Interactions of Colletotrichum musae and Lasiodiplodia theobromae and their biocontrol by Pantoea agglomerans and Flavobacterium sp. in expression of crown rot of “Embul” banana

Previous research has shown that two local isolates of bacteria (Pantoea agglomerans and Flavobacterium sp.) are capable of biocontrol of the two main pathogens (Colletotrichum musae and Lasiodiplodia theobromae) known to cause crown rot on “Embul” (Musa, AAB) banana. In this investigation an attempt was made to elucidate the comparative virulence of these pathogens and to determine the underlying biocontrol mechanisms. L. theobromae was more virulent, causing faster spread of the disease, whereas C. musae was more resistant to the bacterial antagonists. Viable cells of the antagonists were more effective at suppressing conidial germination than cell-free culture media. It seemed that antifungal compounds acting on conidial germination may be heat stable and those acting on mycelia may include heat labile compounds also. Considering the specialized roles observed for each pathogen in terms of causing the disease, and in modes of control by antagonists, future field investigations on biocontrol should consider the roles played by the pathogens and the antagonists.

Characterization of mutants of Caulobacter crescentus defective in surface attachment of the paracrystalline surface layer.

Strains of Caulobacter crescentus express a paracrystalline surface layer (S-layer) consisting of the protein RsaA. Mutants of C. crescentus NA1000 and CB2, isolated for their ability to grow in the absence of calcium ions, uniformly no longer had the S-layer attached to the cell surface. However, RsaA was still produced, and when colonies grown on calcium-sufficient medium were examined, large two-dimensional arrays of S-layer were found intermixed with the cells. Such arrays were not found in calcium-deficient medium even when high levels of magnesium ions were provided. The arrays could be disrupted with divalent ion chelators and more readily with the calcium-selective ethylene glycol-bis (beta-aminoethyl ether)N,N,N',N'-tetraacetic acid (EGTA). Thus, the outer membrane surface was not needed as a template for self-assembly, but calcium likely was. The cell surface and S-layer gene of assembly-defective mutants of NA1000 were examined to determine the basis of the S-layer surface attachment defect. Mutants had no detectable alteration in the rough lipopolysaccharide (LPS) or a characterized capsular polysaccharide, but another polysaccharide molecule was greatly reduced or absent in all calcium-independent mutants. The molecule was shown to be a smooth LPS with a core sugar and fatty acid complement identical to those of the rough LPS and an O polysaccharide of homogeneous length, tentatively considered to be composed of 4,6-dideoxy-4-amino hexose, 3,6-dideoxy-3-amino hexose, and glycerol in equal proportions. This molecule (termed SLPS) was detectable by surface labeling with a specific antiserum only when the S-layer was not present. The rsaA genes from three calcium-independent mutants were cloned and expressed in an S-layer-negative, SLPS-positive strain. A normal S-layer was produced, ruling out defects in rsaA in these cases. It is proposed that SLPS is required for S-layer surface attachment, possibly via calcium bridging. The data support the possibility that calcium binding is required to prevent an otherwise lethal effect of SLPS. If true, mutations that eliminate the O polysaccharide of SLPS eliminate the lethal effects of calcium-deprived SLPS, at the expense of S-layer attachment.     

Insecticide resistance spectra and resistance mechanisms in populations of Japanese encephalitis vector mosquitoes, Culex tritaeniorhynchus and Cx. gelidus, in Sri Lanka

Culex tritaeniorhynchus Giles and Cx. gelidus Theobald (Diptera: Culicidae), both vectors of Japanese encephalitis, were collected in 1984 and 1998 from two disease endemic localities in Sri Lanka: Anaradhapura and Kandy. Using wild-caught adult mosquitoes from light traps, log dosage-probit mortality curves for insecticide bioassays were obtained for three insecticides: malathion (organophosphate), propoxur (carbamate) and permethrin (pyrethroid). LD50 values showed that, in 1998, Cx. tritaeniorhynchus was -100-fold more resistant to malathion and 10-fold more resistant to propoxur than was Cx. gelidus. This difference was attributed to Cx. tritaeniorhynchus breeding mostly in irrigated rice paddy fields, where it would have been exposed to pesticide selection pressure, whereas Cx. gelidus breeds in other types of aquatic habitats less prone to pesticide applications. Resistance in Cx. tritaeniorhynchus increased between 1984 and 1998, whereas Cx. gelidus remained predominantly susceptible. Propoxur inhibition of acetylcholinesterase (AChE) activity (the target site of organophosphates and carbamates) indicated that in 1998, frequencies of insensitive AChE-based resistance were 9% in Cx. gelidus and 2-23% in Cx. tritaeniorhynchus, whereas in 1984 this resistance mechanism was detected only in 2% of the latter species from Anaradhapura. The AChE inhibition coefficient (ki) with propoxur was 1.86+/-0.24 x 10(5) M(-)1 min(-1) for Cx. tritaeniorhynchus from Anaradhapura in 1998. Both species were tested for activity levels of detoxifying glutathione S-transferases (GSTs) and malathion-specific as well as general carboxylesterases. High activities of GSTs and carboxylesterases were detected in Cx. tritaeniorhynchus but not Cx. gelidus. Malathion-specific carboxylesterase was absent from both species. Native polyacrylamide gel electrophoresis resolved two elevated general carboxylesterases, CtrEstbeta1 and CtrEstalpha1, from Cx. tritaeniorhynchus and none from Cx. gelidus. CtrEstbeta1 was the most intensely staining band. Gel inhibition experiments showed that both elevated esterases were inhibited by organophosphates and carbamates but not by pyrethroids. The major elevated esterase CtrEstbeta1 was partially purified (15-fold) by sequential Q-Sepharose and phenyl Sepharose column chromatography. The bimolecular rate constant (ka) and the deacylation rate constant (k3) for the malaoxon/enzyme interaction were 9.9+/-1.1 x 10(3) M(-1) min(-1) and 3.5+/-0.05 x 10(-4) M(-1) min(-1), respectively, demonstrating that the role of this enzyme in organophosphorus insecticide resistance is sequestration.     

Structural investigation of the capsular polysaccharide of Klebsiella serotype K80

The use of methylation, periodate oxidation, β-elimination, and selective hydrolysis enabled the structure of the K80 polysaccharide to be determined. The nature of the anomeric linkages was established by ¹H-n.m.r. spectroscopy, and confirmed by the results of oxidation of the fully acetylated polysaccharide with chromic acid. The K80 polysaccharide is comprised of repeating units of the pentasaccharide shown, and contains a pyruvic acetal on each repeating unit. This pattern constitutes the first instance, in this series of polysaccharides, of a pyruvic acetal attached to a side-chain rhamnosyl group.

     

Management of blood system in disasters

Managing the blood system in disasters is one of the main challenges for any blood transfusion service exposed to natural hazards such as earthquakes, floods and tsunamis, biological threats such as pandemic influenza as well as manmade disruptions and terrorism. Sri Lanka had to face contrasting types of situations. The most unexpected and dreadful one was the 2004 December 26 tsunami catastrophe which cost >30,000 human lives, leaving more than 23,000 injured. Health services were over-burdened with influx of dead bodies and injured people, most due to drowning. Injuries varied from small lacerations to deeper wounds, broken arms, legs, ribs, damaged organs and head injuries. The National Blood Transfusion Service, had to coordinate its blood supply effectively and to manage large number of blood donors during the first few days following tsunami.In contrast to the acute destruction of tsunami, Sri Lanka also faced a “chronic disaster” of 3 decades due to the separatist war waged by the Tamil Tigers, until it was completely terminated in 2009. There were large scale terrorist attacks using suicide bombers, land mines and claymore mines resulting in frequent influxes of dead bodies and injured patients to hospitals.The experiences of Sri Lanka blood system in the face of two contrasting types of disastrous situations are presented.     

Efficacy of Curcuma aeruginosa rhizome and Adhatoda vasica plant extracts, on red spider mite, Tetranychus urticae in Livistona rotundifolia

Queen palm, Livistona rotundifolia foliage contributes greatly in export industry. Red spider mite (RSM) (Tetranychus urticae) infests on the foliage and reduces its affordable market quality. T. urticae is found in dry environment and is one of the phytophagous mite belongs to family Tetranychidae. Different chemicals such as 80% sulphur + Diazinon @ (50g+12ml/10L) are recommended against red spider mite, but these have lesser effect on this tiny mite. Since these chemicals are not environment friendly, Green Farms Ltd., in Sri Lanka prefers to use biological agents for mite management. Extracts of Curcuma aeruginosa rhizome and Adhatoda vasica plant parts were studied separately causing mortality on T. urticae. Field experiments were conducted to study the efficacy of C. aruginosa extract for controlling RSM on L. rotundifolia leaves. Curcuma aruginosa was tested at concentrations of 2, 5, 10, 15, 20 and 25 g/L and a control with equal amount of water. C aruginosa extracts of different concentrations were treated six times at five days interval on the palms separately. Living spider mites and eggs were pre-counted in marked leaves before applying C. aruginosa extracts. Next count was taken a day prior to next spraying. The result revealed that all the concentrations except 2 g/L were found to be effective compared to control. However there was no difference between the concentrations from 5 to 25 g/L. Hence C. aruginosa rhizome extract at its lowest concentration of 5 g/L is equally effective for the control of RSM on L. rotundifolia leaves. In another experiment extracts of Adhatoda vasica bark, leaves, and flower and water as control were applied thrice with three days interval. Pre treatment counting of living spider mites and eggs were taken in marked leaves. Post count was taken a day prior to next spraying. Third and forth counting were done after three days and four weeks from final spraying respectively. The results revealed that bark, flowers were found to be more effective compared to control. Flowers and bark were the best and hence there is no need of third sprayings as almost all the spider mites population were eradicated after second spraying. Flower extraction showed best performance until three months since final spraying. Flower and bark extracts showed higher acaricidal property and leaf showed moderate acaricidal property.