Two lysin motif extracellular (LysMe) proteins are deployed in rice to facilitate arbuscular mycorrhizal symbiosis.

During arbuscular mycorrhizal (AM) symbiosis, plant innate immunity is modulated to a prime state to allow for fungal colonization. The underlying mechanisms remain to be further explored. In this study, two rice genes encoding LysM extracellular (LysMe) proteins were investigated. By obtaining OsLysMepro:GUS transgenic plants and generating oslysme1, oslysme2 and oslysme1oslysme2 mutants via CRISPR/Cas9 technique, OsLysMe genes were revealed to be specifically induced in the arbusculated cells and mutations in either gene caused significantly reduced root colonization rate by AM fungus Rhizophagus irregularis. Overexpression of OsLysMe1 or OsLysMe2 dramatically increased the colonization rates in rice and Medicago truncatula. The electrophoretic mobility shift assay and dual-luciferase reporter assay supported that OsLysMe genes are regulated by OsWRI5a. Either OsLysMe1 or OsLysMe2 can efficiently rescue the impaired AM phenotype of the mtlysme2 mutant, supporting a conserved function of LysMe across monocotyledonous and dicotyledonous plants. The co-localization of OsLysMe proteins with the apoplast marker SP-OsRAmy3A implies their probable localization to the periarbuscular space (PAS) during symbiosis. Relative to the fungal biomass marker RiTEF, some defense-related genes showed disproportionately high expression levels in the oslysme mutants. These data support that rice plants deploy two OsLysMe proteins to facilitate AM symbiosis, likely by diminishing plant defense responses.

The Rice Qa-SNAREs in SYP13 Subfamily Are Involved in Regulating Arbuscular Mycorrhizal Symbiosis and Seed Fertility.

Qa-SNARE gene SYP132 (isoform α) was previously reported to affect arbuscular mycorrhizal (AM) symbiosis in the legume species Medicago truncatula. In non-legumes especially monocots, it remains unknown whether certain SNARE genes are also involved in AM symbiosis. In this work, we studied a rice orthologous gene OsSYP132, which showed induced expression in mycorrhizal roots and two paralogous genes OsSYP131a and OsSYP131b, which were not induced by the AM fungus Rhizophagus irregularis. After employing CRISPR/Cas9 technique to generate their mutants, the Ossyp131a homozygous mutant T0 plants exhibited a dwarf phenotype and produced no fertile seeds, indicating a required role of this gene in seed fertility. Unlike the case in legume, the Ossyp132 mutants exhibited normal mycorrhizal phenotype, so did the Ossyp131b mutants. In the Ossyp131b Ossyp132 double mutants, however, the colonization rate and arbuscule abundance level decreased markedly, indicating an impaired fungal proliferation ability in rice roots. Such a defect was further confirmed by the reduced expression levels of AM marker genes. Our results in rice therefore demonstrated that while SYP13II members showed evolutionary and induction patterns specific to symbiosis, AM symbiosis is in fact controlled by the combined action of both SYP13I and SYP13II clades, revealing a functional redundancy among SYNTAXIN genes in mutualism.

Dual Roles of OsGH3.2 in Modulating Rice Root Morphology and Affecting Arbuscular Mycorrhizal Symbiosis.

Several angiosperm GRETCHEN HAGEN 3 (GH3) genes, including tomato SlGH3.4 and rice OsGH3.2 are induced during arbuscular mycorrhizal (AM) symbiosis, but their functions remain largely unclear. Recently, tomato SlGH3.4 was suggested to negatively regulate arbuscule incidence via decreasing auxin levels in colonized cells. In this study, by acquiring rice OsGH3.2pro:ß-glucuronidase (GUS) transgenic plants and generating Osgh3.2 mutants via CRISPR/Cas9 technique, the roles of OsGH3.2 in modulating rice root morphology and affecting AM symbiosis were investigated through time course experiments. Unlike SlGH3.4, OsGH3.2 showed asymbiotic expression in rice young lateral roots, and its mutation resulted in a "shallow" root architecture. Such root morphological change was also observed under symbiotic condition and it likely promoted AM fungal colonization, as the mutants exhibited higher colonization levels and arbuscule incidence than wild-type at early stages. Similar to SlGH3.4, OsGH3.2 showed symbiotic expression in cortical cells that have formed mature arbuscules. At late stages of symbiosis, Osgh3.2 mutants showed elongated cortical cells and larger arbuscules than wild-type, indicating elevated auxin level in the colonized cells. Together, these results revealed both asymbiotic and symbiotic roles of OsGH3.2 in modulating rice root architecture and controlling auxin levels in arbusculated cells, which further affected colonization rate and arbuscule phenotype.

Unique bacterial communities associated with components of an artificial aquarium ecosystem and their possible contributions to nutrient cycling in this microecosystem.

In order to better understand the bacterial distribution characteristics in a whole microecosystem, the bacterial communities in different components of an artificial aquarium (i.e., plants, fishes, sand and water) were characterized using high throughput sequencing of bacterial 16S rRNA genes. Across all samples, 2873 operational taxonomic units were identified and assigned to 771 genera in 36 phyla. In a principle coordinate analysis, samples clustered according to their origin, indicating that bacterial communities from the same component were most similar. Further taxonomic analysis revealed that most dominant genera, even those with the similar functions, were biased to one component: Nitrospira and Rhodobacter were mainly abundant in plant samples; Rhodococcus, Serratia, Ralstonia, Sphingobacterium and Pseudomonas were most common in sand samples; Cetobacterium and Aeromonas dominated fish samples; and Flavobacterium, Alpinimonas and Limnobacter were especially common in water samples. Functional predictions performed by PICRUSt and the dominant genera exhibited that bacteria detected in each component could participate in all nutrient cycles in the aquarium. However, those involved in carbon and nitrogen cycling were most common in plant and fish samples, while phosphate metabolism-related pathways were more abundant in sand and water samples. Moreover, the aquarium plants, in association with their bacterial communities might be the most important component in the aquarium, as indicated by their highest bacterial richness and diversity. This study adds to our understanding on the differences in the microbiome of different components and their possible contributions to nutrient cycling in a self-sustaining aquarium.

OsADK1, a novel kinase regulating arbuscular mycorrhizal symbiosis in rice.

Arbuscular mycorrhizal (AM) symbiosis relies on the formation of arbuscules for efficient nutrient exchange between plants and AM fungi. In this study, we identified a novel kinase gene in rice named OsADK1 (Arbuscule Development Kinase 1) that is required for arbuscule development. By obtaining OsADK1pro::GUS transgenic rice plants and also generating Osadk1 mutants via CRISPR/Cas9 technique, OsADK1 was revealed to be specifically induced in the arbusculated cortical cells and mutations in OsADK1 resulted in an extremely low colonisation rate (c. 3%) of rice roots by AM fungus Rhizophagus irregularis. In the mutant roots, the very few observed arbuscules nearly all arrested at an early 'trunk-forming' phase without forming any branches. Increasing the inoculum strength of AM fungus or cocultivation with a wild-type nurse plant did not result in the rescue of the arbuscule phenotype. Transcriptome sequencing of both nursed and un-nursed Osadk1 mutants then revealed that the mutation of OsADK1 could greatly affect the AM symbiotic programme, including many key transcription factors such as RAM1 and WRI5. OsADK1 therefore represents a new rice kinase that is required for arbuscule branching. Its identification opens a new window to explore the elaborate signal transduction pathway that determines arbuscule development during plant-fungus symbiosis.

Effect of L3MBTL3/PTPN9 polymorphisms on risk to alcohol-induced ONFH in Chinese Han population.

PURPOSE: Alcohol-induced osteonecrosis femoral head necrosis (ONFH) is a disease that seriously affects human health. Abnormal expression of L3MBTL3/PTPN9 gene can cause a variety of human diseases. The purpose of this study is to investigate the effect of L3MBTL3/PTPN9 gene polymorphism on the susceptibility of alcohol-induced ONFH in Chinese Han population. METHODS: A total of 308 alcohol-induced ONFH patients and 425 healthy controls were enrolled in this case-control study. Alleles, genotypes, genetic models, haplotypes, and multifactor dimensionality reduction analyses (MDR) based on age-corrected by using odds ratio (OR) and 95% confidence interval (CI) were performed. RESULTS: Our result revealed rs2068957 in the L3MBTL3 gene increased the risk of alcohol ONFH under the recessive model after correction. Besides, we also found that rs75393192 in the PTPN9 gene was a protective site in stratification over 40 years of age and stage. In stratified analysis of necrotic sites, we only found that rs2068957 was associated with increased susceptibility of alcohol-induced ONFH under the co-dominant model and recessive model. Haplotype "GC" in the block (rs76107647|rs10851882 in PTPN9 gene) significantly decreased the susceptibility of alcoholic ONFH. CONCLUSIONS: Our results provide evidence that L3MBTL3/PTPN9 polymorphisms are associated with alcohol-induced ONFH risk in Chinese Han population.

An assessment of the genomics, comparative genomics and cellulose degradation potential of Mucilaginibacter polytrichastri strain RG4-7.

In this study, whole genome sequencing and comparative genomic analyses were performed for Mucilaginibacter polytrichastri RG4-7 and its carboxymethyl cellulose degradation potential was assessed. The results showed that the genome of strain RG4-7 was 5.84 Mb and contained 5019 predicted genes, in which a high proportion of strain-specific genes were related to carbohydrate metabolism. The carboxymethyl cellulose (CMC) degradation and cellulase activity tests revealed the strong cellulose degradation ability, CMCase and ß-glucosidase activity in strain RG4-7. Real-time RT-PCR testing of most cellulose degradation related glycoside hydrolase (GH) families showed that GH9 (OKS85969), GH1 (OKS85832), GH3 (OKS89331 and OKS85615) were significantly up-regulated when strain RG4-7 was inoculated with CMC-Na, which suggested that GH9, GH1 and GH3 might determine its cellulose degradation ability. Certainly, further research need to be done to elucidate cellulose degradation mechanisms in strain RG4-7 in order to develop its industrial application value in lignocellulosic biomass degradation and waste management.

Sphingobacterium athyrii sp. nov., a cellulose- and xylan-degrading bacterium isolated from a decaying fern (Athyrium wallichianum Ching).

Assessment of the bacterial diversity associated with a decaying fern, Athyrium wallichianum Ching, revealed the presence of a novel bacterial strain named M46T. It was Gram-stain-negative, rod-shaped, non-motile and aerobic with cellulose and xylan degradation abilities. Phylogenetic analysis based on 16S rRNA gene sequences showed that strain M46T was affiliated to the genus Sphingobacterium, exhibiting the highest sequence similarity of 97.9 % to Sphingobacterium ginsenosidimutans THG 07T, Sphingobacterium canadense CR11T and Sphingobacterium detergens6.2 ST. Multilocus sequence analysis (MLSA) based on concatenated sequences of the rpoB, cpn60 and 16S rRNA genes showed that strain M46T clustered together with S. canadense CR11T. The genome of strain M46T had a G+C content of 40.6 mol% and chromosome of 6 853 865 bp. Average nucleotide identity (ANI) between strain M46T and S. detergens 6.2 ST and S. siyangense SY1T was 85.1 and 78.1 %, respectively. DNA-DNA relatedness values among strain M46T and other closely related Sphingobacterium species were <70 %. ANI and DNA-DNA relatedness findings strongly supported M46T as a putative novel strain of Sphingobacterium. The predominant fatty acids of strain M46T were iso-C15 : 0, summed feature 3 (C16 : 1ω7c and/or C16 : 1ω6c) and iso-C17 : 0 3-OH, and MK-7 was the dominant isoprenoid quinone. The polar lipid profile of strain M46T contained phosphatidylethanolamine as the dominant component, while minor amounts of phosphoglycolipid, one unidentified aminophospholipid, two unidentified phospholipids and four unidentified lipids were also detected. Based on 16S rRNA gene sequence similarities, MLSA results, genomic characteristics, and phenotypic and biochemotaxonomic analyses, strain M46T is considered to represent a novel species in the genus Sphingobacterium, for which the name Sphingobacterium athyrii sp. nov. is proposed. The type strain is M46T (=CGMCC 1.13466T=JCM 32543T).