Cloning and sequencing of beta toxin gene of Clostridium perfringens type C

A gene encoding beta toxin was amplified by polymerase chain reaction from C. perfringens type C isolate and cloned in pUC 19 vector. The nucleotide sequence was identical with C. perfringens type B beta toxin gene sequence. The Southern hybridization using labelled beta toxin gene probe revealed the presence of positive signals only in beta producing C. perfingens.     

Pax6 heterozygous eyes show defects in chamber angle differentiation that are associated with a wide spectrum of other anterior eye segment abnormalities

The development of the chamber angle was studied in the eyes of heterozygous Pax6(lacZ/+) mutant mice (Nature 387 (1997) 406). Mutations in PAX6 cause aniridia, a condition that is frequently associated with glaucoma, a blinding disease that may be associated with chamber angle defects. Mesenchymal cells were seen in the chamber angle at P1-P5. In wild-type mice, these cells differentiated into typical trabecular meshwork (TM) cells next to Schlemm's canal. In Pax6(lacZ/+) mice, TM cells remained undifferentiated and Schlemm's canal was absent. From P1 to P4, staining for beta-galactosidase and immunoreactivity for Pax6 were observed in chamber angle mesenchyme, but were absent later. Cultured murine TM cells expressed Pax6. The defects in chamber angle and TM differentiation were associated with a wide spectrum of other anterior eye defects, which included various degrees of iris hypoplasia and corneal haze, isolated iridocorneal adhesions and atypical coloboma, and a vascularized cornea in all adult animals. A third of the animals showed Peters' anomaly including corneal opacity and iridocorneal adhesions. The separation of the lens from the cornea was incomplete, and epithelial layers of lens and cornea were continuous. Pax6 activity is directly required for differentiation of the chamber angle. Variations in phenotype of Pax6(lacZ/+) mice appear not to involve direct dominant-negative or dose-dependent effects.     

Volume Regulation and Renal Function at High Altitude across Gender

Aims

We investigated changes in volume regulating hormones and renal function at high altitudes and across gender.

Methodology

Included in this study were 28 subjects (n = 20 males; n = 8 females. ages: 19 – 65 yrs), who ascended to a height of 3440m (HA1), on the 3^rd day and to 5050m (HA2), on the 14^th day. Plasma and urinary creatinine and urinary osmolality as well as plasma levels of plasma renin activity (PRA), Aldosterone, antidiuretic hormone (ADH), and atrial natriuretic peptide (ANP) were measured. The plasma volume loss (PVL) was estimated from plasma density and hematocrit. Glomerular filtration rate (GFR) was measured based on nocturnal (9 hour) creatinine clearance; this was compared with various methods for estimation of GFR.

Results

The mean 24-hour urine production increased significantly in both sexes across the expedition. But PVL reached significance only in males. No changes in Na⁺ in plasma, urine or its fractional excretion were seen at both altitudes. Urinary osmolality decreased upon ascent to the higher altitudes. ADH and PRA decreased significantly at both altitudes in males but only at HA2 in females. However, no changes in aldosterone were seen across the sexes and at different altitudes. ANP increased significantly only in males during the expedition. GFR, derived from 9-h creatinine clearance (CreaCl), decreased in both sexes at HA1 but remained stable at HA2. Conventional Crea[p]-based GFR estimates (eGFR) showed only poor correlation to CreaCl.

Conclusions

We report details of changes in hormonal patterns across high altitude sojourn. To our knowledge we are not aware of any study that has examined these hormones in same subjects and across gender during high altitude sojourn. Our results also suggest that depending on the estimation formula used, eGFR underestimated the observed decrease in renal function measured by CreaCl, thus opening the debate regarding the use of estimated glomerular filtration rates at high altitudes.     

An Efficiently Cleaved HIV-1 Clade C Env Selectively Binds to Neutralizing Antibodies

An ideal HIV-1 Env immunogen is expected to mimic the native trimeric conformation for inducing broadly neutralizing antibody responses. The native conformation is dependent on efficient cleavage of HIV-1 Env. The clade B isolate, JRFL Env is efficiently cleaved when expressed on the cell surface. Here, for the first time, we report the identification of a native clade C Env, 4-2.J41 that is naturally and efficiently cleaved on the cell surface as confirmed by its biochemical and antigenic characteristics. In addition to binding to several conformation-dependent neutralizing antibodies, 4-2.J41 Env binds efficiently to the cleavage-dependent antibody PGT151; thus validating its native cleaved conformation. In contrast, 4-2.J41 Env occludes non-neutralizing epitopes. The cytoplasmic-tail of 4-2.J41 Env plays an important role in maintaining its conformation. Furthermore, codon optimization of 4-2.J41 Env sequence significantly increases its expression while retaining its native conformation. Since clade C of HIV-1 is the prevalent subtype, identification and characterization of this efficiently cleaved Env would provide a platform for rational immunogen design.     

Molecular phylogeny of C1 inhibitor depicts two immunoglobulin-like domains fusion in fishes and ray-finned fishes specific intron insertion after separation from zebrafish

C1 inhibitor (C1IN) is a multi-facet serine protease inhibitor in the plasma cascades, inhibiting several proteases, notably, regulates both complement and contact system activation. Despite huge advancements in the understanding of C1IN based on biochemical properties and its roles in the plasma cascades, the phylogenetic history of C1IN remains uncharacterized. To date, there is no comprehensive study illustrating the phylogenetic history of C1IN. Herein, we explored phylogenetic history of C1IN gene in vertebrates. Fishes have C1IN with two immunoglobulin like domains attached in the N-terminal region. The RCL regions of CIIN from fishes and tetrapod genomes have variations at the positions P2 and P1′. Gene structures of C1IN gene from selected ray-finned fishes varied in the Ig domain region with creation of novel intron splitting exon Im2 into Im2a and Im2b. This intron is limited to ray-finned fishes with genome size reduced below 1 Gb. Hence, we suggest that genome compaction and associated double-strand break repairs are behind this intron gain. This study reveals the evolutionary history of C1IN and confirmed that this gene remains the same locus for ∼450 MY in 52 vertebrates analysed, but it is not found in frogs and lampreys.     

Effect of temperature and arsenic on Aeromonas hydrophila growth,a modelling approach

Aeromonas hydrophila is frequently reported from arsenic affected areas. Present study was aimed to determine the effect of arsenic and temperature on growth of A. hydrophila. The bacteria were isolated from naturally infected fish from a water body in Birbhum, West-Bengal, India, which is reported to be an arsenic-free area. Arsenic concentration in natural aquatic reservoirs (e.g., pond, lake or river) varies from 0–6 mg/L. No significant change in bacterial growth was observed within this range of arsenic exposure. However, variation in temperature impacted the growth of A. hydrophila. A single dimension model was constructed using simple logistic equation. Rate parameters of the model were derived from the experimental observations. Comparison of model results and laboratory observations gives a good conformity regarding the effect of variation of arsenic concentration and temperature change on growth of this bacterium. From the analysis of this model we further get the idea that the maximum growth of A. hydrophila is supposed to be at 31.4°C in absence of arsenic, whereas at 477 mg/L arsenic concentration, the growth of the bacteria totally stops at 30°C.     

Genetic relatedness among fish species of Genus Channa using mitochondrial DNA genes

The family Channidae is represented by 26 species, out of which 23 species are found in Asia. However, the taxonomy and phylogeny of the Channid fishes found in India are poorly understood. In the present study, eight species of Channa (Channa striata, Channa punctatus, Channa marulius, Channa gachua, Channa stewartii, Channa aurantimaculata, Channa barca and Channa bleheri) were investigated using partial sequences of 16S rRNA and Cytochrome c Oxidase subunit I (COI) of mitochondrial genes to differentiate among the eight species and study their relationships. The sequence analysis of the genes revealed two distinct groups, which are genetically distant from each other and exhibit identical phylogenetic resolution. The partial sequences of both the genes provided sufficient phylogenetic information to distinguish all the eight species of Channa.