Invasion by Cenchrus spinifex changes the soil microbial community structure in a sandy grassland ecosystem.

Continuous nitrogen deposition increases the nitrogen content of terrestrial ecosystems and alters the soil nitrogen cycling process. Invasive plants have strong environmental adaptability, which can not only affect the composition and diversity of soil microbial community but also significantly affect the transformation process of soil nitrogen, leading to successful invasion. Currently, research on invasive plant soil ecosystems mainly focused on changes in soil nutrients and soil microorganisms. As an invasive annual grass weed with strong ecological adaptability, the impact of Cenchrus spinifex at different growth periods on soil environment and soil microbial structure composition and diversity in sandy grassland ecosystems is still unclear. In this study, soil samples were collected from four habitats with different degrees of invasion in situ during the vegetation and reproductive growth periods of Cenchrus spinifex. High-throughput sequencing and qPCR technology were used to analyze the changes in the composition, structure and diversity characteristics of the soil microbial communities during Cenchrus spinifex invasion. The results indicated that Cenchrus spinifex invasion had different effects on the soil environment at different growth periods, and Cenchrus spinifex had a preference for the utilization of ammonium nitrogen during vegetation growth period. Moreover, Cenchrus spinifex invasion significantly changed the composition and structure of soil bacterial communities, and the response of soil bacterial and fungal communities to the invasion was inconsistent. Additionally, the bacterial network was more stable than the fungal network. At different growth periods, Cenchrus spinifex had a significant impact on the key microbial communities of soil nitrogen cycling. The invasion increased the abundance of nifH and AOA-amoA, while decreased the abundance of AOA-amoB. Alkaline hydrolyzed nitrogen, total nitrogen and total phosphorus content were key factors that affect vegetation growth period and change the key microbial communities of nitrogen cycling. Alkaline hydrolyzed nitrogen, total phosphorus and organic carbon were key factors in reproductive growth period that alter the nitrogen cycling of key microbial communities.

Different responses of soil fungal and bacterial communities to nitrogen addition in a forest grassland ecotone.

Introduction: Continuous nitrogen deposition increases the nitrogen content of terrestrial ecosystem and affects the geochemical cycle of soil nitrogen. Forest-grassland ecotone is the interface area of forest and grassland and is sensitive to global climate change. However, the structure composition and diversity of soil microbial communities and their relationship with soil environmental factors at increasing nitrogen deposition have not been sufficiently studied in forest-grassland ecotone. Methods: In this study, experiments were carried out with four nitrogen addition treatments (0 kgN·hm-2·a-1, 10 kgN·hm-2·a-1, 20 kgN·hm-2·a-1 and 40 kgN·hm-2·a-1) to simulate nitrogen deposition in a forest-grassland ecotone in northwest Liaoning Province, China. High-throughput sequencing and qPCR technologies were used to analyze the composition, structure, and diversity characteristics of the soil microbial communities under different levels of nitrogen addition. Results and discussion: The results showed that soil pH decreased significantly at increasing nitrogen concentrations, and the total nitrogen and ammonium nitrogen contents first increased and then decreased, which were significantly higher in the N10 treatment than in other treatments (N:0.32 ~ 0.48 g/kg; NH4+-N: 11.54 ~ 13 mg/kg). With the increase in nitrogen concentration, the net nitrogen mineralization, nitrification, and ammoniation rates decreased. The addition of nitrogen had no significant effect on the diversity and structure of the fungal community, while the diversity of the bacterial community decreased significantly at increasing nitrogen concentrations. Ascomycetes and Actinomycetes were the dominant fungal and bacterial phyla, respectively. The relative abundance of Ascomycetes was negatively correlated with total nitrogen content, while that of Actinomycetes was positively correlated with soil pH. The fungal community diversity was significantly negatively correlated with nitrate nitrogen, while the diversity of the bacterial community was significantly positively correlated with soil pH. No significant differences in the abundance of functional genes related to soil nitrogen transformations under the different treatments were observed. Overall, the distribution pattern and driving factors were different in soil microbial communities in a forest-grassland ecotone in northwest Liaoning. Our study enriches research content related to factors that affect the forest-grassland ecotone.

Changes in grassland soil types lead to different characteristics of bacterial and fungal communities in Northwest Liaoning, China.

Introduction: Soil microbial communities are critical in regulating grassland biogeochemical cycles and ecosystem functions, but the mechanisms of how environmental factors affect changes in the structural composition and diversity of soil microbial communities in different grassland soil types is not fully understood in northwest Liaoning, China. Methods: We investigated the characteristics and drivers of bacterial and fungal communities in 4 grassland soil types with 11 sites across this region using high-throughput Illumina sequencing. Results and Discussion: Actinobacteria and Ascomycota were the dominant phyla of bacterial and fungal communities, respectively, but their relative abundances were not significantly different among different grassland soil types. The abundance, number of OTUs, number of species and diversity of both bacterial and fungal communities in warm and temperate ecotone soil were the highest, while the warm-temperate shrub soil had the lowest microbial diversity. Besides, environmental factors were not significantly correlated with soil bacterial Alpha diversity index. However, there was a highly significant negative correlation between soil pH and Shannon index of fungal communities, and a highly significant positive correlation between plant cover and Chao1 index as well as Observed species of fungal communities. Analysis of similarities showed that the structural composition of microbial communities differed significantly among different grassland soil types. Meanwhile, the microbial community structure of temperate steppe-sandy soil was significantly different from that of other grassland soil types. Redundancy analysis revealed that soil total nitrogen content, pH and conductivity were important influencing factors causing changes in soil bacterial communities, while soil organic carbon, total nitrogen content and conductivity mainly drove the differentiation of soil fungal communities. In addition, the degree of connection in the soil bacterial network of grassland was much higher than that in the fungal network and soil bacterial and fungal communities were inconsistently limited by environmental factors. Our results showed that the microbial community structure, composition and diversity of different grassland soil types in northwest Liaoning differed significantly and were significantly influenced by environmental factors. Microbial community structure and the observation of soil total nitrogen and organic carbon content can predict the health changes of grassland ecosystems to a certain extent.

B-A Chromosome Translocations Possessing an A Centromere Partly Overcome the Root-Restricted Process of Chromosome Elimination in Aegilops speltoides.

Some eukaryotes exhibit dramatic genome size differences between cells of different organs, resulting from the programmed elimination of chromosomes. Aegilops speltoides is an annual diploid species from the Poaceae family, with a maximum number of eight B chromosomes (Bs) in addition to its inherent seven pairs of standard A chromosomes (As). The Bs of this species undergo precise elimination in roots early in embryo development. In areal parts of the plant, the number of Bs is stable. To affect the root restricted process of B chromosome elimination, we employed X-ray mutagenesis, and different types of restructured Bs were identified. Standard Bs were observed in all analyzed shoots of mutagenized plants, while B-A translocations were only observed in 35.7% of F1 plants. In total 40 different B variants inconsistently escaped the elimination process in roots. As a result, mosaicism of B chromosome variants was found in roots. Only a small B chromosome fragment fused to an A chromosome was stably maintained in roots and shoots across F1 to F3 generations. The absence of B-A translocation chromosomes possessing a derived B centromere in root cells implies that the centromere of the B is a key component of the chromosome elimination process.

Characterization of a wheat-tetraploid Thinopyrum elongatum 1E(1D) substitution line K17-841-1 by cytological and phenotypic analysis and developed molecular markers.

BACKGROUND: Tetraploid Thinopyrum elongatum (2n = 4x = 28) is a promising source of useful genes, including those related to adaptability and resistance to diverse biotic (Fusarium head blight, rust, powdery mildew, and yellow dwarf virus) and abiotic (cold, drought, and salt) stresses. However, gene transfer rates are low for this species and relatively few species-specific molecular markers are available. RESULTS: The wheat-tetraploid Th. elongatum line K17-841-1 derived from a cross between a hexaploid Trititrigia and Sichuan wheat cultivars was characterized based on sequential genomic and fluorescence in situ hybridizations and simple sequence repeat markers. We revealed that K17-841-1 is a 1E (1D) chromosomal substitution line that is highly resistant to stripe rust pathogen strains prevalent in China. By comparing the sequences generated during genotyping-by-sequencing (GBS), we obtained 597 specific fragments on the 1E chromosome of tetraploid Th. elongatum. A total of 235 primers were designed and 165 new Th. elongatum-specific markers were developed, with an efficiency of up to 70%. Marker validation analyses indicated that 25 specific markers can discriminate between the tetraploid Th. elongatum chromosomes and the chromosomes of other wheat-related species. An evaluation of the utility of these markers in a F2 breeding population suggested these markers are linked to the stripe rust resistance gene on chromosome 1E. Furthermore, 28 markers are unique to diploid Th. elongatum, tetraploid Th. elongatum, or decaploid Thinopyrum ponticum, which carry the E genome. Finally, 48 and 74 markers revealed polymorphisms between Thinopyrum E-genome- containing species and Thinopyrum bessarabicum (Eb) and Pseudoroegneria libanotica (St), respectively. CONCLUSIONS: This new substitution line provide appropriate bridge-breeding-materials for alien gene introgression to improve wheat stripe rust resistance. The markers developed using GBS technology in this study may be useful for the high-throughput and accurate detection of tetraploid Th. elongatum DNA in diverse materials. They may also be relevant for investigating the genetic differences and phylogenetic relationships among E, Eb, St, and other closely-related genomes and for further characterizing these complex species.

FISH-Based Markers Enable Identification of Chromosomes Derived From Tetraploid Thinopyrum elongatum in Hybrid Lines.

Tetraploid Thinopyrum elongatum, which has superior abiotic stress tolerance characteristics, and exhibits resistance to stripe rust, powdery mildew, and Fusarium head blight, is a wild relative of wheat and a promising source of novel genes for wheat improvement. Currently, a high-resolution Fluorescence in situ hybridization (FISH) karyotype of tetraploid Th. elongatum is not available. To develop chromosome-specific FISH-based markers, the hexaploid Trititrigia 8801 and two accessions of tetraploid Th. elongatum were characterized by different repetitive sequences probes. We found that all E-genome chromosomes could be unambiguously identified using a combination of pSc119.2, pTa535, pTa71, and pTa713 repeats, and the E-genome chromosomes of the wild accessions and the partial amphiploid failed to exhibit any significant variation in the probe hybridization patterns. To verify the validation of these markers, the chromosome constitution of eight wheat- Th. elongatum hybrid derivatives were analyzed. We revealed that these probes could quickly detect wheat and tetraploid Th. elongatum chromosomes in hybrid lines. K16-712-1-2 was a 1E (1D) chromosome substitution line, K16-681-4 was a 2E disomic chromosome addition line, K16-562-3 was a 3E, 4E (3D, 4D) chromosome substitution line, K15-1033-8-2 contained one 4E, two 5E, and one 4ES⋅1DL Robertsonian translocation chromosome, and four other lines carried monosomic 4E, 5E, 6E, and 7E chromosome, respectively. Furthermore, the E-genome specific molecular markers analysis corresponded perfectly with the FISH results. The developed FISH markers will facilitate rapid identification of tetraploid Th. elongatum chromosomes in wheat improvement programs and allow appropriate alien chromosome transfer.

Cytogenetics and stripe rust resistance of wheat-Thinopyrum elongatum hybrid derivatives.

BACKGROUND: Amphidiploids generated by distant hybridization are commonly used as genetic bridge to transfer desirable genes from wild wheat species into cultivated wheat. This method is typically used to enhance the resistance of wheat to biotic or abiotic stresses, and to increase crop yield and quality. Tetraploid Thinopyrum elongatum exhibits strong adaptability, resistance to stripe rust and Fusarium head blight, and tolerance to salt, drought, and cold. RESULTS: In the present study, we produced hybrid derivatives by crossing and backcrossing the Triticum durum-Th. elongatum partial amphidiploid (Trititrigia 8801, 2n = 6× = 42, AABBEE) with wheat cultivars common to the Sichuan Basin. By means of cytogenetic and disease resistance analyses, we identified progeny harboring alien chromosomes and measured their resistance to stripe rust. Hybrid progenies possessed chromosome numbers ranging from 40 to 47 (mean = 42.72), with 40.0% possessing 42 chromosomes. Genomic in situ hybridization revealed that the number of alien chromosomes ranged from 1 to 11. Out of the 50 of analyzed lines, five represented chromosome addition (2n = 44 = 42 W + 2E) and other five were chromosome substitution lines (2n = 42 = 40 W + 2E). Importantly, a single chromosome derived from wheat-Th. elongatum intergenomic Robertsonian translocations chromosome was occurred in 12 lines. Compared with the wheat parental cultivars ('CN16' and 'SM482'), the majority (70%) of the derivative lines were highly resistant to strains of stripe rust pathogen known to be prevalent in China. CONCLUSION: The findings suggest that these hybrid-derivative lines with stripe rust resistance could potentially be used as germplasm sources for further wheat improvement.

Cytogenetic study and stripe rust response of the derivatives from a wheat - Thinopyrum intermedium - Psathyrostachys huashanica trigeneric hybrid.

To transfer multiple desirable alien genes into common wheat, we previously reported a new trigeneric hybrid synthesized by crossing a wheat - Thinopyrum intermedium partial amphiploid with wheat - Psathyrostachys huashanica amphiploid. Here, the meiotic behavior, chromosome constitution, and stripe rust resistance of F5 derivatives from the wheat - Th. intermedium - P. huashanica trigeneric hybrid were studied. Cytological analysis indicated the F5 progenies had chromosome numbers of 42-50 (average 44.96). The mean meiotic configuration was 1.28 univalents, 21.74 bivalents, 0.04 trivalents, and 0.02 tetravalents per pollen mother cell. In 2n = 42 lines, the average pairing configuration was 0.05 I + 19.91 II (ring) + 1.06 II (rod) + 0.003 IV, suggesting these lines were cytologically stable. Most lines with 2n = 43, 44, 46, 48, or 50, bearing a high frequency of univalents or multivalents, showed abnormal meiotic behavior. Genomic in situ hybridization karyotyping results revealed that 25 lines contained 1-7 Th. intermedium chromosomes, but no P. huashanica chromosomes were found among the 27 self-pollinated progenies. At meiosis, univalents (1-5) possessing Th. intermedium hybridization signals were detected in 19 lines. Bivalents (1-3) expressing fluorescence signals were observed in 12 lines. Importantly, 21 lines harbored wheat - Th. intermedium chromosomal translocations with various alien translocation types. Additionally, two homozygous lines, K13-668-10 and K13-682-12, possessed a pair of wheat - Th. intermedium small fragmental translocations. Compared with the recurrent parent Zhong 3, most lines showed high resistance to the stripe rust (Puccinia striiformis f. sp. tritici) pathogens prevalent in China, including race V26/Gui22. This paper reports a highly efficient technical method for inducing alien translocation between wheat and Th. intermedium by trigeneric hybridization. These lines might be potentially valuable germplasm resources for further wheat improvement.

Divergent Development of Hexaploid Triticale by a Wheat - Rye -Psathyrostachys huashanica Trigeneric Hybrid Method.

Hexaploid triticale is an important forage crop and a promising energy plant. Some forms were previously reported for developing the hexaploid triticale, such as crossing tetraploid wheat or hexaploid wheat with rye, crossing hexaploid triticale and/or hexaploid wheat with octoploid triticale, and spontaneously appearing in the selfed progenies of octoploid triticale. In the present study, we developed an effective method for production of diverse types of hexaploid triticale via wheat-rye-Psathyrostachys huashanica trigeneric hybrid. Genomic in situ hybridization (GISH) and fluorescence in situ hybridization (FISH) karyotyping revealed that D genome chromosomes were completely eliminated and the whole A, B, and R genome chromosomes were retained in three lines. More interestingly, the composite genome of the line K14-489-2 consisted of complete A and B genomes and chromosomes 1D, 2R, 3R, 4R, 5R, 6R, and 7R, that of line K14-491-2 was 12 A-genome (1A-6A), 14 B-genome (1B-7B), 12 R-genome (1R-3R, 5R-7R), and chromosomes 1D and 3D, and that of the line K14-547-1 had 26A/B and 14R chromosomes, plus one pair of centric 6BL/2DS translocations. This finding implies that some of D genome chromosomes can be spontaneously and stably incorporated into the hexaploid triticale. Additionally, a variety of high-molecular-weight glutenin subunits (HMW-GS) compositions were detected in the six hexaploid triticale lines, respectively. Besides, compared with its recurrent triticale parent Zhongsi828, these lines showed high level of resistance to stripe rust (Puccinia striiformis f. sp. tritici, Pst) pathogens prevalent in China, including V26/Gui 22. These new hexaploid triticales not only enhanced diversification of triticale but also could be utilized as valuable germplasm for wheat improvement.

Molecular cytogenetic characterization of a new wheat-rye 1BL•1RS translocation line expressing superior stripe rust resistance and enhanced grain yield.

MAIN CONCLUSION: A new wheat-rye 1BL•1RS translocation line, with the characteristics of superior stripe rust resistance and high thousand-kernel weight and grain number per spike, was developed and identified from progenies of wheat-rye- Psathyrostachys huashanica trigeneric hybrids. The wheat-rye 1BL•1RS translocation line from Petkus rye has contributed substantially to the world wheat production. However, due to extensive growing of cultivars with disease resistance genes from short arm of rye chromosome 1R and coevolution of pathogen virulence and host resistance, these cultivars successively lost resistance to pathogens. In this study, a new wheat-rye line K13-868, derived from the progenies of wheat-rye-Psathyrostachys huashanica trigeneric hybrids, was identified and analyzed using fluorescence in situ hybridization (FISH), genomic in situ hybridization (GISH), acid polyacrylamide gel electrophoresis (A-PAGE), and molecular markers. Cytological studies indicated that the mean chromosome configuration of K13-868 at meiosis was 2n = 42 = 0.14 I + 18.78 II (ring) + 2.15 II (rod). Sequential FISH and GISH results demonstrated that K13-868 was a compensating wheat-rye 1BL•1RS Robertsonian translocation line. Acid PAGE analysis revealed that clear specific bands of rye 1RS were expressed in K13-868. Simple sequence repeat (SSR) and rye 1RS-specific markers ω-sec-p1/ω-sec-p2 and O-SEC5'-A/O-SEC3'-R suggested that the 1BS arm of wheat had been substituted by the 1RS arm of rye. At the seedling and adult growth stage, compared with its recurrent wheat parent SM51 and six other wheat cultivars containing the 1RS arm in southwestern China, K13-868 showed high levels of resistance to stripe rust (Puccinia striiformis f. sp. tritici, Pst) pathogens prevalent in China, which are virulent to Yr10 and Yr24/Yr26. In addition, K13-868 expresses higher thousand-kernel weight and more grain number per spike than these controls in two growing seasons, suggesting that this line may carry yield-related genes of rye. This translocation line, with significant characteristics of resistance to stripe rust and high thousand-kernel weight and grain number per spike, could be utilized as a valuable germplasm for wheat improvement.

Cytogenetic Behavior of Trigeneric Hybrid Progeny Involving Wheat, Rye and Psathyrostachys huashanica.

Trigeneric hybrids are commonly used as bridges to transfer genes from some wild species to cultivated wheat and to measure the genomic interaction between donor species. We previously reported that trigeneric germplasms were produced by crossing wheat-Psathyrostachys huashanica amphiploids (PHW-SA, 2n = 8x = 56, AABBDDNsNs) with hexaploid triticale (Zhongsi 828, 2n = 6x = 42, AABBRR). In the present study, chromosome pairing behavior and the genome constitution of the F4 progenies of wheat-rye-P. huashanica trigeneric hybrids were studied. Cytological analysis showed that the chromosome number of F4 progenies ranged from 39 to 46, and 57.5% of them had 42 chromosomes. The mean meiotic configuration of F4 lines was 1.71 univalents, 20.26 bivalents, 0.04 trivalents, and 0.001 quadrivalents per pollen mother cell. Among the lines with 2n = 42, the average pairing configuration was 1.21 univalents, 16.22 ring bivalents, 4.16 rod bivalents, and 0.01 trivalents. This result indicated that these lines were cytologically stable. Other lines with 2n = 39, 40, 41, 43, 44, 45, and 46, bearing a high number of univalents or multivalents, showed abnormal meiotic behavior. Genomic in situ hybridization (GISH) revealed that all F4 lines had 11-14 rye chromosomes, but no P. huashanica chromosomes. The complete set of 14 rye chromosomes was found in 19 lines. At meiosis, GISH detected 1-6 univalents with hybridization signals of rye in 13 lines. Bivalents with fluorescence signals were identified in each line, ranging from 3 to 7. A quadrivalent with hybridization signals was observed in only 1 line, K13-714-8. Lagging chromosomes, chromosome bridges, micronuclei, and chromosome fragments hybridizing with the probe were not discovered in any of the lines. These results inferred that the behavior of rye chromosomes was normal during meiosis. In addition, 21 lines of 2n = 42 (91.3%) with 12 or 14 rye chromosomes, always contained 6 or 7 bivalents bearing fluorescence signals. This suggested that the rye chromosomes exhibiting complete pairing in these lines were cytologically stable during meiosis and may therefore be considered as new hexaploid triticales. Thus, these lines might be potential materials for further hexaploid triticale improvement.

Characterization of wheat - Psathyrostachys huashanica small segment translocation line with enhanced kernels per spike and stripe rust resistance.

Psathyrostachys huashanica Keng (2n = 2x = 14, NsNs), a distant wild relative of common wheat, possesses rich potentially valuable traits, such as disease resistance and more spikelets and kernels per spike, that could be useful for wheat genetic improvement. Development of wheat - P. huashanica translocation lines will facilitate its practical utilization in wheat breeding. In the present study, a wheat - P. huashanica small segmental translocation line, K-13-835-3, was isolated and characterized from the BC1F5 population of a cross between wheat - P. huashanica amphiploid PHW-SA and wheat cultivar CN16. Cytological studies showed that the mean chromosome configuration of K-13-835-3 at meiosis was 2n = 42 = 0.10 I + 19.43 II (ring) + 1.52 II (rod). GISH analyses indicated that chromosome composition of K-13-835-3 included 40 wheat chromosomes and a pair of wheat - P. huashanica translocation chromosomes. FISH results demonstrated that the small segment from an unidentified P. huashanica chromosome was translocated into wheat chromosome arm 5DS, proximal to the centromere region of 5DS. Compared with the cultivar wheat parent CN16, K-13-835-3 was highly resistant to stripe rust pathogens prevalent in China. Furthermore, spikelets and kernels per spike in K-13-835-3 were significantly higher than those of CN16 in two growing seasons. These results suggest that the desirable genes from P. huashanica were successfully transferred into CN16 background. This translocation line could be used as novel germplasm for high-yield and, eventually, resistant cultivar breeding.