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1.
Microbiol Immunol ; 63(9): 350-358, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31407393

RESUMO

Kenya is endemic for cholera with different waves of outbreaks having been documented since 1971. In recent years, new variants of Vibrio cholerae O1 have emerged and have replaced most of the traditional El Tor biotype globally. These strains also appear to have increased virulence, and it is important to describe and document their phenotypic and genotypic traits. This study characterized 146 V. cholerae O1 isolates from cholera outbreaks that occurred in Kenya between 1975 and 2017. Our study reports that the 1975-1984 strains had typical classical or El Tor biotype characters. New variants of V. cholerae O1 having traits of both classical and El Tor biotypes were observed from 2007 with all strains isolated between 2015 and 2017 being sensitive to polymyxin B and carrying both classical and El Tor type ctxB. All strains were resistant to Phage IV and harbored rstR, rtxC, hlyA, rtxA and tcpA genes specific for El Tor biotype indicating that the strains had an El Tor backbone. Pulsed field gel electrophoresis (PFGE) genotyping differentiated the isolates into 14 pulsotypes. The clustering also corresponded with the year of isolation signifying that the cholera outbreaks occurred as separate waves of different genetic fingerprints exhibiting different genotypic and phenotypic characteristics. The emergence and prevalence of V. cholerae O1 strains carrying El Tor type and classical type ctxB in Kenya are reported. These strains have replaced the typical El Tor biotype in Kenya and are potentially more virulent and easily transmitted within the population.


Assuntos
Cólera/epidemiologia , Cólera/microbiologia , Surtos de Doenças , Vibrio cholerae O1/classificação , Vibrio cholerae O1/genética , Vibrio cholerae O1/isolamento & purificação , Proteínas de Bactérias/genética , Técnicas de Tipagem Bacteriana/métodos , Toxina da Cólera/genética , DNA Bacteriano/genética , Genótipo , Técnicas de Genotipagem , Humanos , Quênia/epidemiologia , Testes de Sensibilidade Microbiana , Fenótipo , Polimixina B/farmacologia , Vibrio cholerae O1/efeitos dos fármacos , Virulência/genética , Fatores de Virulência/genética
2.
Rev Soc Bras Med Trop ; 52: e20190095, 2019 Jul 18.
Artigo em Inglês | MEDLINE | ID: mdl-31340369

RESUMO

INTRODUCTION: Staphylococcus aureus is a major nosocomial pathogen that is associated with high virulence and the rapid development of drug resistance. METHODS: We analyzed and compared the antimicrobial resistance, virulence profiles, and molecular epidemiology of 67 S. aureus strains, including 36 methicillin-sensitive (MSSA) and 31 methicillin-resistant (MRSA) strains recovered from a public hospital located in south-eastern Brazil. RESULTS: The clones circulating in this hospital presented a great diversity, and the majority of the strains were related to clones responsible for causing worldwide epidemics: these included USA100 (New York/Japan clone), USA300, and USA600. The 31 MRSA (22 SCCmecII and 9 SCCmecIV) and 36 MSSA strains exhibited low resistance against gentamicin and trimethoprim/sulfamethoxazole. No MRSA strain showed resistance to tetracycline. Virulence gene carriage was more diverse and abundant in MSSA than in MRSA. Of the evaluated adhesion-related genes, ebpS was the most prevalent in both MSSA and MRSA strains. The genes bbp and cna showed a strong association with MSSA strains. CONCLUSIONS: Our findings reinforce the idea that MSSA and MRSA strains should be carefully monitored, owing to their high pathogenic potential.


Assuntos
Antibacterianos/farmacologia , Resistência a Meticilina , Staphylococcus aureus Resistente à Meticilina/genética , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/genética , Fatores de Virulência/genética , Brasil/epidemiologia , Hospitais Públicos , Humanos , Testes de Sensibilidade Microbiana , Epidemiologia Molecular , Infecções Estafilocócicas/epidemiologia , Staphylococcus aureus/classificação , Staphylococcus aureus/isolamento & purificação , Centros de Atenção Terciária , Virulência/genética
3.
Plant Dis ; 103(9): 2252-2262, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31298990

RESUMO

Spot blotch, caused by the fungal pathogen Cochliobolus sativus, is a limiting factor for barley (Hordeum vulgare) production in northeast China, which causes significant grain yield losses and kernel quality degradation. It is critical to determine the virulence diversity of C. sativus populations for barley resistance breeding and the judicious grouping of available resistance varieties according to the predominant pathotypes in disease epidemic regions. With little information on the barley pathogen in China, this study selected 12 typical barley genotypes to differentiate the pathotypes of C. sativus isolates collected in China. Seventy-one isolates were grouped into 19 Chinese pathotypes based on infection responses. Seventeen isolates were classified as pathotype 3, which has only been identified in China, whereas most (52 of 71) were classified as pathotype 1. All of the tested isolates had low virulence on the North Dakota (ND) durable, resistant line ND B112. Using 22 selected amplified fragment-length polymorphism (AFLP) primer combinations, genetic polymorphism was used to analyze 68 isolates, which clustered into three distinct groups using the unweighted pair group method average with the genetic distance coefficient. No relationship was found between the virulence of isolates and their origins. Isolates of the same pathotype or those collected from the same location did not group into clusters based on the AFLP analysis.


Assuntos
Ascomicetos , Variação Genética , Hordeum , Virulência , Análise do Polimorfismo de Comprimento de Fragmentos Amplificados , Ascomicetos/classificação , Ascomicetos/genética , China , Hordeum/microbiologia , Doenças das Plantas/microbiologia , Polimorfismo Genético , Virulência/genética
4.
Plant Dis ; 103(9): 2451-2459, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31322491

RESUMO

Stripe rust, caused by Puccinia striiformis f. sp. tritici, is one of the most devastating wheat diseases in Ethiopia. To study virulence genetics of the pathogen, 117 progeny isolates were produced through sexual reproduction of an Ethiopian isolate of the stripe rust pathogen on Berberis holstii plants under controlled conditions. The parental and progeny isolates were characterized by phenotyping on wheat lines carrying single Yr genes for resistance and genotyped using 10 polymorphic simple sequence repeated (SSR) markers. The progeny isolates were classified into 37 virulence phenotypes and 75 multilocus genotypes. The parental isolate and progeny isolates were all avirulent to resistance genes Yr5, Yr10, Yr15, Yr24, Yr32, YrTr1, YrSP, and Yr76 but virulent to Yr1 and Yr2, indicating that the parental isolate was homozygous avirulent or homozygous virulent at these loci. The progeny isolates segregated for virulence to 12 Yr genes. Virulence phenotypes to Yr6, Yr28, Yr43, and Yr44 were controlled by a single dominant gene; those to Yr7, Yr9, Yr17, Yr27, Yr25, Yr31, and YrExp2 were each controlled by two dominant genes; and the virulence phenotype to Yr8 was controlled by two complementary dominant genes. A linkage map was constructed with seven SSR markers, and 16 virulence loci corresponding to 11 Yr resistance genes were mapped with some loci linked to each other. These results are useful in understanding host-pathogen interactions and selecting resistance genes to develop wheat cultivars with highly effective resistance to stripe rust.


Assuntos
Basidiomycota , Berberis , Ligação Genética , Recombinação Genética , Virulência , Basidiomycota/genética , Basidiomycota/patogenicidade , Berberis/genética , Etiópia , Doenças das Plantas , Triticum/microbiologia , Virulência/genética
5.
BMC Vet Res ; 15(1): 235, 2019 Jul 08.
Artigo em Inglês | MEDLINE | ID: mdl-31286947

RESUMO

BACKGROUND: Enterococcus is an important component of normal flora in human and animals, but in recent years, the pathogenicity of Enterococcus has been confirmed in clinical medicine. More and more animal infections have been reported in veterinary clinics. For the last decades, outbreaks of encephalitis in lambs have become much more common in Northern Xinjiang, China. Consequent studies have confirmed that these affected lambs had been commonly infected with E. faecalis. More than 60 E. faecalis were isolated from the brain of infected lambs, A highly virulent strain entitled E. faecalis 2A (XJ05) were selected, sequenced and analyzed. RESULT: Using whole genome sequence and de novo assembly, 18 contigs with NGS and annotation were obtained. It is confirmed that the genome has a size of 2.9 Mb containing 2783 protein-coding genes, as well as 54 tRNA genes and 4 rRNA genes. Some key features of this strain were identified, which included 7 predicted antibiotic resistance genes and 18 candidate virulence factor genes. CONCLUSION: The E. faecalis 2A (XJ05) genome is conspicuous smaller than E.faecalis V583, but not significantly different from other non-pathogenic E. faecalis. It carried 7 resistance genes including 4 kind of antibiotics which were consistent with the results of extensive drug resistance phenotypic, including aminoglycoside, macrolide, phenicol, and tetracycline. 2A (XJ05) also carried 18 new virulence factor genes related to virulence, hemolysin genes (cylA, cylB, cylM, cylL) may play an important role in lamb encephalitis by E. faecalis 2A (XJ05).


Assuntos
Farmacorresistência Bacteriana/genética , Encefalite/veterinária , Enterococcus faecalis/genética , Enterococcus faecalis/patogenicidade , Genoma Bacteriano/genética , Doenças dos Ovinos/microbiologia , Virulência/genética , Animais , Resistência a Múltiplos Medicamentos/genética , Encefalite/microbiologia , Ovinos
6.
Microbiol Res ; 226: 48-54, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31284944

RESUMO

The Burkholderia pseudomallei complex consists of six phylogenetically related Gram-negative bacterial species that include environmental saprophytes and mammalian pathogens. These microbes possess multiple type VI secretion systems (T6SS) that provide a fitness advantage in diverse niches by translocating effector molecules into prokaryotic and eukaryotic cells in a contact-dependent manner. Several recent studies have elucidated the regulation and function of T6SS-2, a novel contact-independent member of the T6SS family. Expression of the T6SS-2 gene cluster is repressed by OxyR, Zur and TctR and is activated by GvmR and reactive oxygen species (ROS). The last two genes of the T6SS-2 gene cluster encode a zincophore (TseZ) and a manganeseophore (TseM) that are exported into the extracellular milieu in a contact-independent fashion when microbes encounter oxidative stress. TseZ and TseM bind Zn2+ and Mn2+, respectively, and deliver them to bacteria where they provide protection against the lethal effects of ROS. The TonB-dependent transporters that interact with TseZ and TseM, and actively transport Zn2+ and Mn2+ across the outer membrane, have also been identified. Finally, T6SS-2 provides a contact-independent growth advantage in nutrient limited environments and is critical for virulence in Galleria mellonella larvae, but is dispensable for virulence in rodent models of infection.


Assuntos
Proteínas de Bactérias/genética , Burkholderia pseudomallei/genética , Burkholderia pseudomallei/metabolismo , Manganês/metabolismo , Sistemas de Secreção Tipo VI/genética , Sistemas de Secreção Tipo VI/metabolismo , Zinco/metabolismo , Animais , Proteínas de Bactérias/metabolismo , Burkholderia pseudomallei/classificação , Regulação Bacteriana da Expressão Gênica , Genes Reguladores/genética , Homeostase , Larva , Proteínas de Membrana Transportadoras/genética , Metiltransferases , Família Multigênica , Oxirredução , Estresse Oxidativo , Espécies Reativas de Oxigênio/metabolismo , Virulência/genética
7.
Microbiol Res ; 226: 55-64, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31284945

RESUMO

Functional association between genomic loci and specific biological traits remains lacking in many fungi, including the African tree pathogen Ceratocystis albifundus. This is mainly because of the absence of suitable transformation systems for allowing genetic manipulation of this and other fungi. Here, we present an optimized protocol for Agrobacterium tumefaciens-mediated transformation of C. albifundus. Strain AGL-1 of A. tumefaciens and four binary T-DNA vectors (conferring hygromycin B or geneticin resistance and/or expressing the green fluorescent protein [GFP]) were used for transforming germinated conidia of three isolates of C. albifundus. Stable expression of these T-DNA-encoded traits was confirmed through sequential sub-culturing of fungal transformants on selective and non-selective media and by using PCR and sequence analysis. Single-copy integration of the respective T-DNAs into the genomes of these fungi was confirmed using Southern hybridization analysis. The range of experimental parameters determined and optimised included: (i) concentrations of hygromycin B and geneticin required for inhibiting growth of the wild type fungus and (ii) the dependence of transformation on acetosyringone for inducing the bacterium's virulence genes, as well as (iii) the duration of fungus-bacterium co-cultivation periods and (iv) the concentrations of fungal conidia and bacterial cells used for the latter. The system developed in this study is stable with a high-efficiency, yielding up to 400 transformants per 106 conidia. This is the first report of a transformation protocol for C. albifundus and its availability will be invaluable for functional studies in this important fungus.


Assuntos
Agrobacterium tumefaciens/genética , Ascomicetos/genética , Transformação Genética , Ascomicetos/citologia , Ascomicetos/efeitos dos fármacos , Ascomicetos/crescimento & desenvolvimento , Southern Blotting , Carbenicilina/farmacologia , Técnicas de Cocultura , DNA Bacteriano , Regulação Fúngica da Expressão Gênica , Gentamicinas/farmacologia , Proteínas de Fluorescência Verde/genética , Higromicina B/farmacologia , Canamicina/farmacologia , Reação em Cadeia da Polimerase , Análise de Sequência , Virulência/genética
8.
Rev Inst Med Trop Sao Paulo ; 61: e36, 2019 Jul 22.
Artigo em Inglês | MEDLINE | ID: mdl-31340248

RESUMO

During the last years, Brazilian government control programs have detected an increase of Salmonella Heidelberg in poultry slaughterhouses a condition that poses a threat to human health However, the reasons remain unclear. Differences in genetic virulence profiles may be a possible justification. In addition, effective control of Salmonella is related to an efficient epidemiological surveillance system through genotyping techniques. In this context, the aim of this study was the detection of 24 virulence-associated genes in 126 S. Heidelberg isolates. We classified the isolates into 56 different genetic profiles. None of the isolates presented all the virulence genes. The prevalence of these genes was high in all tested samples as the lowest number of genes detected in one isolate was 10/24. The lpfA and csgA (fimbriae), invA and sivH (TTSS), and msgA and tolC (intracellular survival) genes were present in 100% of the isolates analyzed. Genes encoding effector proteins were detected in the majority of SH isolates. No single isolate had the sefA gene. The pefA gene was found in only four isolates. We have also performed a screening of genes associated with iron metabolism: 88.9% of isolates had the iroN geneand 79.4% the sitC gene . Although all the isolates belong to the same serotype, several genotypic profiles were observed. These findings suggest that there is a diversity of S. Heidelberg isolates in poultry products. The fact that a single predominant profile was not found in this study indicates the presence of variable sources of contamination caused by SH. The detection of genetic profiles of Salmonella strains can be used to determine the virulence patterns of SH isolates.


Assuntos
Doenças das Aves Domésticas/microbiologia , Produtos Avícolas/microbiologia , Salmonelose Animal/microbiologia , Salmonella/genética , Salmonella/patogenicidade , Fatores de Virulência/genética , Virulência/genética , Animais , Microbiologia de Alimentos , Genótipo , Reação em Cadeia da Polimerase
9.
J Med Microbiol ; 68(9): 1320-1323, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31329091

RESUMO

The recent increase in pertussis cases observed in some countries may have several causes, including the evolution of Bordetella pertussis populations towards escape of vaccine-induced immunity. Most genomic studies of B. pertussis isolates performed so far are from countries that use acellular vaccines. The objective was to analyse genomic sequences of isolates collected during the 2014 whooping cough epidemic in Tunisia, a country where whole-cell vaccines are used. Ten Tunisian isolates and four vaccine strains were sequenced and compared to 169 isolates from countries where acellular vaccines are used. Phylogenetic analysis showed that Tunisian isolates are diverse, demonstrating a multi-strain 2014 epidemic peak, and are intermixed with those circulating in other world regions, showing inter-country transmission. Consistently, Tunisian isolates have antigen variant composition observed in other world regions. No pertactin-deficient strain was observed. The Tunisian B. pertussis population appears to be largely connected with populations from other countries.


Assuntos
Bordetella pertussis/genética , Variação Genética , Genoma Bacteriano/genética , Filogenia , Coqueluche/microbiologia , Antígenos de Bactérias/genética , Proteínas da Membrana Bacteriana Externa/genética , Bordetella pertussis/classificação , Bordetella pertussis/imunologia , Bordetella pertussis/isolamento & purificação , DNA Bacteriano/genética , Humanos , Lactente , Recém-Nascido , Epidemiologia Molecular , Vacina contra Coqueluche/administração & dosagem , Vacina contra Coqueluche/genética , Análise de Sequência de DNA , Tunísia/epidemiologia , Virulência/genética , Fatores de Virulência de Bordetella/genética , Coqueluche/epidemiologia , Coqueluche/transmissão
10.
Int J Food Microbiol ; 306: 108269, 2019 Oct 02.
Artigo em Inglês | MEDLINE | ID: mdl-31330452

RESUMO

Salmonella enterica serovar Enteritidis strain SE86 has been associated with several foodborne diseases occurring in Southern Brazil, becoming and important causative agent of human salmonellosis. In this work, the complete genome of the bacterium Salmonella Enteritidis SE86 was sequenced using the Illumina MiSeq platform. An in silico analysis of the SE86 genome was performed in order to compare it with different Salmonella strains as well as to investigate the presence of stress-resistance and virulence genes. This strain showed a variety of genes that can be involved in antimicrobial and biocide resistance, acid and thermal resistance as well as virulence and adhesion. These genetic features could explain its increased resistance and the prevalence of this strain in foodborne outbreaks in Southern Brazil.


Assuntos
Antibacterianos/farmacologia , Intoxicação Alimentar por Salmonella/epidemiologia , Salmonella enteritidis , Brasil/epidemiologia , Surtos de Doenças , Genoma Bacteriano/genética , Humanos , Testes de Sensibilidade Microbiana , Intoxicação Alimentar por Salmonella/microbiologia , Salmonella enteritidis/efeitos dos fármacos , Salmonella enteritidis/genética , Salmonella enteritidis/patogenicidade , Virulência/genética , Fatores de Virulência/genética
11.
Vet Res ; 50(1): 43, 2019 Jun 04.
Artigo em Inglês | MEDLINE | ID: mdl-31164171

RESUMO

Riemerella anatipestifer is a major pathogenic agent of duck septicemic and exudative diseases. Genetic analyses suggest that this pathogen has a novel protein secretion system, known as the "type IX secretion system" (T9SS). We previously reported that deletion of the AS87_RS08465 gene significantly reduced the bacterial virulence of the R. anatipestifer strain Yb2, but the mechanism remained unclear. The AS87_RS08465 gene is predicted to encode the gliding motility protein GldM (GldM) protein, a key component of the T9SS complex. In this study, Western blotting analysis demonstrated that R. anatipestifer GldM was localized to the cytomembrane. Further study revealed that the adhesion and invasion capacities of the mutant strain RA2281 (designated Yb2ΔgldM) in Vero cells and the bacterial loads in the blood of infected ducks were significantly reduced. RNA-Seq and PCR analyses showed that six genes were upregulated and five genes were downregulated in the mutant strain Yb2ΔgldM and that these genes were mainly involved in the secretion of proteins. Yb2ΔgldM was also found to be defective in gliding motility and protein secretion. Liquid chromatography-tandem mass spectrometry analysis revealed that nine of the proteins had a conserved T9SS C-terminal domain and were differentially secreted by Yb2ΔgldM compared to Yb2. The complementation strain cYb2ΔgldM recovered the adhesion and invasion capacities in Vero cells and the bacterial loads in the blood of infected ducks as well as the bacterial gliding motility and most protein secretion in the mutant strain Yb2ΔgldM to the levels of the wild-type strain Yb2. Taken together, these results indicate that R. anatipestifer GldM is associated with T9SS and is important in bacterial virulence.


Assuntos
Aderência Bacteriana/genética , Expressão Gênica , Riemerella/genética , Riemerella/patogenicidade , Sistemas de Secreção Tipo IV/genética , Mutação , Peptídeo Hidrolases/biossíntese , Riemerella/enzimologia , Sistemas de Secreção Tipo IV/metabolismo , Virulência/genética , Fatores de Virulência/genética
12.
Microbiol Res ; 223-225: 63-71, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31178053

RESUMO

The phosphoenolpyruvate (PEP):carbohydrate phosphotransferase system (PTS) catalyzes the translocation of sugar substrates with their concomitant phosphorylation in bacteria. In addition to its intrinsic role in sugar transport and metabolism, numerous recent studies report the versatility of the PTS to interconnect energy and signal transduction in response to sugar availability. In this study, the role of PTS in Salmonella virulence regulation was explored. To decipher the regulatory network coordinated by the PTS during Salmonella infection, a transcriptomic approach was applied to a transposon insertion mutant with defective expression of ptsI and crr, which encode enzyme I and enzyme IIAGlc of the PTS, respectively. There were 114 differentially expressed genes (DEGs) exhibiting two-fold or higher expression changes in the transposon mutant strain, with 13 up-regulated genes versus 101 down-regulated genes. One-third of the DEGs were associated with energy production and carbohydrate/amino acid metabolism pathways, implicating the prominent role of the PTS in carbohydrate transport. With regard to regulation of virulence, the tested mutant decreased the expression of genes associated with quorum sensing, Salmonella pathogenicity islands, flagella, and the PhoPQ regulon. We investigated the possibility of PTS-mediated regulation of virulence determinants identified in the transcriptomic analysis and proposed a regulatory circuit orchestrated by the PTS in Salmonella infection of host cells. These results suggest that Salmonella divergently controls virulence attributes in accordance with the availability of carbohydrates in the environment.


Assuntos
Proteínas de Bactérias/genética , Regulação Bacteriana da Expressão Gênica , Fosfoenolpiruvato/metabolismo , Fosfotransferases/metabolismo , Salmonella/genética , Salmonella/metabolismo , Fatores de Virulência/genética , Transporte Biológico , Elementos de DNA Transponíveis , Flagelos/genética , Perfilação da Expressão Gênica , Proteínas de Membrana/genética , Mutação , Fosforilação , Regulon , Salmonella/patogenicidade , Infecções por Salmonella , Salmonella typhimurium/genética , Transdução de Sinais , Transcriptoma , Sistemas de Secreção Tipo III/genética , Virulência/genética
13.
Microbiol Res ; 223-225: 72-78, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31178054

RESUMO

Pseudomonas syringae pathovars are known to produce N-acyl-homoserine lactones (AHL) as quorum-sensing molecules. However, many isolates, including P. syringae pv. tomato DC3000 (PtoDC3000), do not produce them. In P. syringae, psyI, which encodes an AHL synthase, and psyR, which encodes the transcription factor PsyR required for activation of psyI, are convergently transcribed. In P. amygdali pv. tabaci 6605 (Pta6605), there is one nucleotide between the stop codons of both psyI and psyR. However, the canonical stop codon for psyI in PtoDC3000 was converted to the cysteine codon by one nucleotide deletion, and 23 additional amino acids extended it to a C-terminal end. This resulted in overlapping of the open reading frame (ORF) for psyI and psyR. On the other hand, stop codons in the psyR ORF of P. syringae 7 isolates, including pv. phaseolicola and pv. glycinea, were found. These results indicate that many pathovars of P. syringae have genetically lost AHL production ability by the mutation of their responsible genes. To examine whether PtoDC3000 modulates the gene expression profile in a population-dependent manner, we carried out microarray analysis using RNAs prepared from low- and high-density cells. We found the expressions of rsmX and rsmY remarkably activated in high-density cells. The activated expressions of rsmX and rsmY were confirmed by Northern blot hybridization, but these expressions were abolished in a ΔgacA mutant of Pta6605. These results indicate that regardless of the ability to produce AHL, P. syringae regulates expression of the small noncoding RNAs rsmX/Y by currently unknown quorum-sensing molecules.


Assuntos
Proteínas de Bactérias/genética , Regulação Bacteriana da Expressão Gênica , Genes Bacterianos/genética , Pseudomonas syringae/genética , Pseudomonas syringae/metabolismo , Pequeno RNA não Traduzido/genética , Acil-Butirolactonas/metabolismo , Doenças das Plantas/microbiologia , Pseudomonas syringae/enzimologia , Pseudomonas syringae/patogenicidade , Percepção de Quorum/genética , Análise de Sequência de DNA , Fatores de Transcrição/genética , Transcriptoma , Virulência/genética
14.
Microbiol Res ; 223-225: 79-87, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31178055

RESUMO

Vibrio parahaemolyticus is a seafood-borne Gram-negative bacteria causing diarrheal diseases in humans world wide. ToxR is a membrane-associated transcriptional factor which plays an important role in acid stress tolerance and regulates the expression of virulence genes including type III secretion system 1 (T3SS1) and type VI secretion system 1 (T6SS1) in V. parahaemolyticus. However, possible mechanisms of ToxR mediating virulence gene expression have not been fully understood. In this study, we demonstrated that ToxR is essential for V. parahaemolyticus to tolerate acid stress by constructing a ToxR deletion mutant (ΔtoxR) and its complemented strain (toxR+). Quantitative PCR showed that the expression of toxR was up regulated under acid stress condition. RNA-seq analysis showed that ompU encoding one of outer membrane proteins was dramatically down regulated in ΔtoxR. Furthermore, the mutation of ompU also led to a significant reduction in tolerating acid stress indicating that ToxR mediated acid stress through regulating ompU expression. RNA-seq results further confirmed that acid stress condition could alter multiple signaling pathways either depending on ToxR (e.g., quorum sensing, fatty acid metabolism) or independent of ToxR (e.g., biosynthesis of secondary metabolites, microbial metabolism in diverse environment, biosynthesis of antibiotics, biosynthesis of amino acids and carbon metabolism pathways). We also for the first time demonstrated that ToxR positively regulated the expression of T6SS2 gene and the interbacteria killing activity. Our study provides comprehensive understanding of signaling pathways which are regulated by both acid stress and ToxR.


Assuntos
Proteínas de Bactérias/metabolismo , Proteínas de Ligação a DNA/metabolismo , Regulação Bacteriana da Expressão Gênica , Estresse Fisiológico , Fatores de Transcrição/metabolismo , Vibrio parahaemolyticus/metabolismo , Adesinas Bacterianas/genética , Proteínas da Membrana Bacteriana Externa/genética , Proteínas da Membrana Bacteriana Externa/metabolismo , Proteínas de Bactérias/genética , Proteínas de Ligação a DNA/genética , Ácidos Graxos/metabolismo , Percepção de Quorum , Metabolismo Secundário , Análise de Sequência de RNA , Deleção de Sequência , Fatores de Transcrição/genética , Transcriptoma , Sistemas de Secreção Tipo III/genética , Sistemas de Secreção Tipo VI/genética , Vibrio parahaemolyticus/genética , Vibrio parahaemolyticus/crescimento & desenvolvimento , Virulência/genética
15.
Microbiol Res ; 223-225: 99-109, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31178057

RESUMO

Streptococcus suis has received increasing attention for its involvement in severe infections in pigs and humans; however, their pathogenesis remains unclear. ClpX and ClpP, two subunits of the ATP-dependent caseinolytic protease Clp, play key roles in bacterial adaptation to various environmental stresses. In this study, a virulent S. suis serotype 2 strain, ZY05719, was employed to construct clpX and clpP deletion mutants (ΔclpX and ΔclpP, respectively) and their complementation strains. Both ΔclpX and ΔclpP displayed significantly reduced adaptability compared with the wild-type strain, evident through several altered phenotypes: formation of long cell chains, tendency to aggregate in culture, and reduced growth under acidic pH and H2O2-induced oxidative stress. ClpP and ClpX were required for the optimal growth during heat and cold stress, respectively. An in vitro experiment on RAW264.7 macrophage cells showed significantly increased sensitivity of ΔclpX and ΔclpP to phagocytosis compared with the wild-type strain. Mouse infection assays verified the deletion of clpX and clpP led to not only fewer clinical symptoms and lower mortality but also to a marked attenuation in bacterial colonization. These virulence-related phenotypes were restored by genetic complementation. Furthermore, the deletion of clpX or clpP caused a significant decrease in the expression of sodA, tpx, and apuA compared with the wild-type strain, suggesting that these genes may be regulated by ClpX and ClpP as downstream response factors to facilitate the bacterial tolerance against various environmental stresses. Taken together, these results suggest that ClpX and ClpP play important roles in stress tolerance for achieving the full virulence of S. suis serotype 2 during infection.


Assuntos
ATPases Associadas a Diversas Atividades Celulares/metabolismo , Proteínas de Bactérias/metabolismo , Endopeptidase Clp/metabolismo , Chaperonas Moleculares/metabolismo , Streptococcus suis/metabolismo , Streptococcus suis/patogenicidade , Animais , Proteínas de Bactérias/genética , Biofilmes , Resposta ao Choque Frio , Endopeptidase Clp/genética , Deleção de Genes , Regulação Bacteriana da Expressão Gênica , Resposta ao Choque Térmico , Peróxido de Hidrogênio/metabolismo , Concentração de Íons de Hidrogênio , Camundongos , Chaperonas Moleculares/genética , Pressão Osmótica , Estresse Oxidativo , Fagocitose , Células RAW 264.7 , Infecções Estreptocócicas/microbiologia , Streptococcus suis/genética , Streptococcus suis/crescimento & desenvolvimento , Transcriptoma , Virulência/genética , Fatores de Virulência/genética , Fatores de Virulência/fisiologia
16.
J Microbiol Biotechnol ; 29(6): 962-972, 2019 Jun 28.
Artigo em Inglês | MEDLINE | ID: mdl-31216839

RESUMO

Non-typhoidal Salmonella (NTS) is one of the most frequent causes of bacterial foodborne illnesses. Considering that the main reservoir of NTS is the intestinal tract of livestock, foods of animal origin are regarded as the main vehicles of Salmonella infection. In particular, poultry colonized with Salmonella Typhimurium (S. Typhimurium), a dominant serotype responsible for human infections, do not exhibit overt signs and symptoms, thereby posing a potential health risk to humans. In this study, comparative genomics approaches were applied to two S. Typhimurium strains, ST1539 and ST1120, isolated from a duck slaughterhouse and a pig farm, respectively, to characterize their virulence and antimicrobial resistance-associated genomic determinants. ST1539 containing a chromosome (4,905,039 bp; 4,403 CDSs) and a plasmid (93,876 bp; 96 CDSs) was phylogenetically distinct from other S. Typhimurium strains such as ST1120 and LT2. Compared to the ST1120 genome (previously deposited in GenBank; CP021909.1 and CP021910.1), ST1539 possesses more virulence determinants, including ST64B prophage, plasmid spv operon encoding virulence factors, genes encoding SseJ effector, Rck invasin, and biofilm-forming factors (bcf operon and pefAB). In accordance with the in silico prediction, ST1539 exhibited higher cytotoxicity against epithelial cells, better survival inside macrophage cells, and faster mice-killing activity than ST1120. However, ST1539 showed less resistance against antibiotics than ST1120, which may be attributed to the multiple resistanceassociated genes in the ST1120 chromosome. The accumulation of comparative genomics data on S. Typhimurium isolates from livestock would enrich our understanding of strategies Salmonella employs to adapt to diverse host animals.


Assuntos
Farmacorresistência Bacteriana , Genoma Bacteriano/genética , Aves Domésticas , Salmonelose Animal/microbiologia , Salmonella typhimurium/genética , Salmonella typhimurium/patogenicidade , Matadouros , Animais , Antibacterianos/farmacologia , Proteínas de Bactérias/genética , Sobrevivência Celular , Farmacorresistência Bacteriana/genética , Feminino , Células HeLa , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Filogenia , Plasmídeos/genética , Prófagos/genética , República da Coreia , Salmonella typhimurium/classificação , Salmonella typhimurium/isolamento & purificação , Sorogrupo , Suínos , Virulência/genética
17.
Plant Physiol Biochem ; 141: 332-342, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31207494

RESUMO

Plant-parasitic nematodes cause major agricultural losses worldwide. Examining the molecular mechanisms underlying plant-nematode interactions and how plants respond to different invading pathogens is attracting major attention to reduce the expanding gap between agricultural production and the needs of the growing world population. This review summarizes the most recent developments in plant-nematode interactions and the diverse approaches used to improve plant resistance against root knot nematode (RKN). We will emphasize the recent rapid advances in genome sequencing technologies, small interfering RNA techniques (RNAi) and targeted genome editing which are contributing to the significant progress in understanding the plant-nematode interaction mechanisms. Also, molecular approaches to improve plant resistance against nematodes are considered.


Assuntos
Interações Hospedeiro-Parasita , Nematoides/patogenicidade , Raízes de Plantas/parasitologia , Plantas/parasitologia , Animais , Mapeamento Cromossômico , Biologia Computacional/métodos , Feminino , Genoma de Planta , Masculino , Doenças das Plantas/parasitologia , Fenômenos Fisiológicos Vegetais , Raízes de Plantas/genética , Plantas/genética , Plantas Geneticamente Modificadas/parasitologia , Locos de Características Quantitativas , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Transcriptoma , Virulência/genética
18.
J Vet Sci ; 20(3): e26, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-31161744

RESUMO

Enterococcus spp. are opportunistic pathogens that cause lameness in broiler chickens, resulting in serious economic losses worldwide. Virulence of Enterococcus spp. is associated with several putative virulence genes including fsr, efm, esp, cylA, cad1, ace, gelE, and asa1. In this study, multiplex polymerase chain reaction (PCR) for the simultaneous detection of these virulence genes in Enterococcus spp. was developed, and detection limits for E. faecium, E. faecalis, and E. hirae were 64.0 pg/µL, 320.0 pg/µL, and 1.6 ng/µL DNA, respectively. Among 80 Enterococcus isolates tested, efm and cad1 were detected in all 26 E. faecium samples, and only cad1 was observed in E. hirae. Additionally, the presence of virulence genes in 25 E. faecalis isolates were 100% for cad1, 88.0% for gelE, 64.0% for fsr, 44.0% for asa1, 16.0% for cylA, and 4.0% for esp. No virulence genes were found in E. gallinarum isolates. A total of 49 isolates were resistant to tigecycline and to at least 2 different classes of antibiotics. The most prevalent resistance was to ciprofloxacin (73.5%), quinupristin/dalfopristin (55.1%), and tetracycline (49.0%). No strains were resistant to vancomycin or linezolid. This is the first multiplex PCR assay to simultaneously detect eight virulence genes in Enterococcus spp., and the method provides diagnostic value for accurate, rapid, and convenient detection of virulence genes. Additionally, we report the prevalence of virulence genes and antimicrobial resistance in Enterococcus isolates from commercial broiler chickens suffering lameness.


Assuntos
Farmacorresistência Bacteriana/genética , Enterococcus/genética , Genes Bacterianos/genética , Reação em Cadeia da Polimerase Multiplex/veterinária , Virulência/genética , Animais , Galinhas , Resistência Microbiana a Medicamentos/genética , Limite de Detecção , Reação em Cadeia da Polimerase Multiplex/normas
19.
J Appl Microbiol ; 127(3): 897-910, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31173435

RESUMO

AIMS: To elucidate the antibiotic resistance and virulence genes of nisin-resistant Enterococcus faecalis isolated from raw buffalo milk and to study the effect of nisin-sensitive and -resistant E. faecalis on the innate immunity of rats. METHODS AND RESULTS: Slanetz-Bartley agar plates containing nisin were used to isolate nisin-resistant E. faecalis. The virulence factors were ascertained using quantitative real-time polymerase chain reaction. Cell viability, phagocytosis, intracellular survival and enzyme assays were performed to investigate the interaction of E. faecalis with rat macrophages. Nisin-resistant E. faecalis was less prone to phagocytosis and survived longer inside the macrophages, due to reduced production of reactive oxygen species and nitric oxide. The viability and activation of macrophages was also reduced in the presence of resistant E. faecalis, as observed by enhanced lactate dehydrogenase production and reduced ß-galactosidase. CONCLUSIONS: Nisin-resistant E. faecalis and its virulence factors were reported in raw buffalo milk. This study shows that nisin-resistant variants exhibited cross resistance to antibiotics and suppressed the innate immune responses of rats by directly affecting macrophage activity. SIGNIFICANCE AND IMPACT OF THE STUDY: This study elucidated the contamination of raw buffalo milk by nisin-resistant E. faecalis, which may pose food safety risk.


Assuntos
Enterococcus faecalis/genética , Macrófagos/efeitos dos fármacos , Leite/microbiologia , Animais , Búfalos , Farmacorresistência Bacteriana/genética , Enterococcus faecalis/efeitos dos fármacos , Enterococcus faecalis/isolamento & purificação , Macrófagos/metabolismo , Macrófagos/fisiologia , Testes de Sensibilidade Microbiana , Nisina/farmacologia , Ratos , Espécies Reativas de Oxigênio/metabolismo , Virulência/genética , Fatores de Virulência/genética
20.
Vet Microbiol ; 233: 190-195, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31176407

RESUMO

This study describes the prevalence of capsule biosynthesis genes, LPS genotypes, virulence associated genes and the analysis of the outer membrane protein (ompA) sequence of Pasteurella multocida isolates (n = 180) from different locations in Hungary, from various host species, including humans. When combining capsular types with LPS genotypes, eight capsule - LPS genotype combinations were detected. A: L3 was the most dominant in bovine and porcine isolates, A: L1 in feline and human isolates, while D: L3 was the most common among strains from small ruminants. The P. multocida toxin encoding gene toxA was highly prevalent among small ruminant and porcine strains, while in human, feline and bovine isolates it could not be detected. Combination of the tested virulence associated genes (hgbA, nanH, hgbB, tbpA, pfhA, hsf1, hsf2, tadD, ptfA) classified our P. multocida isolates into 13 different virulence gene profiles (VGPs). These VGPs showed an association with host species. Analysis of the ompA sequence data confirmed this distribution by host species, which may indicate that host adaptation is taking place. The typing scheme used in this study may be useful in epidemiological investigations.


Assuntos
Proteínas da Membrana Bacteriana Externa/genética , Infecções por Pasteurella/veterinária , Pasteurella multocida/genética , Fatores de Virulência/genética , Animais , Gatos , Bovinos , Perfilação da Expressão Gênica , Genes Bacterianos , Variação Genética , Especificidade de Hospedeiro , Humanos , Hungria/epidemiologia , Tipagem Molecular , Infecções por Pasteurella/epidemiologia , Pasteurella multocida/patogenicidade , Reação em Cadeia da Polimerase , Ruminantes/microbiologia , Suínos , Virulência/genética
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