Atul Singh

Marker-assisted improvement of bacterial blight resistance in parental lines of Pusa RH10, a superfine grain aromatic rice hybrid

Pusa RH10, the widely cultivated superfine grain aromatic rice hybrid, and its parental lines Pusa6B and PRR78 are susceptible to bacterial blight (BB) disease caused by Xanthomonas oryzae pv. oryzae. Pusa1460, a Basmati rice variety, was utilized as the donor for introgressing BB resistance genes xa13 and Xa21 into Pusa6B and PRR78 using a marker-assisted backcross breeding program. The markers RG136 and pTA248 linked to BB resistance genes xa13 and Xa21, respectively, were used for foreground selection. Seventy-four STMS markers polymorphic between Pusa6B and Pusa1460, and 54 STMS markers polymorphic between PRR78 and Pusa1460, were utilized for background selection to recover the recurrent parent genome ranging from 85.14 to 97.30% and 87.04 to 92.81% in the 10 best BC2F5 families of Pusa6B and PRR78, respectively. RM6100, an STMS marker linked to fertility restorer gene (Rf), was used for marker-assisted selection of Rf gene in an improved version of PRR78. The extent of donor segments in the improved version of Pusa6B was estimated to be <0.97 and <2.15xa0Mb in the genomic regions flanking xa13 and Xa21, respectively, whereas in improved PRR78, it was estimated to be <2.07 and <3.45xa0Mb in the corresponding genomic regions. Improved lines of Pusa6B and PRR78 showed yield advantages of up to 8.24 and 5.23%, respectively. The performance of the BB-resistant version of Pusa RH10 produced by intercrossing the improved parental lines was on a par with or superior to the original Pusa RH10.

Methane production in rice soil is inhibited by cyanobacteria

The present study was aimed at understanding the role of cyanobacteria and Azolla in methane production and oxidation in laboratory simulation experiments using soil samples from rice fields. All the seven cyanobacterial strains tested effected a significant decrease in the headspace concentration of methane in flooded soil, incubated under light. Synechocystis sp. was the most effective in retarding methane concentration by 10-20 fold over that in controls without cyanobacteria. The decrease in the headspace concentration of methane was negligible in nonsterile soil samples, inoculated with Synechocystis sp. and then incubated under dark. Moist soil cores (0-5 cm depth), collected from rice fields that had been treated with urea in combination with a cyanobacterial mixture, Azolla microphylla, or cyanobacterial mixture plus A. microphylla, effected distinctly more rapid decrease in the headspace concentration of methane added at 200 microl(-1) than did the soil cores from plots treated with urea alone (30, 60, 90 and 120 kg N ha(-1)), irrespective of the rate of chemical nitrogen applied to rice fields. Besides, soil cores from plots treated with urea alone at 60, 90 and 120 kg N ha(-1) oxidised methane more rapidly than did the core samples from plots treated with urea alone at 30kg N ha(-1). Cyanobacteria and A. microphylla, applied to flood water, appear to play a major role in mitigation of methane emission from rice fields-through enhanced methane oxidation.

Molecular breeding for the development of multiple disease resistance in Basmati rice

Marker assisted backcross breeding for combining three resistance genes (xa13 and Xa21 for Bacterial Blight, Pi54 for blast) and a major QTL (qSBR11-1 for resistance to Sheath blight) in Basmati rice.

Prediction of hybrid performance based on the genetic distance of parental lines in two-line rice (Oryza sativa L.) hybrids

A total of six TGMS (thermosensitive genic male sterile lines) and nine pollinator lines were subjected to molecular characterization using 48 genome-wide SSR (simple sequence repeat) markers. Cluster analysis revealed clear differentiation among the TGMS lines according to their source of origin. The SSR-based genetic distance between the hybrids of the parental lines ranged from 0.36 to 0.79 suggesting a high degree of genetic divergence. Among a set of 54 hybrids generated using parental lines, 32 showed better parent heterosis (+21.7%) while 19 showed mid-parent heterosis (+15.0%). For the trait yield per plant genetic distance (GD) was negatively correlated with F1 performance (r = −0.202), mid-parent heterosis (r = −0.325*; P < 0.05), and better parent heterosis (r = −0.261), while it was positively correlated with specific combining ability (r = 0.042). Based on the grouped genetic distance (GGD), the hybrid combinations were divided into four groups. The GGD showed linear correlation with hybrid performance within the group (GGD = 40–50: r = −0.07; GGD = 70–80: r = 0.32). This information can be utilized in the development of higher yielding, two-line rice hybrids through selection of intermediately diverse parental lines using GGD.

Pathological findings of experimental Aeromonas hydrophila infection in Golden Mahseer (Tor putitora)

a) Introduction: nMahseers, belong to the family Cyprinidae, are well renowned for the excellent game as well as food i¬x81shes. Various aspects simultaneously considered towards the domestication of this fish species also include the study of fish health issues, preventive and remedial programmes for sustainable aquaculture. Experimental studies can be conducted in the control condition to study the pathological symptoms during bacterial infection. Considering the bacterial pathogens as a major constraint for aquaculture, the present study was carried out to standardize the bacterial concentration of A. hydrophila for the experimental challenge of Golden Mahseer for the first time, and various clinical changes were also monitored during the infection. Materials and Methods: A total of 140 live fishes were acclimatized to the laboratory conditions for experimental study. A constant water quality was monitored during the acclimation period and throughout the experiment. The test group was injected with A. hydrophila and various physiological, biochemical and tissue level changes were observed during the experiment in comparison to phosphate buffer saline (PBS) injected control group. nb) Results: LD50 value of A. hydrophila for Golden Mahseer was successfully standardized. Clinical signs including weakness, slower movement, swimming closer to the surface, fin haemorrhages and red patches at the gut region were observed. Enlargement of spleen followed by tissue necrosis along with signs of haemorrhagic septicaemia was also seen in infected fishes. The necrosis of hepatocytes was observed in the histological section of liver tissue. The bacterial infection increases the superoxide dismutase activity and cortisol level in Golden Mahseer. nc) Conclusion: The LD50 value of A. hydrophila for experimental challenge studies on Golden Mahseer is 1.74 × 105 cfu per 100 g of body weight. The symptoms of severe bacterial haemorrhage were observed. Necrosis of hepatocytes was observed in infected fishes. The superoxide dismutase activity and cortisol level also increased in infected fishes. A. hydrophila was confirmed to be the etiological agent which was re-isolated using spread plate method and confirmed by 16s rRNA sequencing.

Determination of genetic relationships among elite thermosensitive genic male sterile lines (TGMS) of rice (Oryza sativa L.) employing morphological and simple sequence repeat (SSR) markers

A set of morphological traits and SSR markers were used to determine the genetic relationship among 12 elite thermosensitive genic male sterile (TGMS) lines developed at three different research institutions of India. Agro-morphological data recorded on 20 morphological traits revealed a wide base of genetic variation and a set of four morphological traits could distinguish most of the TGMS lines. Analysis with 30 SSR markers (20 EST-SSRs and 10 genomic SSRs) revealed 27 markers to be polymorphic, amplifying a total of 83 alleles. Each SSR marker amplified 2–6 alleles with an average of 2.76 alleles per marker and a PIC value varying from 0.54 to 0.96. Cluster analysis based on SSR and morphological data clearly differentiated the lines according to their source of origin. Correlation analysis between morphological and molecular data revealed a very poor association (ru2009=u20090.06), which could be attributed to selection pressure, genetic drift, sampling error and unknown relationship among related lines. The SSR markers discriminated the genotypes distinctly and quantified the genetic diversity precisely among the TGMS lines. Data on the yield per plant indicated that the genotypes grouping under a similar cluster showed same heterotic behaviour as compared to the genotypes from different clusters when crossed to similar pollinators.

Introgression of multiple disease resistance into a maintainer of Basmati rice CMS line by marker assisted backcross breeding

Globally, blast incited by Magnaporthe oryzae and sheath blight (ShB) by Rhizoctonia solani Kuhn forms two major fungal diseases that cause significant yield loss in rice. Pusa 6B, the Basmati quality maintainer line of the popular superfine grain aromatic rice hybrid Pusa RH10, is highly susceptible to both the diseases. The rice cultivar Tetep was used as the donor for transferring a major blast resistant gene, Pi54 and three ShB resistant quantitative trait loci (QTLs) namely, qSBR11-1, qSBR11-2 and qSBR7-1 into Pusa 6B using a marker assisted backcross breeding scheme with restricted number of backcrosses. Plants heterozygous for the alleles of interest and phenotypic similarity to the recurrent parent were used for generating BC1F2 population by selfing selected BC1F1 plants. Selected BC1F2 plants homozygous for Pi54 were selfed to generate BC1F3 families that were subjected to a step-wise reductive screening for the three ShB resistant QTLs. Final selections were advanced to BC1F5 generation through selfing while subjecting to stringent phenotypic selection. The advanced selections carrying blast and ShB resistant genes (Pi54, qSBR11-1, qSBR11-2, qSBR7-1) in the background of Pusa 6B were resistant to highly virulent strains of rice blast as well as ShB isolates without compromising the grain and cooking quality of Pusa 6B. Marker assisted transfer of blast and ShB resistance into Pusa 6B will aid in developing CMS lines with inbuilt resistance to these diseases. When combined with restorer lines possessing resistant genes/QTLs for these diseases, the improved Pusa6A lines will aid in development of improved Pusa RH10 and other novel aromatic hybrids with resistance to blast and ShB diseases. The present study demonstrates successful use of a restricted backcrossing strategy for introgression of multiple loci conferring resistance to two important fungal diseases in rice.

Complete mitochondrial genome organization of Tor tor (Hamilton, 1822)

Abstract The complete mitochondrial genome of Tor tor, a threatened “Mahseer” was sequenced for the first time. The mitochondrial genome size determined to be 16,554u2009bp in length and consisted of 13 protein-coding genes (PCGs), 22 tRNAs, 2 rRNA genes and a control region or displacement loop (D-Loop) region, resembling the typical organizational pattern of most of the teleost. The overall base composition found was A: 31.8%, T: 25%, G: 15.7% and C: 27.4%; Au2009+u2009T: 56.9% and Gu2009+u2009C: 43.1%. The phylogenetic tree constructed using 11 other cyprinids’ total mtDNA datasets confirmed the location of present species among mahseers. The total sequence data could support further study in molecular systematics, species identification, evolutionary and conservation genetics.

Molecular characterization of rainbow trout, Oncorhynchus mykiss (Walbaum, 1792) stocks in India

In India, rainbow trout was introduced by British more than 100 years ago (Agarwal 2006) for recreational purposes. The fish is being cultured in both government and private farms of different coldwater states of India, mainly in Jammu and Kashmir, Himachal Pradesh, Sikkim, Arunachal Pradesh and Uttarakhand, and also in Nilgiri hills of south India for breeding and rearing purposes. In recent past, growth and production rate of this fish in various farms has been reduced. Microsatellite markers in combination with recent statistical approaches represent a useful tool for genetic characterization which ultimately supports the management of cultured stocks. These markers have been successfully used to evaluate the wild and farm stocks of rainbow trout in western Australia (Ward et al. 2003); resident and anadromus forms in the Walla Walla river (Narum et al. 2004); domesticated strains of rainbow trout in USA (Silverstein et al. 2004); strains in northern and eastern Europe (Gross et al. 2007) and three groups of different origin in north of Iran (Yousefian et al. 2012). Hence, the present study was carried out to assess the genetic variability in different stocks of rainbow trout in India using microsatellite markers.