Molecular genotyping of Indian blood group antigens amongst regular voluntary blood donors of Surat city, Gujarat, India.

BACKGROUND: There is paucity of data related to the prevalence of the rare blood group antigens amongst South Gujarat blood donor population due to unavailability and high cost of antisera. Therefore it is difficult to screen donors for such rare antigens by gold standard haemagglutination assay. The single nucleotide polymorphism (SNPs) of Ina and Inb antigens is the base of the PCR based detection methods that help to detect these alleles in regular voluntary blood donors. MATERIALS & METHODS: Blood samples of 200 unrelated regular voluntary blood donors wee collected. DNA was extracted using phenol-chloroform method and genotyped for Indian (Ina/IN*01, Inb/IN*02) blood group alleles by Sequence Specific PCR. Ina antigen positivity was confirmed by serology test. RESULTS: Four donors were found heterozygous for Ina antigen i.e. In (a + b+) by SS-PCR and their Ina positivity were confirmed by in-house polyclonal Anti-Ina reagent. SS-PCR was standardized using known heterozygous sample of a blood donor. The frequency of Ina antigen (2.0 %) was higher than Caucasians, lower than Iranians and Arabs while comparable to those reported among Indians of Mumbai city. CONCLUSION: In absence or unavailability of antisera particularly for low frequency alleles like Ina, such PCR based method would be extremely helpful to prepare rare donor registry by screening blood donors' at large scale. Red cells of Ina positive donors can be used as in-house reagent red cells for screening and identification of corresponding antibody.

Assessment of Long-Term in vitro Multiplied Human Wharton's Jelly-Derived Mesenchymal Stem Cells prior to Their Use in Clinical Administration.

The quantity of mesenchymal stem/stromal cells (MSCs) required for a particular therapy demands their subsequent expansion through ex vivo culture. During in vitro multiplication, they undergo replicative senescence which may alter their genetic stability. Therefore, this study was aimed to analyze cellular, molecular, and chromosomal alterations in Wharton's jelly-derived MSCs (WJ-MSCs) during their in vitro sequential passages, where WJ-MSCs were sequentially passaged up to P14 and cells were evaluated at an interval of P2, P6, P10, and P14. They were examined for their morphology, tumorigenicity, surface markers, stemness markers, DNA damage, chromosomal aberration, and telomere length. We have processed five full-term delivered human umbilical cord samples to obtain WJ-MSCs. Morphological appearance observed at initial stages was small fine spindle-shaped WJ-MSCs which were transformed to flat, long, and broader cells in later passages. The cell proliferation rate was gradually decreased after the 10th passage. WJ-MSCs have expressed stemness markers OCT-4 and NANOG, while they showed high expression of positive surface markers CD90 and CD105 and lower expression of CD34 and CD45. They were non-tumorigenic with slow cellular aging during subsequent passages. There was no chromosomal abnormality up to the 14th passage, while increase in comet score and decrease in telomere length were observed in later passages. Hence, our study suggests that early and middle passaged (less than P10) WJ-MSCs are good candidates for clinical administration for treatment.

Red cell alloimmunization among antenatal women attending tertiary care center in Jamnagar, Gujarat, India.

BACKGROUND: The following study was conducted to measure the presence of alloantibodies of Rh and other blood group antigens produced due to fetomaternal hemorrhage in all antenatal women as well as those leading to hemolytic disease of fetus and newborn; presenting to a tertiary care center, G.G. Government Hospital, Jamnagar, Gujarat, India, between April 2014 and March 2016 (2 years). MATERIALS AND METHODS: All multiparous women irrespective of their period of gestation or obstetrics history were included whereas those having taken anti-D immunoprophylaxis or with a history of blood transfusion were excluded. Antibody screening and identification were done using Bio-Rad ID microtyping system. RESULTS: Out of total 8920 multigravida females, 8488 were D-antigen positive whereas 432 were D-antigen negative. A total of 126 antibodies among 117 females (1.31%) were found; out of them, 33 were found in D-antigen positive females (0.39%) and 84 in D-antigen negative ones (19.44%) looking at overall frequency of other antibodies such as anti-C: 9, anti-c: 9, anti-E: 13, anti-Cw: 1, anti-M: 5, anti-S: 8, anti-Fya: 3, and anti-D: 78; it was found that anti-D is the most common. CONCLUSION: The rate of alloimmunization in D-antigen negative women was found to be very high as compared to other studies in western region; hence, strict follow-up of immunoprophylaxis of all Rh D-negative women needs to be taken care of. Apart from this, D-antigen-positive women also show alloimmunization against various antigens giving the prevalence of 0.39%; hence, it should be mandatory that there should be one standard universal protocol for screening of all antenatal women.

Risk of hepatitis-E virus infections among blood donors in a regional blood transfusion centre in western India.

BACKGROUND: Hepatitis-E virus (HEV) is an emerging infectious threat to blood safety. The enormity of the transmission of HEV and its clinical consequence are issues currently under debate. This study aimed to evaluate the prevalence of HEV-RNA in blood donors in western India. MATERIALS AND METHODS: We screened 13 050 blood donors for HEV using HEV-RNA screening of 10 mini-pools using RealStar HEV RT-PCR Kit (95% limit of detection (LOD): 4.7 IU/ml). Furthermore, all HEV-RNA-positive donors were investigated for the presence of IgM/IgG antibody along with liver function tests. RESULTS: Of the 13 050 blood donations, 7 (0.53%) were found to be HEV-RNA positive, and the prevalence of HEV nucleic acid testing yield cases among blood donors was 1 in 1864. All seven HEV-RNA-positive samples were tested with anti-HEV IgM and anti-HEV IgG antibodies; this resulted in two (28.5%) positive anti-HEV IgM and two (28.5%) positive anti-HEV IgG antibodies. Hepatic activity was measured, with two of seven HEV-RNA-positive donors demonstrating abnormal serum glutamic oxaloacetic transaminase (SGOT) andserum glutamic pyruvic transaminase (SGPT). Two HEV-RNA-positive blood donors who had abnormal SGOT and SGPT were found to have a high HEV viral load. Furthermore, we were able to follow up two HEV-RNA donors, and both were HEV-RNA positive and had anti-HEV IgM and anti-HEV IgG antibodies; moreover, their liver function tests were also abnormal. One of the HEV-RNA donors with high viral load did show hepatitis-E-like virus on electron microscopy. CONCLUSION: Our studies indicate that there is a significant risk of blood-borne transmission of HEV. This finding may help to provide a direction towards the safety of blood transfusions in clinical settings in countries like India, which fall under the endemic category for HEV infection.

Seroprevalence of HBV, HCV and HIV-1 and Correlation with Molecular Markers among Multi-Transfused Thalassemia Patients in Western India.

BACKGROUND: Multitransfused ß-thalassemia major patients are always at high risk of having Transfusion Transmitted Infections (TTIs). This study was aimed to determine the seroprevalence of HBsAg, Anti-HIV-1/2, and Anti-HCV among these patients and to correlate the same with NAT testing. METHODS: A total of 196 patients with ß-thalassemia were included in the study. Patients were screened for the presence of viral markers by third-generation ELISA test as well as for viral DNA/RNA by NAT test. RESULTS: Among 196 multi-transfused Beta-thalassemia patients, the seroprevalence of anti-HCV was very high 100 (51.1%), however, anti-HIV1/2 was 6 (3.1%), and HBsAg were 3 (1.5%). Surprisingly similar patterns were observed in the prevalence of molecular markers, as HCV-RNA were 66 (33.7%) of the patients along with HIV-1 RNA were 8 (4.1%), and HBV-DNA were 5 (2.5%) patients. Overall eight (4.1%) patients were found to have coinfections, where two were positive for HBsAg/anti-HCV by ELISA along with 3 (1.5%) were positive for HBV-DNA/ HCV-RNA, 1 (0.5%) was positive for HIV-RNA/HBV-DNA, and 2 (1%) had coinfection of HIV-RNA/ HCV RNA by NAT testing. CONCLUSION: The prevalence of HCV infection among multi-transfused ß-thalassemia patients is significantly higher than that of the HBV and HIV infections. This scenario should be controlled and monitored by doing regular follow-up testing schedules of such patients and also the administration of the booster dose of the HBV vaccine along with HCV treatment with antiviral DAAs.