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Objective@#To compare the pathogenicity of isolates of sequence type 7 (ST-7) ( ) belonging to four different serogroups (A, B, C, and X).@*Methods@#Four ST-7 isolates serogrouped as A, B, C, and X and characterized by different capsule structures, were examined for their adhesion and invasion properties, and their ability to induce cytokine release and apoptosis in the host cell (the A549 cell line).@*Results@#Among the four ST-7 isolates, the serogroup A isolate possessed the strongest adhesion and invasion ability. This isolate also induced the release of the highest levels of the pro-inflammatory mediators interleukin-6, interleukin-1β, and interferon, and the highest apoptosis rate in the host cells. However, there was no significant difference in interleukin-8 and tumor necrosis factor-α secretion between the four isolates. Based on the findings, the serogroup X isolate had the weakest pathogenicity, whereas there was almost no difference in the pathogenicity of the isolates from serogroups B and C.@*Conclusions@#The differences in the capsular structure of the four isolates of ST-7 affected their pathogenic capacities. The findings also imply that the hyperinvasive ST-7 lineage may include hypoinvasive isolates.
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The carriage rate and serotype distribution of Streptococcus pneumoniae (S. pneumoniae) in a healthy population in China remains unclear. In this study, we collected the oropharyngeal swabs from 513 individuals in Xinjiang, China. Real-time PCR targeting the lytA gene and 12 serotypes were assessed to identify S. pneumoniae carriage. The total carriage rate of S. pneumoniae was 70.4% (361/513). The most prevalent serotypes were 19B/F, 18B/C, 5, and 6A/B. The highest carriage rate of S. pneumoniae was noted in children aged 6-10 years (88.6%), which merits further attention. The co-colonization rate of two or more S. pneumoniae serotypes was 79.8% (264/331). This study aimed to investigate the baseline pneumococcal carriage rate among healthy individuals in China to improve our understanding of the epidemiology of S. pneumoniae.
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Adolescent , Adult , Child , Child, Preschool , Female , Humans , Infant , Male , Middle Aged , Young Adult , Carrier State , Epidemiology , Microbiology , China , Epidemiology , Cross-Sectional Studies , Pneumococcal Infections , Epidemiology , Microbiology , Prevalence , Real-Time Polymerase Chain Reaction , Serogroup , Streptococcus pneumoniae , Classification , GeneticsABSTRACT
Objective To analyze the levels of human serum antibody against Neisseria meningitidis serogroup C measured by serum bactericidal assay (SBA) and ELISA.Methods SBA and a modified ELISA were applied to measure the serum bactericidal titer and the specific concentration of immunoglobulin G (IgG) against meningoeoccal serogroup C in sera samples.Seventy-five sera were from healthy adults without undertaking vaccination while another 429 and 388pre- and post- vaccinated sera were from 143 infants and 194 young children immunized with conjugate vaccine or polysaccharide vaccine,respectively.Correlation between serum bactericidal titer and the concentration of specific IgG against meningococcal serogroup C was analyzed.Results The concentration of meningococcal serogroup C specific IgG in healthy adults showed a strong correlation (r=0.814 33,P＜0.001 ) with serum bactericidal titer through linear regression analysis.Weak correlation was observed between SBA titers and IgG concentration in pre vaccinated sera of infants and children ( conjugate/polysaccharide vaccine ) ( infants:r =0.140 64,P ＞ 0.100/r =0.2899,P＜0.05; children:r=0.540 40,P＜0.05/r=0.194 36,P＜0.05).After immunization with 2-dose conjugate vaccine in infants and 1-dose in children,a strong correlation between the two panels of results was observed (r=0.809 38,P＜0.001 and r=0.837 23,P＜0.001 respectively).However after immunization with polysaccharide vaccine,the correlation between serum bactericidal titer and concentration of specific IgG was weak (r＜0.500 00).Conclusion Among healthy adults and post vaccinated infants or young children immunized with conjugate vaccine,the concentration of specific IgG was comparable to the serum bactericidal titer against meningococcal serogroup C.However,it was not unfavorable to use ELISA as the principal means of measuring serum antibody responses to polysaccharide vaccine for infants under 1 year old.
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Objective To analyze the molecular types of Legionella (L.) pneumophila strains isolated in China,and to develop the PulseNet-China Database of L.pneumophila.Methods Pulsed field gel electrophoresis (PFGE) was used to analyze 262 L.pneumophila strains collected from 11 provinces between 2004 and 2009 in China.Different kinds of genomic DNA in different L.pneumophila strains were isolated and separated after digesting with Asc Ⅰ.BioNumerics software was used to analysis the PFGE fingerprints.Results L.pneumophila strains isolated in China were quite different regarding their PFGE patterns.There were 108 PFGE types among the 262 strains tested in this study.The similarity value of these strains was in the range of 16%-100% and the same types were discovered in different provinces and years.Conclusion L.pneumophila strains isolated in China were with high genetic variations.There might be different clones existed in China.The development of PulseNet China Database was thus of great significance in monitoring the L.pneumophila strains in the future.
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<p><b>OBJECTIVE</b>To analyze the characteristics of Sequence-based Typing (SBT) of the Serotype 1 Legionella pneumophila (Lp1) isolated from environmental water in China, and then create a preliminary database.</p><p><b>METHODS</b>A total of 82 strains of Lp1 isolated from environmental water in 9 provinces of China between 2005 and 2008 were genotyped by SBT method and Pulsed-field Gel Electrophoresis (PFGE) method. The results of the two different typing methods were then compared by cluster analysis, adopting BioNumerics version 5.1 software.</p><p><b>RESULTS</b>By SBT method, the 82 strains of Lp1 were divided into 22 ST types, of which 17 new types and one new allele was discovered. The dominant type was ST-1 type, found in 8 provinces, accounting for 46.3% (38/82). ST-1, ST-150, ST-154, ST-159, ST-160 and ST-630 types were found in more than 2 isolated-sites; while more than 2 different ST types were found in 5 isolated-sites, as site B4, B5, B6, S3 and S8. In cluster analysis, 15 ST types were grouped into three complexes (ST-1 complex, ST-154 complex and ST-149 complex); and the other 7 ST types were not assigned complex. By PFGE method, 46 banding patterns were observed. As a result of the combination of the two methods, the 82 isolates strains could be divided into 54 molecular types, which showed a reliable accordance in the cluster analysis between the two methods.</p><p><b>CONCLUSION</b>The SBT of the Lp1 in environmental water in China was unique. From the study, a preliminary SBT database was set up.</p>
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China , Cluster Analysis , Electrophoresis, Gel, Pulsed-Field , Genotype , Legionella pneumophila , Classification , Genetics , Serotyping , Methods , Water PollutionABSTRACT
Objective To type Klebsiella pneumonia through methods including pulse-field gel electrophoresis (PFGE) in combination with multilocus sequence typing. Methods Four selected different Eps, referring to the Standard Operating Procedure of PulseNet China, were used. The single colony of Klebsiella pneumonia was quantified after enriched culture. Embedding organisms in agarose and genome DNA were lysed with Proteinase K and then digested by restriction endonuclease Xba Ⅰ , to produce agarose gel. Fingerprint was obtained through PFGE and bands were marked with their molecular weights and then analyzed by BioNumerics software. Using MLST to analyze the strains that were highly similar, by PFGE typing Results By comparing the four results from each Eps, fk3 (switch time from 6s to 36s,total run time is 18.5 hours) seemed to be better than the others.59 strains of Klebsiella pneumonia were divided into 47 PFGE types and 19 PFGE clusters. The highly similar strains could be typed into ST-340、ST-342、ST-343、ST-344、ST-345 by MLST. Among them, ST-342、 ST-343、 ST-344、 ST-345 types were all new MLST types that were reported in China.Conclusion Highly similar Klebsiella pneumonias typed by PFGE could also be typed by MLST.
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Objective To optimize the serum bactericidal assay (SBA) , detect and analyze the bactericidal antibody level against Neisseria meningitidis serogroup C strains after divalent polysaccharide (A plus C) vaccine immunization. Methods Two Neisseria meningitidis serogroup C strains, vaccine candidate strain (C11) and epidemic strain (053442), were selected as targets. The national Neisseria meningitidis standardized serum was used as reference serum. Pel-Freez infant rabbit complements was available. The optimized SBA method was used to detect bactericidal antibody against strain C11 and 053442 for 122 pairs of sera before and after immunization with a divalent polysaccharide (A and C) vaccine. Results The strain C11 and 053442 both could be used as targeted strain for SBA. The optimized concentration of targeted strain was achieved when a whole-cell suspension of 0.35 A at 600 nm was diluted 4×104 times. Before immunization, SBA geometric mean titers(GMT) of 122 sera against strain C11 and 053442 were 1: 1.75 and 1:2.63 respectively, and the protective rates were 9.8% and 17.2% respectively. After immunization, the GMTs and the protective rates of 122 sera both rose significantly (P<0.01), the GMTs against strain C11 and 053442 were 1:483.73 and 1:412.57 respectively. The protective rates against strain C11 and 053442 were 100% and 95.9% respectively. Conclusion Immunization with a divalent polysaccharide (A and C) vaccine could elevate remarkably the population SBA titer against Neisseria meningitidis serogroup C strains of different subtypes, but the surveillance of vaccine effect against different targeted strains remains necessary.
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Objective To research the distribution and molecular epidemiology of insertion sequence IS1301 in Neisseria (N.) meningitidis strains in China, so as to provide scientific and available evidence for a new method of genotyping in N.meningitidis strains with IS1301. Methods Examined the IS1301 by PCR in 219 N.meningitidis strains from 16 provinces and 3 cities during 2007 and 2008 in China, productions of amplification were sent for sequencing. The positive N.meningitidis strains were analyzed by pulse field gel electrophoresis (PFGE) and nucleic acid blotting hybridization(Southern blot) by electrophoresis. Results The positive rates with IS1301 were 15.53%, 11.11%, 20.75%, 6.17% and 28.57% for four serotypes (A, B, C, N) respectively. The sequence comparability between the amplification productions and No.Z49092.1 N.meningitidis which registered in GenBank was 94%-100%. There were two types of clusters devided by cladogram analysis. There appeared large IS1301 sequence difference between the serotype C and others. The number of IS1301 replica ranged from 6-17 per strain at least. The number of IS1301 replica changed in the same type of PFGE N.meningitidis respectively. Conclusion Typing by IS1301 combined with PFGE could comprehend the homology and genetic polymorphism of N.meningitidis epidemic strains at the molecular level.
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Objective To establish TaqMan Real-Time PcR method for detection and identification of Neisseria meningitidis.Methods Seven sets of primers and FAM-labeled probes targeting different genes of Neisseria meningitidis were designed and synthesized.ctrA gene was used for identification of N.meningitidis species.Six serogruops(A,B,C,X,Y,W135)of N.meningitidis were detected with following genes:sacB(A),siaD(B),siaD(C),xcbB(X),synF(Y)and synG(W135)respectively.Sensitivity and specificity of Real-Time PCR were assessed for different primers and probes.121cerebrospinal fluid(CSF)specimens from suspected N.meningitidis invasive meningitis cases were detected by latex agglutination test and Real-Time PCR assay simultaneously.Resuits 79 N.meningitidis isolates of different serogroups could be detected and identified by seven sets of primers and probes in this study.Real-Time PCR seemed more sensitive than standard PCR bv 101-103 times.The respective sensitivities for ctrA,sacB,siaD(B),siaD(C),xcbB,synF and synG were 8,8,80,8,8,80,8 genomeDNA copies in each reaction.Of the 121 CSF specimens,11 were positive for Real-Time PCR and 6 for latex agglutination test.Conclusion Real-Time PCR could rapidly detect and identify N.meningitidis of different serogroups and seemed more sensitive.It could be widely used for diagnose of invasive meningitis caused bv N.meningitidis.
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<p><b>OBJECTIVE</b>To develop a rapid method for detecting Haemophilus influenzae by multiplex polymerase chain reaction (M-PCR).</p><p><b>METHODS</b>Primers (Hi) were designed for amplification of p6 gene coding P6 protein of Haemophilus influenzae, which was used to identify Haemophilus influenzae species. Primers (Hi-cap) were designed for amplification of bexA gene which coding capsular polysaccharide (cap) synthesis was used for detecting whether Haemophilus influenzae isolates possess bexA gene relating to cap synthesis. Twelve primers (Hia-Hif) were designed for amplification of cap synthesis gene to identify the cap-type of Haemophilus influenzae. Other relative enteric pathogenic bacteria were amplified by M-PCR to serve as controls. 200 strains isolated from patients were identified. Results from M-PCR were compared to two methods including V and X factors grow requirement test and standard slide agglutination serotyping (SAST).</p><p><b>RESULTS</b>The results indicated that the M-PCR assay was high specificity and sensitivity and might be valuable for differential diagnosis of Haemophilus influenzae. The sensitivity of detection was 0.935 pg. 189 strains out of the 200 belonged to Haemophilus influenzae isolates, and one isolate was cap-type f. An agreement results were seen among the V and X factors grow requirement test, SAST and M-PCR methods.</p><p><b>CONCLUSION</b>M-PCR method showed satisfactory sensitivity, specificity and stability for detecting and identifying Haemophilus influenzae, and could be used in clinic diagnosis, surveillance and rapid diagnosis for plague of Haemophilus influenzae.</p>