[Show abstract][Hide abstract] ABSTRACT: Legionella pneumophila serogroup 1 causes Legionnaires' disease. Water systems contaminated with Legionella are the implicated sources of Legionnaires' disease. This study analyzed L. pneumophila serogroup 1 strains in China using sequence-based typing. Strains were isolated from cooling towers (n = 96), hot springs (n = 42), and potable water systems (n = 26). Isolates from cooling towers, hot springs, and potable water systems were divided into 25 sequence types (STs, index of discrimination [IOD] = 0.711), 19 STs (IOD = 0.934), and 3 STs (IOD = 0.151), respectively. The genetic variation among the potable water isolates was lower than that among cooling tower and hot spring isolates. ST1 was the predominant type, accounting for 49.4% of analyzed strains (n = 81), followed by ST154. With the exception of two strains, all potable water isolates (92.3%) belonged to the ST1. By contrast, 53.1% (51/96) and only 14.3% (6/42) of cooling tower and hot spring isolates belonged to the ST1, respectively. There were differences in the distribution of clone groups among the water sources. The comparisons among L. pneumophila strains isolated in China, Japan, and South Korea revealed that similar clones (ST1 complex and ST154 complex) exist in these countries. In conclusion, in China, STs had several unique allelic profiles and ST1 was the most prevalent sequence type of environmental L. pneumophila serogroup 1 isolates, which was similar to that in Japan and South Korea.
Applied and Environmental Microbiology 01/2014; · 3.95 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Legionella is the causative agent of Legionnaires' disease, and hot springs are a major source of outbreaks of this disease. It is important from a public health perspective to survey hot spring environments for the presence of Legionella.
Prospective surveillance of the extent of Legionella pollution was conducted at three hot spring recreational areas in Beijing, China in 2011. Pulsed-field gel electrophoresis (PFGE) and sequence-based typing (SBT) were used to describe the genetic polymorphism of isolates. The intracellular growth ability of the isolates was determined by interacting with J774 cells and plating the dilutions onto BCYE agar plates.
Overall, 51.9% of spring water samples showed Legionella-positive, and their concentrations ranged from 1 CFU/liter to 2,218 CFU/liter. The positive rates of Legionella were significantly associated with a free chlorine concentration of ≥0.2 mg/L, urea concentration of ≥0.05 mg/L, total microbial counts of ≥400 CFU/ml and total coliform of ≥3 MPN/L (p<0.01). The Legionella concentrations were significantly associated with sample temperature, pH, total microbial counts and total coliform (p<0.01). Legionella pneumophila was the most frequently isolated species (98.9%), and the isolated serogroups included serogroups 3 (25.3%), 6 (23.4%), 5 (19.2%), 1 (18.5%), 2 (10.2%), 8 (0.4%), 10 (0.8%), 9 (1.9%) and 12 (0.4%). Two hundred and twenty-eight isolates were analyzed by PFGE and 62 different patterns were obtained. Fifty-seven L. pneumophila isolates were selected for SBT analysis and divided into 35 different sequence types with 5 main clonal groups. All the 57 isolates had high intracellular growth ability.
Our results demonstrated high prevalence and genetic polymorphism of Legionella in springs in Beijing, China, and the SBT and intracellular growth assay results suggested that the Legionella isolates of hot spring environments were pathogenic. Improved control and prevention strategies are urgently needed.
PLoS ONE 01/2013; 8(3):e59018. · 3.53 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Legionella are prevalent in human-made water systems and cause legionellosis in humans. Conventional culturing and polymerase chain reaction (PCR) techniques are not sufficiently accurate for the quantitative analysis of live Legionella bacteria in water samples because of the presence of viable but nonculturable cells and dead cells. Here, we report a rapid detection method for viable Legionella that combines ethidium monoazide (EMA) with quantitative real-time PCR (qPCR) and apply this method to detect Legionella in a large number of water samples from different sources. Results yielded that samples treated with 5 μg/ml EMA for 10 min and subsequently exposed to light irradiation for 5 min were optimal for detecting Legionella. EMA treatment before qPCR could block the signal from approximately 4 log(10) of dead cells. When investigating environmental water samples, the percent-positive rate obtained by EMA-qPCR was significantly higher than conventional PCR and culture methods, and slightly lower than qPCR. The bacterial count of Legionella determined by EMA-qPCR were mostly greater than those determined by culture assays and lower than those determined by qPCR. Acceptable correlations were found between the EMA-qPCR and qPCR results for cooling towers, piped water and hot spring water samples (r = 0.849, P < 0.001) and also found between the EMA-qPCR and culture results for hot spring water samples (r = 0.698, P < 0.001). The results indicate that EMA-qPCR could be used as a complementary tool for the detection and monitoring of Legionella in water systems, especially in hot spring water samples.
World Journal of Microbiology and Biotechnology (Formerly MIRCEN Journal of Applied Microbiology and Biotechnology) 05/2012; 28(5):1881-90. · 1.35 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Here, we describe four cases of laboratory-confirmed Legionella infection. Case 1 was a culture-confirmed case of Legionella infection in a patient with liver cirrhosis. Following this, three other liver cirrhosis cases (cases 2-4) were diagnosed with Legionella infection as confirmed by quantitative real-time PCR. The cause of the pneumonia was determined as Legionella pneumophila by positive direct fluorescence assay and isolation of the causative agent. The infections were successfully treated by administering appropriate antibiotics. These cases highlight the importance of considering Legionella as a cause of pneumonia in patients with liver disease and lung infections. The strain of L. pneumophila isolated from Case 1 was characterized as being closely related to strain Philadelphila-1 (ATCC 33152(T)), which is the type strain of the species, belonging to serogroup 1 and sequence type 36 (ST36), and is known to be distributed worldwide. To our knowledge, this is the first report of Legionella infection on the Chinese mainland for a decade and highlights the need to raise awareness of diagnostic methods for Legionnaires' disease in China and the requirement for further epidemiological surveillance strategies to monitor this disease.
Journal of Medical Microbiology 03/2012; 61(Pt 7):1023-8. · 2.30 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Legionella pneumophila grows in amoebae and has achieved the ability to grow at various temperatures, although the mechanisms controlling this ability remain poorly understood. The Icm/Dot type IVB secretion system is composed of more than 25 proteins and is known to be essential for intracellular growth. The role of the icmN gene in intracellular multiplication and the effects of culture temperatures on it are not precisely understood. We conducted our investigation using an icmN mutant made by gene replacement mutagenesis. Intracellular growth of the mutant was impaired both in mammalian macrophages and amoeba at 37 °C. In particular, intracellular growth in amoebae was completely impaired at 25 °C. It was found that genes from icmN to icmC formed an operon, i.e., icmN, -M, -L, -E, -G, -C,, and the promoter activity of the icmN operon was stronger at 25 than at 37 °C. It was suggested that icmM and its downstream genes had a secondary promoter that enables icmN mutant grow in amoebae at lower temperatures and macrophages at 37 °C. These results show that the icmN promoter has a low temperature inducible nature, and gene products of the icmN operon require high expression for bacterial proliferation at low temperatures within amoeba.
Canadian Journal of Microbiology 03/2012; 58(4):490-501. · 1.20 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Legionella (Fluoribacter) dumoffii is one of the agents causing Legionnaires' disease. Here, we used Illumina second-generation sequencing technology to decipher for the first time the whole-genome sequences of two strains of this species, TEX-KL and NY-23. The assembly results for both strains consist of one chromosome and two plasmids.
Journal of bacteriology 03/2012; 194(5):1251-2. · 3.94 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Despite extensive childhood immunization, pertussis remains one of the world's leading causes of vaccine-preventable deaths. Incidence of pertussis in adolescents and adults has increased in many countries despite high vaccination coverage. In China, booster vaccinations against diphtheria, tetanus and pertussis are not used in adults, and little is known about pertussis incidence in the age group. The aim of this study was to determine seroprevalence of IgG antibodies to pertussis toxin (PT) and diphtheria among adults in China.
Blood samples were obtained from 210 healthy adults aged 18-50 years in Weifang city, China during the period of May and June 2010. Serum IgG antibodies against PT (anti-PT IgG) and diphtheria were determined by the commercial ELISA kits, respectively. According to the kit, concentration of anti-PT IgG higher than 30 IU/mL was considered positive. An antibody concentration of ≥ 0.1 IU/mL was defined as evidence of seroprotection against diphtheria.
The mean concentration of anti-PT IgG antibodies was 9.95 IU/mL (95% confidence interval (CI) 8.45-11.44). Eleven (5.24%) of the studied subjects were proved to be seropositive to pertussis. Of the 210 subjects, 161 (76.6%) had anti-diphtheria antibody concentration ≥ 0.1 IU/mL and 49 (23.3%) had the antibody concentration between 0.01 and 0.099 IU/mL.
Our study indicated that about 5% of adults aged 18-50 years had positive anti-PT IgG antibodies, suggesting that adult pertussis is not uncommon in China. Although a high proportion of studied subjects had a protective level of immunity against diphtheria, the antibody level decreased with the increasing age of adults. Booster vaccinations against pertussis should be considered in adults in China.
The Journal of infection 08/2011; 63(6):441-6. · 4.13 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: A total of 32 strains of Legionella pneumophila were used to optimize pulsed-field gel electrophoresis (PFGE) for subtyping of L. pneumophila. Twenty-six isolates of L. pneumophila with various origins and 11 isolates from five different water systems were used as the panels. For optimization of electrophoretic parameters (EPs) of SfiI PFGE, 26 isolates were analyzed with SfiI digestion, using four EPs yielding the same D value. The EP of a switch time of 5 to 50 s for 21 h had the smallest similarity coefficients and was declared the optimal EP for SfiI PFGE of L. pneumophila. By software analysis and pilot study, AscI was chosen as another PFGE enzyme. AscI PFGE could cluster the isolates from each water system into the same or very similar patterns and had a high degree of typing concordance with other molecular methods. In evaluating the discriminatory power of AscI with the panel of 26 isolates, AscI PFGE gave one single pattern and a D value of 100%. AscI PFGE had a high discriminatory power and a high degree of consistency with epidemiological data and other molecular typing methods for L. pneumophila subtyping, and hence, AscI could be used as a restriction enzyme in PFGE subtyping of L. pneumophila.
Applied and Environmental Microbiology 03/2010; 76(5):1334-40. · 3.95 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Two hundred and seventy-three Haemophilus influenzae strains isolated from pediatric pneumonia patients in China were studied. We used Multilocus Sequence Typing (MLST) to analyze genotypic characteristics. All strains were biotyped and serotyped. Relatedness and patterns of genes among isolates were determined by the analysis of MLST and eBURST. H. influenzae primarily causes acute pneumonia in children under 1 year old. Nontypeable H. influenzae was responsible for most cases of pediatric pneumonia. All 273 strains were classified into eight biotypes. They mostly belonged to the I, II, and III biotypes (17.6%, 43.6%, and 22.7%, respectively). 62 strains (22.7%) produced beta-lactamase. We found 28 novel alleles. Fifty different STs were found by MLST, of which 39 were novel. These were ST477 through ST508 and ST521 through ST527. Group 17 and predicted founders 503 were new groups in this study. No STs correlated with strains from Korea, which is adjacent to China. The H. influenzae strains from China appeared to have heterogeneous ST types patterns which may be the reason no outbreaks or epidemics of H. influenzae infections have occurred in Chengdu city, Sichuan, China.
The Journal of Microbiology 08/2009; 47(4):494-7. · 1.28 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Outbreaks of a new serogroup C meningococcal disease emerged during 2003-04 (five outbreaks with 43 cases) and in 2004-05 (five outbreaks with 29 cases), all in Anhui province, China. We describe the molecular epidemiology and features of the causative bacterial strains.
We used pulsed-field gel electrophoresis (PFGE) and multi-locus sequence typing (MLST) to analyse the strains.
Of 34 strains of Neisseria meningitidis cultured during 2003-04 from Anhui province, 31 were group C meningococci, 28 of which were associated with three of five outbreaks; one from a patient and 27 from close contacts of eight patients. Of 30 strains isolated from Anhui province during 2004-05, 17 were identified as serogroup C meningococci, ten of which were associated with four of five outbreaks. In a nationwide survey, 542 strains were isolated during 2004-05; 58 were serogroup C meningococci interspersed among 11 other provinces where no serogroup C outbreak occurred. Of the 106 serogroup C strains analysed, 89 had identical PFGE patterns, designated AH1. Of 28 strains selected for MLST analyses, 25 were sequence type 4821 (ST-4821), which did not belong to any of the previously reported sequence types that can form a new hypervirulent lineage.
ST-4821 seems to be unique and caused the serogroup C meningitis outbreaks during the two seasons from 2003 to 2005 in Anhui province. The emergence of this sequence type has epidemiological importance that should be monitored for future spread in China and the rest of the world.
The Lancet 03/2006; 367(9508):419-23. · 39.06 Impact Factor