OBJECTIVES: Brucellosis is a major zoonotic disease that poses a significant public health threat worldwide. The classical bacteriological detection process used to identify Brucella spp. is difficult and time-consuming. This study aimed to develop a novel molecular assay for detecting brucellosis. METHODS: All complete sequences of chromosome 1 with 2.1-Mbp lengths were compared among all available Brucella sequences. A unique repeat sequence (URS) locus on chromosome 1 could differentiate Brucella abortus from Brucella melitensis. A primer set was designed to flank the unique locus. A total of 136 lymph nodes and blood samples were evaluated and classified by the URS-polymerase chain reaction (PCR) method in 2013–2014. RESULTS: Biochemical tests and bacteriophage typing as the golden standard indicated that all Brucella spp. isolates were B. melitensis biovar 1 and B. abortus biovar 3. The PCR results were the same as the bacteriological method for detecting Brucella spp. The sensitivity and specificity of the URS-PCR method make it suitable for detecting B. abortus and B. melitensis. CONCLUSION: Quick detection of B. abortus and B. melitensis can provide the most effective strategies for control of these bacteria. The advantage of this method over other presented methods is that both B. abortus and B. melitensis are detectable in a single test tube. Furthermore, this method covered 100% of all B. melitensis and B. abortus biotypes. The development of this URS-PCR method is the first step toward the development of a novel kit for the molecular identification of B. abortus and B. melitensis.
Bacteria ; Bacteriophage Typing ; Brucella abortus* ; Brucella melitensis* ; Brucella* ; Brucellosis ; Chromosomes, Human, Pair 1 ; Lymph Nodes ; Methods ; Polymerase Chain Reaction* ; Public Health ; Sensitivity and Specificity ; Zoonoses

Cyptococcosis is generally caused by Cryptococcus neoformans, the opportunistic agent which has two species such as C. neoformans and C. gattii. Both C. neoformans and C. gattii species contain a number of genetically diverse subgroups that can be differentiated by various molecular typing methods. We conducted a molecular epidemiological analysis of 30 clinical isolates of the C. neoformans from cryptococcosis patients who had been hospitalized between 2008 and 2010 in medical centers located in Seoul and Busan in Korea. To determine the genetic diversity, 30 strains of C. neoformans were typed using PCR fingerprinting with the microsatellite specific primer of the phage M13 and the restriction fragment length polymorphism (RFLP) of orotidine monophosphosphate pyrophosphorylase (URA5) gene. All isolates were identified as serotype A, mating type MATa and molecular type VNI. The random amplified polymorphic DNA (RAPD) profiles obtained by using two primers revealed a single pattern. Our study shows that 30 strains of clinical C. neoformans are genetically homogeneous, with all of the isolates were molecular type VN1, serotype A, mating type MATa.
Bacteriophage M13 ; Cryptococcosis ; Cryptococcus ; Cryptococcus neoformans ; Dermatoglyphics ; DNA ; Genetic Variation ; Humans ; Korea ; Microsatellite Repeats ; Molecular Typing ; Polymerase Chain Reaction ; Polymorphism, Restriction Fragment Length ; Uridine

OBJECTIVETo conduct an etiological molecular epidemiological survey and laboratory test on a foodborne disease epidemic outbreak to make clear of the cause and implement effective prevention and control on it.

METHODSOn May 12th 2012, 135 kindergarten children were sent to Xuzhou City People's Hospital and Children's Hospital with gastrointestinal infection disease. A total of 34 anus swab samples and 4 vomit samples were collected from the patients. Real-time PCR rapid detection, strains separation and cultivation, phage lysis experiments, ATB automated identification system were used to make etiological detection and identification. The genomic DNA of salmonella enteritidis were typed with the pulsed-field gel electrophoresis (PFGE), cluster analysis were carried out together with the patterns of local Salmonella infections.

RESULTSChildren in 20 classes were suffered from the gastrointestinal infection among the 21 classes. There were no significant aggregation of class distribution. Among the 135 patients, 76 were boys (56.3%) and 59 were girls (43.7%). The main symptoms were fever (above 38°C), diarrhea and bellyache. Through real-time PCR detection and strains separation, 19 salmonella enteritidis were isolated from 34 anus swab samples of suspected cases and the detection rate was 56%. There were no strains detected from vomit samples. All of the 19 salmonella enteritidis showed the same serological subtype, biochemical reaction, drug sensitivity and phage lysis pattern. The salmonella enteritidis had the identical PFGE pattern (100% similarity), and were different from the pattern of local sporadic infection cases.

CONCLUSIONIt was confirmed that this was an epidemic outbreak of foodborne disease caused by homologous salmonella enteritidis by epidemiological survey, clinical information, lab etiological test and molecular typing.

Bacteriophage Typing ; Child, Preschool ; China ; epidemiology ; Disease Outbreaks ; Electrophoresis, Gel, Pulsed-Field ; Female ; Humans ; Male ; Molecular Epidemiology ; Salmonella Food Poisoning ; epidemiology ; microbiology ; Salmonella enteritidis ; classification ; isolation & purification

BACKGROUND: Salmonella I 4,[5],12:i:- was first reported as a new serovar by the Spanish National Reference Laboratory in 1997. Thereafter, several outbreaks caused by this serovar have been reported, indicating worldwide transmission. MATERIALS AND METHODS: Stool samples and patient data were collected from diarrhea cases in an outbreak at Daegu city in 2008. Salmonella isolates were characterized by phage typing, antibiotic resistance profile and flagella gene deletion. Deposited isolates in the EnterNet-Korea, acute diarrheal surveillance system, were also screened for this serovar. RESULTS: Isolates from diarrhea patients in the Daegu outbreak (2008) were identified as Salmonella I 4,[5],12:i:-. Screening the deposited isolates in the EnteroNet-Korea revealed that an unidentifed isolate in 2001 was the Salmonella I 4,[5], 12:i:-. CONCLUSIONS: Salmonella I 4,[5],12:i:-. was the causative pathogen of the 2008 foodborne outbreak of salmonellosis in Daegu City. Retrospective screening revealed that Salmonella I 4,[5],12:i:- was present in Korea as early as 2001.
Bacteriophage Typing ; Diarrhea ; Disease Outbreaks ; Drug Resistance, Microbial ; Flagella ; Gene Deletion ; Humans ; Korea ; Mass Screening ; Retrospective Studies ; Salmonella ; Salmonella Infections

OBJECTIVETo investigate the molecular characteristics of phage-type 6b isolates emerging in 1998-2001 cholera epidemics in Sichuan province.

METHODSIsolates were analyzed by phage-typing, pulsed field gel electrophoresis (PFGE) and ompW gene sequencing.

RESULTSAll phage-type 1b and 6b isolates in Sichuan province from 1998 to 2001 were toxigenic. A total of 24 patterns were identified after PFGE analysis, and one predominant pattern consisted of 13 isolates. Several 1b and 6b isolates from Sichuan and isolates of the 1b from other provinces showed the same PFGE pattern. Mutation in ompW gene was found in 6b isolates.

CONCLUSIONV.cholerae O1 6b isolates in Sichuan province from 1998 to 2001 have special genetic markers, and might genetically correlate with contemporaneous 1b isolates.

Bacterial Typing Techniques ; Bacteriophage Typing ; China ; epidemiology ; Cholera ; epidemiology ; microbiology ; DNA, Bacterial ; genetics ; Electrophoresis, Gel, Pulsed-Field ; Genes, Bacterial ; Genotype ; Vibrio cholerae ; classification ; genetics ; isolation & purification

OBJECTIVEUsing pulsed field gel electrophoresis (PFGE) and polymerase chain reaction (PCR) typing to analyze strains isolated from two outbreaks caused by Shigella sonnei and to trace the source of infection.

METHODSVirulence genes ipaH and ial were detected by PCR and PFGE was used to subtype the isolates. Patterns were compared, using the software BioNumerics.

RESULTSWithin the 54 isolates, all were ipaH positive with 48 as ial positive. Strains from the Chongzhou outbreak were clustered into 4 PFGE patterns, with the predominant pattern accounted for 72% of the analyzed strains. The pattern of strains isolated from the cold pork with sauce was identical to the predominant pattern. The strains from Dayi outbreak were clustered into 8 PFGE patterns and the predominant pattern accounted for 56% of the test strains.

CONCLUSIONStrains from the two outbreaks were quite different and the 'cold pork with sauce' seemed to be the major source of infection, causing the outbreak of diarrhea in Chongzhou. The sources of infection of the Dayi outbreak might be complicated whereas PFGE showed a discriminatory and reproducible laboratory tool in the epidemiologic investigation on outbreaks of diarrhea.

Bacteriophage Typing ; China ; epidemiology ; Disease Outbreaks ; Dysentery, Bacillary ; epidemiology ; microbiology ; Electrophoresis, Gel, Pulsed-Field ; Food Microbiology ; Foodborne Diseases ; epidemiology ; microbiology ; Humans ; Shigella ; classification ; isolation & purification

OBJECTIVETo investigate the serologic type, phage-biotype and toxic factor of Vibrio cholerae isolated from different sea products, analyze the relation between the Vibrio cholerae in sea products and cholera epidemiology, and provide references for forecasting cholera epidemic situation and drawing out a preventing plan.

METHODThe biotype of strains isolated was analyzed by using type and phage-biotype serological methods. The toxic gene was detected by PCR.

RESULTSThe constituent ratio of V. cholerae O139, Ogawa and Inaba were, respectively, 48.44%, 20.31% and 31.25% in 64 strains of V. cholerae. The result of phage-biotype showed that the 26 strains of V. cholerae O1 were all non-epidemic strains. The result of toxic gene detecting showed that positive rate of V. cholerae O139 was higher than those of Ogawa and Inaba.

CONCLUSIONThe positive rate of toxic gene in V. cholerae O139 was high and the V. cholerae O139 was mainly in turtle, breed aquatics water and crustacean, so these sea products were the important sectors in cholera prevention and control.

Animals ; Bacteriophage Typing ; DNA, Bacterial ; genetics ; Seafood ; microbiology ; Serotyping ; Vibrio cholerae ; classification ; genetics ; isolation & purification ; Vibrio cholerae O1 ; isolation & purification ; Vibrio cholerae O139 ; isolation & purification

OBJECTIVETo explore and establish fast and sensitive typing diagnostic methods on intestinal pathogen.

METHODSAll 29 isolates of Shigella. sonnei and 12 strains of Escherichia coli O157:H7 (EHEC O157:H7) isolated in 1999 were carried out for typing by using pulse-field gel electrophoresis (PFGE) and phage typing.

RESULTSPFGE results of 4 outbreak incidents caused by Shigella. sonnei showed that the PFGE patterns of the strains isolated from the same incident were almost the same. It was also the same during analysing the 2 outbreak incidents caused by EHEC O157:H7, however, the PFGE patterns of the 3 individual strains of EHEC O157:H7 were different. Moreover, the phage typing method showed its fine typing ability while its being used in the analysing 12 strains of EHEC O157:H7.

CONCLUSIONThe fine typing ability and reliability of PFGE was helpful for finding out the heredity background of intestinal pathogen and the infectious resource of the outbreak. And the epidemic situation might thus be effectively prevented from spreading.

Bacteriophage Typing ; methods ; Disease Outbreaks ; Electrophoresis, Gel, Pulsed-Field ; Enterobacteriaceae Infections ; epidemiology ; Escherichia coli O157 ; classification ; Humans ; Japan ; epidemiology ; Shigella sonnei ; classification

Bacteriophages were induced from staphylococcus intermedius isolates from dogs and used for a phage typing. Mitomycin C induction was performed on 60 strains of S. intermedius and all phages were reacted with the strains. Twenty-eight strains (46.7%) were found to be lysogenic. Based on host ranges, eight phages were selected. By using the eight phages, 129 strains isolated from canine clinical specimens were subjected to the phage typing at the routine test dilution (RTD) and 100xRTD. Typability of the phage set was 78.3%, yielding 40 phage patterns. The phage set was considered useful for differentiating S. intermedius strains isolated from dogs. None of 50 strains of S. aureus, 2 (3.6%) of 56 S. simulance strains, 13 (24.5%) of 53 S. chromogenes strains and 46 (28.1%) of 164 S. hyicus strains were typable by the phages at 100xRTD.
Animals ; Bacteriophage Typing* ; Bacteriophages* ; Dogs ; Host Specificity ; Mitomycin ; Staphylococcus intermedius* ; Staphylococcus*

The frequency of antibiotic resistance among Salmonella enterica serovar Typhimurium has increased due to the transfer of multiple resistance factors. We detected the 13 antibiotic resistance genes by multiplex-PCR and compared with the results of phage typing and antibiotic disk diffusion for 49 S. typhimurium isolated from food-poisoning outbreaks in Seoul from 1999 to 2002. Resistance genes for tetracycline, streptomycin, ampicillin, sulfonamide, amino-glycoside-modifying enzyme, chloramphenicol, kanamycin, and trimethoprim were detected in 67.3%, 57.1%, 26.5%, 8.1%, 8.1%, 5%, 2.0%, and 0% of isolates, respectively. Overall 28 isolates (57.1%) possessed two or more antibiotic resistance genes. Class 1 integron carrying multidrug resistace genes, ant(3")-IaB, blaPSE, qacE delta1/sul, and tet G were amplified especially in only DT104 isolates. Among the related resistance genes for same antibiotics, strA and strB for streptomycin resistance were simultaneously detected but tetA and tetB for tetracycline were sporadically detected. DT 104 isolates contained only aadA2 and tetG.
Ampicillin ; Anti-Bacterial Agents ; Bacteriophage Typing ; Chloramphenicol ; Diffusion ; Disease Outbreaks ; Drug Resistance, Microbial ; Humans ; Integrons ; Kanamycin ; R Factors ; Salmonella enterica* ; Salmonella* ; Seoul* ; Streptomycin ; Tetracycline ; Trimethoprim