Serologic profiling of D variants in donor routine: unveiling the impact on false-negative results and alloimmunization.

The high number of D variants can lead to the unnecessary use of Rh immune globulin, overuse of D- RBC units, and anti-D allommunization. D variant prevalence varies among ethnic groups, and knowledge of the main variants present in a specific population, their behavior in serologic tests, and their impact on clinical practice is crucial to define the best serologic tests for routine use. The present study aimed to explore the serologic profile of D variants and to determine which variants are most associated with false-negative D typing results and alloimmunization. Donor samples were selected in two study periods. During the first period, D typing was performed on a semi-automated instrument in microplates, and weak D tests were conducted in tube or gel tests. In the second period, D typing was carried out using an automated instrument with microplates, and weak D tests were performed in solid phase. Samples from patients typed as D+ with anti-D were also selected. All samples were characterized by molecular testing. A total of 37 RHD variants were identified. Discrepancies and atypical reactivity without anti-D formation were observed in 83.4 percent of the samples, discrepant D typing results between donations were seen in 12.3 percent, and D+ patients with anti-D comprised 4.3 percent. DAR1.2 was the most prevalent variant. Weak D type 38 was responsible for 75 percent of discrepant samples, followed by weak D type 11, predominantly detected by solid phase. Among the D variants related to alloimmunization, DIVa was the most prevalent, which was not recognized by serologic testing; the same was true for DIIIc. The results highlight the importance of selecting tests for donor screening capable of detecting weak D types 38 and 11, especially in populations where these variants are more prevalent. In pre-transfusion testing, it is crucial that D typing reagents demonstrate weak reactivity with DAR variants; having a serologic strategy to recognize DIVa and DIIIc is also valuable.

Characterization of Novel RHD Allele Variants and Their Implications for Routine Blood Group Diagnostics.

The Rh system, including the highly immunogenic D antigen, is one of the clinically most important blood group systems in transfusion medicine. Numerous alleles of the RHD gene are associated with variant RhD phenotypes. In case of Rh incompatibility, some of them can induce hemolytic transfusion reactions and hemolytic disease of the fetus and newborn. Thus, accurate blood group diagnostics are critical for safe transfusion therapy. We characterized phenotypes of four individuals revealing weakened D expression during routine pre-transfusion testing. Standard gel card matrix techniques with monoclonal and polyclonal anti-D antibodies were used for serological typing, complemented using D epitope and antigen density analysis. Genotyping employing PCR with sequence-specific primers, genomic and allele-specific Sanger sequencing and in silico protein analysis were performed. Four novel RHD alleles associated with weak D or partial D phenotypes were identified. One of the mutations is predicted to disrupt the terminal stop codon and result in an elongated translation of the mutant D protein that phenotypically exhibits a loss of D epitopes. Furthermore, a hybrid gene formed with the homologue RHCE gene is described. The presented data enhances the understanding of the Rh system and may contribute to continued advances in blood group diagnostics.

A Korean family with RHD*DNT only detectable after anti-D alloimmunization.

OBJECTIVES: Identification of DNT, a rare partial D, can be challenging, as it is difficult to distinguish from D+. This study aimed to identify DNT individuals by analyzing the DNT proband's family members, characterize DNT, and propose management strategies. METHODS: Family members of the first Korean DNT proband were recruited. RHD genotyping was conducted, and weak D tests were carried out using several anti-D reagents. RESULTS: Three DNT individuals were identified among 6 family members, including 1 with an anti-D alloantibody. As DNT red cells exhibited strong reactivity with all anti-D clones, DNT was serologically indistinguishable from D+. Moreover, unusual serologic findings in DNT individuals only became apparent after anti-D alloimmunization. CONCLUSIONS: We recommend DNT individuals as candidates for Rh immune globulin prophylaxis during the perinatal period and transfusions with D- blood components. An anticipatory RHD genotyping is suggested for partial D family members to prevent potential partial D individuals from becoming alloimmunized.

Switching antipsychotics to partial dopamine D2-agonists in individuals affected by schizophrenia: a narrative review.

OBJECTIVE: The aim of this review is to analyse the literature regarding studies centred on the clinical outcome of individuals affected by schizophrenia and treated with various antipsychotics, and then switched to orally administered partial D2-dopamine agonists (PD2A): Aripiprazole (ARI), brexpiprazole (BREX) or cariprazine (CARI). METHOD: A PubMed literature search was performed on 16 February 2021, and updated on Jan 26, 2022 for literature on antipsychotic switching in individuals affected by schizophrenia. Literature was included from 2002 onward. Six strategies were defined: Abrupt, gradual and cross-taper switch, and 3 hybrid strategies. The primary outcome was all-cause discontinuation rate per switch strategy per goal medication. RESULTS: In 10 reports on switching to ARI, 21 studies with different strategies were described, but there were only 4 reports and 5 strategies on switching to BREX. Only one study about CARI was included, but it was not designed as a switch study. The studies are difficult to compare due to differences in methodology, previous antipsychotic medication, doses of the introduced P2DA and study duration. CONCLUSION: This analysis did not reveal evidence for a preferable switching strategy. A protocol should be developed which defines optimal duration, instruments to be used, and the timing of the exams.KEY MESSAGESMost switch studies on partial D2-agonists focus on ARI, with only a few on BREX, while little is known about the clinical outcome of switching individuals to CARIThere is a wide variation of possible switch methods: Abrupt switch - gradual switch - cross-tapering switch - hybrid strategies including plateau switchThe protocols used differ considerably between the studies. A strict comparison between the studies is difficult, for which reason the present evidence does not support an unambiguous preference for a particular switch strategy.From a methodological point of view, a standardised clinical protocol should be developed to allow comparisons between studies regarding the clinical outcome of individuals switched from one antipsychotic drug to another.

Overview of the serologic and molecular basis of D variants with a focus on D variants in the Indian population.

Complexities of D within the Rh blood group system have long been recognized, initially using basic serologic testing and, more recently, using advanced and sensitive typing reagents. Discrepancies may arise when an individual carries a D antigen showing altered D antigen expression. These D variants are clinically important, since they may lead to production of anti-D in the carrier and induce alloimmunization in D- recipients, making their correct identification imperative. For clinical purposes, D variants can be classified into three groups: weak D, partial D, and DEL. The problem surrounding proper characterization of D variants exists because routine serologic tests are sometimes inadequate to detect D variants or resolve discrepant or ambiguous D typing results. Today, molecular analysis has revealed more than 300 RH alleles and is a better method for investigating D variants. Global distribution of variants differs, as observed in European, African, and East Asian populations. Discovery of the novel RHD*01W.150 (weak D type 150) with a nucleotide change of c.327_487-4164dup is proof. This variant, the result of an insertion of a duplicated exon 3 between exons 2 and 4 in the same orientation, was detected in more than 50 percent of Indian D variant samples in a 2018 study. The outcome of studies worldwide has led to the recommendation to manage D variant individuals as D+ or D- according to RHD genotype. The policies and workup with respect to D variant testing in donors, recipients, and prenatal women differ among blood banks, depending on type of variants predominantly encountered. Thus, a general genotyping protocol cannot be followed globally, and an Indian-specific RHD genotyping assay (multiplex polymerase chain reaction) designed to detect D variants frequently found in the Indian population was developed to save time and resources. This assay is also helpful for detecting several partial and null alleles. Identification of D variants by serology and characterization by molecular testing need to go hand-in-hand for better and safer transfusion practices.

Transfusion management of Africans with RHD variants in China.

The presence of D variant among minorities could produce a higher rate of alloimmunization observed in patients from this group. This is partly due to the ethnic and racial specificity of RHD variants and the limited availability of Rh-matched blood donors. Approximately half a million African migrants in China carrying distinct Rh blood type composition have presented to the health care system with an imperative safety requirement of blood transfusion among 1.3 billion Chinese individuals. We depict the clinically significant RHD alleles among African migrants living in China and identify the genetic similarities and disparities to Chinese. We discussed practical strategies to manage the unique transfusion needs of African migrants in China.

Seventy Years of Treating Delusional Disorder with Antipsychotics: A Historical Perspective.

For many decades, delusional disorder (DD) has been considered a treatment-resistant disorder, with antipsychotics acknowledged as the best, though imperfect, treatment. It is possible that the discovery of the right drug could turn treatment resistance into treatment response. The goal of this narrative review is to provide a historical perspective of the treatment of DD since the introduction of antipsychotics 70 years ago. The following search terms were used to scan the literature: antipsychotics AND "delusional disorder". Findings were that therapy for DD symptoms has changed over time. Initial reports suggested that the drug of choice was the antipsychotic pimozide, and that this drug was especially effective for the somatic subtype of DD. Subsequent studies demonstrated that other antipsychotics, for instance, risperidone and olanzapine, were also highly effective. Treatment response may vary according to the presence or absence of specific symptoms, such as cognitive defect and depression. Clozapine, partial D2 agonists, and long-acting injectable drugs may be more effective than other drugs, but the evidence is not yet in. Because of the absence of robust evidence, treatment guidelines for the optimal management of DD are not yet available.

Systematic RHD genotyping in Brazilians reveals a high frequency of partial D in transfused patients serologically typed as weak D.

BACKGROUND: The discrimination between weak D types and partial D can be of clinical importance because carriers of partial D antigen may develop anti-D when transfused with D-positive red blood cell units. The aim of this study was to determine by molecular analysis the type of D variants among Brazilian patients requiring transfusions with serologic weak D phenotypes. MATERIAL AND METHODS: Samples from 87 patients (53 with sickle cell disease, 10 with thalassemia and 24 with myelodysplastic syndrome), serologic typed as weak D by manual tube indirect antiglobulin test or gel test were first RHD genotyped by using the RHD BeadChip Kit (BioArray, Immucor). Sanger sequencing was performed when necessary. RESULTS: RHD molecular analysis revealed 32 (36.8 %) variant RHD alleles encoding weak D phenotypes and 55 (63.2 %) alleles encoding partial D antigens. RHD variant alleles were present in the homozygous state or as a single RHD allele, one variant RHD allele associated with the RHDΨ allele, or two different variant RHD alleles in compound heterozygosity with each other in 70 patients, 4 patients and 13 patients, respectively. Alloanti-D was found in 9 (16.4 %) cases with RHD alleles predicting a partial D. DISCUSSION: The frequency of partial D was higher than weak D types in Brazilian patients serologically typed as weak D, showing the importance to differentiate weak D types and partial D in transfused patients to establish a transfusion policy recommendation.

Molecular Characteristics of the Serological Weak D Phenotype in Koreans.

Serological weak D is a reaction of 2+ or less to anti-D reagent and includes weak D and partial D phenotypes. Although identifying the RhD subtype is important for transfusion safety, serological tests are insufficient for defining the RhD subtype, and molecular tests are needed. To analyze the molecular characteristics of D variants in Koreans to facilitate the formulation of individualized transfusion strategies, molecular tests such as RhD genotyping using real-time polymerase chain reaction (PCR) and partial-D and/or weak-D sequence-specific amplification (SSP) were performed on 105 Korean Rare Blood Program (KRBP) patients exhibiting serological weak D. In total, 58 out of 68 serologically determined weak D KRBP patients were typed as having weak D or partial D phenotypes via RhD genotyping. In detail, eight (13.8%) were typed as partial DVa or DBS, nine (15.5%) as weak D type 15, and four others (6.8%) as partial DVI, partial DVII, weak D type 2, or weak D type 41 or 45, whereas the rest (n = 37, 63.8%) was typed as having either weak D or partial D. This suggests that serological weak D Koreans who require transfusion should be treated as D-negative.

Updated Evaluation of RhD Status Among Women of Child-Bearing Age in Detroit, Michigan.

OBJECTIVES: The Rh blood group system is one of the most important and immunogenic blood group systems after the ABO blood group system and, like other blood group antigens, it follows ethnic and racial trends. However, when it comes to D variants-partial D and weak D-most of the cohorts studied in the literature have been of European descent. This study aimed to discover the variant D trends in Detroit, Michigan, with an emphasis on Black communities. METHODS: From 2016 to 2018, there were 102 patients (women of childbearing potential: < 50 years) at Henry Ford Hospital that had serologic D discrepant testing. These patients were sent out for molecular RHD determination. RESULTS: In total, 12.7% of patients were characterized as RhD positive and 87.3% of patients were characterized as RhD variants (nominated as RhD negative at our institution). CONCLUSIONS: Our predominantly Black cohort sheds light on the diversity of the RhD antigen. The majority of Blacks were classified as RhD variants (RhD negative nomination at our institution). Therefore, molecular testing for this patient population with serologic RhD discrepancies is paramount to properly manage their obstetric care.

Molecular characterization of RhD variant phenotypes among blood donors: A study from the coastal region of India.

BACKGROUND: RhD expression varies with population and ethnicity. Accurate typing of RhD antigen among blood donors is important to prevent development of anti-D among recipients of blood transfusion. We aimed to screen blood donors for variant D phenotypes and accurately characterize them by genotyping. MATERIAL AND METHODS: We have done prospective study on blood donors by performing RhD typing using three different commercial monoclonal anti-D reagents by both column agglutination and conventional tube techniques. Samples that showed ambiguous results were further screened with the Bio-Rad Partial RhD typing kit. Minor phenotyping for C, c, E, e antigens was performed. Multiplex PCR and Sequencing of all RHD exons with Sanger's sequencing was performed for molecular characterization of variant D. RESULTS: A total of 16,974 blood donors were screened during the study period. Among them, 31 (0.18 %) donors were found to have a RhD variant phenotype. The male to female ratio was 10:1. The presence of 'C' antigen was noted among all RhD variant samples. Serological typing identified two samples as DV phenotype and the rest could not be characterized. Molecular genotyping characterized 90.3 % of the samples as Indian specific weak D type 150 variants. Three samples were subjected to Sangers sequencing and showed wild type pattern. CONCLUSION: The present study showed that the most common variant in this population was Weak D type 150. This study highlights that serological methods may serve as a screening tool, however, molecular techniques are essential for characterization of RhD variants.

Frequency and characterization of RHD variant alleles in a population of blood donors from southeastern Brazil: Comparison with other populations.

BACKGROUND: The correct determination of D antigen could help to avoid alloimmunization in pregnant women and patients receiving blood transfusions. However, there are limitations in the identification of D variants as the partial and weak D phenotypes make the determination of D antigen a great challenge in the transfusion routine.' STUDY DESIGN AND METHODS: The molecular characterization of D variants was performed on blood donors from southeastern Brazil with atypical D typing. Furthermore, the serological profile of all RHD variant alleles identified was analyzed using different Anti-D clones. The prevalence of RHD alleles and genotypes found was compared with those described in other countries and in other regions from Brazil. RESULTS: Atypical serologic D typing occurred in 0.79 % of blood donors. The majority of RHD variant alleles (88 %) were first characterized by multiplex PCR and PCR-SSP as RHD*weak partial 4 (47 %), followed by RHD*weak D type 3 (29.9 %), RHD*weak D type 2 (3.9 %) and RHD*weak D type 1 (3.1 %). Genomic DNA sequencing characterized the RHD*weak partial 4 variants found in RHD*DAR1.2 (weak 4.2.2) (22 %), RHD*DAR3 (weak 4.0.1) (2.4 %), RHD*DAR3.1 (weak 4.0) (22 %) and RHD*DAR4 (weak 4.1) (0.8 %). RHD variant alleles associated with partial D, such as, RHD*DAU-4 (1.6 %), RHD*DAU-5 (2.4 %), RHD*DAU-6 (1.6 %), RHD* DIII type 8 (1.6 %), RHD*DVII (3.9 %) and RHD* DMH (0.8 %) were also observed. CONCLUSION: The prevalence of RHD variant alleles observed in this cohort differ from those found in other populations, including Brazilians from other regions. RHD allele distribution in specific regions should be considered for implementation of algorithms and genotyping strategies aiming at a more effective and safe transfusion.

Variable reactivity of Rh D antigen and its serological characterization.

Background: Variation in the reactivity on Rh D typing may pose challenges in interpretation and ambiguity in further patient management.Materials and Methods: A prospective study was conducted in the department of transfusion medicine for a period of 18 months. Blood grouping was performed by fully automated equipment employing column agglutination technique. All the samples with Rh D negative or discrepant reactions were subjected to weak D testing by the antihuman globulin testing method. Samples that tested positive were categorized as serological weak D type or Variant D and were further phenotyped with Partial D typing set with 6 monoclonal anti D antisera.Results: A total of 82,824 samples were tested for Rh D type during the study period. Of the study population, 65.7% were males. On Rh D type majority were Rh D positive (93%), 6.9% were negative, and the result was discrepant in 0.1% (70) samples. The overall prevalence of variant D was 1.28% (75) of the Rh D negative population and 0.09% of the total study population. The detection rate of variant D phenotype was significantly higher by the Column agglutination technique. Upon testing with Partial D kit, the partial D variant in the majority reacted wil all the 6 antisera and hence we could not rule out DIII(60%), in rest it was inconclusive. In 43% of subjects with Rh D discrepancy 'C' antigen was found in a homozygous state.Conclusion: The introduction of partial D typing kit alone may not help in the absolute characterization of variant D. Extended serological testing and selective integration of molecular testing is the need of the hour.

Frequency of RHD variants in serologically weak D Turkish blood donors.

BACKGROUND: RhD typing has remained of primary importance, as being the leading cause of hemolytic disease of the newborn. Among Rh system's 55 blood group antigens, RhD is the most immunogenic. We aimed with this study to determine weak D/partial D variant frequency in blood donors who were admitted to our blood center and have serologically designated blood group weak D. MATERIALS AND METHODS: We screened blood donors who admitted between 2011 and 2017 to our blood center. Sixty-seven serologically weak D phenotyped donors have participated in the study. These donors' samples were studied further by Polymerase Chain Reaction Sequence- Specific Primers (PCR-SSP) for determining D variants. RESULTS: Weak D phenotype was detected in 228(0.12 %) out of 177,554 donors. Sixty-seven of them agreed to take part in the study. The frequency of weak D and partial D was 68.7 % (n = 46), and 22.4 % (n = 15), in order. The most encountered weak D and partial D variant was type 15 and DFR type, respectively. CONCLUSIONS: The prevalence of serologically weak D phenotypes varies by race and ethnicity. Turkey is a country covering a mixture of European and Asian DNA with different ethnic groups. Thus, our research as giving the overall distribution of RHD variants from the largest city of Turkey, which may reflect the general ethnic background of the country, would help to the establishment of a databank for blood banking. This paper is the first molecular study on RHD variants in Turkey. New molecular research would be more reliable and precise.

D variants in the population of D-negative blood donors in the north-eastern region of Croatia.

OBJECTIVES: The aim of this study was to determine RHESUS D GENE (RHD) allelic variants among Croatian D-negative blood donors and compare our results with respective data from other European countries. BACKGROUND: Altered or reduced D antigen expression can result in D variants, which can be mistyped and can lead to the alloimmunisation of the blood recipient. RHD genotyping can distinguish D variants: weak D, partial D and DEL, thus preventing alloimmunisation. MATERIAL/METHODS: A total of 6523 samples obtained from D-negative Croatian donors were screened for the presence of RHD using the real-time polymerase chain reaction (PCR) method. PCR-SSP was performed for D variant genotyping by using commercial genotyping kits (Inno-Train, Kronberg, Germany). Genomic DNA sequencing for all 10 exons of the RHD was performed when the genotyping kits failed to assign a D variant. RESULTS: RHD molecular screening revealed 23 (0.35%) RHD-PCR positive samples, all C/E positive, in decreasing frequency: 11 hybrid RHD-CE (2-9) D-CE variants, 4 weak partial D type 11 and 2 weak D type 2. Six samples remained unresolved and were sequenced. For 12 of 23 samples (excluding large hybrids), an adsorption/elution of anti-D serum was performed, confirming that all 12 were RhD+. The calculated frequency of clinically significant D alleles in RhD-negative blood donors was 1:543 (0.18%) or 1:53 (1.89%) in C/E blood donors. CONCLUSION: Data on the significant frequency of D variants among serologically D-negative blood donors in the north-eastern region of Croatia could help in introducing RHD molecular screening of blood donors in a routine workflow.

Serologic strategy in detecting RHD altered alleles in Brazilian blood donors

ABSTRACT Background: We evaluated different technological approaches and anti-D clones to propose the most appropriate serologic strategy in detecting the largest numbers of D variants in blood donors. Methods: We selected 101 samples from Brazilian blood donors with different expressions of D in our donor routine. The tests were performed in immediate spin (IS) with eleven commercially available anti-D reagents in a tube and microplate. The D confirmatory tests for the presence of weak D included the indirect antiglobulin test (IAT) in a tube, gel and solid-phase red blood cell adherence (SPRCA). All DNA samples were extracted from peripheral blood and the D variants were classified using different molecular assays. Results: The RHD variants identified by molecular analysis included weak D types (1, 2, 3, 11 and 38) and partial Ds (DAR1.2, DAR1, DAR3.1, DAU0, DAU2, DAU4, DAU5, DAU6, DMH and DVII). The monoclonal-monoclonal blend RUM-1/MS26 was the best anti-D reagent used in detecting the D antigen in the IS phase in a tube, reacting with 83.2% of the D variants, while the anti-D blend D175 + 415 was the best monoclonal antibody (MoAb) used in a microplate to minimize the need for an IAT, reacting with 83.2% of the D variants. The D confirmatory tests using SPRCA showed a reactivity (3 - 4+) with 100% of the D variant samples tested. Conclusion: Our results show that, even using sensitive methods and MoAbs to ensure the accurate assignment of the D antigen, at least 17% of our donor samples need a confirmatory D test in order to avoid alloimmunization in D-negative patients.

Investigation of olfactory interactions of low levels of five off-flavour causing compounds in a red wine matrix.

The qualitative sensory perception of individual and of complex mixtures of five compounds, guaiacol ('burnt note'), o-cresol ('phenolic/tar'), 4-ethylphenol (4-EP, 'leather/barnyard'), 2-iso-butyl-3-methoxypyrazine (IBMP, 'green pepper/herbaceous'), and 2,4,6-trichloroanisole (TCA, 'cork taint/ mouldy') were tested in a partially de-aromatised red wine matrix using descriptive analysis by a trained panel of eleven judges. Compounds were characterised at peri- and sub-threshold concentrations using a partial D-optimal statistical design and response surface methodology. Results indicated that complex mixtures in red wine elicit an olfactory response that could not be predicted from the attributes or descriptors of single compounds. Positive sweet/fruity attributes were more intense in solutions containing fewer off-flavour compounds. Novel findings of this study include that IBMP at sub- and peri-threshold levels shows perceptual interaction with volatile phenols at the same levels, and samples containing combinations of these compounds manifested herbaceous and burnt characteristics. Olfactory interactions of this many off-flavour compounds have not been investigated previously in one study. The findings have direct implications for wines made from cultivars that are known to contain these compounds, and add to the understanding of the behaviour and impact of very low levels (peri- and sub-threshold) of volatile phenols, IBMP, and TCA derived from various sources during winemaking.

Serologic strategy in detecting RHD altered alleles in Brazilian blood donors.

BACKGROUND: We evaluated different technological approaches and anti-D clones to propose the most appropriate serologic strategy in detecting the largest numbers of D variants in blood donors. METHODS: We selected 101 samples from Brazilian blood donors with different expressions of D in our donor routine. The tests were performed in immediate spin (IS) with eleven commercially available anti-D reagents in a tube and microplate. The D confirmatory tests for the presence of weak D included the indirect antiglobulin test (IAT) in a tube, gel and solid-phase red blood cell adherence (SPRCA). All DNA samples were extracted from peripheral blood and the D variants were classified using different molecular assays. RESULTS: The RHD variants identified by molecular analysis included weak D types (1, 2, 3, 11 and 38) and partial Ds (DAR1.2, DAR1, DAR3.1, DAU0, DAU2, DAU4, DAU5, DAU6, DMH and DVII). The monoclonal-monoclonal blend RUM-1/MS26 was the best anti-D reagent used in detecting the D antigen in the IS phase in a tube, reacting with 83.2% of the D variants, while the anti-D blend D175 + 415 was the best monoclonal antibody (MoAb) used in a microplate to minimize the need for an IAT, reacting with 83.2% of the D variants. The D confirmatory tests using SPRCA showed a reactivity (3 - 4+) with 100% of the D variant samples tested. CONCLUSION: Our results show that, even using sensitive methods and MoAbs to ensure the accurate assignment of the D antigen, at least 17% of our donor samples need a confirmatory D test in order to avoid alloimmunization in D-negative patients.

Molecular and computational analysis of 45 samples with a serologic weak D phenotype detected among 132,479 blood donors in northeast China.

BACKGROUND: RH1 is one of the most clinically important blood group antigens in the field of transfusion and in the prevention of fetal incompatibility. The molecular analysis and characterization of serologic weak D phenotypes is essential to ensuring transfusion safety. METHODS: Blood samples from a northeastern Chinese population were randomly screened for a serologic weak D phenotype. The nucleotide sequences of all 10 exons, adjacent flanking intronic regions, and partial 5' and 3' untranslated regions (UTRs) were detected for RHD genes. Predicted deleterious structural changes in missense mutations of serologicl weak D phenotypes were analyzed using SIFT, PROVEAN and PolyPhen2 software. The protein structure of serologic weak D phenotypes was predicted using Swiss-PdbViewer 4.0.1. RESULTS: A serologic weak D phenotype was found in 45 individuals (0.03%) among 132,479 blood donors. Seventeen distinct RHD mutation alleles were detected, with 11 weak D, four partial D and two DEL alleles. Further analyses resulted in the identification of two novel alleles (RHD weak D 1102A and 399C). The prediction of a three-dimensional structure showed that the protein conformation was disrupted in 16 serologic weak D phenotypes. CONCLUSIONS: Two novel and 15 rare RHD alleles were identified. Weak D type 15, DVI Type 3, and RHD1227A were the most prevalent D variant alleles in a northeastern Chinese population. Although the frequencies of the D variant alleles presented herein were low, their phenotypic and genotypic descriptions add to the repertoire of reported RHD alleles. Bioinformatics analysis on RhD protein can give us more interpretation of missense variants of RHD gene.