There is evidence that carcinoma in situ (CIS) is the precursor of invasive urothelial carcinoma, a tumour characterized by frequent gene promoter methylation. The timing of altered DNA methylation is unknown in this pathway. Here we investigate gene methylation in 196 consecutive samples of normal urothelium, CIS, and tumours from 104 patients with both CIS and invasive urothelial carcinoma using quantitative methyl-sensitive polymerase chain reaction for six genes (p16, p14, E-cadherin, RARbeta2, RASSF1a, and GSTP1). Control normal urothelial samples from 15 patients with no history of urothelial carcinoma were also analysed. Immunohistochemistry established the expression of well-characterized CIS markers p53 and cytokeratin 20. Promoter methylation occurred frequently in both normal urothelium and CIS samples from patients with urothelial carcinoma, and increased with progression from normal to invasive urothelial carcinoma, at both specific loci (chi2 test: E-cadherin, p=0.0001; RASSF1a, p=0.003, RARbeta2, p=0.007, p16, p=0.024) and in general (methylation indices [t-test, p<0.0001]). Methylation was associated with cytokeratin 20 expression (t-test, p=0.004) and poor prognosis, and with increased progression to tumour death in patients whose CIS samples showed methylation, in comparison with those without methylation (log rank p<0.03). Promoter methylation occurs early in the urothelial carcinogenic pathway and appears to be a good biomarker of the invasive urothelial carcinoma phenotype.

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We report the draft genome of Brevundimonas diminuta strain XGC1, isolated from a tuberculosis-infected patient in Gujarat, India. This study also reveals that the B. diminuta XGC1 strain has acquired mutation to confer resistance to quinolone drugs.

Co-inheritance of sickle cell anemia and β-thalassemia traits require medical attention. Individuals with sickle cell and β-thalassemia disorders produce abnormal form of hemoglobin or decreased synthesis or complete absence of the β-globin chains of hemoglobin. Therefore, affected individuals might require blood transfusions at regular intervals. Prenatal diagnosis of fetal hemoglobinopathy should be offered when the fetus is at risk of being affected. The aim of this study was to assess the applicability of the nucleotide sequencing method in the identification of both diseases and to identify and counsel asymptomatic parents to make reproductive choices. We demonstrate the ability to detect these traits in a family, which was suspected to be carrying β-thalassemia mutations as per the clinician.

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β-Thalassemia is a genetic disease characterized by reduced or non-functionality of β-globin gene expression, which is caused due to a number of variations and indels (insertions and deletions). In this case study, we have reported a rare occurrence of compound heterozygosity of two different variants, namely, HBBc.92G > C and HBBc.92 + 5G > C in maternal amniotic fluid sample. Prenatal β-thalassemia mutation detection in fetal DNA was carried out using nucleotide sequencing method. After analysis, the father was found to be heterozygous for HBBc.92G > C (Codon 30 (G > C)) mutation which is β0 type and the mother was heterozygous for HBBc.92 + 5G > C (IVS I-5 (G > C)) mutation which is β+ type. When amniotic fluid sample was analyzed for β-globin gene (HBB), we found the occurrence of heterozygous allelic pattern for aforesaid mutations. This compound heterozygous state of fetus sample was considered as β+/β0 category of β thalassemia which was clinically and genotypically interpreted as β-thalassemia major. Regular blood transfusions are required for the survival of thalassemia major patients hence prenatal diagnosis is imperative for timely patient management. Prenatal diagnosis helps the parents to know the thalassemic status of the fetus and enables an early decision on the pregnancy. In the present study, we have identified compound heterozygosity for β-thalassemia in the fetus which portrays the importance of prenatal screening.

About 200 causative mutations are characterized in the β-globin gene. Beta thalassemia diagnosis is very complicated due to the genetic diversity of HBB gene across different geographical regions of the world. In the present study, we have analyzed 138 clinical specimens among them 66 were from 21 unrelated families (trio samples which had DNA from father, mother and chorionic villus sample/amniotic fluid sample) and 72 individual specimens using newly developed sequencing and PCR based assay. We observed 11 different HBB gene mutations in 138 samples, which were also cited by literature as the most prevalent mutations in Indian subcontinent population. The most common mutation observed in our study was HBB.C.92+5 G>C (GC+CC genotype was observed to be 44.93%). Few interesting case studies like co-inheritance of sickle cell anemia and β-thalassemia traits, compound heterozygosity of beta thalassemia major mutation in the case of twin pregnancy were also focused briefly. Commercially available molecular diagnostic kits of HBB gene can detect and identify targeted mutations but will not detect novel and non-targeted mutations of beta thalassemia in parental blood and fetal samples. Hence, a screening technique involving complete sequencing of HBB gene (β-globin gene) is required along with gap PCR approach to provide complete diagnosis of beta thalassemia disease.

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