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Analytical methods that allow simultaneous determination of the concentration and enantiomeric composition of small sample amounts and are also compatible with high-throughput multi-well plate technology have received increasing attention in recent years. We now introduce a new class of broadly useful small-molecule probes and a relay sensing strategy that together accomplish these tasks with five classes of compounds including the challenging group of mono-alcohols-a scope that stands out among previously reported UV, fluorescence, and CD assays. Several chlorophosphite probes and aniline indicators have been evaluated and used for on-the-fly CD/UV sensing following a continuous workflow. The wide application range of the readily available sensors is highlighted with almost 30 alcohols, diols, hydroxy acids, amines and amino alcohols, and the accuracy of the stereochemical analysis is showcased with samples covering a wide range of concentrations and enantiomeric ratios., (© 2020 Wiley-VCH GmbH.)

The role of Discoidin Domain Receptors (DDRs) is poorly understood in neurodegeneration. DDRs are upregulated in Alzheimer's and Parkinson's disease (PD), and DDRs knockdown reduces neurotoxic protein levels. Here we show that potent and preferential DDR1 inhibitors reduce neurotoxic protein levels in vitro and in vivo. Partial or complete deletion or inhibition of DDR1 in a mouse model challenged with α-synuclein increases autophagy and reduces inflammation and neurotoxic proteins. Significant changes of cerebrospinal fluid microRNAs that control inflammation, neuronal injury, autophagy and vesicular transport genes are observed in PD with and without dementia and Lewy body dementia, but these changes are attenuated or reversed after treatment with the DDR1 inhibitor, nilotinib. Collectively, these data demonstrate that DDR1 regulates autophagy and reduces neurotoxic proteins and inflammation and is a therapeutic target in neurodegeneration., (© The Author(s) 2020. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

A generally useful palladium-catalyzed method for the asymmetric allylic amination with a large variety of isatins, sulfonamides, imides, amines, and N -heterocycles is introduced. A single protocol with a readily available catalyst accomplishes this reaction at room temperature with high yields and enantioselectivities often exceeding 90%, which is demonstrated with 31 examples.

Asymmetric reaction development within a day or two has been a dream of synthetic chemists for several decades. We now show that such a task is feasible with a highly efficient streamlined screening strategy using the asymmetric allylation of isatins with a chiral boron complex as a case study. Our high-throughput screening (HTS) method is based on fast optical UV/CD analysis of minute amounts of crude reaction mixtures (≈3 mg scale) and it obviates product isolation and the general need for reference compounds which greatly reduces preliminary work and analysis time. The setup, reaction screening, analysis and data processing for 54 asymmetric allylations of nine different isatins in six different solvents was handled by a single operator in less than 20 work hours. One could easily extend this HTS strategy to hundreds of reactions in roughly the same time frame and further reduce the labor with commercially available automated high-throughput experimentation equipment. The effectiveness of this asymmetric reaction development strategy is confirmed with the upscale synthesis of two representative 3-allyl-3-hydroxyisatins in 98-99 % yield and with 91-94 % ee under optimized conditions., (© 2019 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

An efficient organocatalytic method for the synthesis of difluoromethyl and trifluoromethyl substituted β-hydroxynitriles is introduced. The decarboxylative cyanomethylation of fluorinated ketones with readily available cyanoacetic acid gives a variety of tertiary alcohols in high yields and without concomitant water elimination. The reaction occurs in the presence of catalytic amounts of triethylamine, can be upscaled and applied to chlorofluoromethyl ketones and difluoromethyl ketimines.

Click reactions have become powerful synthetic tools with unique applications in the health and materials sciences. Despite the progress with optical sensors that exploit the principles of dynamic covalent chemistry, metal coordination or supramolecular assemblies, quantitative analysis of complex mixtures remains challenging. Herein, we report the use of a readily available coumarin conjugate acceptor for chiroptical click chirality sensing of the absolute configuration, concentration and enantiomeric excess of several compound classes. This method has several attractive features, including wide scope, fast substrate fixation without by-product formation or complicate equilibria often encountered in reversible substrate binding, excellent solvent compatibility, and tolerance of air and water. The ruggedness and practicality of this approach are demonstrated by comprehensive analysis of nonracemic monoamine samples and crude asymmetric imine hydrogenation mixtures without work-up. Click chemosensing addresses increasingly important time efficiency, cost, labor and chemical sustainability aspects and streamlines asymmetric reaction development at the mg scale.

Amphotericin B (AmB) is effective against visceral leishmaniasis (VL), but the renal toxicity of the conventional form, mixed micelles with deoxycholate (M-AmB), is often dose-limiting, while the less toxic lipid-based formulations such as AmBisome ® are very expensive. Two different strategies to improve the therapeutic index of AmB with inexpensive ingredients were evaluated on this work: (i) the heat treatment of the commercial formulation (H-AmB) and (ii) the preparation of an AmB-loaded microemulsion (ME-AmB). M-AmB was heated to 70 °C for 20 min. The resulting product was characterized by UV spectrophotometry and circular dichroism, showing super-aggregates formation. ME-AmB was prepared from phosphate buffer pH 7.4, Tween 80 ® , Lipoid S100 ® and Mygliol 812 ® with AmB at 5 mg/mL. The droplet size, measured by dynamic light scattering, was about 40 nm and transmission electron microscopy confirmed a spherical shape. Rheological analysis showed low viscosity and Newtonian behavior. All the formulations were active in vitro and in vivo against Leishmania donovani (LV9). A selectivity index (CC 50 on RAW/IC 50 on LV9) higher than 10 was observed for ME-AmB, H-AmB and AmBisome ® . Furthermore, no important in vivo toxicity was observed for all the samples. The in-vivo efficacy of the formulations after IV administration was evaluated in Balb/C mice infected with LV9 (three doses of 1 mg/kg AmB) and no significant difference was observed between H-AmB, M-AmB, ME-AmB and AmBisome ® . In conclusion, these two inexpensive alternative formulations for AmB showing good efficacy and selectivity for Leishmania donovani merit further investigation., (Copyright © 2018 Elsevier Inc. All rights reserved.)

A highly diastereoselective organocatalytic method that produces 3-fluoro-3'-hydroxy-3,3'-bisoxindoles and the corresponding 3-fluoro-3'-amino derivatives having two adjacent chirality centers from fluorooxindoles and isatins in high yields is described. The reaction occurs in protic solvents at room temperature, it can be upscaled without compromising yield and stereoselectivity, and chromatographic product purification is not required.

A highly diastereoselective organocatalytic reaction for the synthesis of fluorinated 3,3'-bisindolines exhibiting adjacent tetrasubstituted carbon stereocenters is described. A broad variety of heterochiral bisindolines was prepared in 91-99% yield using 3-fluorooxindoles and isatylidene malononitriles in the presence of catalytic amounts of triethylamine in water or aqueous solution. The reaction can be upscaled without compromising yield and diastereoselectivity and the general usefulness of this method was demonstrated with various Michael acceptors and extended to aldol and Mannich reactions.

Synthetically versatile 3,3-disubstituted fluorooxindoles exhibiting vicinal chirality centers were obtained in high yields and with excellent enantio-, diastereo-, and regioselectivity through catalytic asymmetric fluoroenolate alkylation with allylic acetates. The reaction proceeds under mild conditions and can be scaled up without compromising the asymmetric induction. The unique synthetic usefulness of the products is highlighted by the incorporation of additional functionalities and the formation of 3-fluorinated oxindoles exhibiting an array of four adjacent centers of chirality. A new C-F bond functionalization path that provides unprecedented possibilities for the stereoselective generation of a chiral quaternary carbon center in the alkaloid scaffold is introduced., (© 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Sequential chlorination/fluorination of aromatic trifluoroacetylated ketones gives 1-aryl 2-chloro-2,4,4,4-tetrafluoro-butan-1,3-dione hydrates that are used for the synthesis of ketones and alkenes exhibiting a terminal bromochlorofluoromethyl group. The hydrates undergo detrifluoroacetylative cleavage and subsequent bromination in the presence of a copper(II)bisoxazoline catalyst, K 2 CO 3 and NBS at room temperature. The corresponding bromochlorofluoromethyl ketones can be applied in Wittig and Horner-Wadsworth-Emmons reactions and dibromoalkenylations.

2-n-propylquinoline (2-n-PQ) had shown interesting in vivo antileishmanial activities after administration by oral route on leishmaniasis animal models. However, the lipophilic properties of this compound avoid its use by intravenous route, this route being indicated in cases of severe visceral leishmaniasis with vomiting. Thus, a 2-n-propylquinoline hydroxypropyl beta-cyclodextrin (2-n-PQ-HPC) formulation was set up in this aim. The formulation was active in vitro both on Leishmania donovani axenic and intramacrophage amastigotes with IC50 values at 6.22±0.82μM and 20.01±0.52μM, respectively, without any toxicity on macrophages. 2-n-PQ-HPC exhibited similar activity on WT and drug-resistant parasites. Its in vitro interactions with antimonials, amphotericin B and miltefosine were found as additive both in axenic amastigotes and intramacrophage amastigotes. 2-n-PQ-HPC was not able to generate drug resistance after in vitro drug pressure since the resistance index was less than 4. 2-n-PQ-HPC was also active on the L. donovani/Balb/c mice model with an intravenous treatment regimen at 10mgkg(-1)day(-1) on 10 consecutive days without hepatic, renal and blood toxicity. The pharmacokinetics of 2-n-PQ in rats showed that after an intravenous treatment of the formulation at 10mgkg(-1), the plasma drug concentrations rapidly declined bi-exponentially with a half-life of 58.7min and a total clearance of 18.63lh(-1)kg(-1). The apparent volume of distribution was higher than the blood volume in rats, indicating that 2-n-PQ was well distributed in tissues, allowing parasite elimination. Such a formulation is worth of further antiparasitic and toxicological evaluations., (Copyright © 2015 Elsevier Masson SAS. All rights reserved.)

Background: Postoperative infectious complications in patients undergoing pancreatic surgery are a significant cause for morbidity and mortality. Although synbiotics have beneficial effects on human health, their clinical value in surgical patients remains unclear given a paucity of applicable clinical studies., Aim: To determine the impact of perioperative synbiotic therapy on postoperative infectious complications, morbidity and mortality in patients undergoing pancreatic surgery for chronic pancreatitis., Methods: A trial was conducted in patients with chronic calcific pancreatitis undergoing Frey's procedure. Group A received a specific synbiotic composition, 5 days prior and 10 days after the surgery. Group B received a placebo. Primary study endpoint was the occurrence of postoperative infection during the first 30 days. Secondary outcome measures were mortality, length of hospital stay, days in intensive care unit, and duration of antibiotic therapy. Using previously accrued data, with α of 0.05 and power 80%, the sample size was calculated as 35 patients for each group with a dropout rate of 10%., Results: Of the 79 patients enrolled, 75 completed the trial [group A (n = 39) and group B (n = 36)]. The incidence of postoperative infectious complications (12.8% vs 39%; P < 0.05), duration of antibiotics therapy (P < 0.05), and length of hospital stay (P < 0.05) were significantly lower in the synbiotic group., Conclusions: Synbiotics significantly reduce septic complications, hospital stay, and antibiotic requirement in patients undergoing pancreatic surgery for chronic pancreatitis. Furthermore, basic and clinical research would clarify the underlying mechanisms of their therapeutic effect and define the appropriate conditions for use.

Introduction: Obstructive jaundice as a result of bile duct tumour thrombus (BDTT) is an unusual clinical entity and an uncommon presenting feature of hepatocellular carcinoma (HCC). This study evaluates the outcome of hepatectomy for HCC with obstructive jaundice as a result of BDTT in non-cirrhotic livers., Methods: Between 1997 and 2012, out of 426 patients with HCC in non-cirrhotic livers, 39 patients with BDTT (Group I n = 39), who underwent a hepatectomy, were analysed and compared with the non-BDTT group (Group II n = 387)., Results: The demographic profile and biochemical parameters between Group I and Group II were compared; apart from the presence of jaundice at presentation and an elevated serum bilirubin, there were no significant differences. Post-operative morbidity and mortality were 11 (28.2%) and 2 (5.1%), respectively, in Group I. There were no differences between the groups with regards to the operative variables and short-term outcomes. The 1-, 3- and 5-year survival rates in Group I were 82%, 48% and 10%, respectively, with a median survival of 28.6 months and were significantly poorer than Group II (90%, 55% and 38%, respectively, with a median survival of 39.2 months)., Conclusion: The mere presence of BDTT in HCC does not indicate an advanced or inoperable lesion. When technically feasible, a formal hepatic resection is the preferred first-line treatment option in these patients., (© 2015 International Hepato-Pancreato-Biliary Association.)

Two of the potential bacterial isolates, viz., Pseudomonas fluorescens (VCRC B-426) and Bacillus subtilis (VCRC B-471) whose toxins kill the mosquito pupae/larvae have been identified at our center. As the mode of action of these bacteria are not known, an attempt was made to find out the binding sites of the toxic proteins through immunological methods. Antibodies were raised in BALB/c mice and egg yolk system of chicken layers against the mosquitocidal proteins. The antibodies showed specific binding on to the cephalic and thoracic cuticle of the pupae as well as the paddles of the larvae, indicating the binding of the mosquitocidal proteins.

Aim: To analyze the differences in outcomes and the clinical impact following pancreatoduodenectomy (PD) in patients with and without aberrant right hepatic artery (aRHA)., Methods: All patients undergoing PD between January 2008 and December 2012 were divided into two groups, one with aRHA and the other without. These groups were compared to identify differences in the intraoperative variables, the oncological clearance and the postoperative morbidity, mortality and hospital stay., Results: A total of 225 patients underwent PD, of which 43 (19.1%) patients were found to have either accessory or replaced right hepatic arteries (aRHA group). The aRHA was preserved in 79% of the patients. There was no significant difference in the intraoperative blood loss but operative time was prolonged, reflecting the complexity of the procedure [420 ± 44 (240-540) min vs 480 ± 45 (300-600) min, P < 0.05)]. There were no differences in the incidence of postoperative complications (pancreatic leak, pancreatic fistula, delayed gastric emptying and mortality) and hospital stay. Oncological clearance in the form of positive resection margins [13 (7.1%) vs 3 (6.9%)] and lymph node yield were also similar in the two groups., Conclusion: An aRHA is found in approximately one fifth of patients undergoing PD. Preservation is technically possible in most patients and can increase the operative complexity but does not negatively affect the safety or oncological outcomes of the procedure.

Bacillus sphaericus is a bio-control agent effective against Culex quinquefasciatus, the vector of bancroftian filariasis. Apart from its larvicidal effect, there are reports of reduced infection of filarial parasites in mosquitoes exposed to it. In the present study, adults of Cx. quinquefasciatus emerged from B. sphaericus treated larvae were fed on blood samples positive for microfilariae of Wuchereria bancrofti and examined at various time intervals to assess the infection level. The rate of infection was reduced from 95% on day 1 post-feeding to 75% on day 13, when fed with blood sample containing 41 mf/20 μl. The mean parasite burden was also reduced from 4.9 per mosquito on day 1 to 2.15 on day 13. When fed with another sample (30 mf/20 μl), the infection was reduced from 100% on day 1 to 80% on day 13. Reduction in parasite burden was 4.0 to 1.75. Abnormally developed second-stage larvae of the parasite were seen in treated mosquitoes. Thus, the results indicated adverse effect of B. sphaericus treatment on infection and development of the filarial parasite in mosquitoes. The possible reason for the parasite regulation was studied through the assessment of the carryover of the bacterium as well as its toxins to the surviving mosquitoes. The presence of B. sphaericus was determined through plating of homogenate of survived mosquitoes on NYSM agar. Toxic protein was detected through immunoblotting. The bacterium as well as its 41.9-kDa toxic protein was found to be transmitted from larvae to adults and affected the parasite development, directly by the toxin or indirectly by eliciting humoral immune response of the mosquito.

This documentation explores the facts about the invasion of marine cyanobacteria in to the tsunami affected coastal villages of Nagapattinam district of Tamilnadu and Karaikkal district of Pondicherry Union Territory (UT) in southern India. Water samples were collected from eight tsunami-hit coastal villages in different open water sources. The collected samples were processed for detecting marine cyanobacterial growth. Totally 110 water samples were processed, three samples were positive for the toxic cyanobacteria, Lyngbya sp., and nine for nontoxic species such as Epithemia sp.,, Johannesbaptistia pellucida, Oscillatoria princeps, Phormidium fragile, Synechocystis sp. Besides posing a public health risk because of the toxic cyanobacteria, the bloom formation by the cyanobacterial species such as Anabaena, Microcystis, Lyngbya, Plectonema, Phormidium contaminated the water bodies and deteriorated the water quality in the tsunami affected villages. The study revealed that another kind of public health risk from the invasion of toxic cyanobacteria to the costal ecosystem during the tsunami. It is necessary, in this context, that the surveillance mechanism, which is geared up during or after natural disasters, should have a provision to monitor the transportation of toxic elements/organisms from marine system to coastal/inland ecosystems and to control such organisms.

Bacillus thuringiensis var. israelensis (B. t. i.) is being widely used in mosquito control programs. However, the large-scale production of this bacillus is expensive due to the high cost of the production medium. In this study, we attempted to develop a cost-effective medium, based on a locally available raw material namely coconut water which is available in plenty as waste product from coconut oil industry. The yield of cell mass, sporulation and mosquito larvicidal activity were studied by growing this bacterium in this waste product and in comparison with the conventional medium (NYSM). Cell mass yield of 3.1g/L, spore count of 3.4x10(11)spores/mL and mosquito larvicidal activity (LC(50)) of 14.85ng/mL (against early fourth-instar larvae of Aedes aegypti) were obtained with a 30h old culture of this bacterium grown in coconut water. This is almost similar to that obtained with NYSM medium. Hence, coconut water-based culture medium is economical for the production of B. t. i.

Background & Objective: Treatment of thromboembolic vascular disease has relied on anticoagulants. However, recognition that lysis of preformed fibrin could be accomplished in vivo by a process involving the conversion of inactive plasminogen to active plasmin enzyme led to an alternative enzyme-based approach. The drugs used for this therapy are called the fibrinolytic enzymes. In this study we attempted the production, purification and characterization of fibrinolytic enzyme from Bacillus sphaericus., Methods: The seed was prepared in nutrient yeast salt medium (NYSM) in shake flask and organism was produced in 100 l pilot fermentor. Biomass was separated by centrifugation and crude protein was prepared by ammonium sulphate precipitation. Purification was done by ion exchange chromatography using Q sepharose followed by gel filtration chromatography using Sephacryl S- 300. Molecular weight was determined through HPLC. Fibrinolytic activity was assayed by fibrin plate method., Results: The production method yielded 64 mg/l of the crude enzyme and after purification it was 6.3 mg/l. The molecular weight of the compound was 18.6 kDa., Interpretation & Conclusion: The enzyme exhibited similar fibrinolytic activity as that of streptokinase, on fibrin plates that were devoid of plasminogen, suggesting that its fibrinolytic action is independent of plasminogen and it is not a plasminogen activator.

Transferrin is a defence protein known to be up-regulated upon infection of parasites/pathogens in Aedes aegypti mosquito. However, no information is available on its up-regulation in Culex quinquefasciatus, the vector of bancroftian filarial parasite. In the present study, enhancement of transferrin in C. quinquefasciatus infected with Wuchereria bancrofti is demonstrated through amplification of the specific mosquito transcript, its sequencing, cloning, and expression. By using two oligonucleotide primers, a 950-bp polymerase chain reaction product was obtained from the first strand cDNA made from RNA of C. quinquefasciatus infected with W. bancrofti. A 707-bp sequence encoding the mature portion of transferrin was confirmed by sequencing the product. This is the first report of transferrin expression in C. quinquefasciatus. The deduced amino acid sequence shared 85% homology with A. aegypti transferrin precursor molecule. Western blot analysis of haemolymph proteins of infected C. quinquefasciatus with antibodies raised against recombinant transferrin protein showed binding to a 66-kDa protein, confirming its identity as transferrin. Hence, this molecule also could be added to the list of immune molecules of C. pipiens group, such as the defensin, gambicin, and cecropin, which are already known.

Methanolic extracts of 20 medicinal plants were screened at 1-10 mg/ml for in vitro macrofilaricidal activity by worm motility assay against adult Setaria digitata, the cattle filarial worm. Four plant extracts showed macrofilaricidal activity by worm motility at concentrations below 4 mg/ml and an incubation period of 100 min. Complete inhibition of worm motility and subsequent mortality was observed at 3, 2, 1 and 1 mg/ml, respectively, for Centratherum anthelminticum, Cedrus deodara, Sphaeranthus indicus and Ricinus communis. 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide (MTT) reduction assay was carried out at 1 mg ml(-1) and 4-h incubation period, and the results showed that C. deodara, R. communis, S. indicus and C. anthelminticum exhibited 86.56, 72.39, 61.20 and 43.15% inhibition respectively in formazan formation compared to the control.

The voltage-gated sodium channel (VGSC) is the target site for insecticides such as DDT and synthetic pyrethroids. A single base (A-T) change in the knock-down resistance (kdr) allele leads to an amino acid substitution at position 267 that confers the target-mediated resistance to DDT and synthetic pyrethroids in Anopheles gambiae. A theoretical model of the VGSC domain II that contains the site of mutation was constructed using the K;+ channel protein of Aeropyrum pernix as a template. The validated model with 88.6% residues in the favored region was subjected to the CASTp program that predicted 30 pockets in the modeled domain II for ligand interaction. In the model, at position 267, leucine was manually replaced with phenylalanine. When this altered model was subjected to the CASTp program, the search results showed the same number of pockets. The docking results indicate that DDT interacts with the modeled VGSC domain II at position 275 in the presence of leucine or in the presence of phenylalanine (binding energy =-5.32 kcal/mol, -6.21 kcal/mol). It appears from the results that the mutation at position 267 has no direct influence on the interaction of DDT with the target protein. Therefore, to understand the interaction affinity of DDT with the target and influence of the mutation on the existence of active sites/pockets in relation to ligand binding, a whole VGSC model is necessary.

The efficiency of laboratory colonies of mosquitoes such as Anopheles stephensi Liston, Aedes aegypti (L.) Liverpool strain, Ae. aegypti wild type, Aedes albopictus (Skuse), Culex tritaeniorhynchus Giles, Culex sitiens Wiedemann, and Armigeres subalbatus Coquillett in supporting the development of Wuchereria bancrofti (Cobbold) (Spirurida: Onchocercidae) microfilariae to infective larvae was investigated. The mosquitoes were fed on heparinized microfilaremic human blood by using a membrane-feeding unit with Parafilm as membrane. The rate of infection, parasite development, and parasite burden were compared with that in the known vector mosquito Culex quinquefasciatus Say. Cx. quinquefasciatus showed the highest percentage of infection, followed by Ae. aegypti Liverpool strain and An. stephensi. The rate of development of the parasite was more or less similar in all the three species, and infective larvae were found on day 13. When the larvae were harvested on day 17, Cx. quinquefasciatus yielded the highest numbers, followed by Ae. aegypti Liverpool strain and An. stephensi. The percentage of infection was low, and the development was slow in Cx. tritaeniorhynchus compared with the other susceptible species. The parasite developed to second-stage larvae only by day 22 and to infective larvae by day 28. When 2-wk-old Cx. tritaeniorhynchus were fed on microfilaremic blood, they could develop the parasite to infective larvae by day 13 postfeeding. All other species of mosquitoes tested were found to be refractory to parasite development. It is shown that Cx. quinquefasciatus is the most suitable mosquito host for the production of infective larvae. However, Ae. aegypti Liverpool strain, which is commonly used for Brugia malayi filarial parasite, also can be used for generation of W. bancrofti infective larvae to circumvent the problem of maintaining two mosquito species.

This paper reports a patient with a history of breast cancer, who presented with altered sensation to the right lower lip and chin. An orthopantomogram showed a probable dentigerous cyst associated with an unerupted lower wisdom tooth, which was closely related to the inferior dental canal. The tooth and cyst were enucleated under general anaesthesia. The subsequent histopathology report concluded that the cyst contained metastatic adenocarcinoma from a primary breast tumour.

Several coastal villages of southern India were affected by the 26 December 2004 tsunami, and 10,749 people were killed. Investigation carried out in the affected villages during fourth, fifth, and sixth weeks posttsunami showed that the fly density was in the range of 12-91.8 flies per sweep net. In total, 3,259 flies belonging to eight species, namely, Musca domestica L., Musca vicina Macquart, Musca sorbens Wiedemann, Calliphora erythrocephala Robineau-Desvody, Sarcophaga ruficornis F., Chrysomyia sp. Robineau-Desvody, Chlorops sp., and Fannia sp. Robineau-Desvody, were recorded. M. domestica was the predominant species constituting 78.2% of the total flies collected. Density of flies was the highest in temporary shelters constructed for the victims, followed by centralized kitchens and devastated human settlements. Lack of waste control at centralized kitchens nearer to the shelters might be the reason for the high fly density in relief shelters. Under these circumstances, outbreak of fly-borne diseases is likely to be aggravated. Therefore, it is suggested that the ongoing space spraying be supplemented with effective waste control measures to reduce the high density of flies.

Background & Objective: Cyclosporins are produced by certain species of the filamentous fungi, belonging to the genus Tolypocladium. While there are numerous reports on the use of cyclosporins in clinical studies, reports on the various aspects of their production have been very limited. Therefore, this study was carried to optimize the medium composition for the production of cyclosporin A, produced by a strain of the filamentous fungus, Tolypocladium species by static fermentation., Methods: The effect of different nutrients on the production of cyclosporin A, produced by Tolypocladium species in stationary culture was studied by growing the fungus for 21 days at 25 +/- 2 degrees C under different media composition. Cyclosporin A was extracted by homogenizing the fungal cells with methanol and the cyclosporin A level was analyzed by high performance liquid chromatography (HPLC)., Results: Among the six different media studied for the production of cyclosporin A, medium 'f' containing glucose (8%), casein acid hydrolysate (3%), malt extract (2%), peptone (1%) and DL- alpha-amino butyric acid (0.5%) favoured the maximum production (2.22 +/- 0.02 g/l medium or 5.85 +/- 0.35 g/kg biomass)., Interpretation & Conclusion: This study showed that by optimizing the composition of fermentation media enhanced production of cyclosporin A was obtained. Since the strain Tolypocladium (VCRC F21 NRRL No.18950) produces a high level of cyclosporin A in the identified fermentation medium, it could be exploited for industrial production.

Ever since the discovery of the first Bacillus thuringiensis strain capable of killing mosquito larvae, namely, B. thuringiensis var israelensis, there are several reports from different parts of the world about the occurrence of mosquitocidal strains belonging to different subspecies/serotypes numbering thirty-six. The main sources of these wild type strains are soils/sediments, plants, animal feces, sick/moribund insects and waters. The toxicity of the strains within a subspecies/serotype varied widely. Some of the strains exhibited toxicity to mosquitoes as well as lepidopterans and dipterans (including mosquitoes) as well as plant parasitic nematodes.

A three dimensional structural model of Glutathione-S-transferase (GST) of the lymphatic filarial parasite Wuchereria bancrofti (wb) was constructed by homology modeling. The three dimensional X-ray crystal structure of porcine pi-class GST with PDB ID: 2gsr-A chain protein with 42% sequential and functional homology was used as the template. The model of wbGST built by MODELLER6v2 was analyzed by the PROCHECK programs. Ramachandran plot analysis showed that 93.5% of the residues are in the core region followed by 5.4 and 1.1% residues in the allowed and generously allowed regions, respectively. None of the non-glycine residues is in disallowed regions. The PROSA II z-score and the energy graph for the final model further confirmed the quality of the modeled structure. The computationally modeled three-dimensional (3D) structure of wbGST has been submitted to the Protein Data Bank (PDB) (PDB ID: 1SFM and RCSB ID: RCSB021668). 1SFM was used for docking with GST inhibitors by Hex4.2 macromolecular docking using spherical polar Fourier correlations.

The mosquito Culex quinquefasciatus Say (Diptera: Culicidae) is the vector of the filarial parasite Wuchereria bancrofti (Cobbold) (Spirurida: Onchocercidae), which causes human bancroftian filariasis. Information on the mosquito humoral response against the filarial parasite during the process of its infection and development is important, as it decides the vector competence of the mosquito. Visible changes in the haemocyte population of mosquito, if any, will be an indicator of the possible humoral factors. The present study was aimed at investigating changes in the populations of various types of haemocytes of Cx. quinquefasciatus following infection with W. bancrofti. On day 2 post-feeding on microfilaraemic blood, the haemolymph perfusate of infected mosquitoes with L1 stage of the parasite showed 44.1% granulocytes, 42% prohaemocytes and 13.9% plasmatocytes, whereas that of the control mosquitoes fed on amicrofilaraemic blood showed 63.4% plasmatocytes, 22.2% prohaemocytes and 14.4% granulocytes. Differences in the population numbers of haemocyte types between the infected and control were significant (P > 0.05). However, the mosquitoes examined on day 6 post-feeding, when the parasite was in L2 stage, did not show any such changes. But, similar changes reappeared on day 12 in mosquitoes with L3 stage of the parasite. The observed haemocyte population changes indicate the possibility of some amount of humoral immune response, through the production of certain immune molecules, in Cx. quinquefasciatus infected with W. bancrofti. The nature and exact role of such a response on the filarial parasite development need further investigation.

Background & Objectives: Biological control through the use of parasitoids and pathogens is one of the alternatives to the use of chemical pesticides for control of insects of public health importance. At the Vector Control Research Centre, a liquid formulation developed using the metabolite of a Pseudomonas fluorescens strain was found to be lethal to larvae as well as pupae of vector mosquitoes. The lethal fraction of the metabolite is a protein with a molecular mass of 44 kDa and toxicity studies showed that it is safe to mammals. In the present study, this formulation was evaluated against immatures of the common house fly, Musca domestica, to find out whether it could be developed into a potential biocontrol tool., Methods: Early second instar larvae of house fly were introduced into rearing medium incorporated with the formulation at concentrations of 1, 5, 10, 15, 20 and 25 per cent, which were equivalent to respectively 1.13, 5.63, 11.25, 16.88, 22.50 and 28.13 microg of the toxic protein/ g of rearing medium. Mortality was monitored until the emergence of adult house fly. Net mortality of larvae and pupae were calculated and the LC50 and LC90 values were determined through probit regression analysis., Results: Larval mortality was obtained from day 3 to 6 post-treatment. Net mortality of larvae was higher at the concentration of 20 than at 25 per cent. However, it was higher at 25 per cent on day 5 and continued to day 6 when there was no larval mortality at other concentrations. The net mortality of pupae was higher than that of larvae at all the concentrations except at 20 per cent. The LC50 and LC90 values calculated from the net mortality of larvae and pupae together, from day 1 to 12 post-treatment, were respectively, 8.25 and 51.79 microg protein/g of the fly rearing medium., Interpretation & Conclusion: The formulation prepared from the exotoxin of P. fluorescens was toxic to the house fly. Pupae were more susceptible than larvae and the activity of the toxin might have been through cuticular absorption. The results are indicative of the possibility of development of the mosquitocidal metabolite for house fly control through appropriate field evaluations.

VCRC B426, 0.09% emulsifiable concentrate (EC) formulation developed from a metabolite of Pseudomonas fluorescens was tested for efficacy against Culex quinquefasciatus larvae and pupae. At application rates of 100, 200, 300 ml/m2, the formulation caused 100% elimination of larvae and pupae at day 1 after treatments and >80% reduction in pupal density for periods of 7, 12 and 11 days in cesspits and 5, 9 and 10 days in U-shaped drains. In both the habitats, the efficacy of the formulation against pupae was 1.7 times more at 200 ml/m2 than at 100 ml/m2. An increase in dosage to 300 ml/m2 did not improve the efficacy in cesspits but a marginal increase was observed in drains.

Secondary metabolites produced by Trichoderma viride, a deuteromycetes fungus, under submerged culture condition were formulated and evaluated for oviposition attractancy against gravid females of Culex quinquefasciatus mosquito. At a concentration of 10 g ml-1 the formulation showed remarkable attractancy with an oviposition active index (OAI) of +0.52. When the oviposition attractancy of the formulation was compared with a known oviposition attractant, p-cresol, both at 10 g ml-1, the former was found to be more attractive to result in 70% egg laying than the later with 30% egg laying. Thin layer chromatography fractions of the secondary metabolites showed that a fraction with Rf value of 0.88 was highly active as oviposition attractant with an OAI of +0.65. Further work on identification of the active principle(s) of the microbial formulation might lead to an oviposition attractant useful in mosquito vector management.

A formulation was developed from the metabolite(s) of a novel Pseudomonas fluorescens Migula strain (VCRC B426) and tested against 4th-instar larvae and pupae of three species of vector mosquitoes, Anopheles stephensi Liston, Culex quinquefasciatus Say and Aedes aegypti (L). The larvae and pupae of An. stephensi were the most susceptible to the formulation, followed by those of C. quinquefasciatus and Ae. aegypti, in that order, and the dosage requirement for pupal mortality was less than that required for larval mortality. The LC50 dosage requirements for larvae of these mosquito species were, respectively, 70.4, 511.5 and 757.3 microg protein ml(-1), whereas for pupae they were, respectively, 2.0, 9.4 and 19.2 microg protein ml(-1). The lethal fraction was purified from the culture broth and its molecular mass, as determined by high performance liquid chromatography, was 44kDa. This is the first report of a microbial formulation acting upon mosquito pupae, a non-feeding stage. Its mode of action and efficacy to control mosquitoes under field conditions need to be studied further.

Larvicidal efficacy and residual activity of a floating sustained release formulation of Bacillus thuringiensis ssp. israelensis (Bti) was tested in polluted water habitats against Culex quinquefasciatus, the vector of bancroftian filariasis, breeding in cesspools and cesspits. A total of 25 habitats having 65 m(2) water surface area with C. quinquefasciatus immatures were treated with the formulation at the rate of 15 kg/ha. For comparison, in another area, 18 habitats with a water surface area of 70 m(2) were kept untreated. Evaluation was continued up to 179 days with three rounds of Bti application. The mean number of egg rafts did not differ significantly between treated and untreated habitats during the trial period indicating same level of recruitment. However, abundance of larvae and pupae was significantly lower in Bti treated habitats compared to that in untreated ones. The activity of the formulation, with 2 weeks delay in onset compared to treatment, caused >80% reduction in pupal abundance for a period of 30-34 days. Considering this, it is suggested that monthly application of Bti formulation will considerably reduce the recruitment of pupae of C. quinquefasciatus.

Oviposition attractants could be used for monitoring as well as controlling mosquitoes by attracting them to lay eggs at chosen sites. In the present study, culture filtrates of seven bacterial species were tested for their attractancy against gravid females of Culex quinquefasciatus. When their oviposition active indices (OAI) were studied, the culture filtrates of Bacillus cereus and Pseudomonas fluorescens exhibited oviposition attractancy (OAI = > 0.3) at 100 ppm and the OAI were respectively 0.70 and 0.47. Culture filtrates of B. thuringiensis var. israelensis (wild type), B. t. var. israelensis (mutant) and B. sphaericus showed attractancy at 2000 ppm with OAI of respectively 0.71, 0.59 and 0.68. However, the OAI of B. megaterium as well as Azospirillum brasilense was 0.13 (at 2000 ppm), which was less than 0.3 required to be considered them as attractants. When the oviposition attractancy of the bacterial culture filtrates were compared with that of a known oviposition attractant, p-cresol (at 10 ppm), the culture filtrates of B. t. var. israelensis (wild type) and B. cereus were found to be more active than p-cresol, respectively with 64.2 and 54.3% oviposition.

Alginate encapsulated B. thuringiensis var. israelensis (B. t. i.) self floating type formulations were prepared. Its spore release rate, floating efficacy and larvicidal activity against Culex quinquefasiatus were tested in the laboratory. The larval mortality of 91-100% was induced by the floating formulation with a mean spore release of 3.04 x 10(4)/ml/day from 6th day to 27th day. From day 28 to 33 the mean number of spores released were 1.16 x 10(4)/ml/day which caused 72.2-88.2% mortality. From 34th day to 40th day the mean number of spores released were 4.97 x 10(3)/ml/day which caused 42.2-67.2% mortality. However, the self floating alginate encapsulated beads were intact and found to float upto 40 days.

Ten species of mosquitoes (Diptera: Culicidae) from five genera were exposed to preparasites of the tropical mermithid nematode species Romanomermis iyengari (Welch) (Nematoda: Mermithidae), a strain isolated in 1978 from Pondicherry. By exposing mosquito larvae during the second instar, nematode infection was invariably lethal, the rate being highest in Culex sitiens Wiedemann (95%) followed by Cx. quinquefasciatus Say (90%), Aedes aegypti (L.) (79%), Anopheles subpictus Grassi (64%), Ae. albopictus (Skuse) and Armigeres subalbatus Coquillett (62%), Cx. tritaeniorhynchus Giles (57%), Mansonia annulifera (Theobald) (46%), An. stephensi Liston (40%) and An. culicifacies Giles (36%). When fourth-instar larvae were exposed, the infection was highest in Ar. subalbatus (66%), followed by An. stephensi (52%), Cx. quinquefasciatus (47%), Ae. aegypti and An. subpictus (42%), Ae. albopictus (30%), An. culicifacies (29%), Cx. sitiens (24%), Cx. tritaeniorhynchus (19%) and Ma. annulifera (8%), with 2-45% of infected culicines surviving to adulthood. The parasitic phase of the nematode lasted 5-7 days in all the host species, yielding 1.1-3.2 parasites per II instar and 1.1-2.5 parasites per IV instar. The overall output of parasites per 100 mosquito larvae (infected + uninfected) was highest for Ae. aegypti when mosquitoes were exposed during II instar (2.53 parasites/larva) and for Ar. subalbatus when mosquitoes were exposed during IV instar (1.65/larva), and lowest for Ma. annulifera exposed during IV instar (0.09/larva). For routine laboratory culture of R. iyengari it is convenient to employ Cx. quinquefasciatus as the host yielding 90-190 parasites/100 larva.

Development of Wuchereria bancrofti in Culex quinquefasciatus emerged from the larvae that survived the exposure of sub-lethal dose of Bacillus sphaericus was examined in the laboratory. Third instar larvae of Cx. quinquefasciatus were treated with B. sphaericus at a sub-lethal dose of 11.35 microg/250 ml. The female mosquitoes that emerged from the survived larvae were fed on microfilaraemic human blood and parasite development was monitored in the fed mosquitoes. Both treated and untreated mosquitoes could ingest microfilaria (mF) equally as there was no significant difference in mF density between them. But, density of developmental stages of the parasite in treated group was significantly lower. Since, there was no mortality of mosquitoes, the lower density of the developmental stages could be attributed to the loss of parasites in the treated mosquitoes. Consequently, the proportion of mosquitoes with infective larvae (L3) and number of L3 were also significantly lower in treated females. Delay in parasite development was also noticed in treated mosquitoes. The present study indicates that B. sphaericus, when applied at sub-lethal dose kills larvae, and in addition, inhibits development of the filarial parasite and consequently reduces L3 yield in adult mosquitoes that emerged from the survived larvae.

Beauveria bassiana, an entomopathogenic fungus, was evaluated for its potential against second and third instar larvae of Culex quinquefasciatus, Anopheles stephensi and Aedes aegypti. Conidiospores of this fungus were effective in causing infection leading to mortality of different larval instars. Larvae of Cx. quinquefasciatus were more susceptible to infection than An. stephensi and the second instar larvae of these two species were more susceptible than third instar larvae. Larvae of Ae. aegypti were resistant to infection by B. bassiana.

Earlier attempts to produce different stages of W. bancrofti, such as fourth stage larvae (L4), in small animal models have yielded very low recovery rates. In order to enhance the recovery of L4, two routes of inoculating a small animal, M. unguiculatus, with infective larvae (L3) viz., intraperitoneal and intrathoracic routes, were compared. On day 17 post-inoculation, higher percentage (23-25%) of L4 were recovered from animals inoculated intrathoracically compared to that from animals inoculated intraperitoneally (2-8%). Also, comparatively higher proportion of worms (75-92%) remained within the intrathoracic region, unlike in the intraperitoneal region (50-80%). A few worms (1-4%) could be recovered even on 31 days post-inoculation from animals inoculated intrathoracically. When the L4 produced in animals were cultured in modified Frank's medium, all of them survived for 15 days and 50 per cent survived till the 25th day. The higher yield and ease of recovery from the thoracic cavity makes this route of inoculation a suitable method for production of L4. In vitro maintenance of L4 for prolonged period is significant with respect to excretory/secretory products or for drug screening.

To understand the physico-chemical factors that influence the efficacy of B. sphaericus formulation in the breeding sites of Culex quinquefasciatus, a study was carried out in Mayiladuturai area of Tamil Nadu (India). The factors studied were hydrogen ion concentration (pH), acidity, alkalinity, chlorides, phosphates, total hardness, sulphates, total solids, dissolved solids, suspended solids, nitrate nitrogen, ammoniacal nitrogen, dissolved oxygen, biological oxygen demand (BOD) and chemical oxygen demand (COD). Efficacy of the formulation was assessed in terms of reduction in larval population in the treated habitats. pH of water in the treated sites was around neutral range (mean +/- SD 7.65 +/- 0.23). Phosphate content was low (2.27 +/- 1.34 ppm) whereas chlorides (326.1 +/- 55.8 ppm) and sulphates (38.9 +/- 23.8 ppm) were high. Total hardness ranged from 206 to 462.5 ppm with a mean of 312.1 +/- 80.5 ppm. The chlorides and sulphates, though present in considerable quantity, did not have any influence on the efficacy of B. sphaericus formulation. However, the proportion of insoluble chlorides and sulphates which contribute to total hardness seemed to influence the formulation adversely.

In vivo treatment of intestinal brush border membrane vesicles with solubilized insecticidal crystal proteins (ICP) from the two strains of B. thuringiensis var. israelensis (VCRC B17 and VCRC MB24) caused no adverse effect on gamma glutamyl transpeptidase, Na+K+ATPase, sucrase and lactase enzymes. But, exposure of membrane vesicles to solubilized ICP's in vitro, lead to significant reduction in the activity of Na+K+ATPase, sucrase and lactase enzymes.

Seven types of formulations were prepared as granules using the larvicidal factor of Bacillus sphaericus and different concentrations of calcium alginate which was used as matrix to immobilize and entrap the active ingredient (ai). All formulations were tested in disused wells against Culex quinquefasciatus at the rate of 15 kg ai per hectare. Among the seven types tested, the type 2 which contained 5% calcium alginate as immobilizing agent, exhibited the maximum larvicidal activity. Persistent control in breeding was noticed for 8 weeks with this formulation type.

Effect of intact and alkali solubilized insecticidal crystal protein (ICP) preparations from a mutant strain of B. thuringiensis var. israelensis (VCRC MB24) and the wild type strain (VCRC B17) in vitro on human erythrocytes with respect to lipid peroxidation, osmofragility and membrane bound enzymes was determined. The alkali solubilized ICPs of both B. thuringiensis strains caused increased lipid peroxidation, decreased resistance to hypotonic lysis and reduction in the activity of membrane bound enzymes. On the contrary, the intact ICPs did not produce any such adverse effect on RBCs under the same experimental conditions. It is suggested that the ICPs are safe when they are intact when compared with solubilized ones.