex PCR. A total of 68 strains of E. coli, previously isolated from psittacines in zoos and commercial breeding facilities in Ceará, Brazil, were assessed for the presence of the following genes and/or probes: eae, bfpA (EPEC - Enteropathogenic E. coli), CVD432 (EAEC - Enteroaggregative E. coli); LT gene and ST gene (ETEC - Enterotoxigenic E. coli); ipaH (EIEC - Enteroinvasive E. coli); stx1 and stx2 (STEC - Shiga toxin-producing E. coli); iroN, ompT, hlyF, iss, and iutA (APEC - Avian pathogenic E. coli). Of the 68 E. coli strains analyzed, 61 (98.7 %) were positive for the following genes and/or probes: Stx1 (61/98.7 %), ST gene (54/79.4 %), CVD432 (49/72 %), bfpA (44/64.7 %), eae (42/61.8 %), Stx2 (41/60.3 %), ipaH (34/50 %), LT gene (33/48.5 %), iroN (21/30.9 %), hlyF (11/6.2 %), iss (06/8.8 %) and iutA (06/8.8 %). The following diarrheagenic pathotypes were identified: 66 (97 %) from STEC, 49 (72 %) from EAEC, 35 (52 %) from EIEC, 25 (37 %) from ETEC, and one (1.5 %) from EPEC. Regarding hetero-pathogenicity, 50 (74 %) heterogeneous strains were identified. Positivity for APEC was seen in four (6 %) strains, all characterized as pathogenic hybrids. This study describes significant associations of virulence factors in E. coli strains DEC/DEC and DEC/APEC, which were isolated from psittacines and may be potentially harmful to One Health.

Asunto(s)

Animales de Zoológico , Enfermedades de las Aves , Infecciones por Escherichia coli , Proteínas de Escherichia coli , Escherichia coli , Factores de Virulencia , Animales , Brasil , Infecciones por Escherichia coli/microbiología , Infecciones por Escherichia coli/veterinaria , Animales de Zoológico/microbiología , Enfermedades de las Aves/microbiología , Escherichia coli/genética , Escherichia coli/patogenicidad , Escherichia coli/aislamiento & purificación , Escherichia coli/clasificación , Proteínas de Escherichia coli/genética , Factores de Virulencia/genética , Virulencia/genética , Escherichia coli Enteropatógena/genética , Escherichia coli Enteropatógena/patogenicidad , Escherichia coli Enteropatógena/aislamiento & purificación , Escherichia coli Enteropatógena/clasificación , Reacción en Cadena de la Polimerasa Multiplex , Psittaciformes/microbiología , Cloaca/microbiología , Escherichia coli Shiga-Toxigénica/genética , Escherichia coli Shiga-Toxigénica/aislamiento & purificación , Escherichia coli Shiga-Toxigénica/patogenicidad , Escherichia coli Shiga-Toxigénica/clasificación , Escherichia coli Enterotoxigénica/genética , Escherichia coli Enterotoxigénica/patogenicidad , Escherichia coli Enterotoxigénica/aislamiento & purificación , Escherichia coli Enterotoxigénica/clasificación

RESUMEN

INTRODUCTION AND OBJECTIVES: Recent studies have suggested an association between H. pylori and metabolic dysfunction associated steatotic liver disease (MASLD). We aim to evaluate the association of H. pylori virulence genes with non-invasive markers of liver injury and fibrosis in MASLD subjects. PATIENTS AND METHODS: A total of 362 dyspeptic patients who underwent gastroscopy were selected. Biochemical, clinical parameters, ultrasound, FIB-4 score, liver stiffness measurement (LSM) by vibration-controlled transient elastography (VCTE), gastric biopsies, and H. pylori virulence genes (cagA, vacA) were evaluated. RESULTS: A cohort comprised of 61 % women and 39 % men with a median age of 52 (40-60) years. MASLD was observed in 42 %, and H. pylori-positive in 45 %. No differences were observed regarding H. pylori status at co-morbid metabolic conditions. In MASLD cohort, H. pylori-positive was associated with higher AST, ALT, FIB-4 and LSM. Indeed, carriers of cagA/vacA-s1/m1-positive allelic combination were associated with higher AST, ALT, FIB-4 and LSM but not cagA/vacA-s1/m1-negative. The OR for high-risk of significant/advanced- fibrosis by VCTE (≥8 kPa) with H. pylori-positive was 2.56 (95 % CI, 1.2-5.75) and for cagA/vacA-s1/-m1-positive allelic carriers was 4.01 (95 % CI, 1.38-11.56), but non-significant association in cagA/vacA-s1/-m1-negative. After adjusting for age, gender, diabetes, BMI and hypertension the OR for VCTE ≥8 kPa with H. pylori-positive was 2.43 (95 % CI, 1.88-12.44), and cagA/vacA-s1/m1-positive allelic carriers was 4.06 (95 % CI, 1.22-14.49). CONCLUSIONS: In our cohort of functional dyspepsia (FD) patients with MASLD, H. pylori was associated with non-invasive markers of liver injury and fibrosis. Carriers of cagA/vacA-s1/m1-positive allelic combination showed an independent risk of significant/advanced fibrosis by VCTE.

Asunto(s)

Antígenos Bacterianos , Proteínas Bacterianas , Diagnóstico por Imagen de Elasticidad , Infecciones por Helicobacter , Helicobacter pylori , Cirrosis Hepática , Humanos , Masculino , Femenino , Persona de Mediana Edad , Antígenos Bacterianos/genética , Proteínas Bacterianas/genética , Adulto , Cirrosis Hepática/microbiología , Infecciones por Helicobacter/complicaciones , Infecciones por Helicobacter/microbiología , Factores de Riesgo , Gastroscopía , Enfermedad del Hígado Graso no Alcohólico/microbiología , Enfermedad del Hígado Graso no Alcohólico/complicaciones , Biopsia , Virulencia , Factores de Virulencia/genética

RESUMEN

The collective involvement of virulence markers of Escherichia coli as an emerging pathogen associated with periodontitis remains unexplained. This study aimed to implement an in vitro model of infection using a human epithelial cell line to determine the virulome expression related to the antibiotic and disinfectant resistance genotype and pulse field gel electrophoresis (PFGE) type in E. coli strains isolated from patients with periodontal diseases. We studied 100 strains of E. coli isolated from patients with gingivitis (n = 12), moderate periodontitis (n = 59), and chronic periodontitis (n = 29). The identification of E. coli and antibiotic and disinfectant resistance genes was performed through PCR. To promote the expression of virulence genes in the strains, an in vitro infection model was used in the human epithelial cell line A549. RNA was extracted using the QIAcube robotic equipment and reverse transcription to cDNA was performed using the QuantiTect reverse transcription kit (Qiagen). The determination of virulence gene expression was performed through real-time PCR. Overall, the most frequently expressed adhesion genes among the isolated strains of gingivitis, moderate periodontitis, and chronic periodontitis were fimH (48%), iha (37%), and papA (18%); those for toxins were usp (33%); those for iron acquisition were feoB (84%), fyuA (62%), irp-2 (61%), and iroN (35%); those for protectins were traT (50%), KpsMT (35%), and ompT (28%); and those for pathogenicity islands were malX (45%). The most common antibiotic and disinfectant resistance genes among gingivitis, moderate periodontitis, and chronic periodontitis strains were sul-2 (43%), blaSHV (47%), blaTEM (45%), tet(A) (41%), dfrA1 (32%), marR-marO (57%), and qacEA1 (79%). The findings revealed the existence of a wide distribution of virulome expression profiles related to the antibiotic and disinfectant resistance genotype and PFGE type in periodontal strains of E. coli. These findings may contribute toward improving the prevention and treatment measures for periodontal diseases associated with E. coli.

Asunto(s)

Antibacterianos , Desinfectantes , Farmacorresistencia Bacteriana , Infecciones por Escherichia coli , Escherichia coli , Factores de Virulencia , Humanos , Escherichia coli/genética , Escherichia coli/efectos de los fármacos , Escherichia coli/patogenicidad , Factores de Virulencia/genética , Antibacterianos/farmacología , Infecciones por Escherichia coli/microbiología , Farmacorresistencia Bacteriana/genética , Desinfectantes/farmacología , Periodontitis/microbiología , Virulencia/genética , Células A549 , Células Epiteliales/microbiología , Genotipo , Adulto , Femenino , Masculino , Persona de Mediana Edad , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo , Electroforesis en Gel de Campo Pulsado

RESUMEN

AIMS: Characterize global genomic features of 86 genomes of Salmonella Gallinarum (SG) and Pullorum (SP), which are important pathogens causing systemic infections in poultry. METHODS AND RESULTS: All genomes harbored efflux pump encoding gene mdsA and gold tolerance genes golS and golT. Aminoglycoside (aac(6')-Ib, aadA5, aph(6)-Id, aph(3'')-Ib, ant(2'')-Ia), beta-lactam (blaTEM-1, blaTEM-135), efflux pump (mdsB), fosfomycin (fosA3), sulfonamide (sul1, sul2), tetracycline [tet(A)], trimethoprim (dfrA17), acid (asr), and disinfectant (qacEdelta1) resistance genes, gyrA, gyrB, and parC quinolone resistance point mutations, and mercury tolerance genes (mer) were found in different frequencies. Additionally, 310 virulence genes, pathogenicity islands (including SPI-1, 2, 3, 4, 5, 6, 9, 10, 12, 13, and 14), plasmids [IncFII(S), ColpVC, IncX1, IncN, IncX2, and IncC], and prophages (Fels-2, ST104, 500465-1, pro483, Gifsy-2, 103 203_sal5, Fels-1, RE-2010, vB_SenS-Ent2, and L-413C) were detected. MLST showed biovar-specific sequence types, and core genome MLST showed country-specific and global-related clusters. CONCLUSION: SG and SP global strains carry many virulence factors and important antimicrobial resistance genes. The diverse plasmids and prophages suggest genetic variability. MLST and cgMLST differentiated biovars and showed profiles occurring locally or worldwide.

Asunto(s)

Genoma Bacteriano , Enfermedades de las Aves de Corral , Salmonella enterica , Serogrupo , Salmonella enterica/genética , Salmonella enterica/efectos de los fármacos , Animales , Enfermedades de las Aves de Corral/microbiología , Antibacterianos/farmacología , Islas Genómicas/genética , Salmonelosis Animal/microbiología , Farmacorresistencia Bacteriana Múltiple/genética , Factores de Virulencia/genética , Plásmidos/genética , Pollos/microbiología , Genómica , Pruebas de Sensibilidad Microbiana , Farmacorresistencia Bacteriana/genética

RESUMEN

Respiratory diseases in ruminants are responsible for enormous economic losses for the dairy and meat industry. The main causative bacterial agent of pneumonia in ovine is Mannheimia haemolytica A2. Due to the impact of this disease, the effect of the antimicrobial protein, bovine lactoferrin (bLf), against virulence factors of this bacterium has been studied. However, its effect on biofilm formation has not been reported. In this work, we evaluated the effect on different stages of the biofilm. Our results reveal a decrease in biofilm formation when bacteria were pre-incubated with bLf. However, when bLf was added at the start of biofilm formation and on mature biofilm, an increase was observed, which was visualized by greater bacterial aggregation and secretion of biofilm matrix components. Additionally, through SDS-PAGE, a remarkable band of ~80 kDa was observed when bLf was added to biofilms. Therefore, the presence of bLf on the biofilm was determined through the Western blot and Microscopy techniques. Finally, by using Live/Dead staining, we observed that most of the bacteria in a biofilm with bLf were not viable. In addition, bLf affects the formation of a new biofilm cycle. In conclusion, bLf binds to the biofilm of M. haemolytica A2 and affects the viability of bacteria and the formation a new biofilm cycle.

Asunto(s)

Biopelículas , Lactoferrina , Mannheimia haemolytica , Biopelículas/efectos de los fármacos , Biopelículas/crecimiento & desarrollo , Mannheimia haemolytica/efectos de los fármacos , Mannheimia haemolytica/fisiología , Lactoferrina/farmacología , Animales , Viabilidad Microbiana/efectos de los fármacos , Bovinos , Factores de Virulencia/metabolismo , Ovinos

RESUMEN

AIMS: To evaluate the prevalence, molecular characteristics, antimicrobial susceptibility, and epithelial invasion of Streptococcus agalactiae strains isolated from pregnant women and newborns in Rio de Janeiro, Brazil. METHODS AND RESULTS: A total of 67 S. agalactiae isolates, 48 isolates from pregnant women and 19 from neonates, were analyzed. Capsular type Ia and V were predominant (35.8%/each). The multilocus sequence typing analysis revealed the presence of 19 STs grouped into 6 clonal complexes with prevalence of CC17/40.3% and CC23/34.3%. The lmb and iag virulence genes were found in 100% of isolates. Four S. agalactiae strains, belonging to CC17/ST1249 and CC23/ST23, were able to adhere to A549 respiratory epithelial cells. Antimicrobial resistance was verified mainly to tetracycline (85%), erythromycin (70.8%), and clindamycin (58.3%). Four S. agalactiae isolates were multidrug resistant. The resistance genes tested were found in 92.5% of isolates for tetM, 58.2% for ermB, 28.4% for mefAE, and 10.4% for tetO. CONCLUSION: The study showed a high prevalence of virulence and antimicrobial genes in S. agalactiae strains isolated from pregnant women and newborns, supporting the idea that continued surveillance is necessary to identify risk factors and perform long-term follow-up in pregnant women and neonates in Rio de Janeiro.

Asunto(s)

Antibacterianos , Células Epiteliales , Pruebas de Sensibilidad Microbiana , Tipificación de Secuencias Multilocus , Infecciones Estreptocócicas , Streptococcus agalactiae , Streptococcus agalactiae/genética , Streptococcus agalactiae/efectos de los fármacos , Streptococcus agalactiae/aislamiento & purificación , Femenino , Humanos , Brasil , Embarazo , Infecciones Estreptocócicas/microbiología , Antibacterianos/farmacología , Recién Nacido , Células Epiteliales/microbiología , Farmacorresistencia Bacteriana/genética , Adulto , Factores de Virulencia/genética , Complicaciones Infecciosas del Embarazo/microbiología , Farmacorresistencia Bacteriana Múltiple/genética , Virulencia/genética

RESUMEN

Staphylococcus pseudintermedius, which is part of the skin microbiome of dogs, causes a variety of opportunistic infections. These infections may become more difficult to treat due to the formation of biofilm. The capacity of S. pseudintermedius to form biofilm, as well as the associated genes, has not been elucidated. This study evaluated the production and composition of S. pseudintermedius biofilm. Samples were collected from both infected dogs and asymptomatic dogs. Isolates were identified using mass spectrometry and Multiplex-PCR. Biofilm production and composition were assessed using a quantitative microtiter plate assay. The presence of ica operon genes and sps genes was investigated using conventional PCR. The investigation of Agr type and virulence genes was conducted in silico on 24 sequenced samples. All strains could produce strong biofilms, with most of the isolates presenting a polysaccharide biofilm. 63.6% of the isolates carried the complete ica operon (ADBC). All samples showed the presence of the genes spsK, spsA, and spsL, while the distribution of other genes varied. Agr type III was the most prevalent (52.2%). All sequenced samples carried the cytotoxins hlb, luk-S, luk-F, as well as the exfoliative toxins siet and se_int. No isolate displayed other exfoliative toxins. Only LB1733 presented a set of different enterotoxins (sea, seb, sec_canine, seh, sek, sel, and seq). Our findings suggest that S. pseudintermedius is a strong producer of biofilm and carries virulence genes.

Asunto(s)

Biopelículas , Enfermedades de los Perros , Staphylococcus , Animales , Biopelículas/crecimiento & desarrollo , Perros , Enfermedades de los Perros/microbiología , Virulencia/genética , Staphylococcus/genética , Staphylococcus/aislamiento & purificación , Staphylococcus/patogenicidad , Staphylococcus/clasificación , Staphylococcus/fisiología , Infecciones Estafilocócicas/microbiología , Infecciones Estafilocócicas/veterinaria , Factores de Virulencia/genética , Proteínas Bacterianas/genética , Operón

RESUMEN

This study aimed to determine the prevalence of V. parahaemolyticus in oysters from the northwestern coast of Mexico and to identify the serotypes, virulence factors, and antibiotic resistance of the strains. Oyster samples were collected from 2012 to 2020 from the northwest coast of Mexico; biochemical and molecular methods were used to identify V. parahaemolyticus from oysters; antiserum reaction to determine V. parahaemolyticus serotypes, and PCR assays were performed to identify pathogenic (tdh and/or trh) or pandemic (toxRS/new, and/or orf8) strains and antibiotic resistance testing. A total of 441 oyster samples were collected and tested for V. parahaemolyticus. Forty-seven percent of oyster samples were positive for V. parahaemolyticus. Ten different O serogroups and 72 serovars were identified, predominantly serotype O1:KUT with 22.2% and OUT:KUT with 17.3%. Twenty new serotypes that had not been previously reported in our region were identified. We detected 4.3% of pathogenic clones but no pandemic strains. About 73.5% of strains were resistant to at least one antibiotic, mainly ampicillin and ciprofloxacin; 25% were multi-drug resistant. In conclusion, the pathogenic strains in oysters and antibiotic resistance are of public health concern, as the potential for outbreaks throughout northwestern Mexico is well established.

Asunto(s)

Antibacterianos , Ostreidae , Mariscos , Vibrio parahaemolyticus , Factores de Virulencia , Animales , Vibrio parahaemolyticus/genética , Vibrio parahaemolyticus/efectos de los fármacos , Vibrio parahaemolyticus/aislamiento & purificación , México/epidemiología , Ostreidae/microbiología , Factores de Virulencia/genética , Antibacterianos/farmacología , Mariscos/microbiología , Farmacorresistencia Bacteriana , Serogrupo , Virulencia/genética , Pruebas de Sensibilidad Microbiana

RESUMEN

Alternative strategies for controlling Staphylococcus aureus and other pathogens have been continuously investigated, with nisin, a bacteriocin widely used in the food industry as a biopreservative, gaining increasing attention. In addition to its antimicrobial properties, bacteriocins have significant effects on genome functionality even at inhibitory concentrations. This study investigated the impact of subinhibitory concentrations of nisin on S. aureus. Culturing in the presence of 0.625 µmol l-1 nisin, led to the increased relative expression of hla, saeR, and sarA, genes associated with virulence while expression of the sea gene, encoding staphylococcal enterotoxin A (SEA), decreased. In an in vivo experiment, Galleria mellonella larvae inoculated with S. aureus cultured in the presence of nisin exhibited 97% mortality at 72 h post-infection, compared to over 40% of larvae mortality in larvae infected with S. aureus. A comprehensive understanding of the effect of nisin on the transcriptional response of virulence genes and the impact of these changes on the virulence of S. aureus can contribute to assessing the application of this bacteriocin in food and medical contexts.

Asunto(s)

Antibacterianos , Larva , Mariposas Nocturnas , Nisina , Staphylococcus aureus , Nisina/farmacología , Staphylococcus aureus/efectos de los fármacos , Staphylococcus aureus/genética , Staphylococcus aureus/patogenicidad , Animales , Virulencia/genética , Larva/microbiología , Larva/efectos de los fármacos , Antibacterianos/farmacología , Mariposas Nocturnas/microbiología , Infecciones Estafilocócicas/microbiología , Regulación Bacteriana de la Expresión Génica/efectos de los fármacos , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Factores de Virulencia/genética , Pruebas de Sensibilidad Microbiana