NG_007494.1(RHD):c.[4A>T;5G>C;6_7insG] with an RhD-negative phenotype.
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ABSTRACT:
SUBMITTER: Srivastava K
PROVIDER: S-EPMC9102550 | biostudies-literature | 2020 Nov
REPOSITORIES: biostudies-literature
Publications
NG_007494.1(RHD):c.[4A>T;5G>C;6_7insG] with an RhD-negative phenotype.
Srivastava Kshitij K Doescher Andrea A Wagner Franz F FF Flegel Willy A WA
Transfusion 20201011 11
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Project description:To investigate the genetic basis of the Rh polypeptide gene, we attempted the isolation of cDNA clones for Rh polypeptide from a family with the RhD-positive and RhD-negative phenotypes using the reverse transcription (RT)-PCR method for each reticulocyte RNAs followed by subcloning. The isolated cDNAs showed the existence of another Rh-related clone (RhPII-1 cDNA, tentative designation) besides the RhPI and RhPII cDNA clones reported previously by us. The RhPII-1 cDNA had a single nucleotide substitution with one amino acid substitution compared with the RhPII cDNA:substitution C-->T in nucleotide 380, changing codon 127 from GCG to GTG (Ala-->Val). The RhPI, RhPII, and RhPII-1 cDNA clones were detected in all individuals by the PCR experiment. This suggests that the Rh polypeptide genes have been inherited from parents and might be highly polymorphic. The PCR amplification of an RhPII-specific region from reticulocyte RNA and genomic DNA in all the family proved that the RhPII gene exists in both RhD-positive and RhD-negative individuals. By Southern-blot analysis of the DNAs from the family, two independent polymorphisms concerning the RhC/c and RhD/d phenotypes were observed. These results demonstrate that the RhPI and RhPII genes are also present in the RhD-negative donors, and the RhPII-related cDNAs encode not the RhD, but the RhC/c and/or E/e, polypeptides.
| S-EPMC1138043 | biostudies-other
Project description:BACKGROUND:Blood group genotyping is increasingly utilized for prenatal diagnosis and after recent transfusions, but still lacks the specificity of serology. In whites, the presence of antigen D is predicted, if two or more properly selected RHD-specific polymorphism are detected. This prediction must fail, if an antigen D negative RHD positive allele is encountered. Excluding RHDpsi and CdeS frequent only in individuals of African descent, most of these alleles are unknown and the population frequency of any such allele has not been determined. METHODS:We screened 8,442 antigen D negative blood donations by RHD PCR-SSP. RHD PCR positive samples were further characterized by RHD exon specific PCR-SSP or sequencing. The phenotype of the identified alleles was checked and their frequencies in Germans were determined. RESULTS:We detected 50 RHD positive samples. Fifteen samples harbored one of three new Del alleles. Thirty samples were due to 14 different D negative alleles, only 5 of which were previously known. Nine of the 14 alleles may have been generated by gene conversion in cis, for which we proposed a mechanism triggered by hairpin formation of chromosomal DNA. The cumulative population frequency of the 14 D negative alleles was 1:1,500. Five samples represented a D+/- chimera, a weak D and three partial D, which had been missed by routine serology; two recipients transfused with blood of the D+/- chimera donor became anti-D immunized. CONCLUSION:The results of this study allowed to devise an improved RHD genotyping strategy, the false-positive rate of which was lower than 1:10,000. The number of characterized RHD positive antigen D negative and Del alleles was more than doubled and their population frequencies in Europe were defined.
| S-EPMC37267 | biostudies-literature
Project description:D antigen is the most important and immunogenic antigen of the Rh blood group. The RhD-negative phenotype has different genetic backgrounds with variable distribution in different populations. Hybrid Rhesus box, resulting from RHD gene deletion, is used in genotyping studies of the Rh blood group as a marker to identify the RHD gene deletion. This study for the first time identified genetic mechanisms for the occurrence of RhD-negative phenotype among the Iranian population. 200 RhD-negative blood donors were randomly selected from Tehran Blood Transfusion Center. The phenotype of D, C, ?, e and c antigens was serologically identified, and DNA was extracted from buffy coat. The molecular analysis of hybrid Rhesus box was performed by PCR-SSP and PCR-RFLP. Moreover, the presence of different exons of RHD gene was investigated by real-time PCR on extracted DNA. Hybrid Rhesus box was detected in all samples, and PCR-RFLP confirmed that 198 (99%) were homozygous for an RHD gene deletion and 2 were heterozygous for hybrid Rhesus box in which one (0.5%) had a weak D type 11 and the other one (0.5%) had a RHD-CE (2-9)-D 2 hybrid allele. Similar to Caucasians, the frequency of RHD gene deletion was high among the Iranian population studied in this investigation, so hybrid Rhesus box can be used as an efficient marker to detect RHD gene deletion in our population.
| S-EPMC6369079 | biostudies-literature
Project description:BackgroundA majority of non-invasive prenatal screening studies determining fetal RhD status have been tested on Caucasian and Asian populations, but limited or no studies have been conducted on the Ethiopian population. In the current study, we carried non-invasive prenatal screening of fetal RHD genotype in selected RhD negative Ethiopian pregnant women.MethodsCell-free DNA was extracted from the plasma samples of 117 RhD pregnant women between 9 and 38 weeks of gestation. Fetal RHD genotypes were detected by targeting exons 5, 7 and 10 of the RHD gene by using real-time PCR assay. RHD genotypic results were confirmed by neonatal cord blood serology.ResultsFetal RHD genotyping was conclusive in all 117 subjects. RHD genotype was correctly predicted in 115 of 117 cases, thus the test yielded 98.3% accuracy (95%CI: 97.3-99.1%). Among 115 cases, 105 were genotyped as RHD positive and 12 were genotyped as RHD negative. The sensitivity and specificity of the test were 99.1% (95% CI: 94.8-99.9%) and 91.7% (95%CI: 61.5-99.7%) respectively. The negative and positive predictive values were 99.9% (95%CI: 99.2-99.9%) and 54.0% (95% CI: 15.2-88.4%) respectively. SRY genotyping results were in complete concordance with fetal sex.ConclusionMulti exon targeted non-invasive prenatal screening test for fetal RhD determination exhibited high accuracy and sensitivity. A confirmatory study with a bigger size of study subjects is warranted before enabling clinical implementation.
| S-EPMC8929601 | biostudies-literature
Project description:ObjectivesTo assess the feasibility of applying a high throughput method, with an automated robotic technique, for predicting fetal RhD phenotype from fetal DNA in the plasma of RhD negative pregnant women to avoid unnecessary treatment with anti-RhD immunoglobulin.DesignProspective comparison of fetal RHD genotype determined from fetal DNA in maternal plasma with the serologically determined fetal RhD phenotype from cord blood.SettingAntenatal clinics and antenatal testing laboratories in the Midlands and north of England and an international blood group reference laboratory.ParticipantsPregnant women of known gestation identified as RhD negative by an antenatal testing laboratory. Samples from 1997 women were taken at or before the 28 week antenatal visit.Main outcome measuresDetection rate of fetal RhD from maternal plasma, error rate, false positive rate, and the odds of being affected given a positive result.ResultsSerologically determined RhD phenotypes were obtained from 1869 cord blood samples. In 95.7% (n=1788) the correct fetal RhD phenotype was predicted by the genotyping tests. In 3.4% (n=64) results were either unobtainable or inconclusive. A false positive result was obtained in 0.8% (14 samples), probably because of unexpressed or weakly expressed fetal RHD genes. In only three samples (0.2%) were false negative results obtained. If these results had been applied as a guide to treatment, only 2% of the women would have received anti-RhD unnecessarily, compared with 38% without the genotyping.ConclusionsHigh throughput RHD genotyping of fetuses in all RhD negative women is feasible and would substantially reduce unnecessary administration of anti-RhD immunoglobulin to RhD negative pregnant women with an RhD negative fetus.
| S-EPMC2292334 | biostudies-literature
Project description:Methylotuvimicrobium buryatense 5G strain:5G Genome sequencing and assembly
| PRJNA89087 | ENA
Project description:BACKGROUND:All non-sensitized Rhesus D (RhD)-negative pregnant women in Germany receive antenatal anti-D prophylaxis without knowledge of fetal RhD status. Non-invasive prenatal testing (NIPT) of cell-free fetal DNA in maternal plasma could avoid unnecessary anti-D administration. In this paper, we systematically reviewed the evidence on the benefit of NIPT for fetal RhD status in RhD-negative pregnant women. METHODS:We systematically searched several bibliographic databases, trial registries, and other sources (up to October 2019) for controlled intervention studies investigating NIPT for fetal RhD versus conventional anti-D prophylaxis. The focus was on the impact on fetal and maternal morbidity. We primarily considered direct evidence (from randomized controlled trials) or if unavailable, linked evidence (from diagnostic accuracy studies and from controlled intervention studies investigating the administration or withholding of anti-D prophylaxis). The results of diagnostic accuracy studies were pooled in bivariate meta-analyses. RESULTS:Neither direct evidence nor sufficient data for linked evidence were identified. Meta-analysis of data from about 60,000 participants showed high sensitivity (99.9%; 95% CI [99.5%; 100%] and specificity (99.2%; 95% CI [98.5%; 99.5%]). CONCLUSIONS:NIPT for fetal RhD status is equivalent to conventional serologic testing using the newborn's blood. Studies investigating patient-relevant outcomes are still lacking.
| S-EPMC7006196 | biostudies-literature
Project description:We previously reported that neuroimmune semaphorin (Sema) 4A regulates the severity of experimental allergic asthma and increases regulatory T (Treg) cell numbers in vivo; however, the mechanisms of Sema4A action remain unknown. It was also reported that Sema4A controls murine Treg cell function and survival acting through neuropilin 1 (NRP-1) receptor. To clarify Sema4A action on human T cells, we employed T cell lines (HuT78 and HuT102), human PBMCs, and CD4+ T cells in phenotypic and functional assays. We found that HuT78 demonstrated a T effector-like phenotype (CD4+CD25lowFoxp3-), whereas HuT102 expressed a Treg-like phenotype (CD4+CD25hi Foxp3+). Neither cell line expressed NRP-1. HuT102 cells expressed Sema4A counter receptor Plexin B1, whereas HuT78 cells were Sema4A+. All human peripheral blood CD4+ T cells, including Treg cells, expressed PlexinB1 and lacked both NRP-1 and -2. However, NRP-1 and Sema4A were detected on CD3negativeCD4intermediate human monocytes. Culture of HuT cells with soluble Sema4A led to an upregulation of CD25 and Foxp3 markers on HuT102 cells. Addition of Sema4A increased the relative numbers of CD4+CD25+Foxp3+ cells in PBMCs and CD4+ T cells, which were NRP-1negative but PlexinB1+, suggesting the role of this receptor in Treg cell stability. The inclusion of anti-PlexinB1 blocking Ab in cultures before recombinant Sema4A addition significantly decreased Treg cell numbers as compared with cultures with recombinant Sema4A alone. Sema4A was as effective as TGF-β in inducible Treg cell induction from CD4+CD25depleted cells but did not enhance Treg cell suppressive activity in vitro. These results suggest strategies for the development of new Sema4A-based therapeutic measures to combat allergic inflammatory diseases. ImmunoHorizons, 2019, 3: 71-87.
| S-EPMC6590919 | biostudies-literature
Project description:BackgroundMaternal-fetal RhD antigen incompatibility causes approximately 50% of clinically significant alloimmunization cases. The routine use of prophylactic anti-D immunoglobulin has dramatically reduced hemolytic disease of the fetus and newborn. Recently, fetal RHD genotyping in RhD negative pregnant women has been suggested for appropriate use of anti-D immunoglobulin antenatal prophylaxis and decrease unnecessary prenatal interventions.Materials and methodsIn this prospective cohort study, in order to develop a reliable and non-invasive method for fetal RHD genotyping, cell free fetal DNA (cffD- NA) was extracted from maternal plasma. Real-time quantitative polymerase chain reaction (qPCR) for detection of RHD exons 7, 5, 10 and intron 4 was performed and the results were compared to the serological results of cord blood cells as the gold standard method. SRY gene and hypermethylated Ras-association domain family member 1 (RASSF1A) gene were used to confirm the presence of fetal DNA in male and female fetuses, respectively.ResultsOut of 48 fetuses between 8 and 32 weeks (wks) of gestational age (GA), we correctly diagnosed 45 cases (93.75%) of RHD positive fetuses and 2 cases (4.16%) of the RHD negative one. Exon 7 was amplified in one sample, while three other RHD gene sequences were not detected; the sample was classified as inconclusive, and the RhD serology result after birth showed that the fetus was RhD-negative.ConclusionOur results showed high accuracy of the qPCR method using cffDNA for fetal RHD genotyping and implicate on the efficiency of this technique to predict the competence of anti-D immunoglobulin administration.
| S-EPMC4845531 | biostudies-literature
Project description:The Rh blood group system (ISBT004) is the second most important blood group after ABO and is the most polymorphic one, with 55 antigens encoded by 2 genes, RHD and RHCE This research uses next-generation sequencing (NGS) to sequence the complete RHD gene by amplifying the whole gene using overlapping long-range polymerase chain reaction (LR-PCR) amplicons. The aim was to study different RHD alleles present in the population to establish reference RHD allele sequences by using the analysis of intronic single-nucleotide polymorphisms (SNPs) and their correlation to a specific Rh haplotype. Genomic DNA samples (n = 69) from blood donors of different serologically predicted genotypes including R1R1 (DCe/DCe), R2R2 (DcE/DcE), R1R2 (DCe/DcE), R2RZ (DcE/DCE), R1r (DCe/dce), R2r (DcE/dce), and R0r (Dce/dce) were sequenced and data were then mapped to the human genome reference sequence hg38. We focused on the analysis of hemizygous samples, as these by definition will only have a single copy of RHD For the 69 samples sequenced, different exonic SNPs were detected that correlate with known variants. Multiple intronic SNPs were found in all samples: 21 intronic SNPs were present in all samples indicating their specificity to the RHD*DAU0 (RHD*10.00) haplotype which the hg38 reference sequence encodes. Twenty-three intronic SNPs were found to be R2 haplotype specific, and 15 were linked to R1, R0, and RZ haplotypes. In conclusion, intronic SNPs may represent a novel diagnostic approach to investigate known and novel variants of the RHD and RHCE genes, while being a useful approach to establish reference RHD allele sequences.
| S-EPMC6199663 | biostudies-literature