Plasmodium falciparum circumsporozoite protein: epidemiological variations among field isolates prevalent in India.

OBJECTIVE: To investigate the extent of genetic variations in T-helper-cell epitopic regions of circumsporozoite (CS) protein in Plasmodium falciparum field isolates collected from different regions of India at different phases of malaria transmission. METHODS: Genomic DNA was isolated from 507 P. falciparum wild-parasite isolates obtained from six geographical locations of India at three time points coinciding with malaria transmissions. The T-helper-cell epitopic regions were polymerase chain reaction (PCR)-amplified and the products were purified and then sequenced. RESULTS: Based on sequences, nine variants were found among isolates and they were categorized into nine groups (V-1 to V-9), where V-1 and V-2 were observed in all three time points (TP). The variants V-1 to V-4 in TP-1; V-1, V-2, V-5 to V-8 in TP-2; and V-1, V-2, V-5 and V-9 in TP-3 were present and they showed restricted heterogeneity. During peak transmission (TP-2), parasite populations were more diverse and heterogeneous and the variants regionally unbiased and restricted. However, the alleles of V-6 and V-9 in both Th2R and Th3R showed identical sequence variation with those observed in other geographical regions of the world. The remaining seven groups did not show such similarity. CONCLUSION: The Th2R and Th3R epitopes are implicated in host immune response to P. falciparum. The polymorphism in these epitopic regions indicates antigenic diversity, which may cause adverse outcome of a subunit vaccine including the CS prototype variant. Therefore, the formulation of a vaccine considering the restricted local repertoire parasite populations may be helpful.

Hyper-production of insecticidal crystal protein (delta-endotoxin) by Bacillus thuringiensis var. israelensis is not related to sporulation-specific biochemical functions.

Hypertoxic mutant strains of Bacillus thuringiensis var. israelensis were isolated by mutagenesis of the parent strain. The correlation, if any, between hyper-production of insecticidal crystal protein (delta-endotoxin) by hypertoxic mutant strains of Bacillus thuringiensis var. israelensis and sporulation-specific biochemical functions was studied. No increase in sporulation-specific biochemical markers was observed in the hypertoxic mutant strains. Asporogenous mutants of hypertoxic mutant strains blocked at different stages of sporulation were isolated, and larvicidal activity was studied. The hypertoxic parent strains and the sporulation-deficient, hypertoxic mutant strains showed almost identical larvicidal activity. Therefore, the increased production of toxin is not related to sporulation-specific biochemical changes.

Strong association, but incomplete correlation, between chloroquine resistance and allelic variation in the pfmdr-1 gene of Plasmodium falciparum isolates from India.

The increasing prevalence of chloroquine-resistant Plasmodium falciparum has complicated the control of falciparum malaria. It has been suggested that point mutations at nucleotide positions 754, 1049, 3598, 3622 and 4234 in the parasite's pfmdr-1 gene are associated with such resistance, although this is a matter of controversy. Eighteen chloroquine-sensitive and 22 resistant isolates of P. falciparum from India were investigated, to examine the role of the pfmdr-1 gene in the resistance, and to determine whether any of the point mutations could be used as a marker for the rapid identification of the chloroquine-resistant strains. As this investigation failed to reveal an explicit association between allelic variation in the pfmdr-1 gene and chloroquine resistance, the use of point mutations to identify the resistant strains does not appear feasible.

Activation and germination of spores of Bacillus thuringiensis var israelensis by alkaline pH and larval (Aedes aegypti) gut fluid.

Alkaline activation of the spores of crystalliferous (Cry+) and acrystalliferous (Cry-) strains of Bacillus thuringiensis var israelensis; wild type B. cereus and its transcipient crystalliferous derivatives and wild type B. subtilis was studied. Also the effect of larval (Aedes aegypti) gut fluid on the activation of spores of these strains was studied. Only the spores of the crystal forming strains were found to be activated by 0.1 M K2CO3 (pH 10) and by the larval gut fluid. The process of alkaline activation was independent of whether crystals were present with the spores in the activation solution. This indicates that protoxin in the spore coat is responsible for the alkaline activation process and may have ecological implications for the organism.

Surprisingly little polymorphism in the merozoite-surface-protein-2 (MSP-2) gene of Indian Plasmodium falciparum.

The polymorphism in the merozoite-surface-protein-2 (MSP-2) gene of six Indian Plasmodium falciparum isolates was studied by PCR amplification, cloning and sequencing. One of the isolates showed a deletion of 63 bp and all showed point mutations, although some of these mutations were silent. All the isolates also exhibited 5' and 3' conserved regions, with the two 32-mer amino-acid repeats characteristic of the FC27 family, and none belonged to the IC-1/3D7 family. Although the MSP-2 genes of these isolates represent new allelic sequences, they belong to the FC27 family and show remarkably little variation.

Microbial control of mosquitoes with special emphasis on bacterial control.

There are a number of microbial agents including fungus, protozoa, virus and bacteria which act as mosquitocidal agents. However, among these agents, Bacillus thuringiensis var israelensis and B. sphaericus are the most potent mosquitocidal agents. Bacillus thuringiensis var israelensis and B. sphaericus are gram-positive sporulating bacteria which produce protoxin crystals during sporulation and are highly toxic to susceptible mosquito larvae when they ingest them. These bacterial agents are environmentally safe due to their host specificity, require in very low dosage, easy to prepare commercially in large-scale and are less costly. Field trials with various formulations of B. sphaericus and B. thuringiensis var israelensis have demonstrated their safety and potential for controlling mosquitoes. Moreover, cloning and expression of the toxin genes of these organisms in other environment friendly bacteria have also made them important for further investigation. Recent advances with novel types of recombinant micro-organisms with new cloning strategies and cloning the toxin genes under strong promoter for over expression together with in vitro gene manipulation and site directed mutagenesis of the active sites for increased toxicity have the potential to provide more effective control of mosquitoes by exploiting these two bacteria. But the toxins of B. sphaericus and B. thuringiensis var israelensis in particular, do not persist long in nature and require frequent application which is a limiting factor for these organisms to be most successful and potent biolarvicide. Nevertheless, they are by far the best choice for controlling mosquitoes. In this review article different mosquitocidal agents with particular emphasis on B. thuringiensis var israelensis and B. sphaericus have been described.

Lipid and fatty acid composition of muga silkworm, Antheraea assama, host plants in relation to silkworm growth.

The lipid and fatty acid composition of the leaves (tender, medium and mature) of muga host plants, Machilus bombycina, Litsaea monopetala (primary food plants) and L. cubeba and L. salicifolia (family: Lauraceae) (secondary food plants) was investigated by standard procedures, gas chromatography after saponification and esterification. The total lipid content of M. bombycina and L. monopetala leaves was recorded to be higher (16 and 18 g%), respectively, than that of L. cubeba (10 g%) and L. salicifolia (12 g%). GC analysis identified the presence of eight fatty acids (C14 to C22) and the concentration varied from 0.0297 to 8.1572 g% dry leaf powder. Among the fatty acids, (C14 to C22), polyunsaturated fatty acids were recorded to be highest in concentration in mature leaves of the primary host plants. The concentrations of saturated and polyunsatuated fatty acids were found to be at a minimum level in all the types of leaves of secondary muga host plants.

Termination of midtrimester pregnancies with extraovular 0.1% ethacridine lactate. Accurate method for estimation of blood loss. Role of spartein sulfate.

PIP: This study evaluated the effectiveness of extraovular .1% ethacridine lactate alone and edacridine lactate plus spartein sulfate in midtrimester pregnancy termination. In the 60 cases where ethacridine lactate alone was administered, 50 cases aborted within 48 hours of instillation (83.3% success rate). Of these 50, 25 aborted within 24 hours (41.7%). Abortion was complete in 45 cases. The time of onset of uterine contractions ranged from 1 hour to 16.5 hours, with a mean 21-1/4 hours. The mean time of membrane rupture in the series was 23 hours and the induction-abortion interval averaged 27-1/4 hours. Side effects included vomiting (18.3%), shivering (16.6%), fever (3.3%), cervical injuries (6.6%), and excessive blood loss (1.7%). Blood loss until expulsion of the fetus averaged 54.1 ml, and blood loss up to 4 hours after abortion averaged 115.1 ml in cases of complete abortion and 219 ml in cases of incomplete abortion. In the 90 cases where both ethacridine lactate and spartein sulfate were used, 76 aborted within 48 hours (success rate 84.6%) and 40 aborted within 24 hours (44.4%). Abortion was complete in 75% of cases. The abortion-induction interval ranged from 4 hours to 47-3/4 hours, with a mean of 28-1/2 hours. These results, which are comparable to those obtained in other studies, indicate that extraovular ethacridine lactate alone appears to be a safe, efficient, and relatively inexpensive method of midtrimester abortion. Although there were fewer reports of side effects in the group that received spartein sulfate, use of this compound does not reduce the induction-abortion interval. The relatively low incidence of side effects such as vomiting and diarrhea, the antiseptic properties of ethacridine lactate, and the absence of serious complications such as rupture of the uterus and cervicovaginal fistula are advantages of the ethacridine lactate method that nullify the disadvantage of its slightly prolonged induction-abortion interval.^ieng