The Impact of ABO Incompatibility on the Outcomes of Hematopoietic Stem Cell Transplantation: A Single-Center Study From Pakistan.

Background and objective Allogeneic hematopoietic stem cell transplantation (alloHSCT) provides curative treatment for several hematological illnesses. In this study, we evaluated the impact of ABO compatibility and incompatibility on outcomes and complications related to hematopoietic stem cell transplantation (HSCT) performed for various hematological disorders at our center. Methodology This was a retrospective, single-center, cohort study in which patients were categorized according to the ABO match and mismatch status. The mismatch group was further subcategorized into major, minor, and bidirectional groups. Results A total of 117 patients underwent alloHSCT, out of which 82 (70.1%) were male and 35 (30%) were female. The median age of the patients was 9.5 years (range: 46 years). The most common indications for stem cell transplant were beta-thalassemia major (BTM; n=58, 49%) and aplastic anemia (AA; n=42, 35.8%). However, the outcomes in match and mismatch groups showed significant results for positive direct Coombs test (DCT), indicating the occurrence of hemolysis. Despite the increased need for blood transfusions, ABO blood group incompatibility (ABOi) had no negative impact on the clinical results. Conclusion Based on our findings, ABO incompatibility does not affect the outcomes in patients undergoing alloHSCT. Patient monitoring can aid in early detection and treatment, thereby minimizing the frequency of fatal events.

Management of Severe Hemophilia A: Low-Dose Prophylaxis vs. On-Demand Treatment.

INTRODUCTION: Prophylactic clotting factor infusion regimens to prevent bleeding and joint deformity has become the standard of care in severe hemophilia A patients. AIM: To assess low-dose factor prophylaxis in our population as an alternative approach to managing severe hemophilia A. METHODS: A prospective cohort study that included 68 hemophilia A patients divided into two groups, i.e., Prophylaxis and on-demand. The two groups were compared for annualized bleeding rate (ABR), hospitalization, units of factor VIII (FVIII) infused, or plasma products transfused, i.e., fresh frozen plasma (FFP) and cryoprecipitate (CP), and development of FVIII inhibitors. RESULTS: Of the 68 patients recruited in this study, 25 (36.7%) were in the prophylaxis group, and 43(63.3%) were in the on-demand group. The on-demand group presented a higher median-IQR ABR [8(20-3) vs. 5(10-1.5), p-value 0.024], several hospitalizations (39.7% vs. 0, p-value 0.001), and inhibitor development (9.3% vs. 0, p-value 0.289) compared to the prophylaxis group. The prophylaxis approach demonstrated a significant negative correlation of ABR with FVIII prophylaxis (r=-0484, p=value=0.014). Moreover, no hospitalizations or inhibitor development was observed in the prophylaxis group. The estimated annual consumption of FVIII was 328 IU/kg/year in the on-demand group and 1662.6 IU/kg/year in the prophylaxis group. However, a highly significant difference in plasma product utilization was observed between the two groups, i.e., p-value <0.001 and 0.038 for FFP and CP, respectively. CONCLUSION: Low-dose factor prophylaxis resulted in improved outcomes compared to on-demand treatment in terms of ABR, joint bleeding, hospitalization, and the development of inhibitors. This treatment approach should be adopted as an economically feasible alternative to high-dose Prophylaxis in resource-constrained countries.

Cytogenetic culture failure and its causes in hematological disorders; a single centre perspective.

OBJECTIVE: To highlight the reasons of culture failure in bone marrow aspirate samples sent for Cytogenetic analysis and to identify the associated parameters causing this impact. METHODOLOGY: This is a retrospective cross-sectional study conducted in the Clinical and Molecular Cytogenetics Laboratory of NIBD Hospital, Karachi, Pakistan. The rates of culture failure are assessed from the year 2017-2020 along with their reasons. Bone Marrow aspirate samples of patients with hematological malignancies were cultured for chromosomal analysis, both at the time of diagnosis or relapse. Statistical analysis was performed using SPSS version 25. RESULTS: A total of 1061 bone marrow aspirate samples were assessed for cytogenetic culture failures from the duration of 2017 to 2020. Ratio of males was predominantly higher i.e. 62.7% than female 37.3% with Mean ± SD age was 36.78 ± 18.94. Frequency of culture failure in the year 2020 was relatively high 20% as compared to the preceding years i.e. 8% in 2017, 6% in 2018, 7% in 2019. However, the patients were diagnosed with the following hematological malignancies; ALL 23%, CML 17.1%, AML 16.5% and AA 12.5%. Among the reasons of culture failure, cytogenetic analysis of patients with on-going chemo resulted in significant culture failures with p-value < 0.001 and the hematological malignancy, Acute Promyelocytic Leukemia, significantly impacted the growth of bone marrow aspirate cultures, with p-value < 0.001. CONCLUSION: Significant findings were associated with causative factors of culture failure including on-going treatment and sample issues of clotted bone marrow as well as with the clinical diagnosis. These evaluations facilitated in overcoming the rise in culture failures. As per our knowledge, no such data, discussing the effects of various parameters such as sample quality, diagnosis, effects of treatment etc., has been documented previously.

Frequency of Intron 22 Inversion in Severe Hemophilia A Patients.

OBJECTIVE: The aim of our study was to find the frequency of Intron 22 inversion (Inv22) in severe hemophilia A (HA) patients and to evaluate the association between Inv22 and FVIII inhibitor formation. METHOD: Data analysis was carried out on IBM SPSS Statistics Version 23.00 (IBM Corp, Armonk, NY). Descriptive statistics were applied to measure the frequencies, percentages, and mean ± SD of the clinical and general history of HA patients, including age, family history, inhibitor status, intron22 inversion, and FVIII levels. Chi-square was applied to evaluate the association between Inv22 and F8 inhibitor formation. RESULTS: A total of 62 HA patients were enrolled in the study with mean±SD age of (14.39±13.2) years. A family history of HA was observed in 36 (58.1%) patients. Out of 62 patients, 28(45.2%) were reported as Inv22 positive while inhibitor status was observed as positive in three (4.83%) patients. However, an insignificant association was observed between the inhibitor and Inv22 positive patients with a p-value=0.443. CONCLUSION: In our study, Inv22 was found to be the major cause of severe HA in our patients, i.e., 45.1%. However, no significant relation was computed between Inv22 and inhibitor formation.

Frequency of Specific and Non-specific Inhibitors in Haemophilia A Patients.

OBJECTIVE: To determine the frequency of specific and non-specific inhibitors in haemophilia A patients. STUDY DESIGN: This is a cross-sectional study. PATIENTS AND METHODS: A total of 150 male haemophilia A patients were included in this cross-sectional study at the National Institute of Blood Diseases and Bone Marrow Transplant (NIBD), Karachi, Pakistan, from September 2019 to January 2022. RESULTS: Among 150 patients included in this study, 23 (15.3%) had an inhibitor and 127 (84.6%) did not. All patients had specific inhibitors against Factor VIII (FVIII). Non-specific inhibitors were not identified in our population. Among the patients in the inhibitor group, there were 13 (56.5%) in the severe (<1%) category. There were 10 (43.5%) patients in the moderate (1-5%) category. There were no patients in the mild category. The median inhibitor level was 15.4 Bethesda unit (BU). CONCLUSION: The development of inhibitors has not been identified as a major problem in our population. However, it is noteworthy that only 15.3% of patients with haemophilia A developed inhibitors in this data set. They were essentially treated with plasma and its products.

Thrombocytopenia in Pregnancy: Identification and Management at a Reference Center in Pakistan.

OBJECTIVE: The study aimed to evaluate the causes of thrombocytopenia in pregnancy and its management along with the outcome in the COVID-19 era. METHODS: Recruitment for this prospective, cross-sectional observational study of thrombocytopenia in pregnancy (platelet counts <100x109/L) was done from January 2017 to August 2020 at the National Institute of Blood Diseases (NIBD) after taking the patients' informed consent. Complete clinical and lab profile of patients was also collected. RESULTS: A total of 150 pregnant women with thrombocytopenia were enrolled, with the mean age being 27.3±4.64 years. Mean platelet counts at baseline were 48.0±24. Main clinical manifestations at baseline included: anemia 65.9%, bruises 23.25%, and edema 9.3%. Causes of thrombocytopenia were gestational thrombocytopenia (GT) 72 (48%), acute fatty liver five (3.3%), pre-eclampsia in 11 (7.3%), and eclampsia seven (4.6%). Causes not specific to pregnancy included 30 (20%) cases of ITP, hepatitis C, and nutritional deficiency was reported in nine (6%) patients each. 72/150 received supportive care treatment to manage thrombocytopenia and were closely monitored and given supplements. Twenty (66.6%) ITP patients received treatment with steroids, with complete response in 70% of them seen. Overall, 38 (25.3%) women with bleeding symptoms and platelet count <50x109/L received platelet transfusions. CONCLUSION: The study shows that pre-eclampsia and eclampsia are serious conditions with a high risk for complications, while GT is a benign and the most common cause of thrombocytopenia in pregnancy which requires no active treatment. The other causes such as ITP and infections require individualized management.

Blood Ordering and Transfusion Practices: An Insight Toward Better Utility of Blood Products.

Background An adequate supply of quality blood products is the backbone of any hospital. To maintain it, the utilization and wastage of the products should be closely monitored. Objective To determine the crossmatch to transfusion (C/T) ratio, transfusion probability (%T), and transfusion index (Ti) of packed red blood cells and to review the use of platelets. Materials and methods A total of 6,326 hematological patients receiving packed red blood cells were included in the study. The random donor platelets that were prepared during this period were also included to know the actual utilization of platelets. Results A total of 26,146 crossmatches were requested for these 6,326 patients in three years. Out of these, 26,024 units were issued and transfused to the patients. The CT ratio of our data was calculated to be 1.00, the transfusion probability was found to be 98.1%, and the transfusion index was computed to be 0.99. For random donor platelets, 37,162 were prepared from whole blood during this period, while 30,971 platelets were transfused to the patients. Conclusion The overall results of our analysis showed proper utilization of blood products at our institution. The wastage was considered to be minimal.

Assessment of inducible clindamycin resistance and Hyper Variable Region (HVR) of mecA gene in clinical staphylococci.

OBJECTIVE: To study the prevalence of inducible clindamycin along with vancomycin and methicillin resistance and assessment of hyper variable region (HVR) of mecA gene among different clinical isolates of Staphylococcus spp. METHODS: A total of 176 clinical isolates of Staphylococci were collected from Pakistan Institute of Medical Sciences (PIMS), Islamabad during 2014-2015. The sample sources were pus, blood, urine, sputum, tracheal secretions and tissue fluids. Bacterial identification was done by colony morphology and biochemical tests. Kirby-Bauer disc-diffusion method was carried out to assess the susceptibility against different antibiotics. Minimal inhibitory concentrations (MICs) were done for vancomycin resistance. Double Disk Diffusion test (D-test) was used to detect the clindamycin inducible resistance. PCR was performed to detect erm(C), mecA and HVR genes. RESULTS: Clindamycin inducible resistance among Staphylococcal isolates was found to be 7%, whereas in S. aureus it was 4%, and in coagulase negative Staphylococci (CoNS) it was 11%. The highest resistance was observed against fosfomycin, fusidic acid and cefoxitin. Vancomycin resistance was observed in 23 isolates (13%) of Staphylococci. erm(C), mecA and HVR genes were found in 18%, 50% and 42% respectively. CONCLUSIONS: D-test must be performed routinely to avoid clindamycin failure. A high level of resistance against vancomycin in Staphylococcal isolates is a concern for public health.

Coagulation abnormalities in dengue and dengue haemorrhagic fever patients.

OBJECTIVE: The aim of this study was to assess abnormality of coagulation and anticoagulation parameters in dengue fever patients and the impact of these tests on the management of patients. BACKGROUND: Dengue fever is endemic in Pakistan with seasonal rise in cases. Morbidities and mortalities are proportionately reported to be increasing and associated with disseminated intravascular coagulation resulting in haemorrhagic or thrombotic manifestations in patients having deranged coagulation profiles. METHODS/MATERIALS: This observational and descriptive study was conducted on confirmed Dengue patients at the National Institute of Blood Diseases during the years 2013 to 2016. Patients of all age groups were included in this study. Results were analysed by SPSS version 23. RESULTS: A total of 200 patients were selected with the mean age being 28.68 years (±13.28) and male predominance (147/200). The mean platelet count, haemoglobin and haematocrit at base line for bleeders and non-bleeders showed significant results, where platelet count at baseline for bleeders was 24 000, whereas for non-bleeders it was 29 000 and it showed significant correlation with bleeding (P-value .027). Platelets were transfused to 76 (38%) patients. However, none of the specialised haemostasis parameters beside the platelet count correlated with bleeding, requiring platelet transfusions. CONCLUSION: Our study showed a significant association of platelet counts, haemoglobin and haematocrit with bleeding. It can be concluded that coagulation and anticoagulation profiles will not benefit the management of dengue patients and in countries like Pakistan, it will only add to the economic burden on the patients.

Genotyping of five Pakistani patients with severe inherited factor X deficiency: identification of two novel mutations.

: Congenital factor X deficiency is a rare coagulation defect characterized by variable bleeding tendency. The aim of the study was to give a first insight of F10 gene mutations in Pakistani probands. Direct sequencing and/or next-generation sequencing was performed on the coding regions, boundaries and 5' and 3' untranslated regions of the F10 gene in five severe factor X-deficient patients from Pakistan. All patients were born from consanguineous marriages and displayed FX:C levels below 2%. Sequencing revealed five different substitutions, including three previously reported p.Ala15Asp, p.Gly406Ser, and p.Gly420Arg missense variants, and also two novel variants: p.Cys57Arg and p.Gln175*. Though one genotype could not be characterized, we were able to confirm the inherited nature of the defect using familial studies. As the copy number variations were ruled out, we hypothesized the presence of deep intronic mutants that might have escaped detection from sequencing or abnormalities in epigenetic regulation. Three patients presented with severe clinical symptoms, in the early days of life, whereas two presented only with trauma-provoked bleeds and bruises later in life. Those patients with milder forms bore the p.Gly406Ser at the homozygous state and F10 unknown alleles, respectively. F10 mutation spectrum in Pakistan is heterogeneous as seen in other populations. Identification of the F10 mutations is important for genetic counseling and prenatal diagnosis in subsequent pregnancies.

Complex Karyotype in Hematological Diseases: A 6-Year Single Centre Study from Pakistan.

BACKGROUND: Most of the hematological disorders are heterogenous with regard to morphology, immunophenotype, and genetic rearrangements. Multiple recurrent chromosomal aberrations have been identified by conventional cytogenetic analysis, which is now widely recognized as one of the most important diagnostic and prognostic determinants in these patients. Though rarer, complex karyotype has been associated with worst prognosis. MATERIALS AND METHODS: A total of 1185 bone marrow or peripheral blood cytogenetics samples were taken with different hematological diseases. They included both benign and malignant disease entities. In each case, cells were cultured and conventional cytogenetic analysis was performed. RESULTS: Among 1185 subjects, 41 (3.4%) patients possessed complex cytogenetic abnormalities. Out of these 41, 33 (80%) were males. The mean age was 37 years (median age 39 years). Myelodysplastic syndromes had the most numbers of complex karyotypes (8%), followed by acute myeloid leukemia (7%) and acute lymphoblastic leukemia (4%). Also we found few patients with acute promyelocytic leukemia, aplastic anemia , chronic myeloid leukemia, and diffuse large B cell Lymphoma possessing complex karyotype. Frequencies of different cytogenetic abnormalities were assessed with respect to disease as well as independently. Trisomy 21 was the most common chromosomal abnormality found in 28% of patients. CONCLUSION: Complex karyotype was most frequently associated with myelodysplastic syndromes and acute myeloid leukemia. Trisomy 21 and deletion 5q were the commonest cytogenetic abnormalities found. We also assessed complex karyotype in benign diseases and detected one patient of aplastic anemia with complex karyotype. This is the first study highlighting the presence of complex karyotypes in hematological disorders in our region.

Application of the ISTH bleeding score in hemophilia.

BACKGROUND: Hemophilia is an inherited bleeding disorder. With proper treatment and self-care, persons with hemophilia can maintain an active, productive lifestyle. Hemophilia can be mild, moderate, or severe, depending on the degree of plasma clotting factor deficiency. The aim of the study was to assess the utility of ISTH-BAT in diagnosis, determining severity of the bleeding condition in newly diagnosed and known hemophilia patients, compare the bleeding score (BS) in adult and pediatric groups and investigate its association with plasma factor levels. METHODS: ISTH-BAT was used to assess BS in a total of 115 patients, 78 with hemophilia A, and 37 with hemophilia B and in 100 controls. RESULTS: BS was significantly higher in HA and HB patients as compared to controls, with no significant difference between HA and HB. The BS was very similar in newly diagnosed compared to known hemophilia patients, lower in pediatric compared to adult and higher in severe compared to mild HA patients. CONCLUSION: The ISTH BAT can help identify hemophilia patients. Therefore it is a useful tool to distinguish between affected and unaffected individuals with bleeding. Moreover, an important finding of our study is that there is no major difference between the scores in known and newly diagnosed patients.