Multiplex real-time RT-PCR assay for transfusion transmitted viruses in sero-negative allogeneic blood donors: an experience from Southern Pakistan.

BACKGROUND: Blood transfusion safety commences with healthy donor recruitment. The threat of transfusion transmitted infections is greatly minimized by serological tools but not entirely eliminated. Recently, nucleic-acid testing for blood donor screening has virtually eliminated this jeopardy. METHODS: This prospective study was conducted from February 2015 to February 2016. Samples from seronegative donors were run on multiplex assay (Cobas, S-201 system platform, Roche) in a batch of six [MP-NAT]. In case of reactive pool, tests were run on every individual sample [IDNAT]. RESULTS: Of 16957 donors, 16836 (99.2%) were replacement donors and the remaining 121 (0.7%) were voluntary donors, with a mean age of 29.09 ± 7.04 years. After serologic screening of all 16957 donors, 955 (5.6%) were found to be reactive; 291(1.71%) were reactive for hepatitis-B surface antigen, 361 (2.12%) for antibody to hepatitis C virus (anti-HCV), 14 (0.08%) for antibody to human immunodeficiency virus, 287 (1.69%) for syphilis and 2 (0.01%) for malaria. 14 (0.08%) NAT reactive donors were identified after testing the 16002 seronegative donors, with an overall NAT yield of one reactivity out of 1143 blood donations; 10 donors for HBV-DNA (HBV NAT yield-1:1600) and remaining 4 for HCV-RNA (HCV-NAT yield-1:4000). None were HIV positive. CONCLUSION: NAT has improved the safety attributes in blood products. Although the positivity rate for NAT testing is low but in view of the high prevalence of transfusion transmitted infections in our country, we recommend the parallel use of both serology and NAT screening of all donated blood.

Male breast cancer.

OBJECTIVE: To review cases of male breast cancer. SETTINGS: Histopathology Section, Department of Pathology, The Aga Khan University, Karachi. METHOD: Fifty one cases of male breast cancer specimens, received during a period of 10 years, routinely processed and stained with Haematoxylin and Eosin were analyzed. Special stains and Immunohistochemistry were used in difficult cases. RESULTS: Male breast cancer affected individuals in the sixth and seventh decades of life with a mean age of 56.2 years. Infiltrating ductal carcinoma (IDC) was the predominant type. Skin involvement was seen in 27.45% of the cases. Breast lump was the most common presenting symptom followed by skin ulceration. At the time of presentation 43.13% patients had a tumour size of more than 3 cm. CONCLUSION: Male breast cancer is a rare disease. Most of our findings correspond to the published local and international data.

Characterization of pathogenic mutations in 21-hydroxylase gene of Pakistani patients with congenital adrenal hyperplasia and their family members--a preliminary report.

OBJECTIVE: To characterize specific mutations within the 21-hydroxylase gene (CYP21-B) using ARMS-PCR assay in patients with congenital adrenal hyperplasia (CAH) and to compare it with that reported in other populations. SUBJECTS AND METHODS: Five families, having an index case with CAH diagnosed on the basis of clinical and biochemical findings volunteered to give blood samples for analysis. A strategy, based on ARMS-PCR (Amplified Refractory Mutation System) was employed for the detection of mutations in 21-hydroxylase gene. The products of ARMS-PCR were resolved on agarose gels and the PCR products were visualized over ultra violet illumination. RESULTS: Twenty-six specimens were analyzed for common point mutations in the 21-hydroxlase genes at the nucleotide positions 659, 1004 and 1688. Seven samples belonged to index cases with CAH. Of these 7, the assigned sex was male in 5 and female in 2 cases. However, genotypic sex was 3 males and 4 females. The mean age was 8 months in 5 cases while the median 17-OH Progesterone levels was 273.2 ng/ml. Vomiting, precocious puberty and ambiguous genitalia were the presenting features in 2, 1 and 4 cases respectively. Analysis for mutation at 659, 100 and 1688 was performed on 7 index cases and the family members of 5 index cases. The mutation analysis for the family members of index case 6 and 7 was not performed due to non-availability of their blood specimens. Index case No. 1, 4 and 7 showed homozygosity for splice mutations at nucleotide position 659, intron 2 with a sequence change of A to G, while the index case No. 2 and 6 showed heterozygosity for the same mutation. No mutation was found at 659, 1004 or 1688 in index case No. 3 and 4 at the analyzed nucleotide position. Nineteen family members of Case Nos. 1-5 were also analyzed for the same mutations. (Family No. 1, 2, 3, 4 and 5 included 3, 2, 7, 4 and 5 members respectively). These included 8 males and 11 females. All were asymptomatic. Both the parents of index case 1 and 4 were heterozygous at 659 while the father of index case No. 2 was heterozygous at 659 and mother was normal. CONCLUSION: Our results demonstrated the A to G transition at nucleotide 659 causing aberrant splicing, reported for some other populations as the most commonly identified point mutations. All cases were appropriately assigned to paternal or maternal chromosomes.