Trans-cellular introduction of HIV-1 protein Nef induces pathogenic response in Caenorhabditis elegans

Background

Caenorhabditis elegans has emerged as a very powerful model for studying the host pathogen interactions. Despite the absence of a naturally occurring viral infection for C. elegans, the model is now being exploited experimentally to study the basic aspects of virus-host interplay. The data generated from recent studies suggests that the virus that infects mammalian cells does infect, replicate and accumulate in C. elegans.

Methodology/Principal Findings

We took advantage of the easy-to-achieve protein introduction in C. elegans and employing the methodology, we administered HIV-1 protein Nef into live worms. Nef is known to be an important protein for exacerbating HIV-1 pathogenesis in host by enhancing viral replication. The deletion of nef from the viral genome has been reported to inhibit its replication in the host, thereby leading to delayed pathogenesis. Our studies, employing Nef introduction into C. elegans, led to creation of an in-vivo model that allowed us to study, whether or not, the protein induces effect in the whole organism. We observed a marked lipodystrophy, effect on neuromuscular function, impaired fertility and reduced longevity in the worms exposed to Nef. The observed effects resemble to those observed in Nef transgenic mice and most interestingly the effects also relate to some of the pathogenic aspects exhibited by human AIDS patients.

Conclusions/Significance

Our studies underline the importance of this in vivo model for studying the interactions of Nef with host proteins, which could further be used for identifying possible inhibitors of such interactions.     

Enhanced resistance against bacterial wilt in transgenic tomato (<Emphasis Type="Italic">Lycopersicon esculentum</Emphasis>) lines expressing the <Emphasis Type="Italic">Xa21</Emphasis> gene

To enhance bacterial wilt resistance in tomato plants and simplify the protocol of Agrobacterium tumefaciens mediated gene transfer, parameters affecting transformation efficiency in tomato have been optimized. A. tumefaciens strain EHA101, harboring a recombinant binary expression vector pTCL5 containing the Xa21 gene under the control of the CaMV 35S promoter was used for transformation. Five cultivars of tomato (Rio Grande, Roma, Pusa Ruby Pant Bahr and Avinash) were tested for transformation. Transformation efficiency was highly dependent on preculture of the explants with acetosyringone, acetosyringone in co-cultivation media, shoot regeneration medium and pre-selection after co-cultivation without selective agent. One week of pre-selection following selection along with 400 μM acetosyringone resulted in 92.3% transient GUS expression efficiency in Rio Grande followed by 90.3% in Avinash. The presence and integration of the Xa21 gene in putative transgenic plants was confirmed by polymerase chain reaction (PCR) and Southern blot analyses with 4.5–42.12% PCR-positive shoots were obtained for Xa21 and hygromycin genes, respectively. Transgenic plants of the all lines showed resistance to bacterial wilt. T₁ plants (resulting from self-pollination of transgenic plants) tested against Pseudomonas solanacearum inoculation in glasshouse, showed Mendelian segregation.     

Evaluation of toxic potential of captan: Induction of hsp70 and tissue damage in transgenic Drosophila melanogaster (hsp70-lacZ) Bg9

The study investigated the working hypothesis that a widely used fungicide captan exerts toxic effects on nontarget organisms. Transgenic Drosophila melanogaster (hsp70-lacZ) was used as a model by assaying stress gene expression as an endpoint for cytotoxicity and also to evaluate whether stress gene expression is sufficient enough to protect and to prevent tissue damage against toxic insult of the chemical. The study was further extended to understand the effect of the pesticide on development, life cycle, and reproduction of the organism and finally to evaluate a concentration of the chemical to be nontoxic to the organism. The study showed that (i) captan causes cytotoxicity at and above 0.015 ppm; (ii) at 0.0015 ppm captan, absence of hsp70 expression in the exposed organism was evaluated as the concentration referred to as no observed adverse effect level (NOAEL) for Drosophila; (iii) emergence pattern of flies was affected only at the highest concentration of captan by 4 days, while hatching and survivorship were unaffected even at this concentration; (iv) reproductive performance was significantly affected only at 125.0 and 1250.0 ppm captan, while in the lower dietary concentrations no such deleterious effects were observed; (v) at 1250.0 ppm, hsp70 failed to protect the cells from toxicant assault after 48 h exposure, thus leading to tissue damage as revealed by Trypan Blue staining. The present study shows the cytotoxic potential of captan and further reveals the application of stress genes in determining NOAEL and its expression as bioindicator of exposure to environmental contaminants.     

Bilirubin augments radiation injury and leads to increased infection and mortality in mice: Molecular mechanisms

Our earlier results demonstrated that clinically relevant concentrations of unconjugated bilirubin (UCB) possessed immunotoxic effects. Whole-body irradiation (WBI) with 1 to 6Gy leads to acute radiation syndrome, immunosuppression, and makes the host susceptible to infection. Since hyperbilirubinemia has been shown to be associated with several types of cancer, the present studies were undertaken to evaluate the radiomodifying effects of UCB in radiation-exposed mice having elevated levels of UCB. Pretreatment of splenic lymphocytes with UCB (1-50μM at UCB/BSA ratio <1) augmented radiation-induced DNA strand breaks, MMP loss, calcium release, and apoptosis. Combination treatment of mice with UCB (50mg/kg bw) followed by WBI (2Gy) 0.5h later, resulted in significantly increased splenic atrophy, bone marrow aplasia, decreased counts of peritoneal exudate cells, and different splenocyte subsets such as CD3+ T, CD4+ T, CD8+ T, CD19+ B, and CD14+ macrophages as compared to either UCB or WBI treatment. Hematological studies showed that WBI-induced lymphopenia, thrombocytopenia, and neutropenia were further aggravated in the combination treatment group. UCB pretreatment of mice potentiated WBI-induced apoptosis and decreased WBI-induced loss of functional response of various immune cells leading to augmentation of immunosuppression and infection susceptibility caused by WBI. In an acute bacterial peritonitis model, UCB pretreatment of mice significantly increased WBI-induced proinflammatory cytokines, nitric oxide, and peritoneal bacterial load resulting in increased infection and death. Studies using the pharmacological inhibitor of p38MAPK demonstrated the involvement of p38MAPK activation in the inflammatory cascade of peritonitis. These findings should prove useful in understanding the potential risk to hyperbilirubinemic patients during radiotherapy and victims of acute radiation exposure in the course of radiation accidents.     

Synthesis and characterization of transition metal fluoride complexes with imidazole: X-ray crystal structure reveals short hydrogen bonds between lattice water and lattice fluoride

A new series of complexes of cobalt(II) fluoride, nickel(II) fluoride, copper(II) fluoride and zinc(II) fluoride with imidazole were synthesized and characterized by elemental analysis, molar conductance, magnetic moments, IR and electronic absorption measurements. Based on elemental and spectral data, the complexes were found to be of [M(im)₆]F₂ · XH₂O type, where M is Co(II), Ni(II), Cu(II) and Zn(II) and X 4-5. The magnetic moments and spectral data suggested that all the complexes possessed an octahedral geometry. The crystal structure of the nickel complex, [Ni(im)₆]F₂ · 5H₂O, is also reported in which nickel atom is surrounded by six nitrogen atoms of imidazole. Strong intra- and inter-molecular hydrogen bonding exists between fluoride ions (uncoordinated), nitrogen of imidazole and the -OH of water molecules.     

Diurnal variations in a shallow tropical freshwater fish pond in Shahjahanpur,U.P. (India)

Summary Diurnal variations in water chemistry and plankton in this eutrophic tropical freshwater ponds were quite apparent. Large fluctuations were noted in dissolved oxygen, pH, carbonate and bicarbonate concentrations. The pond water gets oversaturated with oxygen during day (24.2 p.p.m.) and depleted in the night (1.2 p.p.m.). Diurnal migrations of plankton were not in conformity with temperate waters.     

Selection of buffalo bulls: Sexual behavior and its relationship to semen production and fertility

Forty-four buffalo bulls, used for artificial insemination, were studied to develop libido, mating ability and sexual behavior indices for selection purposes. For each index, 5 categories (i.e., excellent, very good, good, fair and poor) were established. The sexual behavior index was found to be more reliable than the libido and mating ability indices. Buffalo bulls in good to excellent categories were considered acceptable sires.

Reaction time, sexual aggressiveness, and scores of libido, mating ability and sexual behavior differed significantly among the various categories of the 3 indices. Libido significantly correlated with mating ability (r=0.89; P<0.001). Sexual behavior expressed significant relationship with age (r=0.41; P<0.01) and body weight (r=0.48; P<0.01), but was nonsignificant with the scrotal circumference (r=0.28; P>0.05) of buffalo bulls. However, these relationships were absent (P>0.05) in the acceptable sires. Semen production was correlated with sexual behavior in only the fair and poor categories of buffalo bulls (r=0.84; P<0.005). Sexual behavior had no relationship with the fertility rate of buffalo bulls (r=0.44; P>0.05). It is concluded that the sexual behavior index can be used successfully for the selection of buffalo bulls. Excellent- to good bulls should be used in an artificial breeding program if they qualify in the other selection indices.     

Mechanisms that promote bacterial fitness in fungal-affected soil microhabitats

Soil represents a very heterogeneous environment for its microbiota. Among the soil inhabitants, bacteria and fungi are important organisms as they are involved in key biogeochemical cycling processes. A main energy source driving the system is formed by plants through the provision of plant-fixed (reduced) carbon to the soil, whereas soil nitrogen and phosphorus may move from the soil back to the plant. The carbonaceous compounds released form the key energy and nutrient sources for the soil microbiota. In the grossly carbon-limited soil, the emergence of plant roots and the formation of their associated mycorrhizae thus create nutritional hot spots for soil-dwelling bacteria. As there is natural (fitness) selection on bacteria in the soil, those bacteria that are best able to benefit from the hot spots have probably been selected. The purpose of this review is to examine the interactions of bacteria with soil fungi in these hot spots and to highlight the key mechanisms involved in the selection of fungal-responsive bacteria. Salient bacterial mechanisms that are involved in these interactions have emerged from this examination. Thus, the efficient acquisition for specific released nutrients, the presence of type-III secretion systems and the capacity of flagellar movement and to form a biofilm are pinpointed as key aspects of bacterial life in the mycosphere. The possible involvement of functions present on plasmid-borne genes is also interrogated.