HLA-DRB typing among polyarticular juvenile idiopathic arthritis and progressive pseudorheumatoid dysplasia patients

Juvenile Idiopathic Arthritis (JIA) is a multi-factorial disease influenced both by environmental and genetic factors. Progressive pseudorheumatoid dysplasia (PPD) is a rare autosomal recessive genetic disorder affecting multiple joints, mimicking JIA. Aim of the work: to reveal the frequency of HLA-DR types among the studied patients and to correlate the different allele variations clinically. Patients and methods: Thirty JIA patients, in addition to 15 molecularly diagnosed PPD patients were subjected to full history taking and clinical examination. HLA-DRB1 typing was performed to 24/30 JIA and 12/15 PPD cases and thirty healthy age and sex matched children who were included as a control group. Results: The JIA patients were 22 females and 8 males with mean age of 15.8 ± 1.96 years and disease duration 5.3 ± 4.4 years. PPD patients were 8 males and 7 females with mean age of 8.7 ± 3.06 years and disease duration 3.95 ± 2.68 years. A significant frequency of HLA-DRB 04 (p = 0.049) among JIA patients was present in comparison to the controls (OR = 2.81, CI:1.02–7.75), other risky alleles were HLA-DRB 10, 13 and 15. However, HLA-DRB 01, 03, 07, 11 and 14 were found to be protective. HLA-DRB 01, 04, 10 and 13 were found to be risky alleles in PPD. However, HLA 03, 07, 11 and 15 were found to be protective alleles among PPD patients. Conclusion: HLA-DRB 04 was found in a higher frequency in JIA patients with a significant difference in comparison to the controls, denoting that it may play a role in the genetic pathogenesis of JIA.     

A 16-plex Y-SNP typing system based on allele-specific PCR for the genotyping of Chinese Y-chromosomal haplogroups

Y-chromosomal SNP (Y-SNP), with its stable inheritance and low mutation, can provide Supplementary information in forensic investigation. While commonly used Y-chromosomal STR haplotypes show their limitations, typing of Y-SNP would become a powerful complement. In this study, a 16-plex Y-SNP typing system based on allele-specific PCR (AS-PCR) was developed to discriminate four dominant Y-chromosomal haplogroups (C-M130, D-CTS3946, N-M231, and O-M175) and 12 predominant sub-haplogroups of O-M175 (O1a-M119, O1a1a1a-CTS3265, O1b-M268, O1b1a2-Page59, O2-M122, O2a1-L127.1, O2a1b-F240, O2a1b1a1-CTS5820, O2a2-P201, O2a2b1a1-M177, O2a2b1a1a1a-Y17728, O2a2b1a2-F114). A series of experimental validation studies including sensitivity, species specificity, male-female mixture and inhibition were performed. The discrimination of the typing system was preliminarily proved with a haplogroup diversity of 0.9239. Altogether, the Y-SNP typing system based on AS-PCR should be capable of distinguishing China’s dominant Y-chromosomal haplogroups in a rapid and reliable manner, thus can be employed as a useful complement in forensic casework.     

Molecular Characteristics of Neisseria meningitidis Isolated during an Outbreak in a Jail: Associatition with the Spread and Distributition of ST-4821 Complex Serogroup C Cltione in China

Objective 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. Methods 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. Results 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. Conclusion 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.     

A case of pneumonia caused by Legionella pneumophila serogroup 12 and treated successfully with imipenem

The patient was an 83-year-old man hospitalized for Haemophilus influenzae pneumonia, who developed recurrent pneumonia after improvement of the initial episode. Legionella pneumophila serogroup 12 was isolated from the sputum, accompanied by increased serum antibody titers to L. pneumophila serogroup 12. Therefore, the patient was diagnosed as having Legionella pneumonia caused by L. pneumophila serogroup 12.Case reports of pneumonia caused by L. pneumophila serogroup 12 are rare, and the case described herein is the first report of clinical isolation of this organism in Japan. When the genotype was determined by the protocol of The European Working Group for Legionella Infections (Sequence-Based Typing [SBT] for epidemiological typing of L. pneumophila, Version 3.1), the sequence type was ST68. Imipenem/cilastatin therapy was found to be effective for the treatment of Legionella pneumonia in this patient.     

Reassessment of MLST schemes for Leptospira spp. typing worldwide

Leptospirosis is a neglected zoonosis of global importance. Several multilocus sequence typing (MLST) methods have been developed for Leptospira spp., the causative agent of leptospirosis.In this study we reassessed the most commonly used MLST schemes in a set of worldwide isolates, in order to select the loci that achieve the maximum power of discrimination for typing Leptospira spp. Global eBURST algorithm was used to detect clonal complexes among STs and phylogenetic relationships among concatenated and individual sequences were inferred through maximum likelihood (ML) analysis. The evaluation of 12 loci combined to type a subset of strains rendered 57 different STs. Seven of these loci were selected into a final scheme upon studying the number of alleles and polymorphisms, the typing efficiency, the discriminatory power and the ratio dN/dS per nucleotide site for each locus. This new 7-locus scheme was applied to a wider collection of worldwide strains. The ML tree constructed from concatenated sequences of the 7 loci identified 6 major clusters corresponding to 6 Leptospira species. Global eBURST established 8 CCs, which showed that genotypes were clearly related by geographic origin and host. ST52 and ST47, represented mostly by Argentinian isolates, grouped the higher number of isolates. These isolates were serotyped as serogroups Pomona and Icterohaemorrhagiae, showing a unidirectional correlation in which the isolates with the same ST belong to the same serogroup.In summary, this scheme combines the best loci from the most widely used MLST schemes for Leptospira spp. and supports worldwide strains classification. The Argentinian isolates exhibited congruence between allelic profile and serogroup, providing an alternative to serological methods.     

中国嗜吞噬细胞无形体分离株多间隔序列分型分析

目的 对我国嗜吞噬细胞无形体分离株进行多间隔序列分型（MST）分型。方法 根据GenBank收录的4株无形体全基因组序列，利用Mauve 2.3.1软件行种内基因组比对，选择具有变异间隔区进行引物设计，通过引物特异性及扩增效率等预实验筛选引物，并对实验菌株进行PCR扩增，分析单核苷酸多态性（SNPs）。将每株菌间隔序列拼接后进行分型分析并构建进化树，分析不同地区、不同动物种类来源菌株遗传变异关系。结果 共筛选出22对引物，用于我国11株无形体分离株 MST 分型，SNPs 分析显示，碱基转换发生率最高（60.2%，251/417），其中 A-G 转换居首（18.9%，79/417），颠换占23.0%（96/417），插入及缺失发生率占16.7%（70/417）。11株菌22条间隔序列拼接后（约11 047 bp）均为独立变异型。进化分析发现山东省莱州地区重症患者分离株（LZ-H1、H2、H3、H4、H5）及当地长角血蜱分离株 LZ-T1 与美国 Webster 株及 HZ 株聚为一簇，而北京分离株BJ-H1 与新疆分离株 XJ-H1 和 XJ-H3 聚为一类。莱州地区另一蜱分离株 LZ-T2 与新疆另外一人源分离株聚为一类，与莱州重症患者分离株遗传关系密切。结论 我国无形体分离株间隔序列存在明显的遗传多态性，MST分型技术可用于无形体疫情暴发时的快速诊断和疫情追踪。     

Retrospective analysis of molecular biology mechanism of ABO blood group typing discrepancy among blood donors in Jinan blood station

BackgroundTo accurately identify ABO blood typing in pre-transfusion testing is very important to ensure blood transfusion safely, which is a major responsibility of blood station.MethodsEighty-one blood donors samples with ABO blood group typing discrepancy was collected among 61952 donor samples in our blood station from January 2019 to July 2020. Blood group serological method was used to detect ABO blood group. DNA Sequencing was used to determine the genotype. The antibody screening test detects antibodies other than ABO.ResultsIn total, 61,952 donor samples were analysed for ABO typing discrepancies. The incidence among blood donors was 0.13% (81/61952). The most common reason of ABO typing discrepancies was due to specific antibody or non-specific agglutination (54.32%, 44/81), mainly anti-M antibody, cold autoantibody, anti-D antibody, anti-N antibody and anti-Lea antibody. The major cause of forward typing discrepancies among blood donors was ABO subgroups (25.93%, 21/81), including 10 cases of A subtype (1 case of A2, 2 cases of A3, 2 cases of Ax, 3 cases of AxB, 1 case of Ael, 1 case of Ahm), 6 cases of B subtype (2 cases of B3, 1 case of Bel, 3 cases of AB3), 2 cases of B subtype (A), 1 case of cisAB, and 2 cases of acquired B. The serum antibody was weakened in 16 cases (19.75%).ConclusionsThe blood types should be correctly identified by combining serology with gene sequencing to ensure the safety of clinical blood transfusion, when the forward and reverse typing discrepancies among the blood donors.     

Evaluation of the ERIC-PCR as a probable method to differentiate Avibacterium paragallinarum serovars

Infectious coryza, an upper respiratory tract disease in chickens, caused by Avibacterium paragallinarum, leads to huge economic losses. The disease is controlled through vaccination; but vaccination efficacy is dependent on correct identification of the infecting serovar, as limited cross-protection is reported amongst some serovars. Current identification methods include the heamagglutination inhibition test, which is demanding and could be subjective. To overcome this, molecular typing methods proposed are the Multiplex polymerase chain reaction (PCR) and Restriction Fragment Length Polymorphism-PCR, but low reproducibility is reported. Enterobacterial Repetitive Intergenic Consensus (ERIC)-PCR has been suggested for molecular groupings of various bacterial species. This study focuses on evaluating the ERIC-PCR as a probable method to differentiate between different Av. paragallinarum serovars by grouping with reference isolates, based on clonal relations. The ERIC-PCR was performed on 12 reference isolates and 41 field isolates originating from South Africa and South America. The data indicate that the ERIC-PCR is not ideal for the differentiation or for molecular typing of Av. paragallinarum serovars, as no correlation is drawn upon comparison of banding patterns of field isolates and reference strains. However, the results do indicate isolates from the same origin sharing unique banding patterns, indicating potential clonal relationship; but when compared to the reference isolates dominant in the specific area, no correlation could be drawn. Furthermore, although the ERIC-PCR serves a purpose in epidemiological studies, it has proved to have little application in differentiating amongst serovars of Av. paragallinarum and to group untyped field strains with known reference strains.