Brevundimonas diminuta is a lactose non-fermenting Gram-negative rod associated with infection in immunocompromised patients. In three patients from two general wards, B. diminuta was isolated in blood culture sample. The clinical features of the patients did not coincide with the blood culture result, and pseudo-outbreak was suspected. These isolated were biochemically identified as Brevundimonas diminuta, and 16S rRNA sequencing confirmed their identification. The PFGE result showed a single pattern, and their clonality was assumed.
Electrophoresis, Gel, Pulsed-Field ; Humans ; Immunocompromised Host ; Lactose ; Patients' Rooms

BACKGROUND: It is important to define a source of infection when outbreak of Legionella infections has occurred. The performance of a molecular strain typing method was evaluated for environmental and clinical isolates of Legionella pneumophila. METHODS: Thirteen environmental strains, eleven clinical isolates and one type strain (ATCC 33152) of Legionella pneumophila were used for the analysis of pulsed field gel electrophoresis. RESULTS: All 25 strains were discriminated into 21 types. Two strains isolated from different locations in a same building showed different types. Each two, four, and two strains were shown as the same PFGE patterns. CONCLUSIONS: Even though PFGE typing of Legionella pneumophila is excellent for strain differentiation, the same pattern does not necessarily indicate the same source of isolates.
Busan* ; Electrophoresis, Gel, Pulsed-Field ; Korea* ; Legionella pneumophila ; Legionella* ; Water*

Objective: To identify a suspected clustered Typhoid fever by whole genome sequencing(WGS) and pulsed field gel electrophoresis (PFGE) subtyping. Methods: The nature of the epidemic was determined by combination of subtyping results of isolates and epidemiological information. Results: Five S. typhimurium isolates showed identical PFGE patterns and almost the same whole genome sequence. Epidemiological survey showed that five cases had dined in the same restaurant on the same day. Conclusion: Combined with the longest incubation period of typhoid fever, molecular subtyping of pathogenic bacteria and the field epidemiological survey, it can be preliminarily determined that the five cases have common infection sources.
Electrophoresis, Gel, Pulsed-Field ; Epidemics ; Humans ; Typhoid Fever/microbiology*

OBJECTIVETo develop fluorescent amplified fragment length polymorphism (AFLP) method and to evaluate the its typing capability with pulsed-field gel electrophoresis (PFGE) in molecular typing of Vibrio cholerae.

METHODSForty-seven strains of V. cholerae, with different PFGE patterns, were selected as the reference group to optimize the selective primers of AFLP analysis. Eighty-three strains including 20 strains from one epidemic episode, isolated from different provinces during 1961 and 2005, were used to compare the typing abilities of AFLP and PFGE. LI-COR4300 DNA sequencing system was used for AFLP electrophoresis. The images were recorded by Saga(MX) software and transferred to BioNumerics for clustering analysis. A standard protocol for V. cholerae from PulseNet was used in PFGE.

RESULTSWhen comparison was made with different selective primers on AFLP based on the 47 strains, results showed that the optimized selective primer pair was EcoR I-G/Mse I-T, and the reproducibility of the tests was 99.2%. Eighty-three isolates showed 52 AFLP patterns and 44 PFGE patterns, with D values as 0.9545 (AFLP) and 0.9251 (PFGE) respectively.

CONCLUSIONThe protocol of fluorescent AFLP on V. cholerae typing was established. AFLP was higher than PFGE in discrimination of V. cholerae which could be used for molecular typing. When combined with PFGE, AFLP became a more insightful tool to identify genome difference of different isolates.

Amplified Fragment Length Polymorphism Analysis ; methods ; Electrophoresis, Gel, Pulsed-Field ; Genotype ; Phylogeny ; Vibrio cholerae ; classification ; genetics

OBJECTIVETo determine the genetic relationships between different Vibrio cholerae isolates in Shenzhen from 1993 to 2002.

METHODSChromosomal DNA from 60 isolates was digested in seakem gold agrose with restriction enzyme Not I and plugs were then analyzed by pulsed-field gel electrophoresis. Pulsed-field gel electrophoresis (PFGE) patterns of V. cholerae isolates were clustered using BioNumerics software.

RESULTS39 distinctive PFGE patterns were identified with each pattern having 20 to 30 bands. Most PFGE patterns were divided into cluster A or cluster B.

CONCLUSIONThe closely related pandemic clone clusters of V. cholerae strains did exist in Shenzhen. PFGE of V. cholerae could be used for active surveillance and tracking for cholerae.

China ; epidemiology ; Cholera ; epidemiology ; microbiology ; Electrophoresis, Gel, Pulsed-Field ; methods ; Humans ; Phylogeny ; Vibrio cholerae ; classification ; genetics

OBJECTIVETo analyse the molecular types of Listeria (L.) monocytogenes, and to construct the standard China L. monocytogenes pulsed-field gel electrophoresis (PFGE) subtyping database, using the international standardized L. monocytogenes-PFGE protocol.

METHODS118 L. monocytogenes strains isolated from 8 provinces or municipalities of China were subtyped according to L. monocytogenes-PFGE protocol.

RESULTS118 strains of L. monocytogenes were divided into 39 subtypes. In the 39 subtypes, 37 strains (31.36%) were GX6A160004 pattern, showing it was the predominant Lm subtype in China.

CONCLUSIONData from molecular typing suggested that the predominant Lm strains were distributed in different regions of China. PulseNet China-Lm database was constructed which was valuable for the molecular subtyping-based surveillance of L. monocytogenes.

China ; Electrophoresis, Gel, Pulsed-Field ; Food Microbiology ; Listeria monocytogenes ; classification ; genetics ; isolation & purification ; Phylogeny

OBJECTIVETo type and group the Shiga-toxin producing Escherichia coli O157:H7 strains isolated recent years in China to understand the epidemiological features caused by the pathogen.

METHODSPulsed field gel electrophoresis of large restriction fragments of bacterial chromosomal DNA was used.

RESULTSThe 51 isolates of E. coli O157:H7 collected in recent years in China could be divided into 8 Pulsed Field Gel Electrophoresis (PFGE) types based on the size and number of restriction fragments and patterns, that were digested by XbaI. Strains isolated from Ningxia province showed only two types- PFGE1 and PFGE2. Strains isolated in Xuzhou of Jiangsu province had 6 PFGE types. Isolates identified between 1986 - 1988 belonged to PFGE7. Strains isolated from patients in 1999 - 2000 were PFGE5 and PFGE3. Strains isolated from stool samples of domestic animal, food and vegetable were PFGE3 - 6, of which the predominant type was PFGE5. All of the 5 strains isolated from patients with diarrhea and hemolytic uremic syndrome (HUS) belonged to PFGE type 5, which was the dominant type of the isolates from stool samples of domestic animal and samples of food and vegetable contaminated.

CONCLUSIONData suggested that the cluster patients with diarrhea and HUS might have been related to the pathogens from domestic animas and contaminated food or vegetables. The distribution of PFGE types also varied in different provinces of China.

Electrophoresis, Gel, Pulsed-Field ; Escherichia coli O157 ; classification ; genetics ; pathogenicity ; Genotype ; Humans ; Shiga Toxin ; biosynthesis

OBJECTIVETo analyze the molecular types of Salmonella paratyphi A strains isolated in the recent years, and to construct the standard S. paratyphi A databank in the laboratory surveillance network PulseNet China.

METHODSS. paratyphi A isolates from 4 provinces were analyzed with the standard pulsed-field gel electrophoresis (PFGE) protocol used in PulseNet and their patterns compared. The databank was constructed with BioNumerics.

RESULTSEleven PFGE patterns were obtained, in which 3 predominant patterns were identifies with a similarity coefficient of 96.3%. The strains of these patterns, accounted for 86.5% of the analyzed strains, appeared in different provinces and years.

CONCLUSIONThe databank of S. paratyphi A was constructed and could be used in laboratory surveillance of S. paratyphi A in PulseNet China. From the analyses on molecular typing of the isolates, data suggested that the predominant strains might cause the epidemics in different regions.

China ; Electrophoresis, Gel, Pulsed-Field ; Salmonella paratyphi A ; classification ; genetics ; isolation & purification

OBJECTIVETo investigate the epidemiological and molecular typing features of the pathogenic Yersinia enterocolitica strains isolated in China,using pulsed field gel electrophoresis(PFGE) and standardized PFGE method as well as typing database of Yersinia enterocolitica.

METHODSPFGE analysis was performed as Laboratory Directions for molecular subtyping of Salmonella by PFGE (PulseNet,USA) with some modifications and the results of PFGE were analyzed by BioNumerics soft (Version 4.0, Applied Maths BVBA, Belium).

RESULTS114 O:3 Yersinia enterocolitica strains were typed by 25 patterns to have found that K6GN11C30012 (50 strains), K6GN11C30015(19 strains) and K6GN11C30016(10 strains) were the major patterns. K6GNllC30012 had 92.2% cluster similarity with K6GN11C30009-K6GN11C30023. This clone included 91.23% strains of 114 0:3 Yersinia enterocolitica strains. 51 0:9 Yersinia enterocolitica strains were typed by 14 patterns; K6GN11C90004 (22 strains) and K6GN11C90010 (13 strains)were the major patterns. K6GN11C90004 had 81.8% cluster similarity with K6GN11C90010 patterns. The major patterns of 0:3 and 0:9 serotypes were quite different.

CONCLUSIONO:3 Yersinia enterocolitica strains might originate from the same clone and had very few variation in different years and provinces but O:9 Yersinia enterocolitica strains from two different clones with some changes.

China ; Electrophoresis, Gel, Pulsed-Field ; Humans ; Yersinia enterocolitica ; classification ; genetics ; isolation & purification

In order to develop a rapid and reliable method for B. cereus genotyping, factors influencing PFGE results, including preparation of bacterial cells embedded in agarose, lysis of embedded cells, enzymatic digestion of intact genomic DNA, and electrophoresis parameters allowing for reproducible and meaningful DNA fragment separation, were controlled. Optimal cellular growth (Luria-Bertani agar plates for 12-18 h) and lysis conditions (4 h incubation with 500 µg/mL lysozyme) produced sharp bands on the gel. Restriction enzyme NotI was chosen as the most suitable. Twenty-two isolates were analyzed by NotI digestion, using three electrophoretic parameters (EPs). The EP-a was optimal for distinguishing between isolates. The optimized protocol could be completed within 40 h which is a significant improvement over the previous methods.
Bacillus cereus ; genetics ; isolation & purification ; Bacteriological Techniques ; DNA, Bacterial ; chemistry ; genetics ; Electrophoresis, Gel, Pulsed-Field ; methods