OBJECTIVETo study the types of subspecies of Francisella tularensis from China and to investigate the genetic relationships between F. tularensis strains from China and from other countries.

METHODSTen strains of F. tularensis isolated from China were amplified by using typing primers C1/C4 and RD1. On the basis of the lengths of the polymerase chain reaction (PCR) products, it was concluded that these strains of F. tularensis belonged to the same subspecies. At the same time, the fopA, tul4, and 16S rRNA genes of the 10 strains were amplified, and a three-gene based phylogenetic analysis was performed using the Molecular Evolutionary Genetics Analysis software version 4.0.

RESULTSThe 10 strains of F. tularensis from China were all identified as belonging to subspecies holarctica (type B). We found no direct relationship between the genotypes of F. tularensis subsp. holarctica and the geographical area from where they were isolated.

CONCLUSIONThe F. tularensis strains isolated from North China mainly belong to subspecies holarctica (type B). The strains of F. tularensis subsp. holarctica from China may have evolved earlier than those from Europe and North America.

China ; Francisella tularensis ; classification ; genetics ; Molecular Sequence Data ; Phylogeny

PURPOSE: Study on the pathogen and the pathogen-related disease require the information at both cellular and organism level. However, lack of appropriate high-quality antibodies and the difference between the experimental animal models make it difficult to analyze in vivo mechanism of pathogen-related diseases. For more reliable research on the infection and immune-response of pathogen-related diseases, accurate analysis is essential to provide spatiotemporal information of pathogens and immune activity to avoid false-positive or mis-interpretations. In this regards, we have developed a method for tracking Francisella tularensis in the animal model without using the specific antibodies for the F. tularensis. MATERIALS AND METHODS: A dual reporter plasmid using GFP-Lux with putative bacterioferritin promoter (pBfr) was constructed and transformed to F. tularensis live vaccine strain to generate F. tularensis LVS (FtLVS)-GFP-Lux for both fluorescence and bioluminescence imaging. For vaccination to F. tularensis infection, FtLVS and lipopolysaccharide (LPS) from FtLVS were used. RESULTS: We visualized the bacterial replication of F. tularensis in the cells using fluorescence and bioluminescence imaging, and traced the spatio-temporal process of F. tularensis pathogenesis in mice. Vaccination with LPS purified from FtLVS greatly reduced the bacterial replication of FtLVS in animal model, and the effect of vaccination was also successfully monitored with in vivo imaging. CONCLUSION: We successfully established dual reporter labeled F. tularensis for cellular and whole body imaging. Our simple and integrated imaging analysis system would provide useful information for in vivo analysis of F. tularensis infection as well as in vitro experiments, which have not been fully explained yet with various technical problems.
Animals ; Antibodies ; Fluorescence ; Francisella tularensis* ; Francisella* ; Immunodeficiency Virus, Feline ; In Vitro Techniques ; Methods ; Mice ; Models, Animal ; Plasmids ; Vaccination ; Whole Body Imaging

Tularemia is a potentially severe zoonotic disease caused by Francisella tularensis. A lack of awareness about tularemia can be embarrassing and could result in delayed treatment because of improper diagnosis. The diagnosis of tularemia is difficult, because the infections are rare and the clinical spectrum is broad. As only 1 adult case has been reported in Korea thus far, pediatricians in Korea may be unfamiliar with tularemia. We report our experience with a 14-year-old male adolescent with tularemia who presented with atypical pneumonia and possible infective endocarditis. Although the infectivity and mortality rates for tularemia are very high if left untreated, we did not suspect tularemia in this case until the incidental isolation of F. tularensis. The present case suggests that clinicians in Korea should be more aware of tularemia. This case also suggests that tularemia should be considered in undetermined cases of atypical pneumonia or acute febrile illness without local signs.
Adolescent ; Adult ; Diagnosis ; Endocarditis* ; Francisella tularensis ; Humans ; Korea* ; Mortality ; Pediatrics ; Pneumonia* ; Tularemia* ; Zoonoses

Tularemia is a high-risk infectious disease caused by Gram-negative bacterium Francisella tularensis. Due to its high fatality at very low colony-forming units (less than 10), F. tularensis is considered as a powerful potential bioterrorism agent. Vaccine could be the most efficient way to prevent the citizen from infection of F. tularensis when the bioterrorism happens, but officially approved vaccine with both efficacy and safety is not developed yet. Research for the development of tularemia vaccine has been focusing on the live attenuated vaccine strain (LVS) for long history, still there are no LVS confirmed for the safety which should be an essential factor for general vaccination program. Furthermore the LVS did not show protection efficacy against high-risk subspecies tularensis (type A) as high as the level against subspecies holarctica (type B) in human. Though the subunit or recombinant vaccine candidates have been considered for better safety, any results did not show better prevention efficacy than the LVS candidate against F. tularensis infection. Currently there are some more trials to develop vaccine using mutant strains or nonpathogenic F. novicida strain, but it did not reveal effective candidates overwhelming the LVS either. Difference in the protection efficacy of LVS against type A strain in human and the low level protection of many subunit or recombinant vaccine candidates lead the scientists to consider the live vaccine development using type A strain could be ultimate answer for the tularemia vaccine development.
Bioterrorism ; Communicable Diseases ; Francisella tularensis ; Humans ; Sprains and Strains ; Stem Cells ; Tularemia ; Vaccination ; Vaccines

Animals ; Animals, Domestic ; China ; epidemiology ; Coinfection ; Coxiella burnetii ; Francisella tularensis ; Tick-Borne Diseases ; epidemiology ; transmission ; veterinary

To use in the identification of Legionella pneumophila isolates, one Legionella genus-specific, an two L. pneumophila species-specific monoclonal antibodies (MAbs) were produced. Reactivities of the MAbs with Legionella species and non-Legionella strains were tested by immunoblot. MAb 7D8-F1, MAb 7D8-A9, and MAb 1G12-C11 reacted only with protein. MAb 7D8-F1 recognized a genus-specific epitope in the protein ranged from 28 to 34 kDa in the tested 40 species with 61 serogroups. MAb 7D8-A9 and MAb 1G12-C11 reacted strongly with 49 kDa and 70 kDa protein from 18 L. pneumophila serogroups but did not react with 39 non-L. pneumophila species, respectively. In addtion, 17 non-Legionella strains, including Coxiella burnetii, Klebsiella pneumoniae, Mycoplasma pneumoniae, Pseudomonas areoginosa, Francisella tularensis, and Staphylococcus pneumoniae did not show cross-reacivities with the three MAbs. The MAbs reacted with 28 environmemtal isolates and three clinical isolates previously identified as L. pneumophila. When the immunoblot patterns were divided into four types (type I~IV) by using MAb 7D8-F1, all the 31 isolates belonged to the type II. These results indicate that the MAbs were highly specific to Legionella and can be used for the identification of L. pneumophila isolates on the genus and species levels.
Antibodies, Monoclonal* ; Coxiella burnetii ; Francisella tularensis ; Klebsiella pneumoniae ; Legionella pneumophila ; Legionella* ; Mycoplasma pneumoniae ; Pneumonia ; Pneumonia, Mycoplasma ; Pseudomonas ; Staphylococcus

Tularemia is a major laboratory acquired zoonoses caused by Francisella tularensis that have high virulence, and usually transmitted to humans from direct contact with infected wild animals like rabbits or insect vectors like ticks. Clinical tularemia can be divided with 6 major syndromes that are delineated by the mode of organism aquisition, in which ulceroglandular type is the most common. F. tularensis have 3 different biogroups which have homogeneous antigenecity, type A (biogroup tularensis), type B (biogroup palearctica) and biogroup novicida, and can be confirmed by serology most frequently. In the domestic area, there was no reports of tularemia in humans or presence of bacteria in the reservoirs. Authers experienced a case of tularemia which is suspected as F. tularensis type B, ulceroglandular type. A healthy 40-year-old man admitted the hospital for lymph node swelling in both axillary and upper arm area and for furuncles in both forearm and palm. He contacted with dead rabbit and eated it after cooking before 20 days from admission day. In laboratory cultures, F. tularensis did not grow in any of the routine or anaerobic culture media except for one blood agar plate at 5 days. After subculturing that to cystine containing chocolate agar plate at 37C degree, 5% CO2 incubator, we could see the accelerating growth of colony. In microbiological test, it was oxidase and urease negative. In acid production in cystine trypticase agar base, it was glucose positive and sucrose, maltose, glycerol negative. In agglutinating test, F. tularensis antiserum titer (Difco, USA) with isolates was 1:160 or over and antibody titer to F. tularensis antigen (Difco, USA) was 1:320 or over. Anti-F. tularensis-IF assay and Anti-F. tularensis-indirect-EIA with isolates were positive.
Adult ; Agar ; Animals ; Animals, Wild ; Arm ; Bacteria ; Cacao ; Cooking ; Culture Media ; Cystine ; Forearm ; Francisella tularensis* ; Francisella* ; Furunculosis ; Glucose ; Glycerol ; Humans ; Incubators ; Insect Vectors ; Lymph Nodes ; Maltose ; Oxidoreductases ; Rabbits ; Sucrose ; Ticks ; Tularemia* ; Urease ; Virulence ; Zoonoses

PURPOSE: Tularemia is an infection caused by Francisella tularensis. Its diagnosis and treatment may be difficult in many cases. The aim of this study was to evaluate treatment modalities for pediatric tularemia patients who do not respond to medical treatment. METHODS: A single-center, retrospective study was performed. A total of 19 children with oropharyngeal tularemia were included. RESULTS: Before diagnosis, the duration of symptoms in patients was 32.15±17.8 days. The most common lymph node localization was the cervical chain. All patients received medical treatment (e.g., streptomycin, gentamicin, ciprofloxacin, and doxycycline). Patients who had been given streptomycin, gentamicin, or doxycycline as initial therapy for 10–14 days showed no response to treatment, and recovery was only achieved after administration of oral ciprofloxacin. Response to treatment was delayed in 5 patients who had been given ciprofloxacin as initial therapy. Surgical incision and drainage were performed in 9 patients (47.5%) who were unresponsive to medical treatment and were experiencing abcess formation and suppuration. Five patients (26.3%) underwent total mass excision, and 2 patients (10.5%) underwent fine-needle aspiration to reach a conclusive differential diagnosis and inform treatment. CONCLUSION: The causes of treatment failure in tularemia include delay in effective treatment and the development of suppurating lymph nodes.
Biopsy, Fine-Needle ; Child ; Ciprofloxacin ; Diagnosis ; Diagnosis, Differential ; Doxycycline ; Drainage ; Francisella tularensis ; Gentamicins ; Humans ; Lymph Nodes ; Retrospective Studies ; Streptomycin ; Suppuration ; Treatment Failure ; Tularemia

OBJECTIVES: Tularemia is a zoonotic disease transmitted by direct contact with infected animals and through arthropod bites, inhalation of contaminated aerosols, ingestion of contaminated meat or water, and skin contact with any infected material. It is widespread throughout the northern hemisphere, including Iran and its neighbors to the north, northeast, and northwest. METHODS: In this paper, the epidemiology of tularemia as a re-emerging infectious disease in the world with a focus on Iran and the neighboring countries is reviewed. RESULTS: In Iran, positive serological tests were first reported in 1973, in wildlife and domestic livestock in the northwestern and southeastern parts of the country. The first human case was reported in 1980 in the southwest of Iran, and recent studies conducted among at-risk populations in the western, southeastern, and southwestern parts of Iran revealed seroprevalences of 14.4, 6.52, and 6%, respectively. CONCLUSIONS: Several factors may explain the absence of reported tularemia cases in Iran since 1980. Tularemia may be underdiagnosed in Iran because Francisella tularensis subspecies holarctica is likely to be the major etiological agent and usually causes mild to moderately severe disease. Furthermore, tularemia is not a disease extensively studied in the medical educational system in Iran, and empirical therapy may be effective in many cases. Finally, it should be noted that laboratories capable of diagnosing tularemia have only been established in the last few years. Since both recent and older studies have consistently found tularemia antibodies in humans and animals, the surveillance of this disease should receive more attention. In particular, it would be worthwhile for clinical researchers to confirm tularemia cases more often by isolating F. tularensis from infected humans and animals.
Aerosols ; Animals ; Antibodies ; Arthropods ; Bacterial Infections ; Communicable Diseases, Emerging* ; Eating ; Epidemiology ; Francisella tularensis ; Humans ; Inhalation ; Iran* ; Livestock ; Meat ; Rodentia ; Seroepidemiologic Studies ; Serologic Tests ; Skin ; Tularemia* ; Water ; Zoonoses

OBJECTIVETo develop a fast, high-throughput screening method with suspension array technique for simultaneous detection of biothreat bacteria.

METHODS16 S rDNA universal primers for Bacillus anthracis, Francisella tularensis, Yersinia pestis, Brucella spp.and Burkholderia pseudomallei were selected to amplify corresponding regions and the genus-specific or species-specific probes were designed. After amplification of chromosomal DNA by 16 S rDNA primers 341A and 519B, the PCR products were detected by suspension array technique. The sensitivity, specificity, reproducibility and detection power were also analyzed.

RESULTSAfter PCR amplification by 16 S rDNA primers and specific probe hybridization, the target microorganisms could be identified at genus level, cross reaction was recognized in the same genus. The detection sensitivity of the assay was 1.5 pg/microl (Burkholderia pseudomallei), 20 pg/microl (Brucella spp.), 7 pg/microl (Bacillus anthracis), 0.1 pg/microl (Francisella tularensis), and 1.1 pg/microl (Yersinia pestis), respectively. The coefficient of variation for 15 test of different probes was ranged from 5.18% to 17.88%, it showed good reproducibility. The assay could correctly identify Bacillus anthracis and Yersinia pestis strains in simulated white powder samples.

CONCLUSIONThe suspension array technique could be served as an opening screening method for biothreat bacteria rapid detection.

Bacillus anthracis ; isolation & purification ; Bioterrorism ; prevention & control ; DNA Primers ; DNA, Bacterial ; analysis ; Francisella tularensis ; isolation & purification ; Oligonucleotide Array Sequence Analysis ; methods ; Polymerase Chain Reaction ; methods ; RNA, Ribosomal, 16S ; genetics ; Yersinia pestis ; isolation & purification