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Objective:To analyze the clinical characteristics of pertussis cases diagnosed by two pathological detection methods: bacterial culture and real-time polymerase chain reaction (RT-PCR), and to explore the applicable value of two pathological detection methods in the diagnosis of pertussis.Methods:Bilateral nasopharyngeal swabs and clinical information of 165 children suspected of pertussis were collected by Hebei Children′s Hospital from April 2019 to January 2020. The bacterial culture and RT-PCR for nasopharyngeal swab specimens were performed in all cases. Chi-square test was used to analyze the cases of pertussis diagnosed by the above two methods.Results:Based on clinical diagnosis, the sensitivity of bacterial culture and RT-PCR for the diagnosis of pertussis was 61.70% (58/94) and 86.17% (81/94), and the specificity was 92.96% (66/71) and 71.83% (51/71), respectively. The positive rate of RT-PCR in children of all ages, seasons and cough courses is higher than that of bacterial culture. Children with pertussis diagnosed by bacterial culture and RT-PCR were basically similar in age, season, and cough course distribution, with the most common cases ≤3 months old, a high incidence trend in summer and autumn, and the course of coughing in children was mostly within 15-21days. The positive rate of bacterial culture in the diagnosis of pertussis in children is affected by the age of the children, and there are statistical differences between children in different age groups (χ2= 11.929, P=0.036). The positive rate of bacterial culture was the highest in children with >3 years old (51.85% [14/27]), followed by children with ≤3 months old (48.72% [19/39]), and the lowest in children with >6-12 months old (15.00% [3/20]). Moreover, the positive rate of bacterial culture in the diagnosis of pertussis in children is also affected by the cough course of the children, and there are statistical differences between children in different cough course groups (χ2=9.841, P=0.020). The positive rate of bacterial culture was the highest in children with cough course 15-21 days (49.23% [32/65]), followed by 43.59% (17/39) in children with cough course 8-14 days, and the lowest in children with cough course of less than 7 days (22.86% [8/35]). Conclusions:Compared with RT-PCR, bacterial culture has lower sensitivity and higher specificity in the detection of pertussis. These two detection methods have their own advantages and limitations. Medical institutions at all levels should comprehensively analyze different laboratory detection methods. Only by combining the two methods can the diagnostic value and level be effectively improved.
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Objective@#To understand the potential viral pathogens other than enteroviruses existing in samples of hand foot and mouth disease (HFMD) patient and study their molecular feature and genotype.@*Methods@#The deep sequencing analysis of a fecal specimen collected from HFMD patient was conducted by metagenomics and bioinformatics.@*Results@#Enterovirus A71 and sapovirus mixed infection was found in this case. The nucleic acid of sapovirus was confirmed positive by RT-PCR and the 7 429 bp complete genome sequence of sapovirus was obtained by assembling sequencing reads which consisted of 3 open reading frames. Phylogenetic analysis revealed that this strain of virus should belong to the genotype 1 of sapovirus having a homology of 99.4% with sapovirus Hu/G1/Zhejiang1/China/2014 strain, which is a currently predominant genotype circulating in China.@*Conclusions@#The sapovirus, which is a predominant strain circulating in China, was a mixed infected causative agent existing in HFMD sample identified by deep sequencing. This study will serve as a reference for pathogen detection of HFMD and diarrheal related diseases, as well as provide a sequence reference for molecular feature study of sapovirus in China in the future.
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Objective To develop a tetra-primer amplification refractory mutation system PCR(T-ARMS-PCR)assay for detecting four single nucleotide polymorphisms(SNPs): rs1801133,rs1801131, rs1805087 and rs1801394 associated with folate metabolism in a single reaction tube.Methods Methodology was developed.Then, it applied the established method to analyze 150 physical examination children′s anticoagulant blood samples admitted to the Department of Clinical Laboratory, Children′s Hospital of Hebei Province from January 2017 to April 2017.Four sets of chimeric primers consisting of a universal Tag sequence and targeting rs 1801133,rs1801131,rs1805087 and rs1801394 fused to the specific sequence were designed according to the T-ARMS-PCR principle and the multiplex chimeric primers strategy.A single tube PCR was then conducted after optimizing the primer concentrations and the reaction conditions.The amplified products were analyzed by QIAxcel capillary electrophoresis, and the corresponding SNP genotypes of 150 samples were identified.Furthermore,all samples were verified by direct sequencing.And the Hardy-Weinberg Equilibrium(HWE)testing of four SNPs from 150 samples were conducted by SPSS17.0 Chi-square test.Results The improved T-ARMS-PCR combined with capillary electrophoresis can accurately verify eight different alleles of the four SNPs associated with folate metabolism at one time in 3 hours.Four SNPs of 150 whole blood samples were accurately classified and the results were completely consistent with direct sequencing.All the genotype frequencies of these four SNPs were in HWE (χ2rs1801133=0.69, Prs1801133=0.40; χ2rs1801131=0.21, Prs1801131=0.64; χ2rs1805087=3.32, Prs1805087=0.07;χ2rs1801394=1.91, Prs1801394=0.17).Conclusions The proposed improved T-ARMS-PCR combined with capillary electrophoresis in this study can accurately verify four SNPs associated with folate metabolism in a single reaction tube.This method might be a valuable tool to specifically guide the folate supplement in general population.