Plant species identification using fecal DNAs from red-eared slider and Reeves’ pond turtle in agricultural canals for rural ecosystem conservation

Fecal DNA samples from the red-eared slider and Reeves’ pond turtle, suspected pests of lotus root paddies, were used to identify the plant species eaten by these turtles in order to develop a strategy for rural ecosystem conservation. The fecal samples were obtained from young and adult individuals (mostly female) of both species living in agricultural canals surrounding lotus root paddies in Tokushima Prefecture, Japan. The samples were screened for the presence or absence of DNA from nine plant species using PCR and plant species-specific primers for the rbcL gene of chloroplast DNA. In the red-eared slider, our analysis identified seven plant species in the fecal DNA samples of adults and three plant species in those of young individuals. In Reeves’ pond turtle, our analysis identified two plant species from adult fecal samples and one species from those of young individuals. Thus, adult red-eared sliders consume a greater range of plants than young red-eared sliders or Reeves’ pond turtles. Both turtle species, independently of age, consumed lotus plants and were likely to cause feeding damage to lotus roots. Considering the plant species detected in adult red-eared sliders and these plant habitats, we suggest that this adult turtle is likely to travel between the agricultural canals and the lotus root paddies. These findings will help the development of strategies for preventing damage to lotus roots by these turtles; furthermore, they indicate that fecal DNA analysis will be applicable to investigation of the feeding habits of other animal species.     

Effect of nitrogen root zone fertilization on rice yield,uptake and utilization of macronutrient in lower reaches of Yangtze River,China

Improper application of nitrogen (N) has led to high N losses and low N use efficiency in the lower reaches of Yangtze River in China. An effective method to solve such problems is the deep fertilized N in root zone (RZF). Limited information is available on the effect of RZF on the uptake of macronutrients (N, P and K) and rice yield. Field experiments, conducted from 2014 to 2015, compared the farmer fertilizer practice (FFP, with 225 kg ha^?1 of N, split into three doses) and RZF using the same rate but placing N 5 cm away from rice roots in holes 10 cm deep (RZF10) or 5 cm deep (RZF5) as a single application. The highest mean yield (10.0 t ha^?1) was obtained in RZF10, which was 19.5% more than that in FFP. Root zone fertilization of urea (whether 10 cm deep or 5 cm deep) resulted in greater accumulation of N, P and K in stem, leaf sheaths, leaf blades and grains compared to that in FFP in sandy and in loam soils. The uptake of N, P and K was the highest in RZF10 (average at 176.7, 66.2 and 179.1 kg ha^?1, respectively), higher than that in FFP by 45.0, 17.0 and 22.6%, respectively. N apparent recovery efficiency was markedly higher in RZF10 (53.1%) than in FFP (27.5%). RZF10 significantly increased the N, P, K uptake compared with FFP under different N rates in both sandy and loam soils. These results suggest that the N, P and K input amount should be re-determined under RZF.     

Modeling the water and nitrogen transports in a soil–paddy–atmosphere system using HYDRUS-1D and lysimeter experiment

Efficient water and fertilizer use is of paramount importance both in rain-fed and irrigated rice cultivation systems to tread off between the crop water demand during the dry spell and the fertilizer leaching. This lysimeter study on paddy in a lateritic sandy loam soil of the eastern India, to simulate the water and solute transports using the HYDRUS-1D model, reveals that this model could very well simulate the soil depth-specific variations of water pressure heads and nitrogen (N) concentrations with the efficiency of >86 and 89%, respectively. The change in the level of water ponding depth did not have a significant effect on the time to peak and the temporal variability of N concentration in the bottom soil layer. The lysimeter-scale water balance analysis indicated that the average deep percolation loss and crop water use were 35.01 ± 2.03 and 39.74 ± 1.49% of the total water applied during the crop growth period, respectively. Similarly, the amount of N stored in the plant and lost through soil storage, deep percolation, and other losses (mineralization, denitrification, and gaseous N loss to the atmosphere through plant leaves) were 1.60 ± 0.16, 0.17 ± 0.04, 12.00 ± 0.48, and 86.23 ± 0.41% of the total applied nitrogen, respectively. The simulation results reveal that a constant ponding depth of 3 cm could be maintained in paddy fields to reduce the N leaching loss to 7.5 kgN/ha.     

Predicting adoption of double cropping in paddy fields of northern Iran: a comparison of statistical methods

Factors affecting the adoption of double cropping were explored in rice farms of Fouman County of Guilan Province in northern Iran using artificial neural networks (ANNs), linear discriminant analysis (LDA), and logistic regression (LGR). Eleven factors (age, education, occupation, family size, type of farm ownership, distance to the agricultural service center, attending agricultural extension courses, use of financial resources and bank loans, number of domestic animals, area under cultivation, and social participation) were examined. An additional objective was to compare the ability of the three models in predicting the adoption of double cropping. ANNs showed an overall predictive power of 89.8%. LDA showed an overall predictive power of 83.2%, with seven of the eleven independent variables being effective on the adoption of double cropping. LGR indicated an overall predictive power of 87.6%, with eight of the eleven independent variables being effective on the adoption of double-rice cropping. ANNs showed higher power than LGR and LDA in predicting the adoption of double cropping. Based on all three methods used for analysis, the most important independent variables were social participation and area under cultivation (positive factors) as well as distance to the agricultural service center and family members (negative factors). Establishment of cooperatives or other kinds of farmers’ associations to foster social participation could motivate adoption of double cropping, particularly among small-scale farmers. To increase agricultural services, more local centers should be created in rural areas. The government should promote double cropping through effective incentives and technology transfer to small-scale farmers.     

Identification of resistant germplasm containing novel resistance genes at or tightly linked to the <Emphasis Type="Italic">Pi2/9</Emphasis> locus conferring broad-spectrum resistance against rice blast

Background

The rice Pi2/9 locus harbors multiple resistance (R) genes each controlling broad-spectrum resistance against diverse isolates of Magnaporthe oryzae, a fungal pathogen causing devastating blast disease to rice. Identification of more resistance germplasm containing novel R genes at or tightly linked to the Pi2/9 locus would promote breeding of resistance rice cultivars.

Results

In this study, we aim to identify resistant germplasm containing novel R genes at or tightly linked to the Pi2/9 locus using a molecular marker, designated as Pi2/9-RH (Pi2/9 resistant haplotype), developed from the 5′ portion of the Pi2 sequence which was conserved only in the rice lines containing functional Pi2/9 alleles. DNA analysis using Pi2/9-RH identified 24 positive lines in 55 shortlisted landraces which showed resistance to 4 rice blast isolates. Analysis of partial sequences of the full-length cDNAs of Pi2/9 homologues resulted in the clustering of these 24 lines into 5 haplotypes each containing different Pi2/9 homologues which were designated as Pi2/9-A5, ?A15, ?A42, ?A53, and -A54. Interestingly, Pi2/9-A5 and Pi2/9-A54 are identical to Piz-t and Pi2, respectively. To validate the association of other three novel Pi2/9 homologues with the blast resistance, monogenic lines at BC₃F₃ generation were generated by marker assisted backcrossing (MABC). Resistance assessment of the derived monogenic lines in both the greenhouse and the field hotspot indicated that they all controlled broad-spectrum resistance against rice blast. Moreover, genetic analysis revealed that the blast resistance of these three monogenic lines was co-segregated with Pi2/9-RH, suggesting that the Pi2/9 locus or tightly linked loci could be responsible for the resistance.

Conclusion

The newly developed marker Pi2/9-RH could be used as a potentially diagnostic marker for the quick identification of resistant donors containing functional Pi2/9 alleles or unknown linked R genes. The three new monogenic lines containing the Pi2/9 introgression segment could be used as valuable materials for disease assessment and resistance donors in breeding program.

     

Updating the elite rice variety Kongyu 131 by improving the <Emphasis Type="Italic">Gn1a</Emphasis> locus

Background

Kongyu 131 is an elite japonica rice variety of Heilongjiang Province, China. It has the characteristics of early maturity, superior quality, high yield, cold tolerance and wide adaptability. However, there is potential to improve the yield of Kongyu 131 because of the relatively few grains per panicle compared with other varieties. Hence, we rebuilt the genome of Kongyu 131 by replacing the GRAIN NUMBER1a (Gn1a) locus with a high-yielding allele from a big panicle indica rice variety, GKBR. High-resolution melting (HRM) analysis was used for single nucleotide polymorphism (SNP) genotyping.

Results

Quantitative trait locus (QTL) analysis of the BC₃F₂ population showed that the introgressed segment carrying the Gn1a allele of GKBR significantly increased the branch number and grain number per panicle. Using 5 SNP markers designed against the sequence within and around Gn1a, the introgressed chromosome segment was shortened to approximately 430 Kb to minimize the linkage drag by screening recombinants in the target region. Genomic components of the new Kongyu 131 were detected using 220 SNP markers evenly distributed across 12 chromosomes, suggesting that the recovery ratio of the recurrent parent genome (RRPG) was 99.89%. Compared with Kongyu 131, the yield per plant of the new Kongyu 131 increased by 8.3% and 11.9% at Changchun and Jiamusi, respectively.

Conclusions

To achieve the high yield potential of Kongyu 131, a minute chromosome fragment carrying the favorable Gn1a allele from the donor parent was introgressed into the genome of Kongyu 131, which resulted in a larger panicle and subsequent yield increase in the new Kongyu 131. These results indicate the feasibility of improving an undesirable trait of an elite variety by replacing only a small chromosome segment carrying a favorable allele.

     

Conferring resistance to pre-harvest sprouting in durum wheat by a QTL identified in <Emphasis Type="Italic">Triticum spelta</Emphasis>

Pre-harvest sprouting (PHS) causes significant yield loss and degrade the end-use quality of wheat, especially in regions with prolonged wet weather during the harvesting season. Unfortunately, the gene pool of Triticum durum (tetraploid durum wheat) has narrow genetic base for PHS resistance. Therefore, finding out new genetic resources from other wheat species to develop PHS resistance in durum wheat is of importance. A major PHS resistance QTL, Qphs.sicau-3B.1, was mapped on chromosome 3BL in a recombinant inbred line population derived from ‘CSCR6’ (Triticum spelta), a PHS resistant hexaploid wheat and ‘Lang’, a PHS susceptible Australian hexaploid wheat cultivar. This QTL, Qphs.sicau-3B.1, is positioned between DArT marker wPt-3107 and wPt-6785. Two SCAR markers (Ph3B.1 and Ph3B.2) were developed to track this major QTL and were used to assay a BC₂F₈ tetraploid population derived from a cross between the durum wheat ‘Bellaroi’ (PHS susceptible) and ‘CSCR6’ (PHS resistant). Phenotypic assay and marker-assisted selection revealed five stable tetraploid lines were highly PHS resistant. This study has successfully established that PHS-resistance QTL from hexaploid wheat could be efficiently introgressed into tetraploid durum wheat. This tetraploid wheat germplasm could be useful in developing PHS resistant durum cultivars with higher yield and good end-use quality.     

Identification of favorable alleles in the <Emphasis Type="Italic">non</Emphasis>-<Emphasis Type="Italic">yellow coloring 1</Emphasis> gene by association mapping in maize

Association mapping was conducted to explore favorable alleles of the chlorophyll-related non-yellow coloring 1 (NYC1) gene under light and dark using an association panel of 146 maize inbred lines. A total of 14 polymorphic sites were identified to be significantly associated with at least one of the chlorophyll-related traits at the seedling stage. Four single nucleotide polymorphisms (SNPs) (S320, S2951, S3901, and S3355) from the NYC1 gene were respectively strongly associated with chlorophyll b (chlb), the ratio of chlorophyll a to chlorophyll b (chl_ratio), chlorophyll a degradation (chla_deg), and total chlorophyll degradation (total_chl_deg). SNPs S320 (C/A) in exon 1, and S2951 (A/G) in intron 8 was related to chlb, with 6.01 and 8.89% of phenotypic variation under light treatment, respectively. Under dark treatment, SNP S3901 (C/T), located in 3′ untranslated region (3′UTR), was associated with chl_ratio, explaining 7.01% of the observed phenotypic variation, whereas SNP S3355 (C/G) in intron 9 explained 6.48 and 5.18% of phenotypic variations in chla_deg and total_chl_deg, respectively. Taken together, these results indicated that the NYC1 gene plays an important role in chlorophyll content and other related traits, and different sites act on chlorophyll metabolism under different light intensities in maize seedlings. Furthermore, these findings improve our understanding of the genetic basis of chlorophyll metabolism under different light conditions.     

Development of male-specific markers and identification of sex reversal mutants in papaya

Papaya is a productive and nutritious fruit grown in tropical and sub-tropical regions worldwide. It is polygamous with three sex types: female, male and hermaphrodite. Sex determination in papaya is controlled by an XY sex chromosome system with two slightly different Y chromosomes, Y for males and Y^h for hermaphrodites. Comparative analysis of the hermaphrodite-specific region of Y^h chromosome (HSY) and male-specific region of Y chromosome (MSY) revealed 99.6% sequence identity, which explains why DNA markers that amplify for both males and hermaphrodites have easily been developed, but not for the male trait specifically. We examined the 0.4% sequence differences, and found 1887 indels and 21,088 SNPs between MSY and HSY. The vast majority of indels are single nucleotide or few base pairs. A large male-specific retrotransposon insertion of 8396 bp was used to develop two papaya male-specific markers, PMSM1 and PMSM2 that amplify 585 and 548 bp fragments, respectively. These two markers were tested in 11 gynodioecious and four dioecious varieties along with autosomal DNA marker 71E and male/hermaphrodite marker W11, and the results showed clear separation of male from hermaphrodite and female. PMSM1 and PMSM2 were also used to test the sex type of six sex male-to-hermaphrodite reversal mutants which are crucial materials for validating candidate genes for sex determination in papaya. Our result showed all six mutants were positive for the male-specific markers. These male-specific markers can be used to distinguish gynodioecious and dioecious cultivars in papaya seed market, and facilitate genetic and genomic research for papaya improvement.