The Soil Bacterial Community Structure in a Lactarius hatsudake Tanaka Plantation during Harvest.

Lactarius hatsudake Tanaka is a mycorrhizal edible mushroom with an appealing taste and rich nutrition. It is also a significant food and has medicinal value. In this study, the plantation of L. hatsudake during the harvest period was taken as the research object, and this article explores which bacteria in the soil contribute to the production and growth of L. hatsudake. The soil of the control (CK) and the soil of the mushroom-producing area [including the soil of the base of the mushroom (JT) and the mycorrhizal root soil (JG)] was collected in the plantation. The three sites' bacterial community structure and soil diversity were analyzed using high-throughput sequencing technology, and a molecular ecological network was built. Soil bacteria in the L. hatsudake plantation had 28 tribes, 74 classes, 161 orders, 264 families, 498 genera, and 546 species. The dominant phyla were Proteobacteria and Acidobacteria, and the dominant genera were Burkholderia_Caballeronia_Paraburkholderia, Acidothermus, Bradyrhizobium, Candidatus_Xiphinematobacter, and Granulicella. The α-diversity of soil bacteria in JT was significantly lower than that in JG and CK, and the ß-diversity in JT samples was significantly different from that in JG and CK samples. The size and complexity of the constructed network were smaller in JT samples than in JG and CK samples, and the stability was higher in JT samples than in JG and CK samples. The positive correlation between species in JT samples was dominant. The potential mycorrhizal helper bacteria (MHB) species of L. hatsudake was determined using correlation and differential group analysis. The results support future research on mycorrhizal synthesis, plantation management, and the function of microorganisms in the soil rhizosphere of L. hatsudake.

Assembly and comparative analysis of the complete multichromosomal mitochondrial genome of Cymbidium ensifolium, an orchid of high economic and ornamental value.

BACKGROUND: Orchidaceae is one of the largest groups of angiosperms, and most species have high economic value and scientific research value due to their ornamental and medicinal properties. In China, Chinese Cymbidium is a popular ornamental orchid with high economic value and a long history. However, to date, no detailed information on the mitochondrial genome of any species of Chinese Cymbidium has been published. RESULTS: Here, we present the complete assembly and annotation of the mitochondrial genome of Cymbidium ensifolium (L.) Sw. The mitogenome of C. ensifolium was 560,647 bp in length and consisted of 19 circular subgenomes ranging in size from 21,995 bp to 48,212 bp. The genome encoded 35 protein-coding genes, 36 tRNAs, 3 rRNAs, and 3405 ORFs. Repeat sequence analysis and prediction of RNA editing sites revealed a total of 915 dispersed repeats, 162 simple repeats, 45 tandem repeats, and 530 RNA editing sites. Analysis of codon usage showed a preference for codons ending in A/T. Interorganellar DNA transfer was identified in 13 of the 19 chromosomes, with plastid-derived DNA fragments representing 6.81% of the C. ensifolium mitochondrial genome. The homologous fragments of the mitochondrial genome and nuclear genome were also analysed. Comparative analysis showed that the GC content was conserved, but the size, structure, and gene content of the mitogenomes varied greatly among plants with multichromosomal mitogenome structure. Phylogenetic analysis based on the mitogenomes reflected the evolutionary and taxonomic statuses of C. ensifolium. Interestingly, compared with the mitogenomes of Cymbidium lancifolium Hook. and Cymbidium macrorhizon Lindl., the mitogenome of C. ensifolium lost 8 ribosomal protein-coding genes. CONCLUSION: In this study, we assembled and annotated the mitogenome of C. ensifolium and compared it with the mitogenomes of other Liliidae and plants with multichromosomal mitogenome structures. Our findings enrich the mitochondrial genome database of orchid plants and reveal the rapid structural evolution of Cymbidium mitochondrial genomes, highlighting the potential for mitochondrial genes to help decipher plant evolutionary history.

Diversity and Network Relationship Construction of Soil Fungal Communities in Lactarius hatsudake Tanaka Orchard during Harvest.

Lactarius hatsudake Tanaka is a mycorrhizal edible mushroom with rich economic and nutritional value. Although it is artificially planted, its yield is unstable. Soil fungi, including L. hatsudake, coexist with many other microorganisms and plants. Therefore, complex microbial communities have an influence on the fruiting body formation of L. hatsudake. L. hatsudake and its interactions with the rest of the fungal community over time are not completely understood. In this study, we performed high-throughput sequencing of microorganisms in the basal soil of the fruiting body (JT), mycorrhizosphere soil (JG), and non-mushroom-producing soil (CK) in a 6-year-old L. hatsudake plantation at harvest. The results showed that the soil of the L. hatsudake plantation was rich in fungal communities and a total of 10 phyla, 19 classes, 53 orders, 90 families, 139 genera, and 149 species of fungi were detected. At the phylum level, the major groups were Basidiomycota and Ascomycota. At the genus level, the dominant groups were Lactarius, Trichoderma, Suillus, and Penicillium. Among them, L. hatsudake had an absolute dominant position in the soil fungal community of the plantation, and was the only group of Lactarius in the plantation soil. Penicillium cryptum and Penicillium adametzii were unique to the JT soil sample. Chaetopsphaeria, Myxocephala, Devriesia, and Psathyrella were positively correlated with L. hatsudake. In the constructed fungal network, the total number of nodes were ranked in descending order as JG (441) > CK (405) > JT (399), while the total number of edges were ranked in descending order as CK (1360) > JG (647) > JT (586). Analysis of the fungal assembly process revealed that groups CK and JG have determinative processes that dominated community building, while the JT group exhibited a dominant random process with a 0.60 probability. The results indicated that L. hatsudake was successfully colonized in the plantation soil. During harvest, the CK group exhibited the largest network size and the most complex fungal interactions, while the fungal community structure in the mushroom cultivation zone (JT and JG) was stable and less susceptible to external environmental interference. L. hatsudake affects the fungal community in the soil surrounding its fruiting body.

PARP inhibition synergizes with CD47 blockade to promote phagocytosis by tumor-associated macrophages in homologous recombination-proficient tumors.

AIMS: PARP inhibitors (PARPi) are known to exert anti-tumor effects in patients with BRCA-mutated (BRCAmut) or homologous recombination (HR)-deficient cancer, but recent clinical investigations have suggested that this treatment may also be beneficial in patients with HR-proficient tumors. In this study, we aimed to investigate how PARPi exerts anti-tumor effects in non-BRCAmut tumors. MAIN METHODS: BRCA wild-type, HR-deficient-negative ID8 and E0771 murine tumor cells were treated in vitro and in vivo with olaparib, a clinically approved PARPi. The effects on tumor growth in vivo were determined in immune-proficient and -deficient mice and alterations of immune cell infiltrations were analyzed with flow cytometry. Tumor-associated macrophages (TAMs) were further investigated with RNA-seq and flow cytometry. In addition, we confirmed olaparib's effect on human TAMs. KEY FINDINGS: Olaparib did not affect HR-proficient tumor cell proliferation and survival in vitro. However, olaparib significantly decreased tumor growth in C57BL/6 and SCID-beige mice (defective in lymphoid development and NK cell activity). Olaparib increased macrophage numbers in the tumor microenvironment, and their depletion diminished the anti-tumor effects of olaparib in vivo. Further analysis revealed that olaparib improved TAM-associated phagocytosis of cancer cells. Notably, this enhancement was not solely reliant on the "Don't Eat Me" CD47/SIRPα signal. In addition, compared to monotherapy, the concomitant administration of αCD47 antibodies with olaparib improved tumor control. SIGNIFICANCE: Our work provides evidence for broadening the application of PARPi in HR-proficient cancer patients and paves the way for developing novel combined immunotherapy to upgrade the anti-tumor effects of macrophages.

Mycena subpiligera sp. nov., a Symbiotic Species from China Associated with the Seed Germination of Gastrodia elata.

Mycena subpiligera, a new taxon in sect. Fragilipedes that can strongly enhance the germination efficiency of Gastrodia elata seeds, was discovered in subtropical areas of China. As revealed by a morphological comparison with related Mycena species as well as maximum likelihood (ML) and Bayesian phylogenetic analyses based on sequences of the internal transcribed spacer (ITS) and the large subunit (LSU) regions of nuclear ribosomal RNA, the new taxon can be distinguished from phenotypically similar and phylogenetically related species. Optimal cultural conditions for M. subpiligera basidiomata are reported, and the germination rate of the new species is compared with that of M. citrinomarginata.

A high-quality genome assembly of Lactarius hatsudake strain JH5.

Lactarius hatsudake is a species of Lactarius commonly found in pine forests, is edible with a delicious and nutritious fruiting body, and exhibits medicinal properties. It is an ideal natural multifunctional food with bioactive components including fungal polysaccharides, crude fiber, unsaturated fatty acids, nucleic acid derivatives, various amino acids, and vitamins. However, biological and genomic analyses of this mycorrhizal mushroom are sparse, thereby hindering large-scale cultivation. Previously, we isolated and screened L. hatsudake JH5 strains and have applied our garnered knowledge to the large-scale cultivation of mycorrhizal seedlings. In this study, we produced a high-quality genome assembly of L. hatsudake JH5 by combining Illumina paired-end and PacBio single molecule real-time sequencing, resulting in PacBio single molecule real-time reads of 7.67 Gb and Illumina Pair-End reads of 1,560 Mb. Based on the distribution of k-mer frequencies, the genome size of this strain was estimated to be 63.84 Mb (1.14% heterozygosity). Based on de novo genome assembly, the final genome size was determined to be 76.7 Mb, with scaffold N50 of 223.2 kb and N90 of 54.5 kb, and a GC content of 54.38%. BUSCO assessment showed that genome completeness was 89.0%. The N50 length of the JH5 genome was 43.6% longer than that of the previously published L. hatsudake MG20 genome. This high-quality L. hatsudake genome assembly will facilitate research on the functional genome, molecular breeding, yield enhancement, and sustainability of L. hatsudake cultivation.

Isolation and characterization of antagonistic Paenibacillus polymyxa HX-140 and its biocontrol potential against Fusarium wilt of cucumber seedlings.

OBJECTIVE: The aim of this study is to evaluate the efficacy of the strain Paenibacillus polymyxa HX-140, isolated from the rhizosphere soil of rape, to control Fusarium wilt of cucumber seedlings caused by Fusarium oxysporum f. sp. cucumerinum. RESULTS: Strain HX-140 was able to produce protease, cellulase, ß-1,3-glucanase and antifungal volatile organic compounds. An in vitro dual culture test showed that strain HX-140 exhibited broad spectrum antifungal activity against soil-borne plant pathogenic fungi. Strain HX-140 also reduced the infection of Fusarium wilt of cucumber seedlings by 55.6% in a greenhouse pot experiment. A field plot experiment confirmed the biocontrol effects and further revealed that antifungal activity was positively correlated with inoculum size by the root-irrigation method. Here, inoculums at 106 107 and 108 cfu/mL of HX-140 bacterial suspension reduced the incidence of Fusarium wilt of cucumber seedling by 19.5, 41.1, and 50.9%, respectively. CONCLUSIONS: Taken together, our results suggest that P. polymyxa HX-140 has significant potential in the control of Fusarium wilt and possibly other fungal diseases of cucumber.

Association between hyperhomocysteinemia and stroke with atherosclerosis and small artery occlusion depends on homocysteine metabolism-related vitamin levels in Chinese patients with normal renal function.

This study was conducted to investigate the role of different homocysteine metabolism-related vitamin (HMRV) levels in the correlation between hyperhomocysteinemia (HHCY) and ischemic stroke (IS) subtypes. Three hundred and forty-eight IS patients manifesting different vascular subtypes were subclassified on the basis of HMRV deficiencies. Correlation between HHCY and IS subtypes was investigated in all the subgroups. In this study, HHCY was significantly correlated with the IS subtypes in large artery atherosclerosis (OR 1.126, 95%CI: 1.051 ~ 1.206, P = 0.001) and small artery occlusion (OR 1.105, 95%CI: 1.023 ~ 1.193, P = 0.012). Subgroup analysis revealed a correlation between HHCY and IS subgroup (OR 1.201, 1.178, 95%CI: 1.081 ~ 1.334, 1.058 ~ 1.313, P = 0.001, P = 0.003, respectively) in HMRV deficiency, but not significantly with the IS subgroup in normal HMRV levels. Serum vitamin B12 concentrations are inversely correlated with both IS subtypes in HMRV deficiency subgroups (OR 0.992, 0.995, 95%CI: 0.987 ~ 0.996, 0.991 ~ 0.999, P < 0.001, P = 0.007, respectively), which may contribute to HHCY incidence in these populations. The correlation between HHCY and IS subtypes is affected by HMRV levels in this case-control study. Our findings are helpful to understand the inconsistency in prior homocysteine studies. Serum vitamin B12 levels may play a critical role in HHCY incidence in this Chinese population.Cerebrovascular disease has emerged as the leading cause of disability and mortality in both urban and rural areas of China (Neurology branch of Chinese Medical Association 2015). Ischemic stroke (IS) constitutes 60% to 80% of all cerebrovascular disease (Neurology branch of Chinese Medical Association 2014). Among a variety of risk factors, hyperhomocysteinemia (HHCY) has been closely correlated with IS due to intracranial small-vessel disease and extracranial large-artery disease (Selhub et al. 1995; Eikelboom et al. 2000; Alvarez et al. 2012; Jeon et al. 2014). However, the failure to lower homocysteine (HCY) via homocysteine metabolism-related vitamin (HMRV, including folic acid and vitamin B12 but not vitamin B6 in this study) supplementation to reduce stroke morbidity questions the role of HCY as a risk factor for stroke (Lonn et al. 2006; Hankey et al. 2010). Theoretically, HMRV supplementation merely lowers the incidence of stroke induced by HHCY resulting from HMRV deficiency, whereas HHCY-induced stroke concomitant with normal HMRV levels may be refractory to treatment. The correlation between HCY varying with HMRV levels and IS subtypes is still unclear. In this study, we investigated the impact of variation in HMRV levels on the correlation between HHCY and IS subtypes in 348 acute IS patients with large and small vessel diseases. We sought to determine the factors underlying the conflicting results associated with lowering HCY by HMRV supplementation to reduce stroke incidence.

KLF4 expression enhances the efficacy of chemotherapy drugs in ovarian cancer cells.

KLF4 is a transcriptional factor that can function either as a tumor suppressor or oncogene in cancer based on its cellular context. We recently demonstrated that KLF4 was a tumor suppressor in ovarian cancer cells by inhibiting the epithelial to mesenchymal transition. Here we report that KLF4 expression was downregulated in ovarian cancer tissue compared to normal ovarian tissue, and low KLF4 expression correlated with high risk ovarian carcinoma and poor patient survival. Enforced KLF4 expression by lentiviral transduction sensitized ovarian cancer cells to the effects of the chemotherapy drugs, paclitaxel and cisplatin. Treatment of ovarian cancer cells with APTO-253, a small molecule inducer of KLF4, enhanced the efficacy of both chemotherapy drugs. KLF4 expression mediated by lentiviral vector or induced by APTO-253 resulted in G1 phase arrest in ovarian cancer cells. Our results demonstrate that for the first time that inducing KLF4 expression with APTO-253 is a novel therapeutic strategy for treating ovarian cancer.

[Analysis of relevant factors for recurrence of ovarian endometriosis after conservative laparoscopic surgery].

OBJECTIVE: To analyze relevant factors for recurrence of ovarian endometriosis after conservative surgery.
 METHODS: A cohort study was performed on 310 patients who had performed conservative surgery for ovarian endometriosis. All patients underwent clinical interview. The relevant factors included: age at surgery, clinical symptom and signs, medical history, gynecologic examination, preoperative gravidity, complication, adenomyosis, American Society for Reproductive Medicine (ASRM) scores, post-operative drug therapy, post-operative gravidity and so on. The logistic regression analysis was performed to determine the predictive factors for recurrence of endometriosis.
 RESULTS: The relevant factors by univariate analysis were determined. The history of endometriosis surgery, history of intrauterine operation, tenderness nodule at cal-de-sal, bilateral endometrioma, multilocular cyst, intraoperative ASRM scores, complication of adenomyosis and operation time were the risk factors; whereas pre- and post-operative gravidity, post-operative drug therapy, and age at surgery were the protective factors. Meanwhile, the relevant factors by multivariate analysis were also confirmed. The history of endometriosis surgery, history of intrauterine operation, tenderness nodule at cal-de-sal, bilateral endometrioma, multilocular cyst, and intraoperative ASRM scores were the risk factors; whereas post-operative gravidity, post-operative drug therapy, pre-operative gravidity, and age at surgery were the protective factors.
 CONCLUSION: The risk factors for recurrence of ovarian endometriosis are history of endometriosis surgery, history of intrauterine operation, tenderness nodule at cal-de-sal, bilateral endometrioma, multilocular cyst, intraoperative ASRM scores, whereas the protective factors are pre- and post-operative gravidity, post-operative drug therapy and age at surgery.

Transcriptome Analysis of Ramie (Boehmeria nivea L. Gaud.) in Response to Ramie Moth (Cocytodes coerulea Guenée) Infestation.

The ramie moth Cocytodes coerulea Guenée (RM) is an economically important pest that seriously impairs the yield of ramie, an important natural fiber crop. The molecular mechanisms that underlie the ramie-pest interactions are unclear up to date. Therefore, a transcriptome profiling analysis would aid in understanding the ramie defense mechanisms against RM. In this study, we first constructed two cDNA libraries derived from RM-challenged (CH) and unchallenged (CK) ramie leaves. The subsequent sequencing of the CH and CK libraries yielded 40.2 and 62.8 million reads, respectively. Furthermore, de novo assembling of these reads generated 26,759 and 29,988 unigenes, respectively. An integrated assembly of data from these two libraries resulted in 46,533 unigenes, with an average length of 845 bp per unigene. Among these genes, 24,327 (52.28%) were functionally annotated by predicted protein function. A comparative analysis of the CK and CH transcriptome profiles revealed 1,980 differentially expressed genes (DEGs), of which 750 were upregulated and 1,230 were downregulated. A quantitative real-time PCR (qRT-PCR) analysis of 13 random selected genes confirmed the gene expression patterns that were determined by Illumina sequencing. Among the DEGs, the expression patterns of transcription factors, protease inhibitors, and antioxidant enzymes were studied. Overall, these results provide useful insights into the defense mechanism of ramie against RM.

[Targeted interruption of COX-2 gene by siRNA inhibits the expression of VEGF, MMP-9, the activity of COX-2 and stimulates the apoptosis in eutopic, ectopic endometrial stromal cells of women with endometriosis].

OBJECTIVE: To investigate the effect of targeted interruption of cyclooxygenase-2 (COX-2) gene by small interference RNA (siRNA) on the expression of COX-2, vascular endothelial growth factor (VEGF) and matrix metalloproteinase-9 (MMP-9) in eutopic and ectopic endometrial stromal cells (ESC) with endometriosis, and the effect on the content of 6-keto-prostaglandin-F1α (6-keto-PGF1α, metabolites of COX) and the apoptosis of eutopic and ectopic ESC with endometriosis. METHODS: Ectopic and eutopic ESC from 30 women with endometriosis were isolated and cultured respectively. Then, ESC were classified into three groups: interference group, negative control group and blank control group. ESC in interference group were injected into siRNA transfection complex while ESC in negative control group were injected into negative control transfection complex. ESC from 10 participants without endometriosis were the normal control group. The mRNA and protein expression of COX-2, VEGF, MMP-9 in pre-transfected and post-transfected eutopic and ectopic ESC were detected through real time reverse transcription PCR and western blot. The content of 6-keto-PGF1α was determined by ELISA, the apoptotic cells were detected by flow cytometry. RESULTS: After interruption of COX-2 gene, there were no significant difference in the mRNA and protein expression of COX-2, VEGF and MMP-9 between the negative control group and blank control group (P > 0.05); the mRNA and protein expression of the three genes in interference group were significantly lower than those in negative control group and blank control group (P < 0.05); the mRNA expression of the three genes in interference group of eutopic ESC were 0.87 ± 0.06, 1.76 ± 0.59, 1.04 ± 0.32, in interference group of ectopic ESC were 0.75 ± 0.12, 1.62 ± 0.47, 0.88 ± 0.25, the protein expression of the three genes in interference group of eutopic ESC were 0.457 ± 0.019, 0.500 ± 0.012, 0.361 ± 0.008, in interference group of ectopic ESC were 0.323 ± 0.018, 0.474 ± 0.016, 0.339 ± 0.009; the mRNA and protein expression of the three genes in ectopic ESC had a more reduction than those in eutopic ESC (P < 0.05). The results from ELISA revealed that the content of 6-keto-PGF1α in the normal control group [(17.7 ± 1.9) pg/ml] were significantly lower than those in the blank control group (P < 0.05), the content of 6-keto-PGF1α in ectopic ESC were significantly higher than that in eutopic ESC (P < 0.05), the content of 6-keto-PGF1α in the blank control group of eutopic and ectopic ESC were (32.4 ± 2.6) pg/ml, (38.2 ± 3.7) pg/ml; there was no significant difference in the content of 6-keto-PGF1α between the negative control group and blank control group (P > 0.05); compared with those of negative control group and blank control group, the content of 6-keto-PGF1α in interference group decreased significantly (P < 0.05), the content of 6-keto-PGF1α in interference group of eutopic and ectopic ESC were (17.1 ± 2.4) pg/ml, (20.9 ± 2.7) pg/ml; the content of 6-keto-PGF1α in eutopic ESC had a slightly more reduction than that in ectopic ESC (P > 0.05). The results from flow cytometry displayed that, there was no significant difference in apoptotic cells between the negative control group and blank control group (P > 0.05); compared with those of negative control group and blank control group, more apoptotic cells were detected in interference group and the difference was significant (P < 0.01); the apoptotic cells in ectopic ESC were significantly more than that in eutopic ESC (P < 0.05); the apoptosis rate in interference group of eutopic and ectopic ESC were (33.76 ± 0.06)%, (47.18 ± 0.12)%. CONCLUSIONS: Our results suggested the targeted interruption of COX-2 gene by siRNA effectively inhibited the mRNA and protein expression of COX-2, VEGF and MMP-9 in both eutopic ESC and ectopic ESC with endometriosis, greatly increased the apoptotic rate of cells and obviously reduced the content of 6-keto-PGF1α by inhibiting the activity of COX-2. And the changes in ectopic endometrium were more evident than those in eutopic endometrium.

Diagnostic value of serum CA125, CA19-9 and CA15-3 in endometriosis: A meta-analysis.

AIM: To evaluate the diagnostic value of serum cancer antigen (CA)125, CA19-9 and CA15-3 concentrations in endometriosis. METHODS: Case-control studies evaluating CA125, CA19-9 and CA15-3 and endometriosis, published between January 2000 and November 2014 were retrieved from PubMed(®) and Google Scholar. Standardized mean differences (SMD) and 95% confidence intervals (CI) were calculated. Subgroup analyses were carried out by ethnicity and disease stage (early, stage I/II; advanced, stage III/IV). RESULTS: The analysis included 12 case-control studies (963 cases, 855 controls). CA125 was associated with endometriosis in the overall population (SMD 0.82, 95% CI 0.72, 0.92), Caucasian subgroup (SMD 1.08, 95% CI 0.96, 1.19), and early (SMD 1.20, 95% CI 0.93, 1.48) or advanced disease (SMD 1.29, 95% CI 1.04, 1.55). CA19-9 was associated with endometriosis in the overall population (SMD 0.48, 95% CI 0.24, 0.72), Caucasian subgroup (SMD 0.31, 95% CI 0.07, 0.55), Asian subgroup (SMD 9.65, 95% CI 7.88, 11.42) and advanced disease (SMD 0.60, 95% CI 0.34, 0.87). CA15-3 was significantly associated with advanced disease (SMD 0.47, 95% CI 0.09, 0.84). CONCLUSIONS: Serum CA125 and CA19-9 may represent useful biomarkers for the noninvasive diagnosis of endometriosis.