Phosphine estimation in fumigated food grains using gas chromatography equipped with FPD detector.

Phosphine, as per several reports, is considered to be an ideal and effective alternative to methyl bromide (MB) fumigant, phased out globally honouring 'Montreal Protocol'. The food grains fumigated with phosphine are perceived to be residue-free; however, estimation of its residues is imperative. In the present study, the phosphine residues were estimated in grains of wheat, rice, green gram, chickpea and yellow pea fumigated with different formulations of phosphine under 7 and 10 days exposure periods at various locations in India. For efficacy studies, the conventional aluminium phosphide (AlP) tablet (56% @2 and 3 tablet/MT) and granular AlP (77.5% @1.0 and 1.5 g phosphine/m3) formulations were applied. Each treatment including untreated control was replicated thrice. The concentration was monitored every 24 h until the end of exposure periods. Food grain samples were drawn before and post fumigation following a certain waiting period for estimation of phosphine residues using gas chromatography equipped with FPD detector. Residues in fumigated food grains were observed in traces in all the experimental stacks, but below the MRL (0.1 ppm), fixed by Codex Alimentarius Commission (CAC). Phosphine can be considered safe for use as an alternate fumigant in quarantine and long-term storage of food grains leaving residues below MRL.

Biochemical Defense Responses in Red Rice Genotypes Possessing Differential Resistance to Brown Planthopper, Nilaparvata lugens (Stål).

The brown planthopper [Nilaparvata lugens (Stål.)] is one of the most destructive insect pests in all the rice-growing regions of the world. The pest is complicated to manage through the blanket application of chemical pesticides. The development of stable, durable N. lugens-resistant rice varieties is the most economical and efficient strategy to manage the pest. Landraces of red rice genotypes possess numerous nutritional and stress-resistant properties, though an exclusive study on the same is yet to be carried out. In the present study, we evaluated 28 red rice genotypes, along with two resistance checks and one susceptibility check, for their resistance to N. lugens. These promising lines revealed differential responses in the defense mechanism against the pest. The resistant accessions showed a greater accumulation of phenols, peroxidase, polyphenol oxidase, catalase, and superoxide dismutase under N. lugens-stressed conditions. However, the concentration of soluble proteins was substantially decreased in all the test genotypes. The concentration of crude silica was at maximum in highly resistant genotypes. Six red rice genotypes, namely Mata Meher, Manipuri Black, Hermonona, Sonahanan, Bavdi, and Bacharya Khuta fall under the highly resistant category, and can be utilized as valuable sources of resistance in breeding programs.

Indigenous biobed to limit point source pollution of imidacloprid in tropical countries.

Point pollution of pesticides originating from the washing of spraying machines could be controlled by biobed system and it is in use in temperate countries. The biobed system is yet to be established in tropical countries. An indigenous biobed system was prepared using local resources like rice straw, farm yard manures (FYM) and paddy field soil to suit the tropical climate. Lowermost 3 cm layer of the biobed system was filled with rice husk biochar to prevent leaching of pesticides from the system. This model system was tested with high doses of imidacloprid (178 mg/column), a commonly used pesticide against number of insect-pests in different crops, for its degradation. The bio-mix trapped a major part of the imidacloprid on the top most layer of the biobed column and only a very small part of imidacloprid recovered from the leachate. The biobed system could degrade 70.13% of applied imidacloprid within 15 days of the experiment and only 5.27% of the total pesticide recovered 90 days after incubation. Addition of biochar layer adsorbed imidacloprid from the outgoing leachate from the biobed column. Biomixture boosted microbial activity more particularly fungal population, which might be responsible for imidacloprid degradation. Microbial biomass carbon, and soil enzymes indicated faster dissipation of imidacloprid from the top layer of the biobed. This simple but efficient biobed system using local resources can fulfill the need of the small and marginal farmers of Asian countries for pesticide decontamination.

Deciphering the associated risk on soil microbes upon use of biopesticides in rice ecosystem.

Plant species, viz Cleistanthus collinus, Lantana camara, and Strychnos nux-vomica are being traditionally used for pest management in rice. However, limited investigation has been carried out to understand the toxic effect of these materials on soil microbes. Hot water extracts of these plants were evaluated for their effects on soil microbial population and enzyme activities along with neem oil and chlorpyrifos as check. Soil microbial population, viz bacteria, fungi, phosphate-solubilizing bacteria (PSB), and asymbiotic nitrogen fixers were unchanged after application of plant extracts. Maximum population of bacteria including PSB and asymbiotic nitrogen fixers were observed in control, whereas, S. nux-vomica, and C. collinus-treated soil had higher number of actinomycetes and fungal population, respectively. Soil microbial biomass did not vary differently among the plant extracts. Application of plant extracts did not alter dehydrogenase, ß-glycosidase, acid phosphatase, alkaline phosphatase, and urease content in soil. Secondary metabolites present in these plant extracts may be responsible for variable effects on soil microbes. Chlorpyrifos had a fleeting negative effect on soil microbes and enzymes in comparison to plant extracts. All the three plants did not have any negative effect on soil microbes and enzymes and can be safely recommended in rice pest management.

Candidate screening of blast resistance donors for rice breeding.

Rice blast is one of the most serious diseases in the world. The use of resistant cultivars is the most preferred means to control this disease. Resistance often breaks down due to emergence of new races; hence identification of novel resistance donors is indispensable. In this study, a panel of 80 released varieties from National Rice Research Institute, Cuttack was genotyped with 36 molecular markers that were linked to 36 different blast resistance genes, to investigate the varietal genetic diversity and molecular marker-trait association with blast resistance. The polymorphism information content of 36 loci varied from 0.11 to 0.37 with an average of 0.34. The cluster analysis and population structure categorized the 80 National Rice Research Institute released varieties (NRVs) into three major genetic groups. The principal co-ordinate analysis displays the distribution of resistant and moderately resistant NRVs into different groups. Analysis of molecular variance result demonstrated maximum (97%) diversity within populations and minimum (3%) diversity between populations. Among tested markers, two markers (RM7364 and pi21_79-3) corresponding to the blast resistance genes (Pi56(t) and pi21) were significantly associated and explained a phenotypic variance of 4.9 to 5.1% with the blast resistance. These associated genes could be introgressed through marker-assisted to develop durable blast resistant rice varieties. The selected resistant NRVs could be good donors for the blast resistance in rice crop improvement research.

Dissipation of chlorantraniliprole in contrasting soils and its effect on soil microbes and enzymes.

An experiment was set up to determine the rate of dissipation of chlorantraniliprole (CTP) from two soils with contrasting properties. The other objective of the study was to find out the effect of CTP on soil microorganisms (population, microbial biomass carbon and soil enzymes) under controlled environment. CTP residues when applied at recommended dose ((RD) (at 40 g a.i./ha)) could not be recovered either from alluvial soil or red soil at 60 days post application of CTP in a microcosm study. Higher clay content led to higher half-life in alluvial soil compared to red soil. CTP could not be recovered from RD treatment at 30 days after pesticide application under controlled environment. Faster dissipation of CTP was observed in rice rhizosphere soil with 23.89 and 34.65 days dissipation half-lives for RD and double the recommended dose (DRD) treatments, respectively. Different doses of chlorantraniliprole did not have considerable negative effect on actinomycetes, fungi, biological nitrogen fixers and phospahte solubilising bacteria except the bacteria population. Among the treatments, DRD recorded the lowest activity of dehyrodeganse, fluoresein diacetate hydrolase, acid and alkaline phosphatases followed by RD treatment. Microbial biomass carbon, ß -glycosidase and urease did not vary significantly among the different doses of CTP. In general, RD did not have negative effcts on soil microbes. Hence, CTP can be recommeded in rice pest managment maintaining existing soil microbes and soil enzymes activity.

Insights into the structure-function relationship of brown plant hopper resistance protein, Bph14 of rice plant: a computational structural biology approach.

Brown plant hopper (BPH) is one of the major destructive insect pests of rice, causing severe yield loss. Thirty-two BPH resistance genes have been identified in cultivated and wild species of rice Although, molecular mechanism of rice plant resistance against BPH studied through map-based cloning, due to non-existence of NMR/crystal structures of Bph14 protein, recognition of leucine-rich repeat (LRR) domain and its interaction with different ligands are poorly understood. Thus, in the present study, in silico approach was adopted to predict three-dimensional structure of LRR domain of Bph14 using comparative modelling approach followed by interaction study with jasmonic and salicylic acids. LRR domain along with LRR-jasmonic and salicylic acid complexes were subjected to dynamic simulation using GROMACS, individually, for energy minimisation and refinement of the structure. Final binding energy of jasmonic and salicylic acid with LRR domain was calculated using MM/PBSA. Free-energy landscape analysis revealed that overall stability of LRR domain of Bph14 is not much affected after forming complex with jasmonic and salicylic acid. MM/PBSA analysis revealed that binding affinities of LRR domain towards salicylic acid is higher as compared to jasmonic acid. Interaction study of LRR domain with salicylic acid and jasmonic acid reveals that THR987 of LRR form hydrogen bond with both complexes. Thus, THR987 plays active role in the Bph14 and phytochemical interaction for inducing resistance in rice plant against BPH. In future, Bph14 gene and phytochemicals could be used in BPH management and development of novel resistant varieties for increasing rice yield.

Toxicological effect of underutilized plant, Cleistanthus collinus leaf extracts against two major stored grain pests, the rice weevil, Sitophilus oryzae and red flour beetle, Tribolium castaneum.

Toxicity and repellency activity of Cleistanthus collinus (Roxb.) (CC) leaf extracts were evaluated against rice weevil, Sitophilus oryzae (L.) and red flour beetle, Tribolium castaneum (Herbst) under laboratory condition. Five concentration(s) (1%, 1.5%, 2%, 2.5% and 4%) with two controls (acetone and water) treatments along with deltamethrin were used for direct and contact residual toxicity. The insect that survived after CC exposure were transferred to an untreated feeding substrate and the population buildup of subsequent two generation were recorded after 30 (F1) and 60 days (F2). In the contact residual toxicity, highest CC concentration (4%) produced 75% mortality in S. oryzae and 62.5% mortality in T. castaneum during 7 days of exposure, whereas in direct toxicity the mortality were 81% and 58% respectively, for S. oryzae and T. castaneum. The long term effect of CC was apparent in both the insect species, where F2 populations were significantly decreased in the CC treatments. CC treatment at 4% produced similar adult mortality in comparison to deltamethrin at 1%. In addition, repellent activity of CC extracts was observed against both S. oryzae and T. castaneum. This is the first step towards assessing the scientific basis for the understanding the effectiveness of CC extracts against stored grain pests and it could be a viable eco-friendly option for stored grain insect pest management.

Effect of Abiotic Factors on Degradation of Imidacloprid.

The role of soil moisture, light and pH on imidacloprid dissipation was investigated. A high performance liquid chromatography (HPLC) based method was developed to quantify imidacloprid present in soil with a recovery of more than 82%. Rate of dissipation of imidacloprid from soil was faster in submerged condition compared to field capacity and air dried condition. Imidacloprid dissipated non-significantly between sterile and non-sterile soils, but at field capacity, the dissipation was faster in non-sterile soil compared to sterile soil after 60 days of incubation. Similarly, under submergence, the dissipation of imidacloprid was 66.2% and 79.8% of the initial in sterile and non-sterile soils, respectively. Imidacloprid was rather stable in acidic and neutral water but was prone to photo-degradation. Therefore, imidacloprid degradation will be faster under direct sunlight and at higher soil moisture.

Imidacloprid application changes microbial dynamics and enzymes in rice soil.

Extensive use of imidacloprid in rice ecosystem may alter dynamics of microorganisms and can change soil biochemical properties. The objective of this study was to assess the effect of imidacloprid on growth and activities of microbes in tropical rice soil ecosystem. Four treatments, namely, recommended dose (at 25g a.i. ha-1, RD), double the recommended dose (at 50g a.i. ha-1, 2RD), five times the recommended dose (at 125g a.i. ha-1, 5RD) & ten times the recommended dose (at 250g a.i. ha-1, 10RD) along with control were imposed under controlled condition. Dissipation half lives of imidacloprid in soil were 19.25, 20.38, 21.65 and 33.00 days for RD, 2RD, 5RD and 10RD, respectively. In general bacteria, actinomycetes, fungi and phosphate solubilising bacteria population were disturbed due to imidacloprid application. Changes in diversity indices within bacterial community confirmed that imidacloprid application significantly affected distribution of bacteria. Total soil microbial biomass carbon content was reduced on imidacloprid application. Except dehydrogenase and alkaline phosphatase activities, all other soil enzymes namely, ß-glycosidase, fluorescien diacetate hydrolase, acid phosphatase and urease responded negatively to imidacloprid application. The extent of negative effect of imidacloprid depends on dose and exposure time. This study concludes imidacloprid application had transient negative effects on soil microbes.

Correction: Use of molecular markers in identification and characterization of resistance to rice blast in India.

[This corrects the article DOI: 10.1371/journal.pone.0176236.].

Use of molecular markers in identification and characterization of resistance to rice blast in India.

Rice blast disease caused by Magnaporthe oryzae is one of the most destructive disease causing huge losses to rice yield in different parts of the world. Therefore, an attempt has been made to find out the resistance by screening and studying the genetic diversity of eighty released rice varieties by National Rice Research Institute, Cuttack (NRVs) using molecular markers linked to twelve major blast resistance (R) genes viz Pib, Piz, Piz-t, Pik, Pik-p, Pikm Pik-h, Pita/Pita-2, Pi2, Pi9, Pi1 and Pi5. Out of which, nineteen varieties (23.75%) showed resistance, twenty one were moderately resistant (26.25%) while remaining forty varieties (50%) showed susceptible in uniform blast nursery. Rice varieties possessing blast resistance genes varied from four to twelve and the frequencies of the resistance genes ranged from 0 to 100%. The cluster analysis grouped the eighty NRVs into two major clusters at 63% level of genetic similarity coefficient. The PIC value for seventeen markers varied from 0 to 0.37 at an average of 0.20. Out of seventeen markers, only five markers, 195R-1, Pi9-i, Pita3, YL155/YL87 and 40N23r corresponded to three broad spectrum R genes viz. Pi9, Pita/Pita2 and Pi5 were found to be significantly associated with the blast disease with explaining phenotypic variance from 3.5% to 7.7%. The population structure analysis and PCoA divided the entire 80 NRVs into two sub-groups. The outcome of this study would help to formulate strategies for improving rice blast resistance through genetic studies, plant-pathogen interaction, identification of novel R genes, development of new resistant varieties through marker-assisted breeding for improving rice blast resistance in India and worldwide.

Effect of Pretilachlor on Soil Enzyme Activities in Tropical Rice Soil.

Pretilachlor treatments, namely, recommended dose at 600 g a.i. ha-1 (RD), double the recommended dose at 1200 g a.i. ha-1 (2RD), ten times of the recommended dose at 6000 g a.i. ha-1 (10RD) along with control, were used to study the effects of pretilachlor on soil enzymes in tropical rice soil. Pretilachlor, at recommended dose completely dissipated 30 days after herbicide application. Twenty days after herbicide application, the dehydrogenase activity was inhibited up to 27 %, 28 % and 40 % of initial values of RD, 2RD and 10RD treatments, respectively. Increase in fluorescein diacetate hydrolase activity was observed during the first 25 days post herbicide application up to 29 %, 36 % and 10 % of initial values of RD, 2RD and 10RD treatments, respectively. ß-Glucosidase activity in the experiment did not provide a specific trend. In general, urease and acid phosphatase activities were not influenced by pretilachlor application. There were significant differences in alkaline phosphatase activities among the treatments until 25 days after herbicide application. Hence, pretilachlor may cause short term transitory changes in soil enzyme parameters. However, it has negative impact on soil enzymes at very high dose.

Non-target effects of pretilachlor on microbial properties in tropical rice soil.

The use of herbicides has been questioned in recent past for their non-target effects. Therefore, we planned to study the effect of pretilachlor on growth and activities of microbes in tropical rice soil under controlled condition at National Rice Research Institute, Cuttack, India. Three pretilachlor treatments, namely, recommended dose at 600 g a.i. ha(-1) (RD), double the recommended dose at 1200 g a.i. ha(-1) (2RD), and ten times of the recommended dose at 6000 g a.i. ha(-1) (10RD) along with control, were imposed. The initial residue (after 2 h of spray) deposits in soil were 0.174, 0.968, and 3.35 µg g(-1) for recommended, double the recommended, and ten times of the recommended doses, respectively. No residue in soil was detected in RD treatment on day 45. The half life values were 16.90, 17.76, and 36.47 days for RD, 2RD, and 10RD treatments, respectively. Application of pretilachlor at 10RD, in general, had significantly reduced the number of bacteria, actinomycetes, fungi, nitrogen fixers, and microbial biomass carbon. Pretilachlor at RD did not record any significant changes in microbial properties compared to control. The results of the present study thus indicated that pretilachlor at RD can be safely used for controlling grassy weeds in rice fields.