Integrated Comparative Transcriptome and Weighted Gene Co-Expression Network Analysis Provide Valuable Insights into the Mechanisms of Pinhead Initiation in Chinese Caterpillar Mushroom Ophiocordyceps sinensis (Ascomycota).

The initiation and formation of the "pinhead" is the key node in growth process of Ophiocordyceps sinensis (Chinese Cordyceps). The research on the mechanism of changes in this growth stage is the basis for realizing the industrialization of its artificial cultivation. Clarifying the mechanisms of pinhead initiation is essential for its further application. Here, we performed a comprehensive transcriptome analysis of pinhead initiation process in O. sinensis. Comparative transcriptome analysis revealed remarkable variation in gene expression and enriched pathways at different pinhead initiation stages. Gene co-expression network analysis by WGCNA identified 4 modules highly relevant to different pinhead initiation stages, and 23 hub genes. The biological function analysis and hub gene annotation of these identified modules demonstrated that transmembrane transport and nucleotide excision repair were the topmost enriched in pre-pinhead initiation stage, carbohydrate metabolism and protein glycosylation were specially enriched in pinhead initiation stage, nucleotide binding and DNA metabolic process were over-represented after pinhead stage. These key regulators are mainly involved in carbohydrate metabolism, synthesis of proteins and nucleic acids. This work excavated the candidate pathways and hub genes related to the pinhead initiation stage, which will serve as a reference for realizing the industrialization of artificial cultivation in O. sinensis.

Selection and validation of reference genes for RT-qPCR in ophiocordyceps sinensis under different experimental conditions.

The Chinese caterpillar mushroom, Ophiocordyceps sinensis (O. sinensis), is a rarely medicinal fungus in traditional chinese herbal medicine due to its unique medicinal values, and the expression stability of reference genes is essential to normalize its gene expression analysis. In this study, BestKeeper, NormFinder and geNorm, three authoritative statistical arithmetics, were applied to evaluate the expression stability of sixteen candidate reference genes (CRGs) in O. sinensis under different stress [low temperature (4°C), light treatment (300 lx), NaCl (3.8%)] and different development stages (mycelia, primordia and fruit bodies) and formation of morphologic mycelium (aeriasubstrate, hyphae knot mycelium). The paired variation values indicated that two genes could be enough to accurate standardization exposed to different conditions of O.sinensis. Among these sixteen CRGs, 18S ribosomal RNA (18S rRNA) and beta-Tubulin (ß-TUB) showed the topmost expression stability in O.sinensis exposed to all conditions, while glutathione hydrolase proenzym (GGT) and Phosphoglucose isomerase (PGI) showed the least expression stability. The optimal reference gene in different conditions was various. ß-TUB and Ubiquitin (UBQ) were identified as the two most stable genes in different primordia developmental stage, while phosphoglucomutase (PGM) with elongation factor 1-alpha (EF1-α) and 18S rRNA with UBQ were the most stably expressed for differentially morphologic mycelium stages and different stresses, respectively. These results will contribute to more accurate evaluation of the gene relative expression levels in O.sinensis under different conditions using the optimal reference gene in real-time quantitative PCR (RT-qPCR) analysis.

Transcriptome and metabonomics combined analysis revealed the energy supply mechanism involved in fruiting body initiation in Chinese cordyceps.

Chinese cordyceps was one of most valuable traditional Chinese medicine fungi. To elucidate the molecular mechanisms related to energy supply mechanism involved in the initiation and formation of primordium in Chinese cordyceps, we performed the integrated metabolomic and transcriptomic analyses of it at pre-primordium period, primordium germination period and after-primordium period, respectively. Transcriptome analysis showed that many genes related to 'starch and sucrose metabolism', 'fructose and mannose metabolism', 'linoleic acid metabolism', 'fatty acids degradation' and 'glycerophospholipid metabolism' were highly up-regulated at primordium germination period. Metabolomic analysis showed many metabolites regulated by these genes in these metabolism pathways were also markedly accumulated at this period. Consequently, we inferred that carbohydrate metabolism and ß-oxidation pathway of palmitic acid and linoleic acid worked cooperatively to generate enough acyl-CoA, and then entered TCA cycle to provide energy for fruiting body initiation. Overall, our finding provided important information for further exploring the energy metabolic mechanisms of realizing the industrialization of Chinese cordyceps artificial cultivation.

Uncovering Gene Expression Profiles of Three Morphologic Mycelium Forms of the Chinese Caterpillar Mushroom Ophiocordyceps sinensis (Ascomycota) Using High-Throughput Sequencing.

The asexual form of Ophiocordyceps sinensis has been controversial, but various morphologic mycelium appeared when O. sinensis was cultured under experimental conditions. To explore the generation mechanism of morphologic mycelium, developmental transcriptomes were analyzed from three kinds of mycelium (aerial mycelium, hyphae knot, and substrate mycelium). The results showed that diameter and morphology of these three kinds of mycelium were obviously different. KEGG functional enrichment analysis showed that the differential expressed genes (DEGs) of substrate mycelium were enriched in ribosomes and peroxisomes, indicating that prophase culture was rich in nutrients and the metabolism of substrate mycelium cells was vigorous in the stage of nutrient absorption. The up-DEGs of hyphae knot were mainly enriched in the oxidative phosphorylation pathway, indicating that oxidative phosphorylation was the main energy source for mycelium formation in the stage of nutrient accumulation and reproductive transformation. The up-DEGs of aerial mycelium were mainly enriched in the synthesis and degradation pathways of valine, leucine, and isoleucine, suggesting that the occurrence of aerial mycelium was related to amino acid metabolism at the later stage of culture, and nutritional stress accelerated the reproduction of asexual spores. In addition, the important roles of mycelium formation related genes were verified by combined analysis of qRT-PCR and transcriptome sequencing. Collectively, this study will provide theoretical guidance for inhibiting the occurrence of aerogenous mycelium and promoting the development of mycelium into pinhead primordia in the culture of O. sinensis in the future.

Transcriptome Sequencing to Identify Candidate Genes for Mycelium Period Metabolism of Chinese Caterpillar Mushroom, Ophiocordyceps sinensis (Ascomycetes), Exposed to Light Treatment.

The Chinese caterpillar mushroom, Ophiocordyceps sinensis, is rare in traditional Chinese herbal medicine. It was reported that the development of ascospores in sexual stage of Chinese Cordyceps requires certain UV irradiation, but whether light has effect on the asexual stage of O. sinensis is unclear. It is important and necessary to identify the candidate genes involved in asexual stages (mycelium period) metabolism of O. sinensis exposed to light treatment. In this study, the isolated and purified monoascospore strains from O. sinensis were treated with 10 days light (L10) and dark as control. Transcriptome sequencing (RNA-seq) was conducted to investigate the effect of light treatment on O. sinensis at the gene level and the changes in various metabolic pathways. The results showed that the colony surface was covered with villous aerial hyphae with a yellow circular mycelium ribbon in the center of the colony, and the content of polysaccharides, urea, adenosine, and cordycepin were significantly enhanced by L10 treatment. Kyoto Encyclopedia of Genes and Genomes (KEGG) functional annotation analysis showed that cell structure and catalytic metabolism were the significant items classified, and crucial genes affecting the anabolic pathways of polysaccharides, urea, adenosine, and cordycepin were also identified. In addition, the important roles of photoreceptor genes CRYD, WC-1, and FRQ were verified by combined analysis of qRT-PCR and transcriptome sequencing. Collectively, this study would be helpful to better understand the influence of light on the asexual stage of Chinese Cordyceps and provide a preliminary light treatment reference for Cordyceps artificial cultivation.

Comparison of the accelerometer-based navigation system with conventional instruments for total knee arthroplasty: a propensity score-matched analysis.

BACKGROUND: The accelerometer-based navigation (ABN) system is an emerging navigation system for total knee arthroplasty (TKA). This study aimed to determine whether the ABN system could improve the accuracy of mechanical alignment, component positioning, and short-term clinical outcomes for TKA when compared to conventional instruments (CON). METHODS: A total of 204 patients were selected and divided into two groups (CON: 135, ABN: 69) after applying the inclusion and exclusion criteria. Then, 1:1 propensity score matching was performed for age, gender, body mass index, hip-knee-ankle angle (HKA), Knee Society Score (KSS), Western Ontario and McMaster Universities (WOMAC) score, and follow-up times. A total of 82 consecutive patients (82 knees) underwent total knee arthroplasty using ABN (n = 41) or CON (n = 41) were enrolled in this study. The postoperative HKA, frontal femoral component (FFC) angle, frontal tibial component (FTC) angle, lateral femoral flexion (LFF) angle, and lateral tibial component (LTC) angle were compared between the two groups to evaluate mechanical alignment and component positioning. Additional clinical parameters, including haemoglobin reduction, the KSS, and the WOMAC score, were assessed at the final follow-up (the mean follow-up period was 20.9 months in the CON group and 21.2 months in the ABN group). RESULTS: The ABN group had a significantly improved mean absolute deviation of HKA (P = 0.033), FFC (P = 0.004), FTC (P = 0.017), LFF (P = 0.023), and LTC (P = 0.031) compared to those of the CON group. The numbers of FFCs and LTCs within 3° were significantly different (P = 0.021, P = 0.023, respectively) between the two groups. However, no differences in the numbers of FTCs within 3° (P = 0.166) and LFF within 3° (P = 0.556) were found. The ABN group had a significantly higher KS function score (P = 0.032), and the pain and stiffness scores were significantly different (P = 0.034, P = 0.020, respectively) between the two groups. Moreover, the ABN system could reduce hidden blood loss postoperatively. However, no difference was found in the KS knee score and the total WOMAC score between the two groups. CONCLUSION: This study demonstrates that ABN system improved TKA mechanical alignment and component positioning and decreased the hidden blood loss postoperatively compared to conventional instruments. However, no significant differences were found in short-term clinical outcomes between ABN and conventional instruments at the final follow-up. However, whether this system contributes to revision rates and long-term clinical outcomes requires further study.

A mild one-step process from graphene oxide and Cd2+ to a graphene-CdSe quantum dot nanocomposite with enhanced photoelectric properties.

Good connections: A graphene-CdSe quantum dot (QD) nanocomposite is prepared through a one-step hydrothermal method using graphene oxide (GO), Cd(CH(3)COO)(2), Na(2) SeSO(3), and aminoethanethiol (AET). The bifunctional AET acts not only as a covalent linker but also as a reductant to transform GO into graphene. The photoactive graphene-QD nanocomposite exhibits a significantly higher photocurrent compared to the QDs, GO or the graphene substrate under illumination.