A rapid pigmentation test for identification ofCryptococcus neoformans

A rapid pigmentation test for identification ofCryptococcus neoformans is described. The method is based on the formation of a characteristic, mousegrey to violaceous-black pigment when shake cultures ofC. neoformans in a phosphate-buffered,l-DOPA — ferric citrate medium are incubated at 37 C for one hour.This work was carried out in part with financial support from the Council of Scientific and Industrial Research, New Delhi.     

Action of griseofulvin on geophilic dermatophytes and related keratinophilic fungi

Summary Growth response to griseofulvin has been studied in 24 strains of 16 species of geophilic dermatophytes and related keratinophilic fungi. Based on their sensitivity to griseofulvin these fungi fall into three groups: (1) Resistant group includesKeratinomyces ajelloi, Trichophyton terrestre andTrichophyton evolceanui which can grow upto 40µg per ml concentration of griseofulvin. (2) Fairly resistant group comprisingMicrosporon cookei, Microsporon vanbreuseghemii, a strain ofMicrosporon gypseum andTrichophyton sp., with growth occurring upto 8–16µg per ml concentration. (3) Sensitive group includes species ofTrichophyton, Keratinophyton, Microsporon, Nannizzia, Chrysosporium andCtenomyces which are completely inhibited at 4–8µg or lower griseofulvin concentrations. Griseofulvin inhibited formation of cleistothecia inK. terreum, delayed pleomorphic growth inT. indicum andN. incurvata, and influenced pigment production in several strains.     

Combined ipilimumab and nivolumab first‐line and after BRAF‐targeted therapy in advanced melanoma

The combination of ipilimumab and nivolumab is a highly active systemic therapy for metastatic melanoma but can cause significant toxicity. We explore the safety and efficacy of this treatment in routine clinical practice, particularly in the setting of serine/threonine‐protein kinase B‐Raf (BRAF)‐targeted therapy. Consecutive patients with unresectable stage IIIC/IV melanoma commenced on ipilimumab and nivolumab across 10 tertiary melanoma institutions in Australia were identified retrospectively. Data collected included demographics, response and survival outcomes. A total of 152 patients were included for analysis, 39% were treatment‐naïve and 22% failed first‐line BRAF/MEK inhibitors. Treatment‐related adverse events occurred in 67% of patients, grade 3–5 in 38%. The overall objective response rate was 41%, 57% in treatment‐naïve and 21% in BRAF/MEK failure patients. Median progression‐free survival was 4.0 months (95% CI, 3.0–6.0) in the whole cohort, 11.0 months (95% CI, 6.0‐NR) in treatment‐naïve and 2.0 months (95% CI, 1.4–4.6) in BRAF/MEK failure patients. The combination of ipilimumab and nivolumab can be used safely and effectively in a real‐world population. While first‐line efficacy appears comparable to trial populations, BRAF‐mutant patients failing prior BRAF/MEK inhibitors show less response.     

Identification and mapping of leaf, stem and stripe rust resistance quantitative trait loci and their interactions in durum wheat

Leaf rust (Puccinia triticina Eriks.), stripe rust (Puccinia striiformis f. tritici Eriks.) and stem rust (Puccinia graminis f. sp. tritici) cause major production losses in durum wheat (Triticum turgidum L. var. durum). The objective of this research was to identify and map leaf, stripe and stem rust resistance loci from the French cultivar Sachem and Canadian cultivar Strongfield. A doubled haploid population from Sachem/Strongfield and parents were phenotyped for seedling reaction to leaf rust races BBG/BN and BBG/BP and adult plant response was determined in three field rust nurseries near El Batan, Obregon and Toluca, Mexico. Stripe rust response was recorded in 2009 and 2011 nurseries near Toluca and near Njoro, Kenya in 2010. Response to stem rust was recorded in field nurseries near Njoro, Kenya, in 2010 and 2011. Sachem was resistant to leaf, stripe and stem rust. A major leaf rust quantitative trait locus (QTL) was identified on chromosome 7B at Xgwm146 in Sachem. In the same region on 7B, a stripe rust QTL was identified in Strongfield. Leaf and stripe rust QTL around DArT marker wPt3451 were identified on chromosome 1B. On chromosome 2B, a significant leaf rust QTL was detected conferred by Strongfield, and at the same QTL, a Yr gene derived from Sachem conferred resistance. Significant stem rust resistance QTL were detected on chromosome 4B. Consistent interactions among loci for resistance to each rust type across nurseries were detected, especially for leaf rust QTL on 7B. Sachem and Strongfield offer useful sources of rust resistance genes for durum rust breeding.     

Identifying potential entry inhibitors for emerging Nipah virus by molecular docking and chemical-protein interaction network

Abstract

Nipah Virus (NiV) is a newly emergent paramyxovirus that has caused various outbreaks in Asian countries. Despite its acute pathogenicity and lack of approved therapeutics for human use, there is an urgent need to determine inhibitors against NiV. Hence, this work includes prospection of potential entry inhibitors by implementing an integrative structure- and network-based drug discovery approach. FDA-approved drugs were screened against attachment glycoprotein (NiV-G, PDB: 2VSM), one of the prime targets to inhibit viral entry, using a molecular docking approach that was benchmarked both on CCDC/ASTEX and known NIV-G inhibitor set. The predicted small molecules were prioritized on the basis of topological analysis of the chemical-protein interaction network, which was inferred by integrating the drug-target network, NiV-human interaction network, and human protein-protein interaction network. A total of 17 drugs were predicted to be NiV-G inhibitors using molecular docking studies that were further prioritized to 3 novel leads???Nilotinib, Deslanoside and Acetyldigitoxin???on the basis of topological analysis of inferred chemical-protein interaction network. While Deslanoside and Acetyldigitoxin belong to an already known class of anti-NiV inhibitors, Nilotinib belongs to Benzenoids chemical class that has not been reported hitherto for developing anti-NiV inhibitors. These identified drugs are expected to be successful in further experimental evaluation and therefore could be used for anti-Nipah drug discovery. Apart, we also obtained various insights into the underlying chemical-protein interaction network, based on which several important network nodes were predicted. The applicability of our proposed approach was also demonstrated by prospecting for anti-NiV phytochemicals on an independent dataset.

Communicated by Ramaswamy H. Sarma     

Evaluation of the Gastrointestinal Tract as Potential Route of Primary Polyomavirus Infection in Mice

Background

Detection of Polyomavirus (PyV) DNA in metropolitan rivers worldwide has led to the suggestion that primary viral infection can occur by the oral route. The aim of this study was to test this notion experimentally.

Methods

Mouse PyV (MPyV) was used to infect C57BL/6J mice by the nasal or intragastric route. Viral load kinetics was studied 3, 7, 10, 14, 21 and 28 days post-infection (dpi) using quantitative PCR.

Results

Following nasal infection, MPyV DNA was readily detected in many organs including lung, heart, aorta, colon, and stool with viral loads in the range of 10³–10⁶ copies/mg wet weight that peaked 7–10 dpi. Complete viral clearance occurred in the serum and kidney by 28 dpi, while clearance in other organs was partial with a 10–100 fold decrease in viral load. In contrast, following intragastric infection peak detection of PyV was delayed to 21 dpi, and viral loads were up to 3 logs lower. There was no detectable virus in the heart, colon, or stool.

Conclusions

The intragastric route of MPyV infection is successful, not as efficacious as the respiratory route, and associated with delayed viral dissemination as well as a lower peak MPyV load in individual organs.     

Ultrastructural studies on nodules induced by pyrimidine auxotrophs of Sinorhizobium meliloti

Twenty three pyrimidine auxotrophs of Sinorhizobium meliloti Rmd201 were generated by random mutagenesis with transposon Tn5. On the basis of biochemical characters these auxotrophic mutants were classified into car, pyrC and pyrE/pyrF categories. All auxotrophs induced white nodules which were ineffective in nitrogen fixation. Light and electron microscopic studies revealed that the nodules induced by pyrC mutants were more developed than the nodules of car mutants. Similarly the nodules induced by pyrE/pyrF mutants had more advanced structural features than the nodules of pyrC mutants. The nodule development in case of pyrE/pyrF mutants was not to the extent observed in the parental strain. These results indicated that some of the intermediates and/or enzymes of pyrimidine biosynthetic pathway of S. meliloti play a key role in bacteroidal transformation and nodule development.     

Symbiotic characterization of isoleucine+valine and leucine auxotrophs of Sinorhizobium meliloti

Ten isoleucine+valine and three leucine auxotrophs of Sinorhizobium meliloti Rmd201 were obtained by random mutagenesis with transposon Tn5 followed by screening of Tn5 derivatives on minimal medium supplemented with modified Holliday pools. Based on intermediate feeding, intermediate accumulation and cross-feeding studies, isoleucine+valine and leucine auxotrophs were designated as ilvB/ilvG, ilvC and ilvD, and leuC/leuD and leuB mutants, respectively. Symbiotic properties of all ilvD mutants with alfalfa plants were similar to those of the parental strain. The ilvB/ilvG and ilvC mutants were Nod-. Inoculation of alfalfa plants with ilvB/ilvG mutant did not result in root hair curling and infection thread formation. The ilvC mutants were capable of curling root hairs but did not induce infection thread formation. All leucine auxotrophs were Nod+ Fix-. Supplementation of leucine to the plant nutrient medium did not restore symbiotic effectiveness to the auxotrophs. Histological studies revealed that the nodules induced by the leucine auxotrophs did not develop fully like those induced by the parental strain. The nodules induced by leuB mutants were structurally more advanced than the leuC/leuD mutant induced nodules. These results indicate that ilvB/ilvG, ilvC and one or two leu genes of S. meliloti may have a role in symbiosis. The position of ilv genes on the chromosomal map of S. meliloti was found to be near ade-15 marker.     

Isolation and symbiotic characterization of aromatic amino acid auxotrophs of Sinorhizobium meliloti

Ten aromatic amino acid auxotrophs of Sinorhizobium meliloti (previously called Rhizobium meliloti) Rmd201 were generated by random mutagenesis with transposon Tn5 and their symbiotic properties were studied. Normal symbiotic activity, as indicated by morphological features, was observed in the tryptophan synthase mutants and the lone tyrosine mutant. The trpE and aro mutants fixed trace amounts of nitrogen whereas the phe mutant was completely ineffective in nitrogen fixation. Histology of the nodules induced by trpE and aro mutants exhibited striking similarities. Each of these nodules contained an extended infection zone and a poorly developed nitrogen fixation zone. Transmission electron microscopic studies revealed that the bacteroids in the extended infection zone of these nodules did not show maturation tendency. A leaky mutant, which has a mutation in trpC, trpD, or trpF gene, was partially effective in nitrogen fixation. The histology of the nodules induced by this strain was like that of the nodules induced by the parental strain but the inoculated plants were stunted. These studies demonstrated the involvement of anthranilic acid and at least one more intermediate of tryptophan biosynthetic pathway in bacteroidal maturation and nitrogen fixation in S. meliloti. The alfalfa plant host seems to provide tryptophan and tyrosine but not phenylalanine to bacteroids in nodules.     

Commentary on de Cock Buning: a United kingdom perspective

de Cock Buning (1998) highlighted the existence of alternative and more favorable options available to xenotransplantation. Clearly, there is a need to emphasize a review of existing organ procurement programs worldwide. A tree interest in the welfare of animals encourages increased liaison between transplant communities throughout the world to discuss the experiences of various procurement programs.