Microconidia of Neurospora crassa

Neurospora crassa produces two types of vegetative spores-relatively small numbers of uninucleate microconidia and very large numbers of multinucleate macroconidia (blastoconidia and arthroconidia). The microconidia can function either as spermatia (male gametes) or as asexual reproductive structures or both. In nature they probably function exclusively in fertilization of protoperithecia. The environmental conditions favoring their formation and the pattern of their development are quite distinct from those of macroconidia. Mutants of N. crassa have been isolated in which macroconidiation is selectively blocked without affecting microconidiation, showing that these two types of conidial differentiation involve distinct developmental pathways. Unlike microconidia of some related ascomycetes, those of Neurospora are capable of germination, providing viable uninucleate haploid cells which are desired in several types of investigations. A technique of selectively removing macroconidia from culture initiated on cellophane overlying agar medium allows pure microconidia to be obtained even from the wild-type strains of Neurospora. The conditional microcyclic strain, mcm, allows either macroconidia or microconidia to be obtained at will, depending on the conditions of culture. The new methods of obtaining pure microconidia from normal laboratory strains will make it quick and easy to purify heterokaryotic transformants following introduction of DNA into multinucleate protoplasts. Moreover, these methods allow the detection of genetic variability that remains hidden within an individual fungus and the estimation of the frequency of nuclear types in laboratory-constructed heterokaryons. The discovery, function, and development of microconidia are described and their research applications are discussed in this review.     

Polymorphisms in vitamin D receptor,toll-like receptor 2 and Toll-Like receptor 4 genes links with Dengue susceptibility

Host genetic factors are known to determine disease susceptibility in dengue virus infection. Therefore, in this study association of gene polymorphisms of Vitamin D Receptor [rs731236 (Taq) and rs7975232 (Apa1)], Toll-like receptor 2 [rs5743708 (Arg735Gln) and rs5743704 (Pro631His)] and Toll-like receptor 4 [rs4986790A/G(Asp299Gly13843) and rs4986791 C/T(Thr399Ile)] were studied in cases with dengue as compared to controls. Total 98 cases of confirmed dengue virus infection and 98 age, sex and geographically matched healthy controls were enrolled and their genetic polymorphisms for the above mentioned regions were studied by Sanger sequencing. Mutant genotypes CC of VDR rs731236 (Taq1) [(OR 3.808, p value =0.02, CI 1.160-12.498)], GG of VDR rs7975232 (Apa1) [(OR 3.485, p value =0.02, CI 1.162-10.45)] and heterozygous genotypes of TLR4 rs4986790 A/G Asp299Gly [OR 2.40, p value= 0.02, CI 1.12-5.14], TLR4 rs4986791 C/T Thr399Ile [OR 2.09, p value=0.02, CI 1.12-5.14] were found to be significantly more in cases with dengue virus infection as compared to the controls. Also, at these positions mutant alleles were observed in significantly higher number of cases than controls. The values for C allele at VDR rs731236 (Taq1) were OR 1.86, p value 0.009, CI 1.162-3.001; for allele G at rs7975232( Apa1) were OR 2.71, p value 0.006, CI 1.196-2.98 for allele G at TLR4s rs4986790 A/G Asp299Gly were OR 2.35, p value 0.009, CI 1.23-4.50 and for allele T at rs4986791 C/T Thr399Ile were OR 2.36, p value=0.006, CI 1.28-4.38. VDR and TLR4 but not TLR2 gene polymorphisms were found to be associated with dengue susceptibility in Indian population.     

Molecular characterization of distinct YMV (Yellow mosaic virus) isolates affecting pulses in India with the aid of coat protein gene as a marker for identification

The present study was carried out to find out the variations present in different isolates of yellow mosaic virus (YMV) causing yellow mosaic disease of pulses in southern parts of India. The coat protein gene of YMV was amplified using gene specific and deng universal primers with DNA isolated from YMV infected samples. Further, cloning and DNA sequencing of CP gene was carried out. CP gene decrypt sequences revealed that YMV infected samples of Black gram, Cowpea and Green gram were similar to the MYMV-Tamil Nadu isolates. Whereas the YMV infected sample of Horse gram was found to be similar with HYMV. Hence, in the present study, two distinct YMV infecting pulses in Tamil Nadu (MYMV and HYMV species) were identified and it was observed that there exists considerable genetic variation among these species. In addition, Cowpea crop which was earlier supposed not to be susceptible for YMV infection also showed the presence of this virus similar to the MYMV. Overall, the findings of the present study indicate that the CP region is efficient enough to provide a simple, rapid, and reliable method for early detection of YMV infections in pulses, which would help to develop proper management strategies to control these viruses.     

Second Generation Inactivated Eastern Equine Encephalitis Virus Vaccine Candidates Protect Mice against a Lethal Aerosol Challenge

Currently, there are no FDA-licensed vaccines or therapeutics for eastern equine encephalitis virus (EEEV) for human use. We recently developed several methods to inactivate CVEV1219, a chimeric live-attenuated eastern equine encephalitis virus (EEEV). Dosage and schedule studies were conducted to evaluate the immunogenicity and protective efficacy of three potential second-generation inactivated EEEV (iEEEV) vaccine candidates in mice: formalin-inactivated CVEV1219 (fCVEV1219), INA-inactivated CVEV1219 (iCVEV1219) and gamma-irradiated CVEV1219 (gCVEV1219). Both fCVEV1219 and gCVEV1219 provided partial to complete protection against an aerosol challenge when administered by different routes and schedules at various doses, while iCVEV1219 was unable to provide substantial protection against an aerosol challenge by any route, dose, or schedule tested. When evaluating antibody responses, neutralizing antibody, not virus specific IgG or IgA, was the best correlate of protection. The results of these studies suggest that both fCVEV1219 and gCVEV1219 should be evaluated further and considered for advancement as potential second-generation inactivated vaccine candidates for EEEV.     

Changes in Phosphorylation Status of Wheat Plastid Polypeptides as Influenced by Light, Calcium and Cyclic AMP

The polypeptides of etioplast and chloroplast fractions, purified on Percoll discontinuous gradient, were phosphorylated in vitro using (γ-³²P)ATP, resolved by SDS-PAGE and autoradiographed. In general, about 15-18 phosphopolypeptides in the range of 14-150 kD were distinctly visible in autoradiograms of both organelle fractions with varying degree of radiolabel incorporation. Although short-term irradiation with red or far-red light did not have any significant effect on phosphorylation status of etioplast polypeptides, in vivo irradiation with 1 h white light, followed by in vitro phosphorylation, decreased phosphorylation of a 116 kD polypeptide and increased the phosphorylation of polypeptides of 38 kD and a doublet around 20 kD. Strikingly, the phosphorylation status of 116 kD etioplast polypeptide was adversely affected by Ca²⁺ as well, and this phosphopolypeptlde was not distinctly visible in the autoradiogram of the chloroplast fraction proteins. However, in vitro phosphorylation of 98, 57 and 50 kD polypeptides of both etioplast and chloroplast fractions was found to be Ca²⁺ dependent. Unlike Ca²⁺, 3′,5′-cyclic AMP down-regulated the phosphorylation of several polypeptides of both etioplasts and chloroplasts, including 98 and 50 kD, and up-regulated the phosphorylation of 32 and 57 kD polypeptides. The significance of these observations on changes in phosphoprotein profile of etioplasts and chloroplasts, as influenced by light, Ca²⁺ and cyclic nucleotides, has been discussed.     

Low infectivity of vesicular stomatitis virus (VSV) particles released from interferon-treated cells is related to glycoprotein deficiency

We have investigated the mechanism for the low infectivity of vesicular stomatitis virus (VSV) released from interferon (IFN) -treated cells. With 10-30 units/ml of IFN there was an approximately 5-30 fold reduction in the production of virus particles, as measured by VSV proteins; however, the infectivity of the VSV released from IFN-treated mouse LB, JLS-V9R, or human GM2504 was drastically reduced (2 to 4 logs). The low infectivity of VSV was directly related to a deficiency in virion glycoprotein (G). IFN treatment did not change the specific infectivity of the VSV particles released by HeLa cells; their G protein was also not reduced. A further effect of IFN to reduce the amount of virion M protein appeared to be secondary and was probably not related to the reduced infectivity of VSV.     

Establishing the role of detoxifying enzymes in field‐evolved resistance to various insecticides in the brown planthopper (Nilaparvata lugens) in South India

The brown planthopper (BPH), Nilaparvata lugens (Stål), is one of the major pests of rice throughout Asia. Extensive use of insecticides for suppressing N. lugens has resulted in the development of insecticide resistance leading to frequent control failures in the field. The aim of the present study was to evaluate resistance in the field populations of N. lugens from major rice growing states of South India to various insecticides. We also determined the activity of detoxifying enzymes (esterases [ESTs], glutathione S‐transferases [GSTs], and mixed‐function oxidases [MFOs]). Moderate levels of resistance were detected in the field populations to acephate, thiamethoxam and buprofezin (resistance factors 1.05–20.92 fold, 4.52–14.99 fold, and 1.00–18.09 fold, respectively) as compared with susceptible strain while there were low levels of resistance to imidacloprid (resistance factor 1.23–6.70 fold) and complete sensitivity to etofenoprox (resistance factor 1.05–1.66 fold). EST activities in the field populations were 1.06 to 3.09 times higher than the susceptible strain while for GST and MFO the ratios varied from 1.29 to 3.41 and 1.03 to 1.76, respectively. The EST activity was found to be correlated to acephate resistance (r = 0.999, P ≥ 0.001). The high selection pressure of organophosphate, neonicotinoid, and insect growth regulator (IGR) in the field is likely to be contributing for resistance in BPH to multiple insecticides, leading to control failures. The results obtained will be beneficial to IPM recommendations for the use of effective insecticides against BPH.     

Alterations in osteoclast morphology following osteoprotegerin administration in the magnesium-deficient mouse

In the present study, we used osteoprotegerin (OPG), which blocks osteoclastogenesis, to correct and thus explain the hypercalcemia that is seen during dietary Mg deficiency in the mouse. Control and Mg-deficient mice received injections for 12 days of either OPG or vehicle only. Serum Ca was similar in Mg-deficient mice treated with OPG and in control mice receiving OPG (9.2±0.3 mg/dl vs. 9.2±0.5). Both groups had significantly higher serum Ca than controls or Mg-deficient animals receiving vehicle alone. Surprisingly, Mg-depleted mice that received OPG in doses that inhibit osteoclastic bone resorption remained hypercalcemic. Because mature osteoclasts still present in the marrow might be hyperactive, we examined osteoclast morphology at the light microscopic and ultrastructural level. Light microscopic examination of trabecular bone showed few osteoclasts in OPG-treated mice. Ultrastructural examination revealed that osteoclasts in OPG-treated mice have decreased contact with the endosteal bone surface and absence of a ruffled border. Because the morphology of the existing pool of mature osteoclasts did not enhance resorption, another mechanism, such as increased intestinal absorption of Ca in Mg-deficient mice, likely contributes to the hypercalcemia observed during Mg deficiency.     

Structure of orthenthose

A new tetrasaccharide, orthenthose, has been isolated from the dried twigs of Orthenthera viminea (Family: Asclepiadaceae). By spectral and chemical procedures, this new tetrasaccharide has been shown to be O-α-l-oleandropyranosyl-(1→4)-O-α-l-oleandropyranosyl-(1→4)-O-α-l-oleandropyranosyl-(1→4)-β-l-oleandropyrano se.