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1.
Int J Food Microbiol ; 326: 108641, 2020 Aug 02.
Artículo en Inglés | MEDLINE | ID: mdl-32371295

RESUMEN

Thermotolerant Campylobacter is the leading bacterial cause of foodborne illness in humans worldwide. The objectives of this study were to estimate prevalence and to identify and characterize potential sources of thermotolerant Campylobacter contamination in broilers on farms and at the slaughterhouse; to evaluate the clonal relationship among thermotolerant Campylobacter isolates from different stages of the broiler meat supply chain, and to analyze the presence of virulence genes in different sources of thermotolerant Campylobacter. A total of 1210 samples were collected from three broiler meat supply chains in Santa Fe, Argentina. At the farms, the sampling collection included broilers one week prior to slaughter, wild-living birds, domestic dogs, wild rodents, farm workers' boots, litter, feed, drinking water, flies, and darkling beetles (Alphitobius diaperinus). At the slaughtering line, the samples taken were from the evisceration zone (broiler cecum, working surfaces, evisceration knives and workers' hands), from the chiller zone (surfaces and direct supply water) and from the packing zone (work surfaces, workers' hands and broiler carcasses). The samples taken along each supply chain were in the same batch. The isolates obtained were identified to the species level (C. jejuni and C. coli) by multiplex PCR and were analyzed using pulsed-field gel electrophoresis to compare different profiles according to the source. Finally, the presence of 11 virulence genes was examined (cadF, cdtA, cdtB, cdtC, ciaB, flaA, flhA, iam, wlaN, virB11, racR). From 254 isolates, 128 (50.4%) were Campylobacter jejuni and 126 (49.6%) Campylobacter coli. C. jejuni was the species most prevalent in farm and C. coli the species most prevalent at the slaughterhouse. We detected thermotolerant Campylobacter in samples of wild birds, darkling beetles, farm workers' boots, flies and litter. At the slaughterhouse, the prevalence varied along the process line. By analyzing PFGE results, C. jejuni showed 21 profiles with three predominant genotypes, while C. coli showed 14 profiles with four predominant genotypes. A high genotype diversity was found; however, relationships between isolates from different stages of the broiler meat chain, between broiler and potential sources of thermotolerant Campylobacter contamination and between strains in the farm and in the slaughterhouse were detected. Furthermore, there was evidence of cross-contamination at the slaughterhouse. FlaA, flhA genes were detected in all strains, and the third most prevalent virulence gene was cadF. Only those strains obtained from flies, wild-living birds and broiler carcass samples harbored 10 of 11 pathogenic genes. The prevalence of some pathogenic genes between C. jejuni and C. coli was different. This evidence should contribute the scientific basis to implement risk management measures in public health.


Asunto(s)
Campylobacter coli/genética , Campylobacter jejuni/genética , Enfermedades Transmitidas por los Alimentos/microbiología , Carne/microbiología , Aves de Corral/microbiología , Mataderos/estadística & datos numéricos , Animales , Argentina , Proteínas de la Membrana Bacteriana Externa/genética , Proteínas Bacterianas/genética , Infecciones por Campylobacter/microbiología , Campylobacter coli/aislamiento & purificación , Campylobacter coli/patogenicidad , Campylobacter jejuni/aislamiento & purificación , Campylobacter jejuni/patogenicidad , Proteínas Portadoras/genética , Ciego/microbiología , Pollos/microbiología , Escarabajos/microbiología , Dípteros/microbiología , Perros , Agua Potable/microbiología , Electroforesis en Gel de Campo Pulsado , Flagelina/genética , Genotipo , Humanos , Industria para Empaquetado de Carne/estadística & datos numéricos , Proteínas de la Membrana/genética , Prevalencia , Roedores/microbiología , Termotolerancia , Virulencia/genética
2.
Mol Genet Genomics ; 295(4): 1001-1012, 2020 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-32307574

RESUMEN

The increasing number of Chromobacterium haemolyticum human infection reports, especially in tropical regions and connected with environmental sources, resulted in an urge to better describe this species. This study aimed to characterize the C. haemolyticum resistome, virulence determinants and genetic platforms related with genome plasticity. A comparative genomic analysis was conducted between clinical C. haemolyticum genomes publicly available and the genome of an environmental isolate obtained in this study. The pangenome of C. haemolyticum was calculated and a total of 3378 core genes were predicted in its core genome, corresponding to 51.7% of the pangenome. Genetic determinants putatively encoding resistance to beta-lactams, fosfomycin, aminoglycosides and trimethoprim were predicted in all genomes, possibly constituting the intrinsic resistome of this species. In terms of resistance to beta-lactams, 4 genes were predicted encoding beta-lactamases of classes A, C and D. Moreover, the analysis of Chromobacterium genomes and C. haemolyticum environmental isolates reinforced the role of this genus as progenitor of the blaKPC gene. Putative virulence factors (VFs) were predicted in all genomes, related to adherence, toxins production, colonization and cell invasion. Secretion systems, including type III, were detected. A significant number of transposases and genomic islands were predicted in C. haemolyticum, in some cases above the average reported for Gram-negative bacterial genomes. We conclude that C. haemolyticum strains, including those of environmental origin, present a noteworthy collection of antibiotic resistance genes and VFs. Furthermore, sequences related to gene mobility and genome plasticity suggest high adaptability potential and a possible role as disseminator of antibiotic resistance.


Asunto(s)
Infecciones Bacterianas/genética , Chromobacterium/genética , Farmacorresistencia Bacteriana Múltiple/genética , Filogenia , Antibacterianos/efectos adversos , Antibacterianos/uso terapéutico , Infecciones Bacterianas/tratamiento farmacológico , Infecciones Bacterianas/microbiología , Chromobacterium/clasificación , Chromobacterium/efectos de los fármacos , Chromobacterium/patogenicidad , Genoma Bacteriano/efectos de los fármacos , Genoma Bacteriano/genética , Genómica , Humanos , Pruebas de Sensibilidad Microbiana , Virulencia/genética
3.
Crit Rev Microbiol ; 46(2): 182-193, 2020 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-32282268

RESUMEN

The last century has witnessed several assaults from RNA viruses, resulting in millions of death throughout the world. The 21st century appears no longer an exception, with the trend continued with escalated fear of SARS coronavirus in 2002 and further concern of influenza H5N1 in 2003. A novel influenza virus created the first pandemic of the 21st century, the pandemic flu in 2009 preceded with the emergence of another deadly virus, MERS-CoV in 2012. A novel coronavirus "SARS-CoV-2" (and the disease COVID-19) emerged suddenly, causing a rapid outbreak with a moderate case fatality rate. This virus is continuing to cause health care providers grave concern due to the lack of any existing immunity in the human population, indicating their novelty and lack of previous exposure. The big question is whether this novel virus will be establishing itself in an endemic form or will it eventually die out? Endemic viruses during circulation may acquire mutations to infect naïve, as well as individual with pre-existing immunity. Continuous monitoring is strongly advisable, not only to the newly infected individuals, but also to those recovered individuals who were infected by SARS-CoV-2 as re-infection may lead to the selection of escape mutants and subsequent dissemination to the population.


Asunto(s)
Betacoronavirus/patogenicidad , Infecciones por Coronavirus/epidemiología , Infecciones por Coronavirus/virología , Neumonía Viral/epidemiología , Neumonía Viral/virología , Betacoronavirus/genética , Infecciones por Coronavirus/inmunología , Infecciones por Coronavirus/mortalidad , Brotes de Enfermedades , Enfermedades Endémicas , Humanos , Mutación , Pandemias , Neumonía Viral/inmunología , Neumonía Viral/mortalidad , Virulencia/genética
4.
PLoS One ; 15(3): e0230220, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-32163505

RESUMEN

Helicobacter pylori is a Gram-negative bacterium that causes chronic atrophic gastritis and peptic ulcers and it has been associated with the development of gastric adenocarcinoma and mucosa-associated lymphoid tissue (MALT). One of the more remarkable characteristics of H. pylori is its ability to survive in the hostile environment of the stomach. H. pylori regulates the expression of specific sets of genes allowing it to survive high acidity levels and nutrient scarcity. In the present study, we determined the expression of virulence associated protein D (VapD) of H. pylori inside adenocarcinoma gastric (AGS) cells and in gastric biopsies. Using qRT-PCR, VapD expression was quantified in intracellular H. pylori-AGS cell cultures at different time points and in gastric mucosa biopsies from patients suffering from chronic atrophic gastritis, follicular gastritis, peptic ulcers, gastritis precancerous intestinal metaplasia and adenocarcinoma. Our results show that vapD of H. pylori presented high transcription levels inside AGS cells, which increased up to two-fold above basal values across all assays over time. Inside AGS cells, H. pylori acquired a coccoid form that is metabolically active in expressing VapD as a protection mechanism, thereby maintaining its permanence in a viable non-cultivable state. VapD of H. pylori was expressed in all gastric biopsies, however, higher expression levels (p = 0.029) were observed in gastric antrum biopsies from patients with follicular gastritis. The highest VapD expression levels were found in both antrum and corpus gastric biopsies from older patients (>57 years old). We observed that VapD in H. pylori is a protein that is only produced in response to interactions with eukaryotic cells. Our results suggest that VapD contributes to the persistence of H. pylori inside the gastric epithelial cells, protecting the microorganism from the intracellular environment, reducing its growth rate, enabling long-term infection and treatment resistance.


Asunto(s)
Proteínas Bacterianas/genética , Gastritis Atrófica/etiología , Helicobacter pylori/genética , Glicoproteínas de Membrana/genética , Estómago/microbiología , Estómago/patología , Adenocarcinoma/etiología , Adenocarcinoma/microbiología , Adenocarcinoma/patología , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Biopsia , Técnicas de Cocultivo/métodos , Femenino , Mucosa Gástrica/microbiología , Mucosa Gástrica/patología , Gastritis Atrófica/microbiología , Gastritis Atrófica/patología , Gastroscopía/métodos , Infecciones por Helicobacter/complicaciones , Infecciones por Helicobacter/patología , Humanos , Intestinos/microbiología , Intestinos/patología , Masculino , Metaplasia/microbiología , Metaplasia/patología , Persona de Mediana Edad , Úlcera Péptica/metabolismo , Úlcera Péptica/patología , Lesiones Precancerosas/etiología , Lesiones Precancerosas/microbiología , Lesiones Precancerosas/patología , Antro Pilórico/microbiología , Antro Pilórico/patología , Neoplasias Gástricas/etiología , Neoplasias Gástricas/microbiología , Neoplasias Gástricas/patología , Virulencia/genética , Adulto Joven
5.
PLoS One ; 15(3): e0229064, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-32214338

RESUMEN

Streptococcus pyogenes is a strict human pathogen responsible for more than 700 million infections annually worldwide. Strains of serotype M28 S. pyogenes are typically among the five more abundant types causing invasive infections and pharyngitis in adults and children. Type M28 strains also have an unusual propensity to cause puerperal sepsis and neonatal disease. We recently discovered that a one-nucleotide indel in an intergenic homopolymeric tract located between genes Spy1336/R28 and Spy1337 altered virulence in a mouse model of infection. In the present study, we analyzed size variation in this homopolymeric tract and determined the extent of heterogeneity in the number of tandemly-repeated 79-amino acid domains in the coding region of Spy1336/R28 in large samples of strains recovered from humans with invasive infections. Both repeat sequence elements are highly polymorphic in natural populations of M28 strains. Variation in the homopolymeric tract results in (i) changes in transcript levels of Spy1336/R28 and Spy1337 in vitro, (ii) differences in virulence in a mouse model of necrotizing myositis, and (iii) global transcriptome changes as shown by RNAseq analysis of isogenic mutant strains. Variation in the number of tandem repeats in the coding sequence of Spy1336/R28 is responsible for size variation of R28 protein in natural populations. Isogenic mutant strains in which genes encoding R28 or transcriptional regulator Spy1337 are inactivated are significantly less virulent in a nonhuman primate model of necrotizing myositis. Our findings provide impetus for additional studies addressing the role of R28 and Spy1337 variation in pathogen-host interactions.


Asunto(s)
Proteínas Bacterianas/genética , Fascitis Necrotizante/microbiología , Infecciones Estreptocócicas/microbiología , Streptococcus pyogenes/genética , Streptococcus pyogenes/aislamiento & purificación , Virulencia/genética , Animales , Modelos Animales de Enfermedad , Fascitis Necrotizante/patología , Regulación Bacteriana de la Expresión Génica , Heterogeneidad Genética , Humanos , Ratones , Polimorfismo Genético , Infecciones Estreptocócicas/patología , Transcriptoma , Factores de Virulencia/genética
6.
PLoS One ; 15(3): e0230031, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-32163464

RESUMEN

We characterised 80 Staphylococcus aureus strains isolated from human patients with SSTIs at a rural hospital in Ethiopia. Susceptibility to antibiotic of all strains was tested. The MLST method was used to type and a phylogenetic analysis was conducted employing the sequences of 7 housekeeping genes. PCR amplification was used to investigate the presence of the following virulence genes in all strains: hla (α-haemolysin), tstH (toxic shock syndrome toxin), luk PV (Panton-Valentine leukocidin), fnbA (fibronectin binding protein A) and mecA (methicillin resistance). Most of the strains were resistant to penicillin and ampicillin, but only 3 strains were resistant to oxacillin, and 1 of them was a true MRSA. The MLST results showed a high diversity of sequence types (ST), 55% of which were new, and ST152 was the most prevalent. A phylogeny study showed that many of the new STs were phylogenetically related to other previously described STs, but bore little relationship to the only ST from Ethiopia described in the database. Virulence gene detection showed a high prevalence of strains encoding the hla, fnbA and pvl genes (98.77%, 96.3% and 72.84%, respectively), a low prevalence of the tst gene (13.58%) and a markedly low prevalence of MRSA (1.25%). S. aureus strains isolated from patients in a rural area in Ethiopia showed low levels of antibiotic resistance, except to penicillin. Moreover, this study reveals new STs in Eastern Africa that are phylogenetically related to other previously described STs, and confirm the high prevalence of the pvl gene and the low prevalence of MRSA on the continent.


Asunto(s)
Antibacterianos/farmacología , Staphylococcus aureus/efectos de los fármacos , Virulencia/genética , Proteínas Bacterianas/clasificación , Proteínas Bacterianas/genética , Toxinas Bacterianas/clasificación , Toxinas Bacterianas/genética , Cefalosporinas/farmacología , Farmacorresistencia Bacteriana/genética , Etiopía , Exotoxinas/clasificación , Exotoxinas/genética , Hospitales Rurales , Humanos , Leucocidinas/clasificación , Leucocidinas/genética , Staphylococcus aureus Resistente a Meticilina/efectos de los fármacos , Staphylococcus aureus Resistente a Meticilina/genética , Staphylococcus aureus Resistente a Meticilina/aislamiento & purificación , Pruebas de Sensibilidad Microbiana , Tipificación de Secuencias Multilocus , Penicilinas/farmacología , Filogenia , Infecciones Estafilocócicas/diagnóstico , Infecciones Estafilocócicas/microbiología , Staphylococcus aureus/clasificación , Staphylococcus aureus/aislamiento & purificación , Staphylococcus aureus/patogenicidad
7.
Gene ; 741: 144566, 2020 May 30.
Artículo en Inglés | MEDLINE | ID: mdl-32171826

RESUMEN

Bacteria of the genusGlutamicibacterare considered ubiquitous because they can be found in soil, water and air. They have already been isolated from different habitats, including different types of soil, clinical samples, cheese and plants. Glutamicibacter creatinolyticus is a Gram-positive bacterium important to various biotechnological processes, however, as a pathogen it is associated to urinary tract infections and bacteremia. Recently,Glutamicibacter creatinolyticusLGCM 259 was isolated from a mare, which displayed several diffuse subcutaneous nodules with heavy vascularization. In this study, sequencing, genomic analysis ofG. creatinolyticusLGCM 259 and comparative analyseswere performedamong 4representatives of different members of genusfromdifferent habitats, available in the NCBI database. The LGCM 259 strain's genome carries important factors of bacterial virulence that are essential in cell viability, virulence, and pathogenicity. Genomic islands were predicted for 4 members of genusGlutamicibacter,showing ahigh number of GEIs,which may reflect a high interspecific diversity and a possible adaptive mechanism responsible for the survival of each species in its specific niche. Furthermore,G. creatinolyticusLGCM 259 sharessyntenicregions, albeit with a considerable loss of genes, in relation to the other species. In addition,G. creatinolyticusLGCM 259 presentsresistancegenes to 6 differentclasses ofantibiotics and heavy metals, such as: copper, arsenic, chromium and cobalt-zinc-cadmium.Comparative genomicsanalysescouldcontribute to the identification of mobile genetic elements particular to the speciesG. creatinolyticuscompared to other members of genus. The presence of specific regions inG. creatinolyticuscould be indicative of their rolesin host adaptation, virulence, and the characterization ofastrain that affects animals.


Asunto(s)
Absceso/genética , Adaptación Fisiológica/genética , Variación Genética , Micrococcaceae/genética , Absceso/microbiología , Absceso/veterinaria , Animales , Genoma Bacteriano , Islas Genómicas/genética , Genómica , Caballos/microbiología , Masculino , Micrococcaceae/patogenicidad , Filogenia , Virulencia/genética
8.
PLoS Negl Trop Dis ; 14(3): e0008166, 2020 03.
Artículo en Inglés | MEDLINE | ID: mdl-32203536

RESUMEN

Flaviviruses such as yellow fever, dengue or Zika viruses are responsible for significant human and veterinary diseases worldwide. These viruses contain an RNA genome, prone to mutations, which enhances their potential to emerge as pathogens. Bamaga virus (BgV) is a mosquito-borne flavivirus in the yellow fever virus group that we have previously shown to be host-restricted in vertebrates and horizontally transmissible by Culex mosquitoes. Here, we aimed to characterise BgV host-restriction and to investigate the mechanisms involved. We showed that BgV could not replicate in a wide range of vertebrate cell lines and animal species. We determined that the mechanisms involved in BgV host-restriction were independent of the type-1 interferon response and RNAse L activity. Using a BgV infectious clone and two chimeric viruses generated as hybrids between BgV and West Nile virus, we demonstrated that BgV host-restriction occurred post-cell entry. Notably, BgV host-restriction was shown to be temperature-dependent, as BgV replicated in all vertebrate cell lines at 34°C but only in a subset at 37°C. Serial passaging of BgV in Vero cells resulted in adaptive mutants capable of efficient replication at 37°C. The identified mutations resulted in amino acid substitutions in NS4A-S124F, NS4B-N244K and NS5-G2C, all occurring close to a viral protease cleavage site (NS4A/2K and NS4B/NS5). These mutations were reverse engineered into infectious clones of BgV, which revealed that NS4B-N244K and NS5-G2C were sufficient to restore BgV replication in vertebrate cells at 37°C, while NS4A-S124F further increased replication efficiency. When these mutant viruses were injected into immunocompetent mice, alongside BgV and West Nile virus chimeras, infection and neurovirulence were enhanced as determined by clinical scores, seroconversion, micro-neutralisation, viremia, histopathology and immunohistochemistry, confirming the involvement of these residues in the attenuation of BgV. Our studies identify a new mechanism of host-restriction and attenuation of a mosquito-borne flavivirus.


Asunto(s)
Infecciones por Flavivirus/virología , Flavivirus/genética , Flavivirus/patogenicidad , Mutación , Proteínas no Estructurales Virales/genética , Animales , Encéfalo/patología , Encéfalo/virología , Línea Celular , Chlorocebus aethiops , Culicidae/virología , Modelos Animales de Enfermedad , Endorribonucleasas/metabolismo , Femenino , Flavivirus/fisiología , Infecciones por Flavivirus/metabolismo , Infecciones por Flavivirus/patología , Células HEK293 , Humanos , Masculino , Ratones , Mosquitos Vectores/virología , Células Vero , Virulencia/genética , Replicación Viral , Virus del Nilo Occidental/genética
9.
PLoS One ; 15(2): e0227749, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-32012177

RESUMEN

Toxoplasma gondii is classified into 16 haplogroups based on a worldwide genotyping study of the parasite. However, only a few isolates from Japan were included in this analysis. To conduct more precise genotyping of T. gondii, we examined the genotypes of Japanese isolates in this study. DNA sequences of 6 loci were determined in 17 Japanese isolates and compared with those of strains of 16 haplogroups. As a result, Japanese isolates were classified into four groups. We investigated the virulence of some Japanese isolates and found a highly virulent strain in mice, comparable to that of RH strain, although this Japanese isolate was sister to strains of haplogroup 2, which show moderate virulence in mice. We further investigated whether this high virulence isolate had different virulence mechanism and strategy to adapt to Japanese host from other strains by comparing the virulence-related genes, ROP5, 18 and the immunomodulatory gene, ROP16 of the isolate with those of archetypical strains (GT1, ME49 and VEG). This analysis indicated the high virulence of the isolate in mice was partly explained by gene sequences of ROP5 and ROP16. These findings lead to the elucidation of biodiversity of T. gondii and have potential to optimize the diagnostic protocol.


Asunto(s)
Variación Genética , Toxoplasma/genética , Toxoplasmosis Animal/genética , Toxoplasmosis/genética , Alelos , Animales , Genotipo , Humanos , Japón , Ratones , Filogenia , Proteínas Tirosina Quinasas/genética , Proteínas Protozoarias/genética , Toxoplasma/patogenicidad , Toxoplasmosis/parasitología , Toxoplasmosis Animal/parasitología , Virulencia/genética
10.
Environ Pollut ; 260: 114041, 2020 May.
Artículo en Inglés | MEDLINE | ID: mdl-32006889

RESUMEN

Infections caused by carbapenem-resistant Enterobacteriaceae are a growing concern worldwide. Raoultella ornithinolytica is a species in the Enterobacteriaceae family which can cause hospital-acquired infections and is sporadically reported as carbapenem-resistant from human and environmental sources. In this study, we firstly report on an NDM-1-producing R. ornithinolytica, Rao166, isolated from drinking water in an animal cultivation area in China. In addition to carbapenem-resistance, Rao166 was resistant to several other antibiotics including gentamicin, sulfamethoxazole-trimethoprim, tetracycline and fosfomycin. Rao166 carried a novel IncFIC-type megaplasmid, 382,325 bp in length (pRAO166a). A multidrug resistance region, 60,600 bp in length, was identified in the plasmid containing an aac(3)-IId-like gene, aac(6')-Ib-cr, blaDHA-1, blaTEM-1B, blaCTX-M-3, blaOXA-1, blaNDM-1, qnrB4, catB3, arr-3, sul1, and tet(D). Results from virulence assays implied that Rao166 has considerable pathogenic potential. Although pRAO166a was found to be non-transmissible, dissemination of the NDM-1 producing strain may occur from well water to humans or animals through cross-contamination during food preparation or directly via drinking water, and potentially lead to difficult-to-treat infections. Thus, contamination of well water by this carbapenem-resistant and presumptively virulent strain of R. ornithinolytica should be considered a potential public health risk.


Asunto(s)
Carbapenémicos , Farmacorresistencia Bacteriana/genética , Enterobacteriaceae/genética , beta-Lactamasas/metabolismo , Animales , Antibacterianos , China , Enterobacteriaceae/aislamiento & purificación , Humanos , Pruebas de Sensibilidad Microbiana , Plásmidos , Virulencia/genética , Agua
11.
Hum Genet ; 139(6-7): 961-980, 2020 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-32067109

RESUMEN

Neisseria meningitidis is a leading cause of bacterial septicaemia and meningitis worldwide. Meningococcal disease is rare but can be life threatening with a tendency to affect children. Many studies have investigated the role of human genetics in predisposition to N. meningitidis infection. These have identified both rare single-gene mutations as well as more common polymorphisms associated with meningococcal disease susceptibility and severity. These findings provide clues to the pathogenesis of N. meningitidis, the basis of host susceptibility to infection and to the aetiology of severe disease. From the multiple discoveries of monogenic complement deficiencies to the associations of complement factor H and complement factor H-related three polymorphisms to meningococcal disease, the complement pathway is highlighted as being central to the genetic control of meningococcal disease. This review aims to summarise the current understanding of the host genetic basis of meningococcal disease with respect to the different stages of meningococcal infection.


Asunto(s)
Predisposición Genética a la Enfermedad , Genética Humana , Infecciones Meningocócicas/genética , Neisseria meningitidis/patogenicidad , Polimorfismo Genético , Virulencia/genética , Factor H de Complemento/genética , Humanos , Infecciones Meningocócicas/inmunología , Infecciones Meningocócicas/microbiología
12.
PLoS One ; 15(2): e0220428, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-32101543

RESUMEN

BACKGROUND: Multidrug-resistant (MDR) E. coli with extended-spectrum ß-lactamases (ESBLs) is becoming endemic in health care settings around the world. Baseline data on virulence and antimicrobial resistance (AMR) of specific lineages of E. coli circulating in developing countries like India is currently lacking. METHODS: Whole-genome sequencing was performed for 60 MDR E. coli isolates. The analysis was performed at single nucleotide polymorphism (SNP) level resolution to identify the presence of their virulence and AMR genes. RESULTS: Genome comparison revealed the presence of ST-131 global MDR and ST410 as emerging-MDR clades of E. coli in India. AMR gene profile for cephalosporin and carbapenem resistance differed between the clades. Genotypes blaCTX-M-15 and blaNDM-5 were common among cephalosporinases and carbapenemases, respectively. For aminoglycoside resistance, rmtB was positive for 31.7% of the isolates, of which 95% were co-harboring carbapenemases. In addition, the FimH types and virulence gene profile positively correlated with the SNP based phylogeny, and also revealed the evolution of MDR clones among the study population with temporal accumulation of SNPs. The predominant clone was ST167 (blaNDM lineage) followed by ST405 (global clone ST131 equivalent) and ST410 (fast spreading high risk clone). CONCLUSIONS: This is the first report on the whole genome analysis of MDR E. coli lineages circulating in India. Data from this study will provide public health agencies with baseline information on AMR and virulent genes in pathogenic E. coli in the region.


Asunto(s)
Farmacorresistencia Bacteriana Múltiple/genética , Escherichia coli/genética , Genómica , Virulencia/genética , Secuenciación Completa del Genoma , Carbapenémicos/farmacología , Cefalosporinas/farmacología , Escherichia coli/aislamiento & purificación , Infecciones por Escherichia coli/epidemiología , Infecciones por Escherichia coli/genética , Infecciones por Escherichia coli/microbiología , Humanos , India , Polimorfismo de Nucleótido Simple
13.
Helicobacter ; 25(2): e12684, 2020 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-32074664

RESUMEN

BACKGROUND: Resistant Helicobacter pylori to commonly used antimicrobial agents are associated with severe upper gastrointestinal disorders. To provide an epidemiological picture of H pylori and characterize the resistance pattern and genetic variation of clinical isolates, stomach biopsies from patients with functional dyspepsia were evaluated in northeast of Iran. MATERIALS AND METHODS: In this study, 80 patients were recruited. Finally, fifty H pylori strains were isolated from antrum and corpus biopsies by culturing on Columbia agar. All strains were identified by standard laboratory procedures. Susceptibility testing of antibiotics was performed using minimum inhibitory concentration test. Allele-specific primer (ASP)-PCR of 23S rRNA which associated with clarithromycin resistance was done among resistant strains. Moreover, cagA gene and polymorphism in vacA were detected. Random amplified polymorphic DNA polymerase chain reaction (RAPD-PCR) was applied to investigate the genetic variations among all strains. RESULTS: Antibiotic resistance pattern of H pylori strains was as follows: 68% (34/50) to metronidazole, 50% (25/50) to rifampicin, 30% (15/50) to amoxicillin, 28% (14/50) to levofloxacin, 22% (11/50) to clarithromycin, and 16% (8/50) to tetracycline. Multidrug-resistant strains were observed in 19 strains (38%). ASP-PCR of 23S rRNA showed four strains had A2143G mutation, six strains had A2142G mutation, and one strain had a Wt+A2143G mutation. Amplification of virulence-associated genes revealed that cagA was present in 27 isolates (54%) and vacA in 36 isolates (72%). The most common genotype of H pylori was vacA s1am2 (40%) followed by vacA s2m2 (14%), vacA s1am1 (12%), vacA s1bm1 (4%), and vacA s1bm2 (2%). DNA fingerprinting pattern indicated a high heterogeneity among isolated strains. CONCLUSION: An alarming level of resistance to metronidazole and rifampicin and high heterogeneity among H pylori isolates highlighted the importance of continued monitoring of antimicrobial susceptibility and epidemiological surveillance of this pathogen.


Asunto(s)
Farmacorresistencia Bacteriana/genética , Farmacorresistencia Bacteriana Múltiple/genética , Infecciones por Helicobacter , Helicobacter pylori , Virulencia/genética , Adulto , Antígenos Bacterianos/genética , Proteínas Bacterianas/genética , Biopsia , Femenino , Variación Genética , Infecciones por Helicobacter/tratamiento farmacológico , Infecciones por Helicobacter/epidemiología , Helicobacter pylori/efectos de los fármacos , Helicobacter pylori/genética , Helicobacter pylori/aislamiento & purificación , Humanos , Irán/epidemiología , Masculino , Pruebas de Sensibilidad Microbiana , Persona de Mediana Edad , ARN Ribosómico 23S/genética , Técnica del ADN Polimorfo Amplificado Aleatorio , Estómago/microbiología
14.
PLoS One ; 15(2): e0228956, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-32040533

RESUMEN

Listeria monocytogenes (L. monocytogenes) is a foodborne pathogen and the etiologic agent of listeriosis, which can be disseminated within the agricultural environment particularly soil and irrigation water, contaminate farm produce and cause high mortality and morbidity among vulnerable individuals. This study assessed the incidence and antibiogram of L. monocytogenes recovered from irrigation water and agricultural soil samples collected from Chris Hani and Amathole District Municipalities (DMs) in Eastern Cape Province, South Africa. The distribution of presumptive L. monocytogenes in irrigation water and agricultural soil samples was done using the standard plate count method, while polymerase chain reaction (PCR) was used to identify the isolates. The confirmed isolates were screened for 9 key virulence markers using PCR after which they were subjected to antibiotic susceptibility testing against 18 antibiotics used for the alleviation of listeriosis using the disk diffusion method. Relevant putative antibiotic resistance genes in the resistant variants were screened for using PCR. The distribution of L. monocytogenes in irrigation water samples was statistically significant (P ≤ 0.05) and ranged from log10 1.00 CFU/100ml to log10 3.75 CFU/100 ml. In agricultural soil samples, the distribution ranged significantly (P ≤ 0.05) from log10 2.10 CFU/g to log10 3.51 CFU/g. Of the 117 presumptive L. monocytogenes recovered from irrigation water samples and 183 presumptive L. monocytogenes isolated from agricultural soil samples, 8 (6.8%) and 12 (6.6%) isolates were confirmed respectively. Nine virulence genes including inlA, inlB, inlC, inlJ, actA, hlyA, plcA, plcB, and iap were detected in all the isolates. The proportion of the isolates exhibiting phenotypic resistance against the test antimicrobials followed the order: tetracycline (90%), doxycycline (85%), cefotaxime (80%), penicillin (80%), chloramphenicol (70%), linezolid (65%), erythromycin (60%) and trimethoprim/sulfamethoxazole (55%). The isolates exhibited multiple antibiotic resistance against 3 or more antibiotics and the MAR indices of all the multidrug isolates were ≥0.2. The isolates harboured antibiotic resistance genes including tetA, tetB, tetC, sulI, sulII, aadA, aac(3)-IIa and ESBLs including blaTEM, blaCTX-M group 9, blaVEB as well as AmpC. None of the isolates harboured the carbapenemases. We conclude that irrigation water and agricultural soil collected from Chris Hani and Amathole District Municipalities (DMs) in Eastern Cape Province of South Africa are reservoirs and potential transmission routes of multidrug-resistant L. monocytogenes to the food web and consequently threat to public health.


Asunto(s)
Listeria monocytogenes/aislamiento & purificación , Microbiología del Suelo , Microbiología del Agua , Riego Agrícola , Reservorios de Enfermedades/microbiología , Farmacorresistencia Bacteriana Múltiple/genética , Granjas , Microbiología de Alimentos , Genes Bacterianos , Humanos , Listeria monocytogenes/efectos de los fármacos , Listeria monocytogenes/genética , Listeriosis/transmisión , Pruebas de Sensibilidad Microbiana , Sudáfrica , Virulencia/genética
15.
PLoS Pathog ; 16(2): e1008313, 2020 02.
Artículo en Inglés | MEDLINE | ID: mdl-32059031

RESUMEN

Many bacteria use quorum sensing (QS) to regulate virulence factor production in response to changes in population density. QS is mediated through the production, secretion, and detection of signaling molecules called autoinducers (AIs) to modulate population-wide behavioral changes. Four histidine kinases, LuxPQ, CqsS, CqsR and VpsS, have been identified in Vibrio cholerae as QS receptors to activate virulence gene expression at low cell density. Detection of AIs by these receptors leads to virulence gene repression at high cell density. The redundancy among these receptors is puzzling since any one of the four receptors is sufficient to support colonization of V. cholerae in the host small intestine. It is believed that one of the functions of such circuit architecture is to prevent interference on any single QS receptor. However, it is unclear what natural molecules can interfere V. cholerae QS and in what environment interference is detrimental. We show here mutants expressing only CqsR without the other three QS receptors are defective in colonizing the host large intestine. We identified ethanolamine, a common intestinal metabolite that can function as a chemical source of QS interference. Ethanolamine specifically interacts with the ligand-binding CACHE domain of CqsR and induces a premature QS response in V. cholerae mutants expressing only CqsR without the other three QS receptors. The effect of ethanolamine on QS gene expression and host colonization is abolished by mutations that disrupt CqsR signal sensing. V. cholerae defective in producing ethanolamine is still proficient in QS, therefore, ethanolamine functions only as an external cue for CqsR. Our findings suggest the inhibitory effect of ethanolamine on CqsR could be a possible source of QS interference but is masked by the presence of the other parallel QS pathways, allowing V. cholerae to robustly colonize the host.


Asunto(s)
Histidina Quinasa/metabolismo , Percepción de Quorum/fisiología , Vibrio cholerae/metabolismo , Proteínas Bacterianas/metabolismo , Biopelículas/crecimiento & desarrollo , Regulación Bacteriana de la Expresión Génica/genética , Histidina Quinasa/genética , Unión Proteica/fisiología , Transducción de Señal/genética , Vibrio cholerae/patogenicidad , Virulencia/genética
16.
PLoS Pathog ; 16(2): e1008355, 2020 02.
Artículo en Inglés | MEDLINE | ID: mdl-32092131

RESUMEN

Genetic studies have shown essential functions of N-glycosylation during infection of the plant pathogenic fungi, however, systematic roles of N-glycosylation in fungi is still largely unknown. Biological analysis demonstrated N-glycosylated proteins were widely present at different development stages of Magnaporthe oryzae and especially increased in the appressorium and invasive hyphae. A large-scale quantitative proteomics analysis was then performed to explore the roles of N-glycosylation in M. oryzae. A total of 559 N-glycosites from 355 proteins were identified and quantified at different developmental stages. Functional classification to the N-glycosylated proteins revealed N-glycosylation can coordinate different cellular processes for mycelial growth, conidium formation, and appressorium formation. N-glycosylation can also modify key components in N-glycosylation, O-glycosylation and GPI anchor pathways, indicating intimate crosstalk between these pathways. Interestingly, we found nearly all key components of the endoplasmic reticulum quality control (ERQC) system were highly N-glycosylated in conidium and appressorium. Phenotypic analyses to the gene deletion mutants revealed four ERQC components, Gls1, Gls2, GTB1 and Cnx1, are important for mycelial growth, conidiation, and invasive hyphal growth in host cells. Subsequently, we identified the Gls1 N-glycosite N497 was important for invasive hyphal growth and partially required for conidiation, but didn't affect colony growth. Mutation of N497 resulted in reduction of Gls1 in protein level, and localization from ER into the vacuole, suggesting N497 is important for protein stability of Gls1. Our study showed a snapshot of the N-glycosylation landscape in plant pathogenic fungi, indicating functions of this modification in cellular processes, developments and pathogenesis.


Asunto(s)
Retículo Endoplásmico/metabolismo , Magnaporthe/genética , Magnaporthe/metabolismo , Estudios de Evaluación como Asunto , Proteínas Fúngicas/metabolismo , Eliminación de Gen , Regulación Fúngica de la Expresión Génica/genética , Genes Fúngicos/genética , Glicosilación , Hifa/genética , Mutación , Oryza/microbiología , Enfermedades de las Plantas/microbiología , Proteómica/métodos , Eliminación de Secuencia , Esporas Fúngicas/crecimiento & desarrollo , Virulencia/genética
17.
Can J Microbiol ; 66(5): 351-358, 2020 May.
Artículo en Inglés | MEDLINE | ID: mdl-32040345

RESUMEN

Iron is a fundamental element required by most organisms, including Brucella. Several researchers have suggested that the iron response regulator (irr) and rhizobial iron regulator (rirA) genes regulate iron acquisition by Brucella abortus, influencing heme synthesis by and virulence of this pathogen. However, little is known about another Brucella species, Brucella melitensis. In this research, we successfully constructed two mutants: M5-90Δirr and M5-90ΔrirA. The adhesion, invasion, and intracellular survivability of these two mutants were evaluated in RAW264.7 cells infected with 1 × 106 CFU of M5-90Δirr, M5-90ΔrirA, or M5-90. We also tested the sensitivity of cells to hydrogen peroxide and their ability to grow. In addition, the virulence of these two mutants was evaluated in BALB/c mice. The results showed that the ability of these two mutants to invade and adhere inside the murine macrophages RAW264.7 was attenuated but their ability to replicate intracellularly was strengthened, enhancing the resistance to hydrogen peroxide. The M5-90Δirr mutant showed stronger growth ability than the parental strain under iron-limiting conditions. No differences were observed in the number of bacteria in spleen between M5-90 and M5-90Δirr at 7 or 15 days postinfection. However, the number of M5-90ΔrirA in spleen reduced significantly at 15 days postinfection. The splenic index of the M5-90Δirr group is evidently lower than that of M5-90. This is the first report that irr and rirA genes of B. melitensis are associated not only with virulence but also with growth ability. Together, our data suggest that M5-90Δirr is a promising Brucella vaccine candidate.


Asunto(s)
Proteínas Bacterianas/genética , Brucella melitensis/genética , Brucella melitensis/patogenicidad , Regulación Bacteriana de la Expresión Génica/fisiología , Hierro/metabolismo , Factores de Transcripción/genética , Animales , Antiinfecciosos Locales/toxicidad , Western Blotting , Vacuna contra la Brucelosis/inmunología , Brucelosis/prevención & control , Recuento de Colonia Microbiana , Femenino , Peróxido de Hidrógeno/toxicidad , Macrófagos/microbiología , Ratones , Ratones Endogámicos BALB C , Bazo/microbiología , Virulencia/genética
18.
BMC Infect Dis ; 20(1): 108, 2020 Feb 07.
Artículo en Inglés | MEDLINE | ID: mdl-32033541

RESUMEN

BACKGROUND: Urinary tract infection (UTI) is a common cause of morbidity worldwide. Uropathogenic Escherichia coli (UPEC) bacteria are the major cause of urinary tract infections. UPEC strains derive from different phylogenetic groups and possess an arsenal of virulence factors that contribute to their ability to overcome different defense mechanisms and cause disease. The objective of this study was to identify phylogroup and virulence genes of UPEC among urinary tract infection patients. METHODS: A cross sectional study was conducted from January 1, 2017 to October 9, 2017. E. coli bacteria were isolated from UTI patients using culture and conventional biochemical tests. Identification of phylogroup and genes that encodes for virulence factors was done using multiplex polymerase chain reaction (PCR). Data was processed and analyzed with SPSS version16.0 and Epi-info version 3.4.1 software. RESULTS: The most common urologic clinical manifestation combinations in this study were dysuria, urine urgency and urgency incontinence. The frequent UPEC virulence gene identified was fimH 164 (82%), followed by aer 109 (54.5%), hly 103 (51.5%), pap 59 (29.5%), cnf 58 (29%), sfa 50 (25%) and afa 24 (12%).There was significant association between pap gene and urine urgency (p-0.016); sfa and dysuria and urine urgency (p-0.019 and p-0.043 respectively); hly and suprapubic pain (p-0.002); aer and suprapubic pain, flank pain and fever (p-0.017, p-0.040, p-0.029 respectively). Majority of E. coli isolates were phylogroup B2 60(30%) followed by D 55(27.5%), B1 48(24%) and A 37(18.5%). There was significant association between E. coli phylogroup B2 and three virulence genes namely afa, pap, and sfa (p-0.014, p-0.002, p-0.004 respectively). CONCLUSION: In this study the most frequent E. coli virulence gene was fimH, followed by aer, hly, pap, cnf, sfa and afa respectively. There was significant association between E. coli virulence genes and clinical symptoms of UTI. The phylogenetic analysis indicates majority of uropathogenic E. coli isolates were phylogroup B2 followed by phylogroup D. Phylogroup B2 carries more virulence genes. Hence, targeting major UPEC phylogroup and virulence genes for potential vaccine candidates is essential for better management of UTI and further research has to be conducted in this area.


Asunto(s)
Infecciones por Escherichia coli/microbiología , Proteínas de Escherichia coli/genética , Genes Bacterianos , Filogenia , Infecciones Urinarias/microbiología , Escherichia coli Uropatógena/patogenicidad , Virulencia/genética , Adulto , Estudios Transversales , Etiopía , Femenino , Humanos , Masculino , Persona de Mediana Edad , Encuestas y Cuestionarios , Escherichia coli Uropatógena/aislamiento & purificación , Factores de Virulencia/genética
19.
Mikrobiyol Bul ; 54(1): 11-25, 2020 Jan.
Artículo en Turco | MEDLINE | ID: mdl-32050875

RESUMEN

The aim of this study was to investigate the frequency of Campylobacter species, to detect the antibiotic resistance profiles and the virulence genes and to determine the clonal proximity of the isolates in the samples of cutting board, slaughterhouse waste water, wall, knife and carcass from three different slaughterhouses in Kayseri region. For this purpose, a total of 150 samples, 10 of each from knife, wall, cutting board, carcass smear sample and slaughterhouse wastewater were collected from each of the three types of slaughterhouses in 2018 in Kayseri. For the isolation of the Campylobacter species, following preenrichment, the suspensions were inoculated onto modified charcoal cefoperazone desoxycholate (CCD) agar and were incubated at 37°C under microaerophilic condition for 48-72 hours. Suspicious colonies with gray-white color were recovered and subjected to phenotypical (Gram staining, oxidase, catalase test, and motion test) tests. Multiplex polymerase chain reaction (mPCR) was used for the molecular identification of the Campylobacter species. Antimicrobial susceptibilities of the isolates identified at the species level were detected by using the disk diffusion test and antibiotic gradient test. Virulence genes (iam, cadF, cdtA, flaA, ceuE, cdtC, cdtB and virB11) among the isolates were evaluated by PCR. The molecular typing of the isolates determined at species level was performed by Enterobacterial Repetitive Intergenic Consensus PCR (ERIC-PCR). In the study, 17 (11.3%) of the 150 samples taken from the slaughterhouse were found to be suspicious in terms of Campylobacter spp. and as a result of phenotypic identification tests, all of the isolates were verified as Campylobacter spp.. As a result of mPCR; eight of the isolates were identified as Campylobacter jejuni, eight as Campylobacter fetus and one as Campylobacter coli. The isolation of the Campylobacter species from different sources was found to be higher in slaughterhouse wastewater than those of others (p<0.001) and the difference in the proportional distribution of the Campylobacter species obtained from various sources was statistically significant (p<0.05). As a result of the disk diffusion test, while, all C.jejuni isolates were resistant to ciprofloxacin, 87.5%, 25%, 25% and 12.5% of C.jejuni isolates were resistant to enrofloxacin, neomycin, amoxicillin/clavulanic acid, and erythromycin, respectively. In addition, 25%, 25% and 12.5% of C.fetus isolates were resistant to amoxicillin/clavulanic acid, neomycin and gentamicin, respectively. C.coli isolate was not resistant to any of the antibiotics tested. Antibiotic gradient test results were found to be compatible with the disc diffusion test results. One of the virulence genes examined, virB11, was not detected in any of the isolates. Moreover, iam gene was not present in C.fetus and C.coli isolates, but only in one C.jejuni isolate. The flaA gene was detected in six C.jejuni isolates. C.coli isolate and seven C.jejuni and seven C.fetus isolates were positive in terms of the cdtC gene. The cdtA, cdtB, ceuE and cadF genes were found to be positive in all C.jejuni isolates. All isolates analyzed in the study demonstrated different ERIC-PCR profiles. In conclusion, it was shown that Campylobacter strains isolated from slaughterhouses were resistant to the most of the current antibiotics. Moreover, the presence of highly virulent Campylobacters in the slaughterhouse environment threatens public health due to the risk of contamination of the humans via carcasses and foods. Therefore, it is recommended that strict hygiene rules should be followed to reduce Campylobacter species contamination in slaughterhouses.


Asunto(s)
Campylobacter , Virulencia , Mataderos , Antibacterianos/farmacología , Campylobacter/efectos de los fármacos , Campylobacter/genética , Campylobacter/patogenicidad , Humanos , Especificidad de la Especie , Virulencia/genética
20.
Mikrobiyol Bul ; 54(1): 40-49, 2020 01.
Artículo en Turco | MEDLINE | ID: mdl-32050877

RESUMEN

Acinetobacter baumannii is a multi-drug resistant (MDR) gram-negative pathogen leading to nosocomial infections. Hospital-acquired infections due to A.baumannii occur especially in patients hospitalized in intensive care units. Important infections related to this bacterium are pneumonia, bacteremia, endocarditis, skin and soft tissue, urinary tract infections and meningitis. Human transmission is usually through the hospital environment or through medical personnel. A.baumannii isolates increases their virulence not only being multiple resistance to antibiotics but as well as the ability to form biofilm. The biofilm formation of A.baumannii isolates were mostly related with genes encoding curli fiber (csgA), the chaperone-usher fimbria (csuE) and the outer membrane (ompA). The aim of this study was to demonstrate biofilm production and virulence genes in MDR invasive A.baumannii isolates. MDR and similarity status previously known invasive A.baumannii (n= 156) isolates were included in the study. Biofilm production was determined by quantitative microplate biofilm method. Virulence genes csgA, csuE, fimH, ompA and blaPER-1 were investigated by polymerase chain reaction (PCR). It was determined that 60.3% (94/156) of all the isolates formed biofilm. Of these 94 isolates, 17 were weak, 33 were medium and 44 were strong. The mean biomass forming capacity of the isolates was found to be 2.23 ± 0.0033. Among the isolates included in the study (n= 156) the frequency of csgA, csuE, ompA, fimH and blaPER-1 genes were 71.2%, 32.1%, 21.8%, 7.1% and 3.2% respectively. The frequency of csgA, ompA, bap, csuE, fimH virulence genes were found to be 41.5%, 24.5%, 20.2% and 5.3% among biofilm positive isolates respectively. Biofilm-forming isolates were most commonly found in pulsotype II 19.1% (18/94), pulsotype IX 17.0% (16/94) and pulsotype VI 12.8% (12/94). In this study, when the distribution of virulence genes were compAred with the isolates that have weak, medium and strong biofilm, all of the studied genes were found to be more abundant in isolates with strong and medium positive biofilm production. This has shown that excluding fimH gene, csgA, csuE and ompA genes have contributed to the biofilm formation in invasive A.baumannii isolates, respectively.


Asunto(s)
Infecciones por Acinetobacter , Acinetobacter baumannii , Biopelículas , Farmacorresistencia Bacteriana Múltiple , Virulencia , Infecciones por Acinetobacter/microbiología , Acinetobacter baumannii/efectos de los fármacos , Acinetobacter baumannii/genética , Antibacterianos/farmacología , Farmacorresistencia Bacteriana Múltiple/genética , Humanos , Virulencia/genética
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