Molecular characterization of the rpoB gene in Brucella species: new potential molecular markers for genotyping

Marianelli, Cinzia; Ciuchini, F.; Tarantino, Michela; Pasquali, Paolo; Adone, Rosanna

doi:10.1016/j.micinf.2005.10.008

Cited by 47 publications

(61 citation statements)

References 33 publications

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“…In contrast, rpoB sequencing, recently proposed as a molecular typing method for the identification and differentiation of Brucella strains at species and biovar levels (22), allowed us to presumptively identify all isolates as B. melitensis biovar 3. All isolates, in fact, carried the molecular marker M1249I.…”

Section: Discussionmentioning

confidence: 99%

“…The rpoB gene was amplified using the primers ϩ1rB and Ϫ4134rB previously described (22). PCR amplifications were carried out in 50-l volumes using GoTaq DNA polymerase (Promega Corporation, Madison, WI) following the enclosed protocol.…”

Section: Methodsmentioning

confidence: 99%

“…Comparing those sequences to the published B. melitensis 16 M rpoB gene (accession number AE009516), all isolates carried nucleotide alterations of ATC to ATA at codon position 1249 (M1249I), previously identified as a molecular marker of B. melitensis biovar 3 strain Ether and classified as B. melitensis genotype 3 (22). Three out of 20 isolates carried an additional CTT-to-TTT mutation at codon position 670, which led to substitution of amino acid L670F, never before observed in the rpoB gene of Brucella strains (22). This mutation was not found to be associated with the RIF resistance phenotype, as demonstrated by the in vitro antimicrobial susceptibility test.…”

Section: During the Study Period From April 2005 To May 2006 20mentioning

confidence: 99%

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Molecular Epidemiological and Antibiotic Susceptibility Characterization of Brucella Isolates from Humans in Sicily, Italy

et al. 2007

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Brucellosis is a serious problem in Sicily. Brucella melitensis was identified as the species most frequently isolated in humans in Italy. No data, however, are available about the molecular epidemiological characterization of Brucella isolates from humans. We have conducted this study to molecularly characterize clinical isolates of Brucella spp. and to evaluate their antimicrobial susceptibilities. Twenty Brucella isolates were studied. Differential growth characteristics and DNA polymorphisms such as the restriction patterns of the PCR-amplified omp2a and omp2b genes, rpoB nucleotide sequencing, and multiple-locus variable-number tandem repeat analysis of 16 loci (MLVA-16) were used to characterize the strains. In vitro antibiotic susceptibility was determined by the E-test method on two different agar media, and the results were compared. All isolates were identified as B. melitensis biovar 3. rpoB nucleotide sequence analysis allowed the identification of two different genotypes of B. melitensis biovar 3. On the other hand, the MLVA-16 typing assay recognized 17 distinct genotypes. All isolates were sensitive to all tested antibiotics (rifampin, doxycycline, ciprofloxacin, ceftriaxone, and trimethoprim-sulfamethoxazole), and the Mueller-Hinton agar plate is recommended for antibiotic susceptibility testing by the E-test method. Our findings identify B. melitensis biovar 3 as the etiological agent isolated in Sicily and encourage the use of both molecular methods, and in particular of the MLVA-16 assay, in epidemiological trace-back analysis. This study represents the first epidemiological data from molecular typing of Brucella strains circulating in Italy and, in particular, in eastern Sicily.Brucellosis is an important health problem affecting animals and humans in many countries in the world and especially in the Mediterranean areas. Brucella infections may cause tremendous economic losses through reproductive failure in animals. The disease is transmitted by humans through the consumption of contaminated foods or by direct/indirect contact with infected animals. Brucella melitensis is the most important zoonotic agent, followed by Brucella abortus and Brucella suis.

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Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: During the Study Period From April 2005 To May 2006 20mentioning

confidence: 99%

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Molecular Epidemiological and Antibiotic Susceptibility Characterization of Brucella Isolates from Humans in Sicily, Italy

et al. 2007

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“…Insertion sequence based typing and polymerase chain reaction-restriction fragment length polymorphisms (PCR-RFLP) are the examples of such methods [7,8] . In recent studies investigating reference and clinical Brucella isolates, the utility of the rpoB gene, encoding the DNA-dependent RNA polymerase β subunit (RNAP), for genotyping Brucella strains via a single nucleotide polymorphism (SNP) based method were examined [9][10][11] .…”

Section: Introductionmentioning

confidence: 99%

Single Nucleotide Polymorphism Analysis of the rpoB Gene Region for Genotyping of Brucella melitensis Strains Isolated from Field in Turkey

Sayan¹

2014

Kafkas Univ Vet Fak Derg

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We previously described the first molecular characterization of Brucella isolates in Turkey that were examined by single nucleotide polymorphisms (SNPs) at the rpoB locus of B. melitensis strains isolated from adult and pediatric patients. However, the molecular typing of B. melitensis strains causing animal infections in Turkey has not been previously investigated. The aim of this study was to evaluate SNP analysis of rpoB gene of B. melitensis from field isolates in Turkey and to try to find out one of the most appropriate methods other than conventional method for long term evaluation of epidemiological studies. Thirty-two B. melitensis strains isolated from Marmara, Aegean, Mediterranean, Central Anatolia and Eastern Anatolia regions of Turkey were investigated together with 3 reference strains. According to rpoB sequencing results, three distinct genotypes (SNP type 1, type variant 2 and type 2) were recognized. SNP technique characterized the strains at the molecular levels independently from B. melitensis biovars. Our study showed that SNP analysis has a better discriminatory capability in identification of B. melitensis strains compared to classical method. In conclusion, it was suggested that SNP analysis could be useful as a molecular epidemiological method to determine relationships between B. melitensis isolates and might aid in effective surveillance and control method for brucellosis particularly in conjunction with a national databases. Keywords: Brucella melitensis, Molecular typing, Single nucleotide polymorphism, Sequence analysis Türkiye'de Sahadan İzole Edilmiş Brucella melitensis Suşlarının Genotiplendirilmesinde rpoB Gen Bölgesi Tek Nükleotit Polimorfizm Analizi ÖzetÜlkemizde ilk kez Brucella izolatlarının moleküler karakterini ortaya çıkardığımız önceki çalışmada erişkin ve çocuk hastalardan izole edilen B. melitensis izolatlarında rpoB geni tek nükleotit polimorfizm (SNP) analizi ile değerlendirilmişti. Ülkemizde daha once hayvan enfeksiyonlarından izole edilen B. melitensis suşlarında SNP analizi ile genotiplendirme değerlendirilmiş değildir. Bu çalışmada, B. melitensis saha suşlarının moleküler tiplendirmesinde rpoB geninin SNP ile analizinin değerlendirilmesi ve epidemiyolojik çalışmalarda kullanılabilecek uygun tiplendirme yönteminin tanımlanması amaçlanmıştır. Araştırmada referans suşlar ile birlikte Marmara, Ege, Akdeniz, Orta Anadolu ve Güneydoğu Anadolu bölgelerimizden izol edilmiş toplam 32 B. melitensis saha suşu çalışıldı. rpoB geni sekanslarının değerlendirilmesi sonrasında B. melitensis saha suşlarında üç moleküler tip tanımlandı; SNP tip 1, SNP tip 2 ve SNP variant tip 2. SNP analiz tekniği B. melitensis suşlarını biyovar özelliklerinden bağımsız bir şekilde moleküler olarak tiplendirmektedir. Çalışmamız, epidemiyolojik olarak SNP analizinin B. melitensis suşlarını tanımlamada klasik yönteme göre yüksek ayrım gücüne sahip olduğunu göstermektedir. Sonuç olarak SNP analizinin B. melitensis izolatları arasındaki ilişkiyi saptayacak faydalı bir moleküler epidemiyolojik metot old...

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“…The rpoB marker has been used for numerous groups such as Enterobacteriaceae (Mollet et al, 1997), Brucella (Marianelli et al, 2006) or Frankia (Bernèche-D'Amours et al, 2011). Furthermore, rpoB has been tested as a substitute for amplicon metagenomics with excellent results.…”

Section: Introductionmentioning

confidence: 99%

Genomics of Specificity in the Symbiotic Interaction between Rhizobium leguminosarum and Legumes

Rubio¹

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. Para el desarrollo de esta Tesis he contado con una beca UPM homologada financiada por el proyecto MICROGEN: Genómica Comparada Microbiana (Programa Consolider), que ha sido también el proyecto financiador del trabajo experimental.Quisiera reconocer la labor de aquellas personas que han contribuido al desarrollo y consecución de esta Tesis.Dr. Juan Imperial, por la dirección y supervisión de esta Tesis. Por ser un gran mentor y por enseñarme todo lo que sé.Dr. Manuel González Guerrero, por enseñarme los entresijos de la Ciencia y del trabajo en el laboratorio.Dra. Gisèle Laguerre, por su participación en el inicio de este proyecto y por facilitarnos el suelo P1 de Dijon.Dr. Paulo Arruda, por la formación que obtuve durante mi estancia en el laboratorio de Estudio de la Regulación y de la Expresión génica de la Universidad Estatal de Campinas.Dra. Belén Brito, por su gran ayuda en la primera fase de este proyecto y por recomendarme, sin ella nunca hubiera empezado esta Tesis.Rosabel Prieto, por su inestimable ayuda para la obtención de los Pool-Seqs y por enseñarme muchas de las técnicas que he necesitado durante la Tesis.Santiago de la Peña, por su valiosa ayuda en el análisis bioinformático de los genomas individuales.Dra. Julia Quintana, por enseñarme los conceptos de genética de poblaciones y por las largas discusiones científicas que tanto me han aportado.Dra. Viviana Escudero, por enseñarme a cuidar las plantas y valorar las cosas realmente importantes. Dra. Carmen Sánchez, por enseñarme las técnicas básicas de microbiología y genética molecular durante mi periodo como estudiante de máster.Dr. David Durán, por enseñarme los conceptos básicos de filogenia. XIII AGRADECIMIENTOSEscribir los agradecimientos es la parte más difícil de una tesis, no porque sea complicado, si no porque es la única parte que se lee todo el mundo y supone una gran presión añadida. Siempre me imaginaba escribir esta parte como algo extremadamente emotivo, pero después de escribir más de 200 páginas me he quedado sin palabras.La ciencia siempre fue lo que quise hacer, pero nunca pensé que pudiera hacerlo, pero… here I am! Ha sido un camino largo y duro, pero no lo cambiaría por nada, me he convertido en lo que quería y he conocido a los mejores. Desde que tengo 13 años sabía que quería estudiar esto, gracias a la inspiración de mi profesora de biología Marisa Alonso. Cuando llegué a la carrera, las cosas no fueron como esperaba, pero aún así descubrí mi pasión por la micro, la genética y las plantas… Y sobre todo conocí a grandes personas. Desde luego la mejor decisión que tomé fue hacer el máster, este fue el punto de inflexión, descubrí que era hacer ciencia… y eso me enganchó. Quiero agradecer a Pepe Palacios por acogerme en su laboratorio durante este periodo, ya que sin duda, mi paso por allí marcó mi vuelta. Por ello también quiero agradecer a todos los Rizobios y Levaduros, y en especial a Carmen Sánchez, por ser mi supervisora en los inicios y enseñarme muchas de las técnicas básicas de micro y genética molecular que he apre...

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Molecular characterization of the rpoB gene in Brucella species: new potential molecular markers for genotyping

Cited by 47 publications

References 33 publications

Molecular Epidemiological and Antibiotic Susceptibility Characterization of Brucella Isolates from Humans in Sicily, Italy

Molecular Epidemiological and Antibiotic Susceptibility Characterization of Brucella Isolates from Humans in Sicily, Italy

Single Nucleotide Polymorphism Analysis of the rpoB Gene Region for Genotyping of Brucella melitensis Strains Isolated from Field in Turkey

Genomics of Specificity in the Symbiotic Interaction between Rhizobium leguminosarum and Legumes

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