The mycorrhizal fungi of Cymbidium promote the growth of Dendrobiumofficinale by increasing environmental stress tolerance.

Dendrobium officinale is a medicinal herbal plant with important health care value and high demand. Due to its slow growth and scarcity in nature, its yield depends on intensified cultivation while biotic and abiotic stresses were important factors that causes production loss. Orchidaceae can form association with rhizoctonias collectively, and studies have found that some orchids showed a high level of strain-species specificity to orchid mycorrhizal fungi (OMF), yet the specificity of OMF on D. officinale needs to explored. In this study, the effects on D. officinale of four OMF isolated from Cymbidium were tested. The obviously higher mass yield of the treated plants in medium and pots indicated the growth promotion effect of the fungi. Furthermore, an abiotic stress test indicated stronger drought tolerance among the treated plants. For the biotic stress test, two root rot pathogens, Fusarium solani and Fusarium graminearum , were isolated and identified from root rot of D. officinale. In an in vitro inhibition test, the four OMF could resist the growth of these pathogens. In vivo studies showed that these four OMF could improve the survival rate and fresh weight and decrease the root rot rate of pathogen-inoculated seedlings. The four OMF namely; Hyphomycete sp., Umbelopsis sp., Ceratorhiza sp. and Ceratorhiza sp. are compatible strains for improving the growth rate of D. officinale by increasing its environmental stress tolerance, providing an effective way to supply resources through artificial reproduction.

Bursaphelenchus xylophilus: An Important Pathogenic Factor of Pine Wilt Disease and Its Relationship with Bursaphelenchus mucronatus.

Pine wilt disease is the most devastating pine disease caused by Bursaphelenchus xylophilus. Bursaphelenchus mucronatus is morphologically similar to B. xylophilus and geographically overlaps in its distribution. Although interspecific hybridization of the two nematodes has been performed in vitro, the dynamic regularity of hybrid formation and its risk in forests has not been well evaluated. In this study, a hybrid of B. xylophilus and Bursaphelenchus mucronatus mucronatus was identified in the laboratory and fields by molecular markers. The heterozygosity of ITS-5.8S loci for identification was unstable in the hybrid population, and the allele inherited from B. m. mucronatus was lost over several generations. We also provided evidence that hybrids existed in some new epidemic areas, while old epidemic areas were usually dominated by B. xylophilus. Hybrids could be generated when B. m. mucronatus was invaded by B. xylophilus, and the pathogenicity of the hybrids was similar to that of B. xylophilus. These findings may improve the understanding of the natural hybridization between B. xylophilus and B. m. mucronatus and pathogenic variation in pine wilt disease, providing new insights for future studies on disease detection, transmission, and quarantine.

Isolation and identification of beneficial orchid mycorrhizal fungi in Bletilla striata (Thunb.) Rchb.f.(Orchidaceae).

Orchid mycorrhizal fungi (OMF) are essential for orchids growth. Bletilla striata (Thunb.) Rchb.f.(Orchidaceae) has high ornamental and medicinal value. Beneficial OMF isolation is crucial to improve the survival rate of B. striata tissue culture and transplanting. In this study, we isolated and identified the beneficial OMF in B. striata from the roots of sterilized wild B. striata seedlings by culturing in four different mediums. The germination states of B. striata seeds inoculated with diverse OMF were classified and calculated. Fresh and dry weight increments of B. striata seedlings inoculated with diverse OMF were recorded after 90 d of culturing on 1/2 MS medium. ITS sequences of beneficial fungi were amplified by PCR and taxonomically identified using BLAST against the GenBank nucleotide database. Ten kinds of OMF strains were isolated from B. striata and named R1 to R10. R6 significantly promoted B. striata seeds germination (p < .01). R3 and R6 significantly promoted both the fresh and dry weight increments of B. striata seedlings (p < .05). The ITS sequence of R6 was most similar to the sequence of Serendipita. R3 was identified as Schizothecium fimbriatum by 100% ITS identity. R6 and R3 were beneficial OMF in B. striata.

Expression and regulation of miR-449a and AREG in cerebral ischemic injury.

Rodent focal ischemia models are widely used to mimic and examine human strokes. To the best of our knowledge, no investigation has systematically examined the expression changes of microRNA (miR)-449a and Amphiregulin (AREG) as well as their biological relationship during middle cerebral artery occlusion (MCAO) and oxygen and glucose deprivation/reperfusion (OGD/R). The present study examined the histological and behavioral outcomes of MCAO and the function of miR-449a and AREG in cerebral ischemic injury. Rats were subjected to 2 h MCAO, which was followed by reperfusion. miR-449a and AREG were examined in the injury tissues of MCAO rats and the OGD/R cell line by reverse transcription-quantitative polymerase chain reaction. Protein expressions of AREG in the injury tissues of MCAO rats was measured using an immunohistochemistry and the protein expression levels of AREG, epidermal growth factor receptor (EGFR), phosphatidylinositol 3-kinase (PI3K)/ protein kinase B (Akt) and the phosphorylation level of Akt (p-Akt) were analyzed by western blotting. Cell apoptosis was examined following the knock down and subsequent overexpression of AREG in a human OGD/R neuronal cell line by small interfering RNAs (siRNAs) and plasmid transfection. Luciferase reporter assays were used to validate the target of miR-449a. The expression changes and regulatory mechanisms of miR-449a and AREG in an ischemia/reperfusion (I/R) injury model were examined in vivo and in vitro. The neurological deficit score, brain edema volume, cerebral infarct area, and the number of apoptosis cells in ischemic rats were all markedly elevated, than that in the control rats. The expression of miR-449a was decreased and AREG was increased in the MCAO rats and human OGD/R neuronal cell line. miR-449a inhibition or AREG overexpression in OGD/R cells resulted in a significant decrease in apoptotic cells, and AREG was revealed to be one of the direct targets of miR-449a. Molecular recovery was observed following transfection with miR-449a mimics and AREG knockdown in an OGD/R model in vitro. The present study demonstrated that miR-449a was downregulated while AREG was upregulated in cerebral ischemic injury, and the recovery of neurological function can be obtained following the overexpression of miR-449a and the knockdown of AREG in an I/R injury model. miR-449a functions in ischemic stroke via directly targeting AREG. These findings suggest a novel mechanism involving in cerebral I/R injury model and may aid investigators in gaining a deeper understanding of strokes in a clinical setting.

Leaf morphology and ultrastructure responses to elevated O3 in transgenic Bt (cry1Ab/cry1Ac) rice and conventional rice under fully open-air field conditions.

BACKGROUND: Elevated tropospheric ozone severely affects not only yield but also the morphology, structure and physiological functions of plants. Because of concerns regarding the potential environmental risk of transgenic crops, it is important to monitor changes in transgenic insect-resistant rice under the projected high tropospheric ozone before its commercial release. METHODOLOGY/PRINCIPAL FINDINGS: Using a free-air concentration enrichment (FACE) system, we investigated the changes in leaf morphology and leaf ultrastructure of two rice varieties grown in plastic pots, transgenic Bt Shanyou 63 (Bt-SY63, carrying a fusion gene of cry1Ab and cry1Ac) and its non-transgenic counterpart (SY63), in elevated O3 (E-O3) versus ambient O3 (A-O3) after 64-DAS (Days after seeding), 85-DAS and 102-DAS. Our results indicated that E-O3 had no significant effects on leaf length, leaf width, leaf area, stomatal length and stomatal density for both Bt-SY63 and SY63. E-O3 increased the leaf thickness of Bt-SY63, but decreased that of SY63. O3 stress caused early swelling of the thylakoids of chloroplasts, a significant increase in the proportion of total plastoglobule area in the entire cell area (PCAP) and a significant decrease in the proportion of total starch grain area in the entire cell area (SCAP), suggesting that E-O3 accelerated the leaf senescence of the two rice genotypes. Compared with SY63, E-O3 caused early swelling of the thylakoids of chloroplasts and more substantial breakdown of chloroplasts in Bt-SY63. CONCLUSIONS/SIGNIFICANCE: Our results suggest that the incorporation of cry1Ab/Ac into SY63 could induce unintentional changes in some parts of plant morphology and that O3 stress results in greater leaf damage to Bt-SY63 than to SY63, with the former coupled with higher O3 sensitivity in CCAP (the proportions of total chloroplast area in the entire cell area), PCAP and SCAP. This study provides valuable baseline information for the prospective commercial release of transgenic crops under the projected future climate.

The composition of root exudates from two different resistant peanut cultivars and their effects on the growth of soil-borne pathogen.

The high incidence of various soil-borne diseases in the monoculture field of peanut is a major production constraint in the red soil regions of southern China. The peanut root exudates are generally thought to play an important role in regulating soil-borne pathogens. The responses of the soil-borne pathogens, Fusarium oxysporum and F. solani to the peanut root exudates were studied using one susceptible cultivar Ganhua-5 (GH) and one mid-resistant cultivar Quanhua-7 (QH) as the test materials. The components and contents of the amino acids, sugars and phenolic acids in the peanut root exudates were determined. The results demonstrated that the root exudates from both susceptible and mid-resistant cultivars significantly promoted the spore germination, sporulation and mycelial growth of soil-borne pathogens, F. oxysporum, F. solani compared with the control. The extent of the stimulation was depended on the strains of the Fusarium tested, and gradually increased with the increased concentrations of peanut root exudates. HPLC analysis showed that the contents of sugars, alanine, total amino acids in the root exudates of GH were significantly higher than that in QH, whereas the contents of p-hydroxybenzoic acid, benzoic acid, p-coumaric acid and total phenolic acids were significantly lower than that in QH. Results of the study suggested that the differences in the root exudates from the different peanut cultivars were considered to regulate the wilt-resistance mechanism in the rhizosphere of peanut. The results are therefore crucial important to illustrate the mechanism of peanut replanted obstacle, and to develop its control techniques in the red soil regions of southern China.

Root exudates of transgenic cotton and their effects on Fusarium oxysporum.

The components of the root exudates from two transgenic insect-resistant cotton lines and their parental cotton lines, and their effects on the growth of Fusarium oxysporum were investigated. The results demonstrated that the resistance of transgenic insect-resistant cotton to F. oxysporum was significantly reduced compared with their parental lines. Likewise, the root exudates from transgenic insect-resistant cotton significantly promoted the spore germination and mycelial growth of cotton F. oxysporum. The types of compounds found in the root exudates of transgenic insect-resistant cotton were similar to those of the parental cotton, but the composition and relative content of the compounds were different. The type and content of the fatty acids and esters were significantly reduced in the root exudates of the transgenic insect-resistant cotton, as were certain specific materials, whereas several alkanes were increased. The inhibition of the soil-borne pathogen F. oxysporum caused by the root exudates from the transgenic insect-resistant cotton was decreased compared with the parental cotton. This result provides a scientific basis for the decline in disease resistance in transgenic insect-resistant cotton.

Field trials to evaluate effects of continuously planted transgenic insect-resistant cottons on soil invertebrates.

Impacts on soil invertebrates are an important aspect of environmental risk assessment and post-release monitoring of transgenic insect-resistant plants. The purpose of this study was to research and survey the effects of transgenic insect-resistant cottons that had been planted over 10 years on the abundance and community structure of soil invertebrates under field conditions. During 3 consecutive years (2006-2008), eight common taxa (orders) of soil invertebrates belonging to the phylum Arthropoda were investigated in two different transgenic cotton fields and one non-transgenic cotton field (control). Each year, soil samples were taken at four different growth stages of cotton (seedling, budding, boll forming and boll opening). Animals were extracted from the samples using the improved Tullgren method, counted and determined to the order level. The diversity of the soil fauna communities in the different fields was compared using the Simpson's, Shannon's diversity indices and evenness index. The results showed a significant sampling time variation in the abundance of soil invertebrates monitored in the different fields. However, no difference in soil invertebrate abundance was found between the transgenic cotton fields and the control field. Both sampling time and cotton treatment had a significant effect on the Simpson's, Shannon's diversity indices and evenness index. They were higher in the transgenic fields than the control field at the growth stages of cotton. Long-term cultivation of transgenic insect-resistant cottons had no significant effect on the abundance of soil invertebrates. Collembola, Acarina and Araneae could act as the indicators of soil invertebrate in this region to monitor the environmental impacts of transgenic plants in the future.

[Effects of transgenic insect-resistant cotton root exudates on the growth of Verticillium dahliae Kleb].

The decline of disease resistance of transgenic insect-resistant cotton has become one of the important factors restricting China cotton production. With two transgenic insect-resistant cotton varieties and their parental conventional cotton lines as test materials, the effects of their root exudates on the spore germination and mycelial growth of Verticillium dahliae Kleb were studied, and the components and contents of the amino acids and sugars in the exudates were determined. Comparing with their parental lines, the two transgenic insect-resistant varieties had a declined resistance against V. dahliae, and their root exudates had promotion effects on V. dahliae spore germination and mycelial growth. The root exudates of transgenic cotton Zhong-41, compared with those of its parent line Zhong-23, contained two additional amino acids Met and Lys, and the contents of Asp, Glu, Ser, Ala, Val, Leu, and Tyr were significantly higher. Though the components of amino acids in the root exudates of Simian-3 and transgenic cotton GK-12 were the same, the Tyr, Val, and Leu contents in the root exudates of transgenic cotton GK-12 were significantly lower. Four kinds of sugar were detected in the root exudates of Zhong-41, while only glucose was detected in the root exudates of Zhong-23. These four kinds of sugar were also detected in the root exudates of Simian-3 and GK12, but their contents differed significantly.

The effect of root exudates from two transgenic insect-resistant cotton lines on the growth of Fusarium oxysporum.

The attenuation of disease resistance in transgenic insect-resistant cotton has become one of the important factors restricting cotton production in China. Two transgenic insect-resistant cotton lines and their parental conventional cotton lines were used as the testing materials. The effects of root exudates of these cotton lines on the spore germination and mycelial growth of Fusarium oxysporum were studied and the components, contents of amino acids and sugars were determined. The results showed that the resistance of the two insect-resistant cotton lines to F. oxysporum was inferior to the parental lines, and that their root exudates promoted fungal spore germination and mycelial growth. Considerable differences in the components and contents of both, amino acids and sugars were found between the root exudates of transgenic cotton lines and their parental lines, where the disease indices were highly correlated with the total amount of sugars in the root exudates.

[Identification of markers linked to resistance locus of Marssonina leaf spot in poplars by bulked segregant analysis(BSA)].

DNA markers linked to resistance locus of Marssonina leaf spot in poplars were found by bulked segregant analysis(BSA). The bulks consisted of individual with a extreme phenotype taken from a population of 91 F1 clones,which is a progeny of Populus deltoides Bartr.cv."Lux"(I-69/55)(Resistance) and P.euramericana cv.I-45(Susceptible). Out of 114 RAPD primers, four markers showed polymorphisms between the resistance-bulk and the susceptible-bulk.By using selective genotype linkage analysis,OPAI17-1550 and OPAI13-900 were found linked to the resistance locus. The genetic distances between the two markers and the resistance locus were 29.9cM and 37.4cM,respectively.