RESUMO
Objective: To find a suitable ecological cultivation measure to solve the problem of root-knot nematode disease of Panax quinquefolium (Panacis Quinquefolii Radix) and the heavy metals accumulating in its roots. Methods: Three-year-old P. quinquefolium was treated with four different combinations of microbial inoculant (MI) and garbage fermentation liquid (GFL) [the joint application of ‘TuXiu’ MI and Fifty potassium MI (TF), the combination use of ‘No. 1′ MI and Fifty potassium MI (NF), ‘Gulefeng’ poly-γ-glutamic acid MI (PGA), GFL], and the untreated control (CK). Here, high-throughput sequencing, ICP-MS and UPLC were employed to systematically characterize changes of microbial diversity and structure composition, heavy metals (As, Cd and Pb) content and ginsenoside content among different treatments. Results: The results revealed that different MIs and GFL could increase the root dry weight of P. quinquefolium, PGA enhanced it by 83.24%, followed by GFL (49.93%), meanwhile, PGA and GFL were able to lessen root-knot nematode disease incidence by 57.25% and 64.35%. The treatment of PGA and GFL can also effectively reduce heavy metals in roots. The As content in GFL and PGA was decreased by 52.17% and 43.48% respectively, while the Cd and Pb contents of GFL and PGA was decreased somewhat. Additionally, the content of total ginsenosides was increased by 42.14% and 42.07%, in response to TF and NF, respectively. Our metagenomic analysis showed that the relative abundance of particular soil microbial community members related to the biocontrol of root-knot nematode disease and plant pathogen (i.e., Chaetomium in NF, Xylari in GFL, and Microascus in PGA), heavy metal bioremediation (Hyphomacrobium in PGA and Xylaria in GFL), and nitrogen fixation (Nordella and Nitrospira in TF) was significantly increased; notably, potential harmful microflora, such as Plectosaphaerella and Rhizobacter, were more abundant in the control group. Conclusion: MI and GFL could improve the quality of P. quinquefolium by modifying its rhizosphere microbial community structure and composition, both of them are beneficial to the development of ecological cultivation of P. quinquefolium.
RESUMO
Objective To study the relationship between nitric oxide within cervical microenvironment and different HPV types as well as the effect of sodium nitroprusside( SNP), a nitric oxide donor, on the proliferation and apoptosis of cervical cancer cell lines. Methods HPV typing test was assessed from 115 women by using high-risk HPV (HR-HPV) 21 typing test and the release of cervical nitric oxide(NO) was assessed as nitrate, nitrite in cervical fluid. Cervical NO was then compared between women showing different HPV types. Proliferation of Caski and HeLa cervical cells was determined by methyl thiazolyl tetrazolium (MTT) assay, cell apoptosis was detected by flow cytometry after 24 hours treated by different final concentration of SNP (0. 125,0. 25,0. 5,1.0 and 2. 0 mmol/L, respectively). The expressions of HPV E6,E7 gene mRNA and p53 protein were detected by SYBR Green Ⅰ quantitative real-time PCR and western blot. Results ( 1 ) The cervical NO release of women with HR-HPV was higher compared to that in HPV negative women [ (47. 6 ± 1.4) μmol/L vs ( 22. 8 ± 0. 3 ) μmol/L; P ＜ 0. 05 ]; but there was no statistical difference between low-risk HPV (LR-HPV) group [ (24. 1 ± 1.2 ) μmol/L] and control group (P ＞0. 05 ). (2)After 24 hours treated by different final concentration of SNP, the results shown that SNP could inhibited the proliferation and increased apoptosis rate in Caski and HeLa cells, in which the concentration of SNP ≥ 1.0 mmol/L , there were significantly different ( P ＜ 0. 05 ), while when SNP ≥2. 0mmol/L, the proliferation of cells inhibited seriously. Treated by SNP ( 1.0 mmol/L ) 24 hours, the expressions of HPV18 E6, E7 mRNA in HeLa cells were reduced from 27. 362 ±0. 191,22. 962 ±0. 053 to19. 181 ±0. 360, 17. 571 ±0. 010 and the protein expression of p53 increased from 1. 17 ±0. 03 to 0. 23 ±0. 05, there were statistically significant differences between adding SNP group and the control group ( P ＜0. 05); but there were no statistically significant differences in HPV16 E6, E7 mRNA and that of p53 in Caski cells( P ＞ 0. 05 ). Conclusions The presence of HR-HPV is associated with an increased release of NO in the human uterine cervix; NO could inhibit the growth and proliferation and enhance the apoptosis of cervical cancer cells, inhibit the expression of HPV18 E6, E7 mRNA in HeLa cells and activate the expression of p53 protein, the mechanism may be due to higher sensitivity of HeLa cells (cervical adenocarcinoma cell) to SNP. The increasing release of NO may play a role in regulating the elimination of HPV in cervical microenvironment, which is a part of mucous membrane immunity.