Species identification and molecular typing of human Brucella isolates from Kuwait

Brucellosis is a zoonotic disease of major concern in Kuwait and the Middle East. Human brucellosis can be caused by several Brucella species with varying degree of pathogenesis, and relapses are common after apparently successful therapy. The classical biochemical methods for identification of Bruc...

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Bibliographic Details
Published in:PloS one 2017-08, Vol.12 (8), p.e0182111-e0182111
Main Authors: Mustafa, Abu S, Habibi, Nazima, Osman, Amr, Shaheed, Faraz, Khan, Mohd W
Format: Article
Language:English
Subjects:
Animals
Biology and Life Sciences
Brucella
Brucella - genetics
Brucella - isolation & purification
Brucella abortus
Brucella melitensis
Brucellosis
Classification
Cluster Analysis
Clusters
Consensus Sequence
Deoxyribonucleic acid
Differentiation
DNA
DNA Fingerprinting
DNA, Intergenic - genetics
Gene Frequency - genetics
Gene sequencing
Genetic Variation
Genomes
Genotypes
Humans
Identification and classification
Identification methods
Identification systems
Infectious diseases
Kuwait
Medicine and Health Sciences
Methods
Molecular Typing - methods
Multilocus Sequence Typing
Pathogenesis
People and Places
Polymerase Chain Reaction
Primers
Real time
Repetitive Sequences, Nucleic Acid - genetics
Research and Analysis Methods
Ribosomal DNA
RNA sequencing
RNA, Ribosomal, 16S - genetics
rRNA 16S
Sequence Analysis, DNA
Species
Species Specificity
Streptococcus infections
Surveillance
Zoonoses
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Summary:Brucellosis is a zoonotic disease of major concern in Kuwait and the Middle East. Human brucellosis can be caused by several Brucella species with varying degree of pathogenesis, and relapses are common after apparently successful therapy. The classical biochemical methods for identification of Brucella are time-consuming, cumbersome, and provide information limited to the species level only. In contrast, molecular methods are rapid and provide differentiation at intra-species level. In this study, four molecular methods [16S rRNA gene sequencing, real-time PCR, enterobacterial repetitive intergenic consensus (ERIC)-PCR and multilocus variable-number tandem-repeat analysis (MLVA)-8, MLVA-11 and MLVA-16 were evaluated for the identification and typing of 75 strains of Brucella isolated in Kuwait. 16S rRNA gene sequencing of all isolates showed 90-99% sequence identity with B. melitensis and real-time PCR with genus- and species- specific primers identified all isolates as B. melitensis. The results of ERIC-PCR suggested the existence of 75 ERIC genotypes of B. melitensis with a discriminatory index of 0.997. Cluster classification of these genotypes divided them into two clusters, A and B, diverging at ~25%. The maximum number of genotypes (n = 51) were found in cluster B5. MLVA-8 analysis identified all isolates as B. melitensis, and MLVA-8, MLVA-11 and MLVA-16 typing divided the isolates into 10, 32 and 71 MLVA types, respectively. Furthermore, the combined minimum spanning tree analysis demonstrated that, compared to MLVA types discovered all over the world, the Kuwaiti isolates were a distinct group of MLVA-11 and MLVA-16 types in the East Mediterranean Region.
ISSN:1932-6203
1932-6203
DOI:10.1371/journal.pone.0182111