Polymerisation by the chromate/arsenite system

The aqueous polymerisation of methyl methacrylate initiated by the chromate/arsenite system has been studied. It is observed that the polymerisation by the above redox system is catalysed by OH^? though the parent reaction between chromate and arsenite is catalysed by H^⊕. It is also observed that traces of Cu^2⊕ inhibit both the polymerization reaction as also the parent reaction. From these observations it is concluded (1) intermediate valency states of chromium has no initiating power in alkali solution and (2) the redox reaction between chromate and arsenite is a chain reaction involving single electron transfer and the intermediate As^4⊕ thus produced is the initiating species.     

Selection for high and low mating propensity inDrosophila ananassae

InDrosophila ananassae, artificial selection was carried out for high and low mating propensity for 15 generations. Response to selection was from about F₅, with rapid divergence in mating frequencies in replicates of both fast and slow lines. To assess the effect of selection on the two sexes, females and males of the selected lines were tested against their respective counterparts of the control line after 15 generations. Significant differences in mating propensity were observed when selected males were tested against the control females, which suggests that males were much more affected by selection than females. After 15 generations the fast and slow lines (both replicates) were crossedinter se and mating frequencies of F₁ hybrids were studied in the same way as during the selection experiment. F₁ flies had a higher mating activity compared to their parental lines when males were derived from fast lines to produce hybrids. On the other hand, F₁ hybrids produced by crossing slow-line males with fast-line females showed mating frequencies similar to those of the slow parental lines. These findings suggest that mating propensity inD. ananassae is under the control of polygenes. Furthermore, the significant differences in mating propensity of hybrids produced by the fast and slow males indicate the possibility of a Y-linked influence on mating propensity inD. ananassae.The present investigation was carried out during the tenure of a senior research fellowship of the CSIR, New delhi, to S.C.     

Transmission of lac by the sex factor E in Erwinia strains from human clinical sources

Lactose-utilizing (Lac(+)) strains of Erwinia spp. from human clinical material transfer lac by conjugation to plant strains of Erwinia herbicola and Erwinia amylovora, to other Erwinia strains from human clinical sources, and also to Escherichia coli, Paracolobactrum arizonae, Salmonella typhimurium, and Shigella dysenteriae. The frequency of this transfer varies with the donor and recipient strains employed. The lac genes appear stable in these exconjugants, and they are not cured by acridine orange. The Lac(+) exconjugants transfer lac to an Escherichia coli F(-) Lac(-) strain; the frequency of this transfer is high with E. herbicola and S. typhimurium exconjugants and relatively low with other exconjugants. The most studied Erwinia donor strain from human clinical material (EH133) and its Lac(+) exconjugants are insensitive to the F-specific phage, M13. P1-mediated transduction of lac, by using a Lac(+) exconjugant of E. coli as the donor and an E. coli F(-) Lac(-) strain as the recipient, revealed that all 50 Lac(+) transduced clones tested also inherited donor ability, suggesting a close linkage between the Erwinia sex factor (designated as E) and the lac genes. The E. coli culture harboring E-lac (E and the lac genes linked to it) does not restrict phages T1, T7, and lambdavir. E-lac is compatible with F'his, R100 drd-56 (F-like), and R64 drd-11 (I-like); cells harboring F'his or one of the R factors do not show super-infection immunity to the incoming E-lac, and E-lac plus one of the other plasmids can coexist stably in the same cell. The fertility of cells harboring F'his or R100 drd-56-as determined by the frequency of conjugal transfer of his or of the resistance determinant (Tet(r) in case of R100 drd-56) and also by sensitivity to F-specific phage (M13)-is not altered by the presence of E-lac, and this suggests that the sex factor E might belong to the fi(-) class.     

Role of a large plasmid of Salmonella typhi encoding multiple drug resistance

Twenty isolates of Salmonella typhi from cases of typhoid during the 1989-1990 epidemic in Calcutta were examined. Most isolates (84% of all isolates in the epidemic) were resistant to chloramphenicol, ampicillin, tetracycline and streptomycin but were sensitive to nalidixic acid and ciprofloxacin. Plasmids of 120 kb and 14 kb were identified amongst the multi-drug resistant isolates of S. typhi. However, there was no plasmid in the antibiotic-sensitive isolates. The 120-kb plasmid was transferable and transconjugants were resistant to chloramphenicol, ampicillin, tetracycline and streptomycin. Restriction endonuclease analysis patterns after EcoRI digestion of the 120-kb antibiotic-resistance plasmids from the S. typhi isolates and transconjugants were similar.     

RT-PCR-RFLP for genetic diversity analysis of Tunisian Grapevine fanleaf virus isolates in their natural host plants

Genetic diversity was characterized in 20 isolates of Grapevine fanleaf virus (GFLV) recovered from naturally infected grapevine plants (Vitis vinifera) in the North of Tunisia. Viral RNAs were isolated by oligoprobe capture, and a 605 bp fragment containing a part of the viral coat protein gene was amplified by RT-PCR. Sequence variation among isolates was characterized by restriction fragment length polymorphism (RFLP) analysis and confirmed by sequencing. The GFLV infections are found as a complex mixture of closely related genomes. In further studies, RFLP analyses of virus isolates using AluI showed that GFLV populations in Tunisian vineyards consist of two restrictotypes corresponding to distinct sub-populations Sp1 and Sp2. The relative field distribution of these sub-populations showed that Sp2 was more abundant. Individual genomes were recovered by cloning the RT-PCR products. The sequences were found to vary from each other by as much as 11%. Cloning from mixed infections showed that Sp2 are also predominant.     

Analysis of immune responses against T- and B-cell epitopes from Plasmodium falciparum liver-stage antigen 1 in rodent malaria models and malaria-exposed human subjects in India

Liver-stage antigen 1 (LSA-1) is a potential vaccine candidate against preerythrocytic stages of malaria. We report here the immunogenicity of linear synthetic constructs delineated as T(H)-cell determinants from the nonrepeat regions of Plasmodium falciparum LSA-1 in murine models and human subjects from areas where malaria is endemic in Rajasthan State, India. Seven peptide constructs (LS1.1 to LS1.7) corresponding to predicted T-cell sites from both the N- and C-terminal regions and peptide LS1R from a repeat region of PfLSA-1 were synthesized to analyze the cellular immune responses. These linear peptides were also tested for humoral responses in order to determine if there were any overlapping B-cell epitopes in the predicted T-cell sites. Most peptides induced cellular responses in peptide-immunized BALB/c and C57BL/6 mice as measured by proliferation and cytokine analysis. Cross-reactive T-cell recognition of P. falciparum-based peptides in Plasmodium berghei-immune animals was evaluated, but only one peptide, LS1.2 (amino acids 1742 to 1760) triggered T-cell proliferation and interleukin-2 and gamma interferon secretion in P. berghei-immune splenocytes of BALB/c and C57BL/6 mice as well as in Thamnomys gazellae (natural host of P. berghei ANKA). In an enzyme-linked immunosorbent assay with the peptides, only one peptide, LS1.1, was recognized by anti-P. berghei liver-stage serum. Three peptides (LS1. 1, LS1.2, and LS1.3) of the eight peptides tested in this study were recognized by a relatively large percentage of P. falciparum-exposed human subjects; the reactivities ranged from approximately 45% for LS1.3 to approximately 60% for LS1.1 and LS1.2. Interestingly, all of the eight putative T-cell determinants were also recognized by the sera collected from malaria patients, although the response was variable in nature. These T(H)- and B-cell epitopes may be of potential value for preerythrocytic antigen-based malaria subunit vaccine formulations.     

A conserved peptide sequence of the Plasmodium falciparum circumsporozoite protein and antipeptide antibodies inhibit Plasmodium berghei sporozoite invasion of Hep-G2 cells and protect immunized mice against P. berghei sporozoite challenge.

Minutes after injection into the circulation, malaria sporozoites enter hepatocytes. The speed and specificity of the invasion process suggest that it is receptor mediated. The region II sequence of Plasmodium falciparum circumsporozoite (CS) protein includes a nonapeptide (WSPCSVTCG) which is highly conserved in all of the CS proteins sequenced to data, including the one from Plasmodium berghei. We have found that two peptides based on the P. falciparum region II sequence, P18 (EWSPCSVTCGNGIQVRIK) and P32 (IEQYLKKIKNS ISTEWSPCSVTCGNGIQVRIK), significantly inhibited P. berghei sporozoite invasion into Hep-G2 cells in vitro. This inhibition was enhanced if either peptide was preincubated with Hep-G2 cells prior to sporozoite invasion. We confirm that region II is a sporozoite ligand for the hepatocyte receptor; moreover, despite the few differences between P. falciparum and P. berghei region II sequences around the nonapeptide sequence (66% homology), the functional characteristics of the motif sequences are not affected. Since the conserved motifs represent a crucial sequence involved in Plasmodium sporozoite invasion of hepatocytes, antibodies to region II should inhibit sporozite invasion into hepatocytes. Indeed, we found that polyclonal antibodies generated to the P. falciparum-based peptide P32 inhibited P. berghei sporozoite invasion of Hep-G2 cells. Furthermore, inbred mice (C57BL/6) immunized with P32 were protected against a lethal challenge of P. berghei sporozoites. Our results suggest that the conserved region II of the CS protein contains crucial B- and T-cell epitopes, that such peptide sequences from the human malaria parasite P. falciparum can be screened in the P. berghei rodent model, and, finally, that region II can be considered useful as one of the components of a malaria vaccine.     

Mechanisms of nitric oxide interplay with Rho GTPase family members in modulation of actin membrane dynamics in pericytes and fibroblasts

Migration of pericytes such as hepatic stellate cells is fundamentally important for diverse biological and pathological processes including tumor invasion and fibrosis. In prototypical migratory cells such as fibroblasts, the small GTPases Rac1 and RhoA govern the assembly of lamellipodia and stress fibers, respectively, cytoskeletal structures that are integral to the cell migration process. The gaseous signaling molecule nitric oxide (NO) influences growth factor chemotactic responses, although this occurs primarily in cell-type-specific ways and through cell biological effects that are poorly characterized. In this study, we use complementary molecular and cell biological approaches to delineate important roles for Rac1, RhoA, and NO in migration of the human hepatic stellate cell line LX2 and primary rat hepatic stellate cells. Both platelet-derived growth factor (PDGF) and Rac1 overexpression drove migration through formation of actin-positive filopodia spikes in LX2 as compared to the formation of lamellipodia in fibroblasts. NO inhibited PDGF- and Rac1-driven migration in LX2 by abrogating filopodia formation and inhibited migration of fibroblasts by attenuating lamellipodial protrusions. Additionally, RhoA conferred resistance to NO inhibition of migration and restored chemotactic responses to PDGF in the absence of functional Rac1 in LX2. In conclusion, these studies identify novel crosstalk between small GTPases, cytoskeletal structures, and NO in pericyte-specific pathways, providing counterbalances in the chemotactic responses to growth factors.