Bacterial Typing Techniques ; Genotype ; Minisatellite Repeats ; Plague ; Yersinia pestis ; genetics

OBJECTIVEHuman Lyme Borreliosis (LB), which is caused by Borrelia burgdorferi sensu lato (B. burgdorferi), has been identified as a major arthropod-borne infectious disease in China. We aimed to develop a multiple locus variable-number tandem repeat (VNTR) analysis (MLVA) assay for the genotyping of Borrelia burgdorferi strains detected in China.

METHODSB. garinii PBi complete 904.246 kb chromosome and two plasmids (cp26 and lp54) were screened by using Tandem Repeats Finder program for getting potential VNTR loci, the potential VNTR loci were analyzed and identified with PCR and the VNTR loci data were analyzed and MLVA clustering tree were constrcted by using the categorical coefficient and the unweighted pair-group method with arithmetic means (UPGMA).

RESULTSWe identified 5 new VNTR loci through analyzing 47 potential VNTR loci. We used the MLVA protocol to analyse 101 B. burgdorferi strains detected in China and finally identified 51 unique genotypes in 4 major clusters including B. burgdorferi sensu stricto (B.b.s.s), B. garinii, B. afzelii, and B. valaisiana, consistent with the current MLSA phylogeny studies. The allele numbers of VNTR-1, VNTR-2, VNTR-3, VNTR-4, and VNTR-5 were 7, 3, 9, 7, and 6. The Hunter-Gaston index (HGI) of five VNTR loci were 0.79, 0.22, 0.77, 0.71, and 0.67, respectively. The combined HGI of five VNTR loci was 0.96. Clustering of the strains of Xinjiang, Inner Mongolia and Heilongjiang was confirmed, and this situation was consistent with the close geographical distribution of those provinces.

CONCLUSIONThe MLVA protocol esytablished in this study is easy and can show strains' phylogenetic relationships to distinguish the strains of Borrelia species. It is useful for further phylogenetic and epidemiological analyses of Borrelia strains.

Borrelia burgdorferi Group ; genetics ; China ; Genotyping Techniques ; Minisatellite Repeats

Animals ; Genotype ; Humans ; Leptospira interrogans ; classification ; genetics ; Minisatellite Repeats ; genetics ; Molecular Epidemiology ; methods

OBJECTIVETo evaluate four candidate variable number tandem repeat (VNTR) loci for genotyping Mycobacterium tuberculosis complex strains.

METHODSGenomic sequences for two M. tuberculosis strains (CCDC5079 and CCDC5180) were generated, and using published sequence data, four candidate VNTR loci were identified. The VNTRs were used to genotype 225 Chinese clinical M. tuberculosis complex strains. The discriminatory power of the VNTRs was evaluated using BioNumerics 5.0 software.

RESULTSThe Hunter-Gaston Index (HGI) for BJ1, BJ2, BJ3, and BJ4 loci was 0.634, 0.917, 0.697, and 0.910, respectively. Combining all four loci gave an HGI value of 0.995, thus confirming that the genotyping had good discriminatory power. The HGI values for BJ1, BJ2, BJ3, and BJ4, obtained from Beijing family strain genotyping, were 0.447, 0.878, 0.315, and 0.850, respectively. Combining all four loci produced an HGI value of 0.988 for genotyping the Beijing family strains. We observed unique patterns for M. bovis and M. africanum strains from the four loci.

CONCLUSIONWe have shown that the four VNTR loci can be successfully used for genotyping M. tuberculosis complex strains. Notably, these new loci may provide additional information about Chinese M. tuberculosis isolates than that currently afforded by established VNTR loci typing.

Cluster Analysis ; Genotyping Techniques ; Humans ; Minisatellite Repeats ; Mycobacterium bovis ; genetics ; Mycobacterium tuberculosis ; genetics

OBJECTIVETo develop a multiple-locus variable-number tandem-repeat (VNTR) analysis (MLVA) assay for Acinetobacter pittii typing.

METHODSPolymorphic VNTRs were searched by Tandem Repeats Finder. The distribution and polymorphism of each VNTR locus were analyzed in all the A. pittii genomes deposited in the NCBI genome database by BLAST and were evaluated with a collection of 20 well-characterized clinical A. pittii strains and one reference strain. The MLVA assay was compared with pulsed-field gel electrophoresis (PFGE) for discriminating A. pittii isolates.

RESULTSTen VNTR loci were identified upon bioinformatic screening of A. pittii genomes, but only five of them showed full amplifiability and good polymorphism. Therefore, an MLVA assay composed of five VNTR loci was developed. The typeability, reproducibility, stability, discriminatory power, and epidemiological concordance were excellent. Compared with PFGE, the new optimized MLVA typing scheme provided the same and even greater discrimination.

CONCLUSIONCompared with PFGE, MLVA typing is a faster and more standardized alternative for studying the genetic relatedness of A. pittii isolates in disease surveillance and outbreak investigation.

Acinetobacter ; classification ; genetics ; DNA Fingerprinting ; methods ; Electrophoresis, Gel, Pulsed-Field ; Minisatellite Repeats ; Polymerase Chain Reaction

Bacterial Typing Techniques ; Leptospira interrogans ; classification ; genetics ; Minisatellite Repeats ; Multilocus Sequence Typing ; methods

OBJECTIVETo study the expression of Aggrecan and the relationship between the variable number of tandem repeats (VNTR) of Aggrecan and lumbar disc herniation (LDH).

METHODSThe disease group comprised of 74 patients already diagnosed with symptomatic LDH. The control group consisted of 15 patients restricted to spinal trauma and 113 healthy blood donors without symptoms of LDH who were not diagnosed with LDH. Disc tissue samples were obtained from surgical operations and blood samples were donated from all participants. The Aggrecan expression in isolated tissues was assessed by western blot using specific antibodies. The Aggrecan gene VNTR region was analyzed by PCR.

RESULTSThe Aggrecan expression positive rate of control group was statistically and significantly higher (control group:86.67%, disease group:13.51%;χ(2) = 34.83, P < 0.05) than that of the disease group. Moreover, there was a statistically significant higher frequency of Allele 25 or Allele 21 in disease group compared to controls (A25disease group = 22.97%, A25control group = 12.11%, χ(2)A25 = 8.20, PA25 = 0.004; A21disease group = 6.76%, A21control group = 0.39%, χ(2)A21 = 14.35, PA21 = 0.000). Compared to the participants with 2 Alleles>25 repeats, subjects with 1 or 2 Alleles ≤ 25 repeats statistically and significantly over represented the disease group without the expression of Aggrecan (χ(2) = 5.69, P = 0.017).

CONCLUSIONSThe findings suggest a relationship between Aggrecan and symptomatic LDH, where symptomatic LDH has a tendency of allele 21 and allele 25 repeats.In addition, an association between the distribution of Aggrecan gene VNTR polymorphism and the expression of Aggrecan is observed in symptomatic LDH.

Aggrecans ; biosynthesis ; Alleles ; Humans ; Intervertebral Disc Displacement ; genetics ; Minisatellite Repeats ; Polymerase Chain Reaction ; Polymorphism, Genetic

Animals ; Cattle ; microbiology ; Microbial Sensitivity Tests ; Minisatellite Repeats ; Mycobacterium bovis ; drug effects ; genetics

OBJECTIVE: To understand the forensic practice of DNA profiling from minimal amount of DNA. METHODS: Serial dilutions of DNA were amplified with the PowerPlex 16 System Kit, then the genotyping of short tandem repeat(STR) was performed by ABI 377 DNA automated Sequencer. RESULTS: When the mount of DNA template was less than 250 pg, allelic drop-out apparently occurred at several loci. Other disturbed peaks, such as artefact bands and imbalanced heterozygote, also presented. CONCLUSION The anomalous results may result in incorrect genotyping. Careful and comprehensive considerations are needed to interpret the STR profile of minute DNA.
Alleles ; DNA/genetics* ; Forensic Medicine ; Genotype ; Humans ; Male ; Minisatellite Repeats/genetics* ; Sequence Analysis, DNA ; Tandem Repeat Sequences/genetics*

OBJECTIVESelecting variable-number tandem-repeat (VNTR) loci for different serogroups of Shigella spp to explore and establish multilocus variable-number tandem-repeat analysis (MLVA) method, in order to study the molecular characteristic of the isolated strains.

METHODSOf the Shigella strains found by dysentery surveillance in Beijing from 2001 to 2009, 180 strains were selected for this study, according to the number and serotypes of the surveillant strains, at the ratio of 15%; including 50 strains of Shigella sonnei and 130 strains of Shigella flexneri. After screening the polymorphism of the 18 VNTR loci, 10 VNTR loci (sh1-sh10) were retained and constructed three groups of multi-PCR methods to detect all he 180 strains and analyze MLVA molecular subtypes using capillary segments.

RESULTSA range of 2 to 11 alleles were found on the 10 VNTR loci among the 180 Shigella strains, with a diversity index value between 0.158 and 0.766. The 10 loci showed diversity in different serogroups, such as only one allele found in sh6 of Shigella flexneri, sh2 and sh3 of Shigella sonnei individually. The isolated 180 strains were divided into 84 MLVA subtypes, with a resolution ratio D value at 0.967 (95%CI: 0.956 - 0.978). The 130 strains of Shigella flexneri were divided into 63 subtypes, named as TF001-TF063; among which TF001, TF002 and TF 005 were the dominant subtypes, accounting to 17, 16 and 15 strains respectively. The 50 strains of Shigella sonnei were divided into 21 subtypes, named as TS001-TS021; among which TS002 (14 strains) and TS001 (7 strains) were the dominant subtypes.

CONCLUSIONMLVA subtyping method including 10 VNTR loci was preliminarily developed. The MLVA cluster analysis revealed that the subtypes of Shigella strains isolated in Beijing were diverse, and suggested the possibility of multiple-clone source.

Alleles ; Bacterial Typing Techniques ; methods ; China ; DNA, Bacterial ; genetics ; Genotype ; Minisatellite Repeats ; Polymorphism, Genetic ; Shigella ; classification ; genetics ; isolation & purification