Alteration of Genetic Make-up in Karnal Bunt Pathogen (Tilletia indica) of Wheat in Presence of Host Determinants.

Alteration of genetic make-up of the isolates and monosporidial strains of Tilletia indica causing Karnal bunt (KB) disease in wheat was analyzed using DNA markers and SDS-PAGE. The generation of new variation with different growth characteristics is not a generalized feature and is not only dependant on the original genetic make up of the base isolate/monosporidial strains but also on interaction with host. Host determinant(s) plays a significant role in the generation of variability and the effect is much pronounced in monosporidial strains with narrow genetic base as compared to broad genetic base. The most plausible explanation of genetic variation in presence of host determinant(s) are the recombination of genetic material from two different mycelial/sporidia through sexual mating as well as through para-sexual means. The morphological and development dependent variability further suggests that the variation in T. indica strains predominantly derived through the genetic rearrangements.

Cloning, in silico characterization and induction of TiKpp2 MAP kinase in Tilletia indica under the influence of host factor(s) from wheat spikes.

In order to understand the molecular mechanism(s) associated with floret specificity, morphogenetic and disease development of Karnal bunt (KB) pathogen in wheat spikes, host factor(s) was isolated from KB prone susceptible stage of wheat spikes. An orthologue of Kpp2 gene involved in pheromone response and fungal development was isolated from Tilletia indica for analyzing its role in fungal development. The maximum expression of TiKpp2 gene was observed at 14th day and decreased thereafter. To investigate whether the fungus alters the expression levels of same kinase upon interaction with plants, T. indica cultures were treated with 1% of host factor(s). Such treatment induced the expression of TiKpp2 gene in time dependent manner. Host factor(s) treatment tends to increase the myelination in fungal cultures by lowering the sporidial production. Increase in myelination led to impose more pathogenicity levels in the host and prolific multiplication of pathogen inside host causing more damage to developing grains. In silico characterization and protein-protein interaction studies further suggests that isolated gene showed similarity with Ustilago maydis Kpp2 and induction of TiKpp2 might further activate a downstream transcription factor Prf1. The results of present study clearly suggest that host factor(s) derived from wheat spikes provide certain signal(s) which activate TiKpp2 gene during morphogenetic development of T. indica and affect the fungal growth and pathogenicity. In turn it also provides a plausible explanation for floret specificity of KB fungus in wheat.

Induction of MAP kinase homologues during growth and morphogenetic development of Karnal bunt (Tilletia indica) under the influence of host factor(s) from wheat spikes.

Signaling pathways that activate different mitogen-activated protein kinases (MAPKs) in response to certain environmental conditions, play important role in mating type switching (Fus3) and pathogenicity (Pmk1) in many fungi. In order to determine the roles of such regulatory genes in Tilletia indica, the causal pathogen of Karnal bunt (KB) of wheat, semi-quantitative and quantitative RT-PCR was carried out to isolate and determine the expression of MAP kinase homologues during fungal growth and development under in vitro culture. Maximum expression of TiFus3 and TiPmk1 genes were observed at 14th and 21st days of culture and decreased thereafter. To investigate whether the fungus alters the expression levels of same kinases upon interaction with plants, cultures were treated with 1% of host factors (extracted from S-2 stage of wheat spikes). Such treatment induced the expression of MAPks in time dependent manner compared to the absence of host factors. These results suggest that host factor(s) provide certain signal(s) which activate TiFus3 and TiPmk1 during morphogenetic development of T. indica. The results also provides a clue about the role of host factors in enhancing the disease potential due to induction of MAP kinases involved in fungal development and pathogenecity.

Characteristics of Staphylococcus aureus colonization in patients with atopic dermatitis in Sri Lanka.

BACKGROUND: Colonization of the skin of patients with atopic dermatitis (AD) by Staphylococcus aureus (SA) is associated with more severe disease. AIM: To determine the association of SA colonization patterns and densities in lesional and nonlesional skin in patients with varying severities of AD, and to determine the antibiotic sensitivity patterns of SA isolates from Sri Lanka. METHODS: Skin and nasal swabs collected from 100 patients with AD and 120 controls were used to investigate the presence of SA. Severity of AD was graded using the Nottingham Eczema Severity Score. Colony counts were obtained for skin samples, and antibiotic sensitivity testing was performed in cases positive for SA. RESULTS: Skin colonization was seen in 57 patients (57%) but in only 10 controls (8%). Lesional skin of most patients (52/57; 91%) had SA densities of > 300 colony-forming units/cm(2) . Colonization rates with SA significantly increased with increasing age (Spearman correlation coefficient R = 0.9, P < 0.05) and increasing duration of lesions in patients with AD (Spearman R = 0.87, P < 0.05). Isolates from eight patients (13.5%) were found to be methicillin-resistant S. aureus (MRSA). Only 6 isolates (10%) were susceptible to penicillin and 22 (37%) to erythromycin, while 28 (47%) isolates had erythromycin-induced resistance to clindamycin. CONCLUSIONS: SA colonization rates were significantly associated with increasing age and severity of AD, and particularly with duration of lesions. Patients with severe disease were also more likely to be colonized with SA strains resistant to conventional antibiotics.

Sporadic encephalitis lethargica.

Three women (aged 21-36 years) developed acute illnesses that were similar to epidemic encephalitis lethargica. Each presented with a neuropsychiatric disturbance that was succeeded by pyrexia, a fluctuating conscious state and involuntary movements including oculogyria. Cerebrospinal fluid examination showed a predominantly lymphocytic pleocytosis (64-120x10(6) cells/L) and oligoclonal bands were detected in two cases. Two patients died, while the third made a gradual recovery. Post-mortem examination in the two fatal cases showed changes of lymphocytic meningitis and focal diencephalic lymphocytic infiltration, although these changes were mild relative to the effects of the clinical illness. The diagnosis of sporadic encephalitis lethargica relies on identifying shared clinical features with the past epidemic disease plus circumstantial evidence of immunological activity from laboratory investigations and some tests of exclusion of other disorders.

Sol-gel materials for the solid phase extraction of metals from aqueous solution.

Silica that was prepared by sol-gel chemistry to have pore widths in the microporous and in the mesoporous domains was evaluated as the host for performing solid phase extraction (SPE). Selective SPE of Ni(II) was accomplished with dimethylglyoxime (DMG)-doped silica, but the pore width was demonstrated to influence the chemistry of the material. With microporous silica as the host, the stoichiometry of the Ni(II)-DMG complex was 1:1 rather than 1:2, which is the value observed in aqueous solution. A green shift in the visible absorption spectrum was the primary evidence for the difference in stoichiometry; the alternative explanation of a rigidochromic effect on the spectrum was eliminated. The capacity of the DMG-doped mesoporous silica was only 9 mumol Ni g(-1) because of leaching of the complexing agent. The microporous material showed no loss of DMG, but low permeability lowered the capacity. An alternative, albeit not selective, approach was to employ a mesoporous host to which a complexing agent, diethylenetriamine (DTA), was covalently bound. In this case, a capacity of 0.156 mmol Cu g(-1), was achieved.

Donnan dialysis preconcentration coupled with ion chromatography and electrocatalytic oxidation for the determination of cyanide.

An electrocatalytic amperometric detector for the ion chromatographic determination of CN(-) is described. A conducting composite that is based on a graphite-loaded sol-gel material comprises the working electrode. The composite is doped with a Ru(II) metallodendrimer which is demonstrated to promote the electrochemical oxidation of CN(-) at potentials positive of 0.5 V vs. Ag/AgCl. In 6 mM NaOH, 0.05 M NaCl flowed at 1.0 ml min(-1), a 5-point calibration curve with the following linear least squares parameters is obtained over the range, 1.0-30 M CN(-): slope, 24.2+/-0.1 nA M(-1); intercept, -6+/-2 nA; and r, 0.9997. The detection limit, 0.7 muM CN(-), compares favorably to that obtained by amperometry at a silver electrode, 0.5 muM CN(-), under comparable experimental conditions. A 60-min preconcentration by Donnan dialysis increases the sensitivity by a factor of 23.6.

Locally acquired cutaneous leishmaniasis in Sri Lanka.

Cutaneous leishmaniasis in a Sri Lankan, who had not left the country, is reported. The diagnosis was confirmed by detecting the parasite in smears under microscopic examination. This is the first case report of locally acquired leishmaniasis in Sri Lanka.

Therapeutic failure with chloroquine in Plasmodium falciparum infections; possibility of high grade resistance.

Three case reports of falciparum malaria not responding to standard chloroquine therapy and necessitating the use of alternative antimalarial drugs for clinical improvement, are documented. In vitro drug sensitivity tests and chloroquine assays were not carried out to conclusively exclude drug failure. However, the clinical course and the persistence of parasitaemia on thick blood film monitoring were suggestive of high grade chloroquine resistance. Such patients require clinical vigilance and the timely use of alternative antimalarials. When rapid clearance of parasitaemia is required quinine is the drug of choice. In malarial chemotherapy it is mandatory that strict therapeutic criteria be followed, to safeguard the efficacy of chloroquine as well as to prevent abuse of alternative drugs. In this context, the availability of thick blood films and in vitro drug sensitivity testing facilities in hospitals would be useful to clinicians.