Rapid detection of Haemophilus influenzae and Haemophilus parainfluenzae in nasopharyngeal swabs by multiplex PCR / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>To establish multiplex PCR-based assays for detecting H.influenzae and H.parainfluenzae. And the PCR-based assays were applied to detect the carriage rates of H.influenzae and H.parainfluenzae in nasopharyngeal swab specimens which were collected from healthy children.</p><p><b>METHODS</b>Multiplex primers for species-specific PCR were designed by using DNAstar soft based on the sequences of 16S rRNA genes from genus Haemophilus to detect H.influenzae and H.parainfluenzae.</p><p><b>RESULTS</b>The sensitivity of the 16S rRNA PCR assay for detecting H.influenzae and H.parainfluenzae was 97.53% and 100% respectively, and the specificity was 95.89% and 96.63% respectively. Youden's Index on the ability to detect H.influenzae and H.parainfluenzae was 0.9342 and 0.9663 respectively. 666 nasopharyngeal swab specimens were collected from healthy children. The detection rates of H.influenzae and H.parainfluenzae were 14.11% and 16.07% respectively by using isolation and culture methods. The detection rates of H.influenzae and H.parainfluenzae were 43.54% and 57.96% respectively by 16S rRNA PCR assays. The carriage rates of serotypes a, b, c, d, e, f and non-typeable isolates were 0% (0/666), 0.15% (1/666), 1.20% (8/666), 0.15% (1/666), 1.20% (8/666), 1.80% (12/666), 95.50% (636/666) respectively.</p><p><b>CONCLUSION</b>The multiplex PCR assays were very rapid, reliable and feasible methods for detection of H.influenzae and H.parainfluenzae in pharyngeal swab specimens which were compared to conventional isolation and culture methods. 95.5% of H.influenzae strains in healthy children were nontypeable. The encapsulated or typable strains were mainly three serotypes which was c, e, and f serotype.</p>

Application of multiple-locus variable-number tandem-repeat analysis in Brucellosis surveillance / 中国地方病学杂志

Objective To establish the standard operating procedures on multiple locus variable number tandem repeat analysis and to evaluate the values in identification of Brucella(B.) melitensis and epidemiological trace-back.Methods Sixteen B.melitensis,22 B.abortus,21 B.suis and 10 B.cnais were investigated by Brucella MLVA-16 genotyping scheme.All data were analyzed using BioNumerics version 5.1 software (AppliedMaths,Belgium).Clustering analysis was based on the categorical coefficient and unweighted pair group method using arithmetic averages(UPGMA) method.Polymorphism at each locus was quantified using Nei's diversity index.Resultant genotypes were compared using the web-based Brucella 2010 MLVA database.Results MLVA methods were successfully established and some strains can be clustered.Bruce06,bruce08,bruce11,bruce12,bruce42,bruce43,bruce45 and bruce55 were useful for species identification of Brucella isolates.Bruce04,bruce07,bruce09,bruce16 and bruce 30 afforded a higher discriminatory power for investigation of strain relatedness in regions of endemicity.Conclusions TheMLVAmethod has proved to be highly discriminatory and epidemiological concordance and is easy for Brucellosis surveillance in province-level lab.

A Flavobacterium lindanitolerans strain isolated from the ascites sample of a Chinese patient with EV71 virus infection / 生物医学与环境科学（英文）

A strain of Flavobacterium lindanitolerans isolated from a sick child's ascites was described. The 16S rRNA gene of the strain was 100% identical to that of Flavobacterium lindanitolerans which was first identified in India in 2008. It was first described that the isolate required X factor (Hemin) for growth in the optimal conditions of 37 °C with 5% CO(2). The isolate produced indole and H(2)S. It did not present hemolytic feature on blood agar.

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.

The Present Status on Classification of Phytoplasmas / 微生物学通报

The history and present status of phytoplasma classification are introduced briefly in this paper.The newly classification methods and rules for the description of Candidatus species are reviewed.The key problems and direction on the classification and identification of phytoplasmas in China are discussed.