Genetic and Phenotypic Variation of Campylobacter jejuni NCTC11168 Caused by flhA Mutation during Laboratory Passage / 生物医学与环境科学（英文）

OBJECTIVE@#Campylobacter jejuni NCTC11168 is commonly used as a standard strain for flagellar biosynthesis research. In this report, two distinguished phenotypic isolates (CJ1Z, flhA mutant strain, lawn; CJ2S, flhA complemented strain, normal colony) appeared during laboratory passages for NCTC11168.@*METHODS@#Phenotypic assessments, including motility plates, transmission electron microscopy, biofilm formation assay, autoagglutination assay, and genome re-sequencing for these two isolates (CJ1Z, flhA mutant strain; CJ2S, flhA complemented strain) were carried out in this study.@*RESULTS@#Transmission electron microscopy revealed that the flagellum was lost in CJ1Z. Phenotypic assessments and genome sequencing of the two isolates were performed in this study. The capacity for biofilm formation, colony auto-agglutination, and isolate motility was reduced in the mutant CJ1Z. Comparative genomic analysis indicated a unique native nucleotide insertion in flhA (nt, 2154) that caused the I719Y and I720Y mutations and early truncation in flhA.@*CONCLUSION@#FlhA has been found to influence the expression of flagella in C. jejuni. To the best of our knowledge, this is the first study to describe the function of the C-terminal of this protein.

Establishment and Modification of Ninety-seven Pneumococcal Serotyping Assays Based on Quantitative Real-time Polymerase Chain Reaction / 生物医学与环境科学（英文）

OBJECTIVE@#To establish and modify quantitative real-time polymerase chain reaction (qPCR)-based serotyping assays to distinguish 97 pneumococcal serotypes.@*METHODS@#A database of capsular polysaccharide ( cps) loci sequences was generated, covering 97 pneumococcal serotypes. Bioinformatics analyses were performed to identify the cps loci structure and target genes related to different pneumococcal serotypes with specific SNPs. A total of 27 novel qPCR serotyping assay primers and probes were established based on qPCR, while 27 recombinant plasmids containing serotype-specific DNA sequence fragments were constructed as reference target sequences to examine the specificity and sensitivity of the qPCR assay. A panel of pneumococcal reference strains was employed to evaluate the capability of pneumococcal serotyping.@*RESULTS@#A total of 97 pneumococcal serotyping assays based on qPCR were established and modified, which included 64 serotypes previously reported as well as an additional 33 serotypes. Twenty-seven novel qPCR serotyping target sequences were implemented in the pneumococcal qPCR serotyping system. A total of 97 pneumococcal serotypes, which included 52 individual serotypes and 45 serotypes belonging to 20 serogroups, could not be identified as individual serotypes. The sensitivity of qPCR assays based on 27 target sequences was 1-100 copies/µL. The specificity of the qPCR assays was 100%, which were tested by a panel of 90 serotypes of the pneumococcal reference strains.@*CONCLUSION@#A total of 27 novel qPCR assays were established and modified to analyze 97 pneumococcal serotypes.

Comparison of the Pathogenicity of Isolates of Hyperinvasive Sequence Type 7 Belonging to Serogroups A, B, C, and X / 生物医学与环境科学（英文）

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.

Comparative analysis of two incision-making methods for manual small incision cataract surgery / 国际眼科杂志(Guoji Yanke Zazhi)

@#AIM:To evaluate the effect of two incision-making methods on operation and postoperative effect in manual small incision cataract surgery(MSICS)for patients with hard nucleus aged cataract and evaluate the advantages and disadvantages of two incision methods.<p>METHODS: A retrospective analysis of 56 patients with senile cataract with hard nucleus from February 2017 to February 2019 in our hospital was made, which was divided into two groups according to the different surgical methods. group A(31 eyes)with long incision(about 7-8mm), long tunnel(central 5mm length 3.5-4mm, internal incision of both sides extending about 1-1.5mm to the back of the side, making the front end of the incision trapezoid), thick scleral flap(about 2/3 film thickness). group B(25 eyes)with short incision(about 5.5mm), short tunnel(long 3mm, regular flush of internal incision, linear), and regular thickness scleral flap(about 1/2 film thickness). The best corrected visual acuity recovery of 1d, 1wk, 1mo and 3mo after operation, central corneal thickness after 1d, 1wk operation and corneal astigmatism degree, corneal endothelial cell loss degree after 3mo operation were compared.<p>RESULTS: The best corrected visual acuity(greater than or equal to 0.5)for 1d, 1wk, 1mo and 3mo after operation in the two groups(77%, 90%,94% and 94% in the A group and 32%, 72%, 88% and 88% in the B group)was statistically significant \〖<i>β</i>=-1.338, Exp<i>(β)</i>=0.262, <i>P</i><0.05\〗. The central corneal thickness of the two groups had time difference and interaction effect before and after operation(<i>P</i><0.05), and there was no difference between the two groups(<i>P</i>>0.05). There was a statistically significant difference in corneal endothelial cell density(2159.84±245.20/mm2 in the group A and 2019.68±203.97/mm2 in the group B)between the two groups after 3mo of operation(<i>t</i>=2.289, <i>P</i><0.05). There was no significant difference in corneal astigmatism between the two groups(group A 1.57±0.74D and group B 1.39±0.71D)after 3mo of operation(<i>t</i>=0.930,<i>P</i>>0.05).<p>CONCLUSION: MSICS with long incision, long tunnel, thick scleral flap and trapezoidal internal incision has less damage, quicker recovery and better effect on patients with hard nucleus aged cataract than short incision, short tunnel and linear internal incision.

A Cross-sectional Survey Assessing Carriage of Streptococcus pneumoniae in a Healthy Population in Xinjiang Uygur Autonomous Region of China / 生物医学与环境科学（英文）

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.

Molecular characteristics of Neisseria meningitidis isolated during an outbreak in a jail: association with the spread and distribution of ST-4821 complex serogroup C clone in China / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>To characterize the meningococcal strains isolated from cases and close contacts with meningococcal disease associated with an outbreak in a jail in May 2010 by investigating the national distribution of hyperinvasive ST-4821 serogroup C clone associated with this outbreak.</p><p><b>METHODS</b>The cases were described based on the clinical symptoms and laboratory results. Pharyngeal swabs were cultured for N. meningitidis from men in the jail. Meningococcal isolates were identified by serogrouping, pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST), respectively. Four hundred and sixteen serogroup C N. meningitidis strains were collected from 27 provinces between 2003 and 2010 for a nationwide survey and analyzed by PFGE and MLST.</p><p><b>RESULTS</b>Three persons in a jail system were infected with invasive N. meningitidis serogroup C. All isolates tested had matching PFGE patterns and belonged to the multilocus sequence type (ST) 4821 clonal complex. All 47 N. meningitidis strains were identified from the pharyngeal swabs of 166 peoples in the jail, and 26 of them belonged to ST-4821 serogroup C clone, and 90.14% (375/416) serogroup C strains identified in the nationwide survey belonged to the ST-4821 complex. The ST-4821 serogroup C clone was spread nationwide, distributed in 24 provinces, especially in eastern provinces between 2003 and 2010.</p><p><b>CONCLUSION</b>Endemic transmission and carriage rate of ST-4821 serogroup C clone are high in this jail system. The ST-4821 serogroup C clone is spreading in China and nationwide distributed despite the existence of some effective vaccines.</p>

Three quantitative methods to continuously monitor Legionella in spring water / 中华预防医学杂志

<p><b>OBJECTIVE</b>To compare the detection effect of Legionella pollution in spring water by three methods, namely traditional plating method, fluorescent quantitation PCR method and ethidium monoazide (EMA) fluorescent quantitation PCR method.</p><p><b>METHODS</b>Every month (except May), we collected 11 water samples from the 5 selected hot spring pools in one hot spring resort in Beijing in 2011. A total of 121 water samples were collected, and then were detected by the above three methods qualitatively and quantitatively.</p><p><b>RESULTS</b>In our study, the Legionella pollution rate was separately 74.4% (90/121), 100.0% (121/121) and 100.0% (121/121) by the above three methods. The quantitative value of Legionella in the 121 water samples detected by the three methods were around 0.10-216.00 colony-forming units (CFU)/ml, 1.47-1557.75 gene units (GU)/ml and 0.20-301.69 GU/ml, respectively. The median (25th and 75th percentiles) was 75.30 (32.51-192.10) GU/ml, 36.46 (16.08-91.21) GU/ml and 5.30 (0.00-33.70) CFU/ml, respectively. The difference in the quantitative value of Legionella detected by the three methods showed statistical significance (χ(2) = 187.900, P < 0.01). The quantitative value of Legionella detected by fluorescent quantitation PCR method was the highest, followed by the value Legionella detected by EMA-fluorescent quantitation PCR method and traditional plating method.</p><p><b>CONCLUSION</b>The sensitivity of the PCR methods was higher than traditional plating method, in detecting Legionella pollution in spring water, especially the EMA- fluorescent quantitation PCR method, which was more suitable for detecting Legionella in water.</p>

Contamination status and molecular biological characteristics of Legionella in cooling water samples from different places in Wuxi city / 中华预防医学杂志

<p><b>OBJECTIVE</b>To investigate the contamination state of Legionella in cooling water samples from different places in Wuxi city and reveal the molecular biological characteristics of Legionella strains.</p><p><b>METHODS</b>112 parallel water samples (500 ml each) were collected from 56 sites in Wuxi city during year 2009 - 2010. The samples were used for Legionella test and quantitative culture. The isolated Legionella strains were used for serotyping, pulsed-field gel electrophoresis (PFGE), sequence-based typing (SBT), and intracellular growth were tested.</p><p><b>RESULTS</b>The positive proportion of Legionella was 39. 3% (22/56) among all sampling sites. A total of 29 Legionella strains were isolated, and the serotypes include LP1, LP3, LP5 and LP6. LP1 serotype was the major one with a proportion of 65.5% (19/29). 29 Legionella strains got 17 PFGE types. There were 10 SBT types among 10 Legionella strains with different PFGE types. Comparing to LP1 strain (ATCC 33152), WX2011062 (LP6) and WX2011067 (LP5) had strong intracellular growth ability in mouse peritoneal macrophages J774 cell line (the amount of intracellular bacteria on day 0 after infection were (5.5 +/- 1.32) x 10(5), (3.9 +/- 0.60) x 10(5), (7.8 +/- 0.76) x 10(5) CFU/ml, respectively; the amount of intracellular bacteria on day 3 after infection were (58.3 +/- 1.61) x 10(5), (2700.0 +/- 655.74) x 10(5), (3066.7 +/- 208.17) x 10(5) CFU/ml, respectively).</p><p><b>CONCLUSION</b>The Legionella contamination existed in cooling water samples from different places in Wuxi city. Legionella strains isolated showed high genetic variation. Some Legionella strains had vigorous intracellular growth ability.</p>

Comparison on the levels of human serum antibody against Neisseria meningitidis serogroup C measured using serum bactericidal assav and ELISA / 中华流行病学杂志

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.

Sequence-based typing of 82 strains of serotype I Legionella pneumophila isolated from 9 provinces in China / 中华预防医学杂志

<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>

Molecular typing of Legionella pneumophila isolates from China with pulsed field gel electrophoresis / 中华流行病学杂志

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.

Genotypic variability and persistence of Legionella pulsed-field gel electrophoresis patterns in 16 cooling towers in Shanghai, China / 中华流行病学杂志

Objective To investigate the genotypic characteristics and persistence of Legionella pulsed-field gel electrophoresis (PFGE) patterns in 16 air-conditioner cooling towers in six different public sites of Shanghai. Methods From May to October, continuous sampling was operated once per month in 2007. Legionella strains isolated from the 16 cooling towers were confirmed by serological and latex agglutination. PFGE was applied for the fingerprinting of the isolates, while the culster results of PFGE were analyzed by BioNumerics software. Results 131 strains of Legionella were isolated, including L. pneumophila, L. bozemanae, L. micdadei and L anisa.52 distinguishable PFGE patterns were differentiated among the 16 cooling towers, with 37 patterns were owned by just one cooling tower, which was not shared with other cooling towers, while 15 patterns were shared by more than 2 cooling towers. All the cooling towers had ≥2 PFGE patterns,while in 13 cooling towers the same PFGE patterns were recovered during the six months. From June to October of 2007, 18 strains of Legionella belonging to the PFGE pattern of LPAs. SH0078 were isolated continuously from 6 cooling towers. Conclusion This study demonstrated great genotypic diversity and complexity of Legionella in cooling towers. Persistence of the PFGE patterns was observed in 81.25% of the cooling towers. The PFGE pattern of LPAs. SH0078 was distributed widely,suggesting it might be the dominate strain in Shanghai.

Molecular typing of Klebsiella pneumonia by pulse-field gel electrophoresis in combination with multilocus sequence typing / 中华流行病学杂志

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.

Establishment and application of PCR method in serotyping of Streptococcus pneumonia / 中华流行病学杂志

Objective To establish a method based on PCR for serotyping of Streptococcus pneumonia isolates. PCR serotyping method was applied for investigating the serotypes of S. pneumonia strains. Methods 12 pairs of primers targeting different serotypes or S. pneumonia were designed and synthesized. After optimizing the PCR amplification reaction,sensitivity and specificity of each pair was performed. We applied the PCR methods for testing the serotypes of the isolated S. pneumonia strains. Results Each pair of primers showed satisfied PCR sensitivity and specificity. Of all 119 S. pneumonia strains tested by PCR serotyping method,113 isolates were identified (3,5,6A/B,9A/V,14,18,19A,19F,23F) with 6 isolates were unable to be serotyped. Conclusion We deveioped a simple,reliabie and economic method for S. pneumonia serotyping which could be used for testing the serotypes of S. pneumonia that had been prevailed among general population.

Proteome analysis of Neisseria meningitidis serogroup strains C associated with outbreaks in China / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>During 2003-2005, an outbreak of meningitis due to Neisseria meningitidis serogroup C occurred in China. With the aim to find strain clues result in the final epidemics, the ancestral strain 053442, a clinical isolate, and a carrier strain 053426 with different gene type were analyzed.</p><p><b>METHODS</b>Clinical strain 053442 and carrier strain 053426 were cultured on GC agar plates under the same condition. Two-dimensional electrophoresis was performed using the pH 3-10 nonlinear IPG strips of 24 cm length, and all the protein spots were identified by matrix-assisted laser desorption/ionization time of flight spectrometry.</p><p><b>RESULTS</b>502 and 380 protein spots were identified in 053426 and 053442 respectively, relating to 266 and 202 different genes covering a wide range of cellular functions. The express volume and number of proteins involved in energy metabolism, protein synthesis and amino acid biosynthesis in 053426 were higher than in 053442. Virulence factor Opa, Opc and a series of proteins involved in pilus assembly and retraction were identified in 053442, which appear to be of primary importance in colonization and invasion of human cells. Compared to 053442, virulence protein species were less in 053426, with lower express volumes too. No Opa and Opc were detected in 053426.</p><p><b>CONCLUSIONS</b>The different protein expression profiles of the clinical strain 053442 and carrier strain 053426 in the present study provide some clues of the different pathogenicity of the two strains, which may account for result in the final epidemics.</p>

Distribution and molecular epidemiologic characterics of insertion sequence IS1301 in Neisseria meningitidis isolated from China / 中华流行病学杂志

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.

Study on the bactericidal antibody against Neisseria meningitidis serogroup C strains after immunization with a divalent polysaccharide (A plus C) vaccine / 中华流行病学杂志

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.

Detection of Haemophilus influenzae by multiplex polymerase chain reaction method / 中华流行病学杂志

<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>

Establishment and application of TaqMan Real-Time PCR in detection and serogrouping of Neisseria meningitidis / 中华流行病学杂志

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.

Molecular typing of Neisseria meningitidis serogroup C strains with pulsed field gel electrophoresis in China / 中华流行病学杂志

<p><b>OBJECTIVE</b>To study the characteristics of epidemiology and molecular typing on Neisseria meningitidis serogroup C strains associated with outbreaks of Anhui province and sporadic cases in China, using pulsed field gel electrophoresis (PFGE).</p><p><b>METHODS</b>212 Neisseria meningitidis serogroup C strains were isolated from invasive meningococcal cases, close contacts and healthy carriers, including 48 strains from Anhui province with 38 strains associated with serogroup C outbreaks. PFGE were performed by genomic DNA digestion with Nhe I restriction enzyme. The results of PFGE were analyzed by BioNumerics software (Version 4.0, Applied Maths BVBA, Belgium).</p><p><b>RESULTS</b>A total number of 212 Neisseria meningitidis serogroup C isolates were typed by 43 patterns, named AH1 to AH43. In China, AH1 pattern was the major PFGE pattern with 69.3% (n = 147) of all strains, distributed in 11 provinces. Three types of PFGE patterns (AH1 to AH3) were found in 48 strains from Anhui province, in which, 93.8% (n = 45) belonged to AH1. 97.4% (n = 37) of 38 strains associated with serogroup C outbreaks in Anhui province showed AH1 pattern. A total of 53 serogroup C strains were isolated from invasive meningococcal cases with 67.9% (36/53) of AH pattern. 71.9% (87/121) of serogroup C strains isolated from contacts of invasive meningococcal cases was AH1 pattern and 63.2% (24/38) of the strains from healthy carriers showed AH1 pattern.</p><p><b>CONCLUSION</b>By PFGE typing and analysis, AH1 pattern of Neisseria meningitidis serogroup C strains was proved to be the main clone which causing the outbreaks in Anhui province and might be responsible for the sporadic serogroup C meningococcal disease epidemics else where in the country.</p>