Polymerase Chain Reaction-sequence Specific Primer Research Articles

Gene polymorphisms of Interleukin 6 (\u2212174 G/C) and transforming growth factor \u03b2-1(+915 G/C) in ovarian cancer patients

BackgroundIn the study on hand, we investigated the effect of IL-6 (−174 G/C; rs 1800795) and TGF-β1 (+915G/C; rs 1800471) gene polymorphisms on the susceptibility to Ovarian Cancer and their effect on plasma levels. IL-6 (−174 G/C) SNP was analyzed using mutagenically separated polymerase chain reaction (MS-PCR) while TGF-β1 +915G/C (codon 25) SNP was investigated by the sequence-specific primer polymerase chain reaction (SSP-PCR). An enzyme-linked immunosorbent assay (ELISA) was used to quantify IL-6 and TGF-β1 plasma levels in 48 ovarian cancer patients and 48 normal controls.ResultsRegarding IL 6 (−174 G/C), a significant increase in CC and GC+CC genotypes parallel with the C allele was considered as risk factors for ovarian cancer; on the other hand, the G allele was considered as a protective factor for ovarian cancer. TGF-β1 (+915G/C) investigations showed a significant elevation in GC and GC+CC genotypes which can be considered as a risk factor for ovarian cancer. Plasma IL-6 and TGF-β1 were higher in ovarian cancer patients compared with controls. No specific genotype or allele could be responsible for the elevation of TGF-β1 in ovarian cancer patients’ plasma, while the highest significant value for IL6 in subjects carrying GG and CC genotypes in comparison with GC genotype.ConclusionsThis study supports an association of IL6 (−174G/C) and TGF-β1 (+915G/C) gene polymorphisms with the susceptibility to ovarian cancer.

Allopurinol-induced severe cutaneous adverse reactions in Vietnamese: the role of HLA alleles and other risk factors.

Aim: To reveal the association of three class I HLA alleles, including HLA-A*33:03, HLA-B*58:01 and HLA-C*03:02, and allopurinol-induced severe cutaneous adverse reactions (SCARs) in Vietnamese patients. Methods: A case-control study on 100 allopurinol-induced SCARs patients, 183 tolerant controls and 810 population controls was performed. The HLA-A*33:03 and HLA-C*03:02 alleles were detected with the nested allele-specific PCR method; the HLA-B*58:01 allele was detected with the sequence-specific primer PCR method. Results: There were strong associations between HLA-B*58:01 and HLA-C*03:02 and allopurinol-induced SCARs. Specific associations were found between HLA-B*58:01 and Stevens-Johnson syndrome/toxic epidermal necrolysis and between HLA-C*03:02 and drug reaction with eosinophilia and systemic symptoms, with a gene dosage effect. The multivariate regression analysis indicated two significant independent risk factors: HLA-B*58:01/HLA-C*03:02 and estimated glomerular filtration rate <60ml/min/1.73m2. The specificity, positive predictive value and negative predictive value of HLA-B*58:01 testing were higher than the HLA-C*03:02 or the multiplex testing, especially in patients with impaired renal function. Conclusion: The results supported pre-treatment HLA-B*58:01 testing in Vietnamese patients with declined renal function to prevent SCARs.

Identification of a rare platelet-specific antigen HPA-10bw allele among ethnic Han Chinese population in Shandong

To study the polymorphism of human platelet antigen (HPA) system 10 among ethnic Han Chinese from Shandong, China so as to supplement the data of platelet donor bank in the region. Peripheral blood samples of platelet donors from the region were genotyped for HPA-10 alleles by PCR-sequence specific primer (PCR-SSP) and direct sequencing. Among 1401 donors, a rare heterozygote carrier of HPA-10w (a+b+) was identified, which gave an allelic frequency of approximately 0.035%. The detection of rare HPA-10bw antigen allele among ethnic Han Chinese from Shandong is useful for the diagnosis and prevention of neonatal alloimmune thrombocytopenia and post-transfusion purpura in the region.

Human Neutrophil Antigen Genotype and Allele Frequencies in Iranian Blood Donors.

Objective Human neutrophil antigens (HNAs) can be targeted by HNA-allo antibodies and cause a variety of clinical conditions such as transfusion-related acute lung injury (TRALI) and neonatal alloimmune neutropenia (NAIN). The current study is aimed at identifying the genotype and allele frequencies of HNAs in Iranian blood donors. Methods A total of 150 blood samples were obtained from healthy blood donors. HNA-1, HNA-3, HNA-4, and HNA-5 were genotyped, using the polymerase chain reaction sequence-specific primer (PCR-SSP) technique. The expression of the HNA-2 antigen on the neutrophil surface was evaluated by flow cytometry. Results The allele frequencies of FCGR3B∗1 (encoding HNA-1a), FCGR3B∗2 (encoding HNA-1b), and FCGR3B∗3 (encoding HNA-1c) were 0.34, 0.63, and 0.03, respectively. For HNA-3, the allele frequencies for SLC44A2∗1 (encoding HNA-3a) and SLC44A2∗2 (encoding HNA-3b) were 0.63 and 0.37, respectively. The frequencies of ITGAM∗1 (encoding HNA-4a) and ITGAM∗2 (encoding HNA-4b) alleles were 0.85 and 0.15, respectively. Furthermore, the frequencies of ITGAL∗1 (encoding HNA-5a) and ITGAL∗2 (encoding HNA-5b) alleles were 0.72 and 0.28, respectively. In the studied population, HNA-2 antigen was present on the neutrophil surface in 97.3% of the individuals, while no detectable HNA-2 expression was observed in 2.7% of the individuals. However, no significant difference in HNA-2 expression between different age groups was found. Conclusion The present study provides the first report of the HNA allele and genotype frequencies among the Iranian population. All HNAs (HNA-1 to HNA-5) were typed using the PCR-SSP and flow cytometer. In the current cohort study, the determined HNA allele frequencies were similar to the previous reports from British, German, and Danish populations. Considering the presence of different Iranian ethnic groups, further studies with a larger sample size are needed to draw a total picture for HNA allele frequencies.

Evaluation of telomeric KIR genes and their association with CMV infection in kidney transplant recipients.

Cytomegalovirus (CMV) infection is a common complication after organ transplantation. Despite theimmunosuppressed state, natural killer (NK) cells remain the major immune defense cells against viral infections in transplanted patients. The present study aimed at elucidating the correlation between the number of inhibitory and activating genes and the incidence of CMV infection in kidney transplanted recipients. Kidney transplanted recipients including 51 CMV+ and 50 CMV- were genotyped for the presence or absence of 4 activating (KIR2DS1, KIR2DS4, KIR2DS5, KIR3DS1) and 2 inhibitory (KIR3DL1, KIR2DL5a) genes using polymerase chain reaction sequence-specific primers (PCR-SSP) assay. Our results showed that CMV infection occurred in 50.49% of kidney allograft recipients. In addition, there was a significant correlation between the presence of theKIR2DS1 activatinggene in the CMV- group compared to the CMV+ group (p = 0.033). The other three activating KIR receptors did not show a correlation with CMV infection. Our results suggest that the prevalence of the KIR activating KIR2DS1 gene may reduce therate of CMV infection after kidney transplantation in our population.

High-resolution melting PCR analysis for genotyping the gene polymorphism of TNF-α, TGF-β1, IL-10, and IFN-γ in lung transplant recipients

High-resolution melting (HRM) analysis is a genotyping method which has the advantages of simple, rapid, low-cost and closed-tube operation.This study evaluated HRM analysis as an option for detecting the single nucleotide polymorphism (SNP) of cytokine, and profiled the distribution of cytokine gene polymorphism in the lung transplant recipients (LTRs).High-resolution melting-polymerase chain reaction (HRM-PCR) assays for genotyping tumor necrosis factor alpha (TNF-α) (-308 A/G), tumor growth factor beta 1 (TGF-β1) (+869 T/C), interleukin 10 (IL-10) (-592 C/A, -819 T/C, -1082 G/A), and interferon gamma (IFN-γ) (+874 T/A) SNPs were developed on the LightCycler® 480. The SNPs of the aforementioned cytokine genes in 322 LTRs and 266 normal controls were detected using HRM-PCR approach. To confirm the accuracy of the HRM-PCR assay, we randomly selected 100 samples from the LTRs and detected the aforementioned SNPs with sequence-specific primer-polymerase chain reaction (SSP-PCR) method, using a commercial kit.The data show that the HRM-PCR assay can distinguish all the cytokine SNPs, and the results of HRM-PCR analysis are in complete concordance to the genotyping results obtained using a commercial kit (κ = 1.0). Our data also show that the allele and genotype frequencies of the abovementioned cytokine are not significantly different between the LTRs and the control groups (p > 0.05). In addition, we found the genotypes of TGF-β1 +869 associated with high expression phenotype were prevalent in the LTRs. On the contrary, for TNF-α -308, IL-10 and IFN-γ, the genotypes associated with low expression phenotype were most common in the LTRs.In this study, we described a rapid, low-cost and high-throughput HRM-PCR technology for genotyping cytokine SNPs. Our data may be utilized for future studies examining the associations of cytokine gene polymorphisms with the prognosis of the LTRs.

Association of HLA-DRB1 and -DQB1 Alleles with Susceptibility to IgA Nephropathy in Korean Patients.

BackgroundAssociations between IgA nephropathy (IgAN) and HLA-DRB1 and -DQB1 alleles have been reported in several ethnic groups. We investigated the association of HLA-DRB1 and -DQB1 alleles with the predisposition for IgAN and disease progression to end-stage kidney disease (ESKD) in Korean patients.MethodsWe analyzed HLA-DRB1 and -DQB1 genotypes in 399 IgAN patients between January 2000 and January 2019 using a LIFECODES sequence-specific oligonucleotide (SSO) typing kit (Immucor, Stamford, CT, USA) or a LABType SSO Typing Test (One Lambda, Canoga Park, CA, USA). Alleles with a significant difference in two-digit resolution were further analyzed using in-house sequence-based typing and sequence-specific primer PCR. As controls, 613 healthy hematopoietic stem cell donors were included. Kidney survival was analyzed in 281 IgAN patients with available clinical and laboratory data using Cox regression analysis. Where needed, P-values were adjusted using Bonferroni correction.ResultsThe allele frequencies of HLA-DRB1*0405 (corrected P [Pc]<0.001), -DQB1*0401 (Pc=0.048), and -DQB1*0302 (Pc=0.021) were significantly higher in IgAN patients than in controls, whereas those of HLA-DRB1*0701, -DRB1*1501, -DQB1*0202, and -DQB1*0602 (Pc<0.001 for all) were significantly lower in IgAN patients than in controls. The allele frequency of HLA-DQB1*0503 (Pc=0.016) was significantly lower in the ESKD group than in the non-ESKD group; however, there was no significant difference for ESKD progression between these groups.ConclusionsWe report novel associations of HLA-DRB1*1501, DQB1*0202, -DQB1*0302, and -DQB1*0401 with IgAN. Further studies of HLA alleles associated with IgAN progression in a larger cohort and in various ethnic groups are needed.

Association between human leukocyte antigen class II (HLA-DRB and -DQB) alleles and outcome of exposure to Mycobacterium tuberculosis: a cross-sectional study in Nairobi, Kenya.

Introductionhuman leukocyte antigen (HLA) class II alleles play an important role in the early immune response to tuberculosis (TB) by presenting antigenic peptides to CD4+ T cells, hence polymorphisms in those genes can influence the efficiency of the immune response to infection and progression to active disease.Methodsan analytical cross-sectional study of adult pulmonary tuberculosis (PTB) patients at Mbagathi County Hospital, Nairobi and their HHCs. Sociodemographic data were captured on questionnaires and clinical data extracted from patient files. Intravenous blood samples were drawn for interferon-gamma release assay (IGRA) to determine latent tuberculosis infection (LTBI) among HHCs, and for extraction of DNA used in typing of HLA-DQB1 and HLA-DRB1 alleles by PCR sequence specific primer amplification. Chi-square and Fisher's exact test were used to compare the HLA type II allele frequencies of LTBI negative HHCs, LTBI positive HHCs and active TB patients. Logistic regression was used to adjust for HIV status.Resultsthe HLA-DQB1 and HLA-DRB1 alleles were analyzed in 17 PTB and 37 HHCs. Nineteen (19) HHCs were LTBI positive, while 18 were LTBI negative. The frequency of DRB3*1 was 0.17-fold lower [95% CI=0.03-0.83] among PTB patients compared to HHCs before adjustment for HIV status (p=0.048). The frequency of the DRB5*2 allele was significantly higher (p=0.013) among PTB patients (23.5%) compared to HHCS (0.00%). After adjusting for HIV status, the frequency of DRB1*14 was 12-fold higher [95% CI=1.11-138.2] among PTB patients compared to HHCs (p=0.040).Conclusionthe higher frequencies of HLA-DRB5*2 and HLA-DRB1*14 alleles in PTB patients suggest a likely association with progression to active PTB. The higher frequency of HLA-DRB3*1 allele among LTBI negative HHCs shows its likely protective role against M. tuberculosis infection in this population.

Interleukin – 6 Serum Level and Single Nucleotide Gene -174 G/C promoter Polymorphism in Patients with Rheumatoid Arthritis / Iraq

Objective: To 1) assess IL-6 levels in the serum of patients with rheumatoid arthritis (RA). 2) study IL-6 promoter -174 G>C “single nucleotide polymorphism (SNP)” as an imminent factor for the disease development. 3) find any relation between the level of serum IL-6 cytokine and other parameters such as age, gender, clinical severity of diseases and “disease activity scores (DAS28)”. Materials & methods: This research was carried out through a case – control approach at “Ibn – Senna Teaching Hospital” in Mosul city between November 2020 and July 2021. It included 61 RA patients diagnosed according to “ACR / EULAR 2010 criteria” and 50 healthy individuals. IL-6 serum levels were ascertained by ELISA and genotyping of IL-6 promoter was accomplished by “sequence-specific primer-polymerase chain reaction (SSP-PCR)”. Results: Mean IL-6 level in RA (69.42 ng /l ± 62.99) was elevated in comparison to healthy people (14.66 ng /l ± 23.58), P < 0.001. No age or gender effects on IL-6 concentration were noted. The ideal cut-off of IL-6 for discrimination of RA with best discriminative utility compared to healthy controls was 22.80 ng/l. At this value the IL-6 sensitivity was 91.8%, specificity 82.0% and accuracy rate 73.80%. G/G genotype was the most pervasive genotype in both RA patients and controls (70.5% in RA and 64% in healthy controls). However, it did not seem to be a risk factor for RA development compared to G/C or C/C genotypes “(OR = 1.3438, 95% CI=0.605-2.984,P=0.469)”. The mean IL-6 level in patients with GG genotype was (73.70 ng / l ± 71.09) compared to (58.37 ng /l ± 37.86) in patients with GC genotype. There was no significant difference in the IL-6 level between patients with GG and patients with GC genotypes (P = 0.2375). Although higher IL-6 mean concentration was reported in severe RA, however, no significant difference was found between patients with mild, moderate and severe RA respectively. No correlation of serum levels of IL-6 with genetic promoter polymorphism, clinical severity of diseases or DAS 28 score were reported. Conclusion: The concentration of serum IL-6 was elevated in RA in regard to healthy controls which confirmed its pivotal role in RA pathogenesis. Our data did not support the role of IL-6 promoter -174 G> C polymorphism as a risk factor for RA, nor seem to play a major role in the increase of IL-6 level among our patients with RA.

Donor HLA Genotyping using Polymerase Chain Reaction- Sequence Specific Primers (PCR-SSP) as Method for Acquiring Donor Panel in Platelet Refractoriness

Background: Evaluation and identification of HLA antibodies in the recipient’s serum is of utmost importance prior to transplantation and transfusion. HLA typing is a steppingstone in proposing a donor panel. In order to obtain the HLA typing, Polymerase Chain Reaction-Sequence Specific Primers (PCR-SSP) can be performed.Materials and method: This is a preliminary study to determine HLA polymorphism by HLA genotyping in 43 blood donors. DNA from the samples was isolated using commercial kits according to the standard protocol. The DNA then was amplified using PCR-SSP methods and analyzed using the provided set in the kit.Results: This study found that the most frequent HLA-A alleles was HLA-A*24 (41.9%). For HLA-B alleles, the most common was HLA-B*15 (28%). Most frequent HLA-A-B haplotypes was HLA-A*24-B*15 (11.3%). The results from this study concurs with that of previous study. However, some alleles might vary due to difference in study population. Determining HLA-typing is of paramount importance in an ethnically diverse country such as Indonesia. In contrast to homogenous caucassian country, difference in ethnicity might cause platelet refractoriness due to incompatibility. HLA-typing would also guide the diagnostic workup and required treatment strategy for platelet refractoriness.Conclusion: From the HLA typing using PCR-SSP in blood donors in Jakarta, we found that the most frequent alleles were HLA-A*24 and HLA-B*15; and the most frequent haplotypes were HLA-A*24-B*15. This study should be upscaled to include larger population and ethnic groups to obtain complete profile of Indonesian population.Keywords: platelet refractoriness, HLA, donor, PCR-SSP, transfusion medicine

Genetic polymorphisms in the miR-372 (rs12983273) and LncRNA HULC (rs7763881) genes and susceptibility to Hepatitis B virus (HBV) infection.

MicroRNAs (miRNAs) and Long non-coding RNAs (lncRNAs) are two major types of non-coding RNAs (ncRNAs) with regulatory roles. The initiation and progression of numerous diseases have been linked to genetic variation in miRNAs and lncRNAs. Many diseases, including hepatitis infection, are thought to be regulated by miRNA-LncRNA interactions. In this study, Single nucleotide polymorphisms (SNPs) in miR-372 (rs28461391 C/T) and HULC (rs7763881 A/C) were believed to play a role in HBV infection risk. Using the Polymerase chain reaction sequence-specific primer technique (PCR-SSP), 100 HBV patients and 100 healthy controls were genotyped for SNPs rs28461391 in miR-372 and rs7763881 in HULC. There was no significant difference in miR-372 rs12983273 genotype distribution between controls and HBV patients, according to our findings. On the other hand, there was a significant increase in HULC rs7763881 CC genotype (P < 0.05) coincides with a significant decrease in AC genotype distribution (P < 0.05) in HBV patients as compared to controls. Our results showed that the AA genotype is protective for HBV infection (OR 0.3; CI 0.13-9.07) while the CC genotype is associated with an increased risk of HBV infection (OR 3.43; CI 1.3-9.07). Our results suggest that HULC rs7763881 A/C might be a biomarker for HBV susceptibility. Larger sample studies are needed to confirm our preliminary data. To the best of our knowledge, the present study was the first to investigate the relevance of miR-372 (rs28461391 C/T) and HULC (rs7763881 A/C) gene polymorphisms to the risk of HBV infection in the Egyptian population.

Contribution of HLA class II genes, DRB4*01:01, DRB1*07:01, and DQB1*03:03:2 to clinical features of Vitiligo disease in Iranian population.

Vitiligo is a multifactorial depigmentation condition, which is due to skin melanocyte destruction. Increased expression of HLA class II genes in patients with pre-lesions of Vitiligo suggests a crucial role for the participation of immune response in Vitiligo development. Recent studies progressively focused on HLA-DRB1 and DQB1 genes. In this study, we have evaluated the association and role of HLA-DRB4*01:01, -DRB1*07:01, and -DQB1*03:03:2 genes in different clinical subtypes of Vitiligo in the Iranian population. First, Genomic DNA from peripheral blood of 125 unrelated Vitiligo patients and 100 unrelated healthy controls were extracted through the salting-out method. Then, HLA class II genotyping was performed using the sequence-specific primer PCR method. Finally, the clinical relevance of the testing for these genotypes was evaluated by applying the PcPPV (prevalence-corrected positive predictive value) formula. Our results indicated the positive associations of DRB4*01:01 and DRB1*07:01 allelic genes with early-onset Vitiligo (p = 0.024 and 0.022, respectively). DRB4*01:01 also showed strong protection against late-onset Vitiligo (p = 0.0016, RR = 0.360). Moreover, our data revealed that the DRB1*07:01 increases the susceptibility to Sporadic Vitiligo (p = 0.030, RR = 1.702). Furthermore, our findings proposed that elevated vulnerability of Vitiligo patients due to DRB4*01:01 and DRB1*07:01 alleles maybe is correlated with the presence of amino acid Arginine at position 71 at pocket 4 on the antigen-binding site of the HLA-DRB1 receptor. Our findings on different subtypes of Vitiligo suggest that, despite a more apparent autoimmune involvement, a non-autoimmune nature for the etiology of Vitiligo should also be considered.

Analysis of two novel variants of FUT1 gene in a Chinese family with para-Bombay phenotype

To study rare para-Bombay blood type Bmh and to investigate the molecular genetic basis of para-Bombay phenotype in a Chinese family. ABO and H phenotype of the proband and her pedigree were determined with serological methods. The ABO genotype was analyzed by polymerase chain reaction-sequence specific primer(PCR-SSP). The full coding region of alpha-l,2 fucosyltransferase (FUT1) gene of the pedigree was analyzed by polymerase chain reaction and direct sequencing of the amplified fragments. The haplotype of the FUT1 gene were analyzed by cloning sequencing. The rare para-Bombay blood type Bmh was identified in the proband, with ABO*B.01/ABO*O.01.01 genotype. Two variants of FUT1 gene, c.508dupT and c.787A>C, were found in the proband. The cloning sequencing revealed that the two variants were on different alleles, and the haplotype of FUT1 gene was h508dupT/h787C. Both of the two variants were predicted to cause inactivation of the enzyme, which is consistent with the result of serological techniques. Two new alleles of FUT1 gene (h508dupT and h787C), which were associated with para-Bombay phenotype, were identified in the Chinese pedigree.

Distinct TNF-alpha and HLA polymorphisms associate with fibrotic and non-fibrotic subtypes of hypersensitivity pneumonitis

IntroductionSince Hypersensitivity Pneumonitis (HP) categorization in fibrotic and nonfibrotic/inflammatory types seems to be more consistent with the distinctive clinical course and outcomes, recent international guidelines recommended the use of this classification. Moreover, fibrotic subtype may share immunogenetic and pathophysiological mechanisms with other fibrotic lung diseases. AimTo investigate HLA -A, -B, -DRB1 and TNF-α –308 gene polymorphisms among fibrotic and nonfibrotic HP patients due to avian exposure, also in comparison with asymptomatic exposed controls. MethodsWe prospectively enrolled 40 HP patients, classified as fibrotic or nonfibrotic/inflammatory, and 70 exposed controls. HLA and TNF-α polymorphisms were determined by polymerase chain reaction–sequence specific primer amplification. ResultsWhile HLA alleles were not associated to HP susceptibility, fibrotic HP patients showed increased frequencies of HLA A*02 (46.7% vs 25.7%; OR=2.53, p = 0.02) and HLA DRB1*14 (10.0% vs 0.7%; OR=15.44, p=0.02) alleles when compared with exposed controls, although not statistically significant after correction for multiple comparisons. TNF-α G/G genotype (associated with low TNF-α production) frequencies were significantly increased among the non-fibrotic/inflammatory HP patients comparatively to fibrotic presentations (88% vs 60%; RR=0.44; p=0.04) and controls (88% vs 63%, OR 4.33, p=0.037). Also, these patients had a significantly increased frequency of the G allele (94.0% vs 73.3%, RR=0.44, p=0.01), while fibrotic HP patients predominantly presented the A allele (26.7% vs 6.0%, RR=2.28, p=0.01). ConclusionsOur results support the hypothesis that fibrotic and non-fibrotic HP subtypes exhibit a distinct profile of TNF-α and HLA polymorphisms, which may be relevant to predict disease course and better define treatment strategies.

HLA-DPB1 E69 genotype and exposure in beryllium sensitisation and disease

ObjectivesHuman leukocyte antigen-DP beta 1 (HLA-DPB1) with a glutamic acid at the 69th position of the ß chain (E69) genotype and inhalational beryllium exposure individually contribute to risk of chronic...

Genotyping for Dombrock blood group alleles in Northern Pakistani blood donors.

Abstract Genotyping can be used to identify rare blood group antigens and to solve suspected blood group discrepancies, particularly when serologic methods are limited. Unfortunately, only a few such studies have been performed in Pakistan. The present study was conducted to determine the frequency of Dombrock blood group alleles by genotyping samples from blood donors from the north of Pakistan. Blood samples were taken with consent from 300 blood donors; DNA was extracted and tested for DO*01 and DO*02 alleles by sequence-specific primer polymerase chain reaction (PCR-SSP), followed by gel electrophoresis. Allele frequencies were calculated. The observed and expected genotype frequencies were compared using the χ 2 test. The allele frequencies for DO*01 and DO*02 were 0.40 and 0.60, respectively. Genotype frequencies were in Hardy-Weinberg equilibrium. This study in Pakistani blood donors provides Dombrock blood group allele frequencies by PCR-SSP. This approach is efficient and economical and can be applied in developing countries. The findings can contribute to the development of in-house red blood cell panels, identification of rare blood types, and establishment of a national rare blood donor program.

HLA-DRB1 alleles as predisposing and resisting factor in women suffering from vulvovaginal candidiasis

BackgroundVulvovaginal candidiasis (VVC) is a vaginal mucosal infection that usually affects women in their reproductive age. When the signs of VVC persist on a daily basis or last for a long time and repeat at least three times per year, the disease is considered chronic and recurrent. ObjectiveThe purpose of this study was to evaluate the frequency of HLA-DRB1 alleles in patients with recurrent vulvovaginal candidiasis (RVVC). Study design120 patients with RVVC and 136 age-matched healthy controls underwent low-resolution HLA-DRB typing performed using the polymerase chain reaction-sequence-specific primers (PCR-SSP) technique. ResultsIn the present work, we studied different genes that encode HLA-DRB (HLA-DRB1 / HLA-DRB3 / HLA-DRB4 / HLA-DRB5) and showed that HLA-DRB1×14, found in 25% of the patients. In the present study, the significant frequency of HLA-DRB1×10 in the control group suggests a resistant role of this allele to RVVC infections ConclusionsIn the HLA-DRB region, the DRB1×14 allele showed a higher frequency in the patients with RVVC than in the controls. Moreover, the higher frequency of DRB1×10 observed in the controls than in the patients with RVVC. These results demonstrate the HLA-DRB1 alleles are in relation with both susceptibility and immunity factors in RVVC infection and possible susceptible role of HLA-DRB1×14

Genotyping of ABO and Duffy blood groups among malaria patients in Thailand.

ABO blood groups have been proposed to influence malaria parasite infection and disease severity in individuals residing in different geographical areas. In Thailand, genetic polymorphisms of blood groups and susceptibility to malaria infection have rarely been investigated. The aim of this study was to assess the genotype frequencies of ABO and Duffy blood groups and susceptibility to malaria infection in two populations residing in malaria-endemic areas of Thailand. 1100 malaria samples and an identical number of samples from healthy subjects were collected from Thai-Malaysian and Thai-Myanmar areas. Genotyping of ABO and Duffy blood groups was performed by sequence specific primer-polymerase chain reaction. The distribution of ABO and Duffy blood groups was similar in malaria-positive and negative subjects. Blood group O was prevalent in both populations followed by blood group B (BO genotype) and A (AO genotype), respectively. In Plasmodium falciparum infections, blood group A frequency was significantly higher in Thai-Malaysian samples (P = 0.042) whereas blood group B frequency was significantly higher in Thai-Myanmar samples (P = 0.022). FY*A/*A frequency was significantly higher in Plasmodium vivax infection (P = 0.036) while FY*A/*B frequency was significantly higher in healthy subjects (P = 0.005). The different ABO blood group frequencies in the two populations may contribute to susceptibility to P. falciparum infection and the high prevalence of FY*A/*A can confer a risk of P. vivax infection. Further research in various ethnic groups is needed to clarify the association between blood groups and pathogenesis of malaria.

Molecular basis of serological weak D phenotypes and RhD typing discrepancies identified in the Korean population.

The molecular basis of RhD blood groups differs with race/ethnicity. This study aimed to investigate the molecular basis of serological weak D phenotypes and RhD typing discrepancies in the Korean population. The RhD status of 188,852 Korean patients was initially determined using the automated microplate method and manual tile method. In case of no agglutination, weak D testing was further performed using the tube and gel methods. Serologically D-negative samples with C+ and/or E+ were tested using polymerase chain reaction-sequence specific primers for four RHD targets and/or exon 9 sequencing. Samples showing a serological weak D phenotype or an RhD typing discrepancy were subjected to full RHD gene sequencing. Of the 32 samples showing a serological weak D phenotype and 191 samples showing a serologically D-negative phenotype with C+ and/or E+, 23 and 50 were genotyped, respectively. Among the weak D samples, the most common alleles were RHD*15 (n=6), RHD*13.01 (n=4), and RHD*01W.25 (n=4), and no variant was found in two samples. RHD*01EL.01 (n=26) accounted for more than half of the D-negative samples. Of the seven samples that were typed as D-positive using the automated microplate method but showed weak reactivity using the tile method, four were genotyped, and the results were as follows: RHD*01W.33 (n=2), RHD*01W.43 (n=1), and no variant found (n=1). In our cohort, various D variant alleles including RHD*15 were identified; however, RHD*01W.1, RHD*01W.2, RHD*01W.3, RHD*09.03.01, and RHD*09.04, accounting for more than 95% of Caucasians with a serological weak D phenotype, were not found. Our study reaffirms that the distribution of D variant alleles differs between East Asians and Caucasians. Our findings also indicate that some D variants including RHD*01W.33 and RHD*01W.43 are at risk of being mistyped as D-positive by a highly sensitive RhD typing method such as an automated microplate method.

Evaluation of HLA class I and HLA class II allele profile and its relationship with clinical features in patients with alopecia areata: a case–control study

Background Alopecia areata (AA) is an autoimmune disease where autoimmune dysregulations along with genetic susceptibility are hypothesized to play a role in pathogenesis. Objective The aim of this study in to evaluate HLA-A, HLA-B, HLA-C, HLA-DQB1, and HLA-DRB1 profile and its relationship with clinical features in AA patients. Materials and methods Ninety-eight patients with AA and 100 healthy controls were included in the study. HLA-A, HLA-B, HLA-C, HLA-DQB1, and HLA-DRB1 frequencies were analyzed using polymerase chain reaction-sequence specific primers (PCR-SSP). Results HLA-B*39 and HLA-HLA-DRB1*15 allele frequencies were increased (p = .022 and p = .023, respectively), HLA-A*11 and HLA-B*35 frequencies were decreased (p = .006 and p = .014, respectively) in AA patients. HLA-B*13 and HLA-DRB1*11 were associated with poor prognostic factors. A class I allele, HLA-B*13 was associated with recurrence (p = .023) and presence of nevus flammeus (p = .022), while the class II allele HLA-DRB1*11 was associated with widespread hair loss (diffuse or universal alopecia) (p = .026), presence of ophiasis (p = .049) and juvenile onset (p = .018). Conclusion Belonging to two different classes of HLA family, HLA-B*13 and HLA-DRB1*11 alleles identified separate set of risk factors. In addition to increasing the risk of AA, HLA alleles may affect the prognosis of the disease.