RHD*weak partial 4.0 is associated with an altered RHCE*ce(48C, 105T, 733G, 744C, 1025T) allele in the Tunisian population.

BACKGROUND AND OBJECTIVES: D is the most immunogenic blood group antigen. About 1% of whites carry an altered RHD allele leading to quantitative or qualitative changes in the antigen D expression. T201R and F223V encoded by 602C>G and 667T>G are specific amino acid substitutions of the weak D type 4 cluster of African origin, comprising the alleles RHD*09.01, RHD*09.02, RHD*09.03, RHD*09.04 and RHD*09.05. The purpose of this study was to estimate the presence of these RHD genotypes in the Tunisian population. MATERIALS AND METHODS: Ethylenediaminetetraacetate blood samples from 907 D+ and 93 D- blood donors were tested for markers 602G and 667G by allele-specific primer-polymerase chain reaction (PCR-ASP). Samples with positive reactions were re-evaluated by DNA sequencing for RHD and RHCE exons 1-10 and adjacent intronic sequences. RESULTS: Among 907 D+ samples, 19 individuals were identified to harbour the RHD*weak partial 4.0 allele. RHCE sequencing post-haplotype-specific extraction (HSE) revealed an altered RHCE*ce(48C, 105T, 733G, 744C, 1025T) in those samples. The linkage of the RHCE polymorphisms to one haplotype was proven by DNA sequencing post-HSE. CONCLUSION: The RHD*weak partial 4.0 allele syn. RHD*09.03 was estimated to occur 1 in 47 among D+ Tunisians. There was no evidence for other RHD alleles included in the weak D type 4 cluster.

Molecular background of D-negative phenotype in the Tunisian population.

BACKGROUND: Most studies of the molecular basis of Rhesus D-negative phenotype have been conducted in Caucasian and African populations. A comprehensive survey of RHD alleles was lacking in people from North Africa (Tunisians, Moroccans and Algerians) which could be very efficient for managing donors and patients carrying an RHD molecular variant. We analyse the molecular background of D-negative population in Tunisia in the present study. MATERIALS AND METHODS: Blood samples were collected from native Tunisians. A total of 448 D-negative donors from different regions of Tunisia were analysed by RHD genotyping according to an adopted strategy using real-time PCR, ASP-PCR and sequencing. RESULTS: Among the 448 D-negative samples, 443 were phenotyped unequivocally as true D-negative including three molecular backgrounds which were RHD gene deletion (n = 437), RHDψ pseudogene (n = 2) and RHD-CE-D hybrid gene (n = 4) with the respective frequencies of 0·9900, 0·0023 and 0·0046. The remaining five samples, in discordance with the serological results, were identified as two weak D type 11, one weak D type 29, one weak D type 4·0 and one DBT-1 partial D. CONCLUSION: This study showed that the Tunisian population gets closer to Caucasians, given that the RHD gene deletion is the most prevalent cause of D-negative phenotype, but it is slightly different by the presence of the RHDψ pseudogene which was found with a very low frequency compared with that described in the African population. Nevertheless, the relative occurrence of weak D variants among studied serologically D-negative samples make necessary the adaptation of RHD genotyping strategy to the spectrum of prevalent alleles.

Minor histocompatibility antigens in Tunisians: could platelet endothelial cell adhesion molecule 1 marker be one of them?

Platelet endothelial cell adhesion molecule 1 (PECAM-1/CD31) is one of the human minor histocompatibility antigens that are the main targets of alloreactive T-cells after hematopoietic stem cells or solid organs transplantation. In order to investigate its polymorphism in Tunisians, three single nucleotide polymorphisms (SNPs) (rs668, rs12953 and rs1131012) were selected to perform an allele and haplotype analysis. Hundred-and-forty-two healthy and unrelated subjects were enrolled in this survey. Genomic DNAs were extracted using salting out method. SNP genotyping assays were performed with home-designed sequence-specific primers polymerase chain reaction (SSP-PCR). As a result, molecular analysis showed that PECAM-1 is one of the most polymorphic markers in the Tunisian population because minor allele frequency was 0.3, and minimum haplotype frequency was 0.03. A low linkage disequilibrium (D' = 0.45) between rs12953 and rs1131012 was noticed, although all other loci were in the Hardy-Weinberg equilibrium (minimum P value = 0.07). The frequencies were close to those reported in African-American and Caucasian groups.

High frequency of autoimmunization among transfusion-dependent Tunisian thalassaemia patients.

BACKGROUND: Limited data are available on the frequency of RBC alloimmunization and autoimmunization in transfusion-dependent Tunisian ß thalassaemia patients. MATERIALS AND METHODS: We analyzed the clinical and transfusion records of 130 patients (57 females and 73 males; mean age 119 months; range 12-11 months) with ß thalassaemia major and who had regular blood transfusions for periods ranging from 12 to 311 months. RESULTS: Of the 130 patients, ten (7.7%) developed RBC alloantibodies. The most common alloantibodies were directed against antigens in the Rh systems. Erythrocyte-autoantibodies as determined by a positive direct antiglobulin Coombs test, developed in 52(40%) patients with and without underlying RBC alloantibodies, thereby causing autoimmune haemolytic anaemia in eleven patients (21%). CONCLUSIONS: Autoimmunization to erythrocyte antigens is a frequent complication in patients with ß thalassaemia major. Several factors might have contributed to the high autoimmunization rate observed in this study, including non phenotypic blood exposure and alloantibody formation prior to positive Coombs test.

HLA class I and class II polymorphism in a population from south-eastern Tunisia (Gabes Area).

The gene frequencies of HLA class I and class II alleles were investigated in 95 healthy Tunisian individuals from Gabes. Our aim was to compare the genetic relationship between Gabesians and Mediterraneans and sub-Sahara Africans using genetic distances, Neighbour-Joining dendrograms, correspondence and haplotypes analysis, thereby providing additional information about evolutionary history of modern-day Tunisians. Subjects were unrelated and of both genders, and HLA class I and class II genes were genotyped using the polymerase chain reaction-sequence specific primer (PCR-SSO) technique. Our data show that south-eastern Tunisians (Gabes area) are related to present-day North Africans (Algerians, Moroccans, Tunisians) and Iberians (Spaniards, Basques), and along with other North Africans, appear to be genetically related to Berbers, an indication that the Arab invasion (7th-11th centuries) of North Africa had minimal contribution on the HLA makeup of North Africans. On the other hand, Iberians including Spaniards and Basques show relatedness to (native Tunisian) Berbers, suggesting that the gene flow of 7th century AD invaders was also low in Iberians. In conclusion, the successive invasions of North Africa in general, and Tunisia in particular, did not modify markedly the genetic makeup of present-day Tunisians. With the exception of Greeks who have a sub-Saharan genetic profile, all Mediterranean populations depict a typical mediterranean substratum.

HLA class I and class II polymorphisms in Tunisian Berbers.

BACKGROUND: The HLA polymorphism is a powerful genetic tool to study population origins. By analysing allele frequencies and haplotypes in different populations, it is possible to identify ethnic groups and establish the genetic relationships among them. AIM: The Berber (endogenous Tunisians) HLA class I and class II genotypes were analysed and compared with those of Mediterranean and Sub-Saharan African communities using genetic distances, Neighbour-Joining dendrograms, correspondence and haplotype analysis. SUBJECTS AND METHODS: One hundred and five unrelated Berbers were typed for HLA class I (A, B) and class II (DRB1, DQB1) gene alleles using reverse dot-blot hybridization. RESULTS: High frequencies of A*0201 (24.76%), A*3402 (22.38%) and B*44 (32.85%) alleles were recorded for Berbers, the highest recorded for Mediterranean and North African populations. This study shows a close relatedness of Tunisian Berbers to other Tunisians, North Africans and Iberians. CONCLUSION: The apparent relatedness of Tunisian Berbers to present-day (North African) Tunisians, Algerians and Moroccans suggests that the Arab invasion of North Africa (7(th)-11(th) centuries AD) did not significantly impact the genetic makeup of North Africans. Furthermore, Tunisian Berbers appear to be closely related to Iberians (Spaniards and Basques), indicating that the 7(th) century AD gene flow of invaders was low in Iberians and that the main part of their genetic pool came after the Northward Saharan migration, when hyper-arid conditions were established in Sahara (before 6000 BC). Other studied populations belong to the old Mediterranean substratum, which has been present in the area since pre-Neolithic times. This study indicates a higher proportion of Iberian than Arab ancestry in Tunisian Berbers, which is of value in evaluating the evolutionary history of present-day Tunisians. Greeks seem to share genetic HLA features (Chr 6) with Sub-Saharans. The relatedness of Greeks to Sub-Saharans has been confirmed by other studies based on chromosome 7 genetic markers.

Transfusion safety in the Maghreb region.

The Maghreb region comprises five countries: Algeria Libya, Morocco, Mauritania, and Tunisia. This is a review aiming at providing an update on the situation of transfusion in the five countries. Three countries have developed regulations covering all transfusion-related activities including policy development. All the countries are running blood safety activities using a National Blood Service as the main entity. Except for Mauritania and Lybia, all the blood safety activities are centralized and conducted regularly. The blood safety indicators are globally better compared to those of sub-Saharan Africa. Despite the efforts of the states of the Maghreb region, and the progress made in the field of transfusion in these countries, shortcomings persist and concern virtually all the key elements of a national blood supply system mainly in the quality management system.

First successful pregnancy outcome after twelve abortions in a Tunisian-woman with the rare p phenotype.

INTRODUCTION: Anti-PP1P k alloantibody, is produced in the serum of individuals with the rare p phenotype. It is associated with severe haemolytic transfusion reactions, recurrent spontaneous early abortions as well as haemolytic disease of the foetus and newborn. Anti-PP1P k alloimmunization in pregnancy differ from others in their physiopathology. It seems that the placenta would be the main target of anti-PP1P k antibody. CASE REPORT: This report concerns a 35 year old female, with a history of a high incidence (12) of early and recurrent miscarriages. She was found to have the extremely rare p phenotype and anti-PP1P k antibody in her serum. Her 13th pregnancy was successfully managed by plasmapheresis. No substitution fluid was added. Oral hydration was recommended before and after the apheresis sessions. 12 plasmapheresis cycles were performed before a healthy term female infant weighing 3kg600g, was delivered by caesarean section at 38 weeks of gestation. CONCLUSION: Plasmapheresis seems to be the treatment of choice in the management of anti-PP1P k fetomaternal incompatibilities. However in this case, we opted for an original and less expensive protocol. We did resort, neither to substitution fluid nor to intravenous immunoglobulin.

The impact of COVID-19 pandemic on blood supplies and transfusion services in Eastern Mediterranean Region.

BACKGROUND: Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) has spurred a global health crisis. The safety and supply of blood during this pandemic has been a concern of blood banks and transfusion services as it is expected to adversely affect blood system activities. We aim to assess the situation in the Eastern Mediterranean Region (EMR) during the first months of the pandemic. MATERIALS AND METHODS: A survey was designed to address blood supply, transfusion demand, and donor management during the coronavirus disease-19 (COVID-19) pandemic. Medical directors of different blood banks were invited to participate. RESULTS: A total of 16 centers participated with representation from 15/19 countries in the region. In total, 75% were from national blood banks. Most centres had a decrease in the blood supply, ranging from 26-50%. Representatives from 14 countries (93.3%) believed that public fear has contributed to a decrease in donations. Most centres (n=12, 75%) had a reduction in transfusion demand, while those who did not, reported heavy involvement in treating patients with underlying haemoglobinopathies and haematological malignancies. Half of the centres activated their contingency plans. Four centres had to alter the blood donor eligibility criteria to meet demands. All centres implemented donor deferral criteria in relation to SARS-CoV-2, but were variable in measures to mitigate the risk of donor and staff exposure. CONCLUSION: Blood services in the region faced variable degrees of blood shortages. We summarize lessons learnt during this pandemic for the blood banks to consider to plan, assess, and respond proportionately to future similar pandemics.

HA-1 and HA-2 minor histocompatibility antigens in Tunisians.

Minor histocompatibility antigens (MiHAgs), such as HA-1 and HA-2, are the main targets of immune responses after allogeneic stem cell transplantation (SCT). HA-1 and HA-2 are two hematopoietic system-restricted antigens encoded, respectively, by HMHA1 and MYO1G genes. In order to estimate their frequencies in Tunisians, we performed a molecular-based allele analysis for 160 healthy and unrelated subjects. Genomic DNAs were extracted mainly by the salting out method. Single nucleotide polymorphism (SNP) genotyping assays for selected sites at HMHA1 gene (rs3764653 and rs1801284) and at MYO1G gene (rs61739531) were performed with a sequence specific primers-polymerase chain reaction (SSP-PCR) method. Statistical analysis of our results showed that the HA-2 antigen is more frequent than the HA-1 antigen in the Tunisian population because their frequencies were 97% and 57%, respectively. Allele analysis for HMHA1 gene showed that the R variant (500T-504G) was predominant in our population (64%). For the MYO1G gene, the C allele was predominant (84%). All loci were in Hardy-Weinberg equilibrium (minimum P value = 0.06). Our frequencies were close to those reported in African and Caucasian groups.

[Polymorphism of human platelet antigens in Tunisian population: Clinical and anthropological interests]. / Polymorphisme des antigènes plaquettaires humains dans la population tunisienne : intérêt clinique et anthropologique.

OBJECTIVES: Human Platelet Antigens (HPA) are of considerable interest in obstetric transfusion medicine and anthropological genetics. This study aims to provide clinicians with a detailed database of HPA antigenic variants, which allows them to estimate the probability of allo-immunisation of each antigen. In addition, it aims to make an interethnic comparison of the Tunisian population with other populations. METHODS: The target population consists of 324 healthy and unrelated Tunisian blood donors recruited from the National Blood Transfusion Center in Tunis. DNA extraction was performed by the Salting Out method and molecular genotyping was performed by the PCR-SSP technique. The statistical analysis was performed using two approaches: manual calculation and computerized calculation. Phylogenetic trees were constructed through the use of Standard Genetic Distances that were calculated from allelic frequencies. RESULTS: With the exception of the HPA-4 system, statistical analysis showed that all HPA systems are polymorphic especially the two systems HPA-3 and HPA-15. The inter-ethnic analysis showed that Tunisians are closer to North Africans and Caucasians than Sub-Saharan and Asian populations, which shows genetic mixing between Tunisians, Arabs, Europeans and Africans. CONCLUSION: The results of this study could be exploited to prepare a ready-to-use genotyping plate dedicated to HPA antigens, with the aim of ensuring better management, especially for polytransfused patients.

Duffy blood group system genotyping in an urban Tunisian population.

BACKGROUND: The Duffy blood group system, besides its relevance in transfusion medicine, is of major interest for population genetics. In fact, the Duffy molecule is the only red cell receptor for Plasmodium vivax, thus the fixation of FY*silent allele in western south-Saharan Africa resulted in the absence of this type of malaria in that area (for a review see Kwiatowski, Am J Hum Genet 77:171-192, 2005). For the Duffy functional role see, for example, Daniels (Vox Sanguinis 93:331-340, 2007). METHODS: Duffy blood group distribution in 115 unrelated Tunisians was determined using the polymerase chain reaction with sequence specific primer (PCR-SSP) method detecting the five allelic versions of the FY gene. The red cell antigenic FY phenotype, for each donor, was deduced through DNA analysis. The blood samples of the positive FY*X alleles were investigated by serological methods, mainly the fixation-elution technique. RESULTS: The following allele frequencies were found (after having excluded FY*X, which had frequency of 0.0174): FY*1 = 0.291 (expressed 0.260; silent 0.031); FY*2 = 0.709 (expressed 0.427; silent 0.282). The most surprising result in this work is the detection of the FY*1 silent allele, usually quite rare, in four samples (1.74%). For FY*2 silent, the predominant allele in Africans, genotyping results showed a prevalence of 29.57%. The FY locus was in Hardy-Weinberg equilibrium in the present sample. CONCLUSION: When compared with European and African data, Tunisian samples demonstrated the presence of the common signs of these two ancestries (FY*2 and FY*X for the first population; and FY*2 silent for the last one). These data confirm the mixed roots of this urban Tunisian population already suggested by numerous studies on other haematological markers.

Peripheral blood stem cell mobilization in multiple myeloma comparison of two consecutive regimens in a limited resources country.

This study compared retrospectively the effectiveness, toxicity and hematopoietic recovery after autologous peripheral blood stem cell transplantation (ASCT) of two consecutive peripheral blood stem cell mobilization regimens in newly diagnosed MM patients. Patients in group 1 (n=178) were treated with 4 g/m2 of cyclophosphamide (CY) plus G-CSF (5 µg/kg/day). Patients in group 2 (n=117) with 750 mg/m2 of VP16 plus G-CSF (10 µg/kg/day). Optimal mobilization, defined by a target number of 8 × 106 CD34+ cells/kg collected, was achieved in 62.4% and 89.7% of patients in groups 1 and 2, respectively (P<10-4). The median number of aphaeresis sessions was reduced from two in group 1 to one in group 2 (P<10-4). Grade4 neutropenia, febrile neutropenia and IV antibiotic use were significantly more frequent in group 1 than in group 2 (P<10-4). Red blood cell transfusion requirements were significantly greater in group 1 (P=0.007). The switch to VP16-G-CSF10 resulted in a significant reduction of the number of hospitalization days (P<10-4). Neutrophil and platelet recovery after ASCT occurred on days 11 and 12, respectively, in the two groups with no significant differences. VP16+G-CSF10 allowed liberation of resources in the clinical and aphaeresis departments and demonstrated a better effectiveness-safety profile than CY+G-CSF5.

HLA genes in Southern Tunisians (Ghannouch area) and their relationship with other Mediterraneans.

South Tunisian HLA gene profile has studied for the first time. HLA-A, -B, -DRB1 and -DQB1 allele frequencies of Ghannouch have been compared with those of neighboring populations, other Mediterraneans and Sub-Saharans. Their relatedness has been tested by genetic distances, Neighbor-Joining dendrograms and correspondence analyses. Our HLA data show that both southern from Ghannouch and northern Tunisians are of a Berber substratum in spite of the successive incursions (particularly, the 7th-8th century A.D. Arab invasion) occurred in Tunisia. It is also the case of other North Africans and Iberians. This present study confirms the relatedness of Greeks to Sub-Saharan populations. This suggests that there was an admixture between the Greeks and Sub-Saharans probably during Pharaonic period or after natural catastrophes (dryness) occurred in Sahara.

[HFE gene mutations in Tunisian major beta-Thalassemia and iron overload]. / Mutations du gène HFE chez des beta-thalassémiques majeurs tunisiens et surcharge en fer.

OBJECTIVES: In this study we have determined the allele frequency of HFE mutations H63D and C282Y in a group of Tunisian beta-thalassemia major patients. These two mutations are implicated in hereditary hemochromatosis among Caucasians. In this study we wanted to correlate these mutations with the iron status in major beta-thalassemia patients. DESIGN AND METHODS: Fifty Tunisian major beta-thalassemia were screening for the C282Y and H63D by digestion of polymerase chain reaction products (RFLP). Serum ferritin level was measured by immunoenzymatic microparticular essay. RESULTS: The allele frequency of H63D mutation was 17%. C282Y mutation was not present in our studied patients. No statistically significant difference of serum ferritin level was found between major beta-thalassemia with and without HFE mutations. CONCLUSION: Our results suggest that H63D mutation is so frequent in Tunisian major beta-thalassemia patients than in the general population and that the coinheritance of H63D mutation does not influence the severity of iron overload in these patients.

Detection of serum hemolysins in autoimmune hemolytic anemia.

BACKGROUND: Some patients with autoimmune hemolytic anemia (AIHA) have in their sera autohemolysins able to hemolyze RBCs in vitro by activation of complement. We describe three autohemolysins in patients with AIHA and we study clinical correlations. STUDY DESIGN AND METHODS: Thirty-two patients with AIHA were explored by immuno-hematological investigations (DAT, elution and serum testing). RESULTS: Three autohemolysins were detected in three patients. All of these autoantibodies were likely IgM and reacted in vitro only with enzyme-treated RBCs. Two warm autohemolysins were detected in patients with warm-type AIHA. The first one was active at neutral pH with low title. The second, having a wide thermal amplitude reacting at 22 degrees C and a title of 16, was acid. The hemolysin detected in patient 3 with cold hemagglutinin disease, was active at 4 and 22 degrees C, at acid pH. The thermal optimum was 4 degrees C and the title 64. It was also detected at 37 degrees C with the same title, but only at neutral pH. CONCLUSION: Although these autohemolysins were incomplete, hemolyzing in vitro only enzyme-treated RBCs, they were associated for the three patients with severe hemolysis.

[Therapeutic and diagnostic difficulties of abdominal actinomycosis: about one case in a Tunisian female patient]. / Difficultés diagnostiques et thérapeutiques de l'actinomycose abdominale : à propos d'une observation chez une patiente tunisienne.

The Abdominal Actinomycosis (AA) is a rare and often unrecognised suppurative chronic illness. The diagnosis is difficult. It can have a variety of clinical manifestations and can mimic a malignancy. It's established by histology of surgical biopsy. We report a case of a 58 years old woman admitted to our institution for a pain of the right iliac fossa with a mass. The diagnosis was established after surgical intervention and histological examination. The treatment was surgical resection and antibiotherapy by amoxcillin during 6 months. The primary diagnosis of abdominal pelvic actinomycosis is difficult. All organs and anatomic structures of the abdomen can be involved. Even with extensive infection, combined operative and antibiotic therapy allows cure in most cases.

Seroepidemiology of hepatitis B, C and delta viruses in Tunisia.

Serum samples from 33,363 healthy people in Tunisia have been tested for serological markers of hepatitis B, C and delta viruses (HBV, HCV and HDV). Hepatitis B surface antigen (HBsAg) was detected in 6.5% of sera. The overall seroprevalence of HBV was 37.5%. Vertical and perinatal transmission of HBV in the first 3 months of life occurred in only 0.4% of 177 mother and child pairs. HBV seroprevalence was 10.7% in infants under 5 years old and increased with age rapidly till 25 years of age and then more slowly in adulthood, reaching 54% for people aged over 40 years. HBsAg seropositivity varied throughout the country, ranging from 3% to 13% with higher prevalences in the south and central-west regions. Overall seroprevalences for HDV and HCV were 17.7% and 0.4%, respectively. HDV superinfection occurred later than HBV and increased with age in parallel with HBV. Overall, HCV and HBV infections had different geographical distributions throughout the country. The study confirmed the high prevalence of HBV infection in Tunisia; it occurs mainly in children and teenagers, and vertical and perinatal transmission of HBV does not appear to be significant. HDV superinfection is quite common in Tunisia and occurs in almost 44% of individuals infected with HBV. In contrast, seroprevalence of HCV in the Tunisian general population was low (0.4%). These results indicate differences in the distribution of the viruses and/or different routes of transmission.

[ABO system polymorphism in the Tunisian population]. / Polymorphisme du système ABO dans la population tunisienne.

A survey carried out on 18820 Tunisian blood donors enabled us to evaluate the distribution of ABO blood group in Tunisia. The genic frequencies in ABO system were as follows: A(0,192), B(0,122), O(0,686).