Development of a PCR assay for typing and subtyping of Brucella species

Huber, Birgit; Scholz, Holger C.; Lucero, Nidia E.; Busse, Hans-Jiirgen

doi:10.1016/j.ijmm.2009.05.002

Cited by 38 publications

(31 citation statements)

References 33 publications

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“…Therefore it is suggested that these primer pairs can also be useful for the direct detection of Brucella species in tissue samples. Also, like other studies such as those conducted by Huber et al and Kazemi et al, this study showed that PCR was a sensitive and suitable method for detecting Brucella species (25, 26). More studies will be needed to improve the specificity and sensitivity of molecular methods before recommending routine laboratory detection.…”

Section: Discussionsupporting

confidence: 82%

Evaluation of Different Primers for Detection of Brucella by Using PCR Method

2016

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IntroductionBrucellosis is a worldwide zoonosis and a significant cause of loss of health in humans and animals. Traditionally, classic diagnosis is carried out by isolation of Brucella, which is time-consuming, technically challenging and potentially dangerous. The aim of this study was to expand a molecular test that would be used for the develop detection of Brucella in a single reaction with high sensitivity and specificity, by targeting IS711element.MethodsThis study was carried out from 2015 to 2016 at the Ayatolla Rohani hospital in Babol, Iran. The present study was designed to develop PCR assay, based on IS711 gene for rapid diagnosis of Brucella spp. and immediate detection of Brucella, with high sensitivity and specificity. Four pairs of oligo-nucleotide primers with sizes of 547, 403, 291 and 127bp respectively, were planned to exclusively amplify the targeted genes of Brucella species.ResultsOur results show that, five PCR primers set up, would be helpful in amplifying the DNAs from the genus Brucella with high specificity and sensitivity so it can be 12 fg, for Brucella species to provide a valuable tool for diagnosis.ConclusionThis method can be more useful than serological and biochemical tests and in addition, this reduces the number of required tests more rapidly and economically.

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Section: Discussionsupporting

confidence: 82%

Evaluation of Different Primers for Detection of Brucella by Using PCR Method

2016

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“…These can be genes that are present only in the specific strains, antibiotic resistance genes, virulence genes, or housekeeping genes if it is possible to select species-specific primers for them [3,[9][10][11]. Often, this analysis requires several or even many genomic loci (Multi-Locus Analysis).…”

Section: Introductionmentioning

confidence: 99%

Короткие Уникальные Последовательности В Бактериальных Геномах Как Штамм- И Видоспецифические Маркеры

Панюков¹,

Panyukov²

2017

Math.Biol.Bioinf.

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Abstract. The paper presents a new approach for phylotyping that can be potentially used for pure cultures and for mixed bacterial populations. It is based on the use of short unique nucleotide sequences (k-mers) that are present in the genomes of all strains of the same species and are absent in bacterial genomes of other taxonomic groups. We show that the number N of such sequences depends on the percentage bias towards A/T or G/C base pairs, increasing for genomes with approximately equal composition. We found that the largest contribution to the set of primarily unique sequences is given by 16-17-mers, while sigmoidal curves reflecting the dependence of N on the length of k-mers showed the maximum slope increment (ΔN/Δk) for k = 17, 18. Unique sequences of the length 16-18 bases can therefore be offered as potential markers. Comparing the sets of unique k-mers in the genomes of four Enterobacter strains, we estimated the level of their intraspecies stability and interspecies plasticity. As a result, we suggest discriminatory subsets as stencils for phylotyping, thereby increasing the list of genotyping markers with signatures of the new type.

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“…This PCR was later improved by another laboratory by adding specific primers for the identification of B. abortus biovars 5, 6, 9 and genotype 3b of biovar 3 [18]. In 2009, Huber et al developed a random amplified polymorphic DNA PCR assay to differentiate all recognized Brucella species, including the marine mammals-infecting species B. ceti and B. pinnipedialis [19]. However, all published methods are developed to distinguish species and biovars mainly by gel-based bar patterns and the majority of described methods were tested with a few strains.…”

Section: Introductionmentioning

confidence: 99%

A novel real-time PCR assay for specific detection of Brucella melitensis

et al. 2017

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BackgroundBrucellosis is a zoonosis that occurs worldwide. The disease has been completely eradicated in livestock in Sweden in 1994, and all cases of confirmed human brucellosis are imported into Sweden from other countries. However, due to an increase in the number of refugees and asylum seekers from the middle-east to Sweden, there is a need to improve the current diagnostic methodology for Brucella melitensis. Whilst culture of Brucella species can be used as a diagnostic tool, real-time PCR approaches provide a much faster result. The aim of this study was to set up a species-specific real-time PCR for the detection of all biovars of Brucella melitensis, which could be used routinely in diagnostic laboratories.MethodsA Brucella melitensis real-time PCR assay was designed using all available genomes in the public database of Brucella (N = 96) including all complete genomes of Brucella melitensis (N = 17). The assay was validated with a collection of 37 Brucella species reference strains, 120 Brucella melitensis human clinical isolates, and 45 clinically relevant non-Brucella melitensis strains.ResultsIn this study we developed a single real-time PCR for the specific detection of all biovars of Brucella melitensis.ConclusionsThis new real-time PCR method shows a high specificity (100%) and a high sensitivity (1.25 GE/μl) and has been implemented in the laboratories of four governmental authorities across Sweden.

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Development of a PCR assay for typing and subtyping of Brucella species

Cited by 38 publications

References 33 publications

Evaluation of Different Primers for Detection of Brucella by Using PCR Method

Evaluation of Different Primers for Detection of Brucella by Using PCR Method

Короткие Уникальные Последовательности В Бактериальных Геномах Как Штамм- И Видоспецифические Маркеры

A novel real-time PCR assay for specific detection of Brucella melitensis

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