Potential Anti-Candida albicans Mechanism of Trichoderma Acid from Trichoderma spirale.

Candida albicans is the main causal pathogen of fungal infections in human beings. Although diverse anti-C. albicans drugs have been explored, the drug resistance and side effects of these drugs are intensifying. Thus, it is urgent to explore new anti-C. albicans compounds from natural products. In this study, we identified trichoderma acid (TA), a compound from Trichoderma spirale with a strong inhibitory effect on C. albicans. Transcriptomic and iTRAQ-based proteomic analyses of TA-treated C. albicans in combination with scanning electronic microscopy and reactive oxygen species (ROS) detection were performed to investigate the potential targets of TA. The most significant differentially expressed genes and proteins after TA treatment were verified through Western blot analysis. Our results revealed that mitochondrial membrane potential, endoplasmic reticulum, ribosomes in the mitochondria, and cell walls were disrupted in TA-treated C. albicans, leading to the accumulation of ROS. The impaired enzymatic activities of superoxide dismutase further contributed to the increase in ROS concentration. The high concentration of ROS led to DNA damage and cell skeleton destruction. The expression levels of Rho-related GTP-binding protein RhoE (RND3), asparagine synthetase (ASNS), glutathione S-transferase, and heat shock protein 70 were significantly up-regulated in response to apoptosis and toxin stimulation. These findings suggest that RND3, ASNS, and supereoxide dismutase 5 are the potential targets of TA, as further demonstrated through Western blot analysis. The combination of transcriptomic, proteomic, and cellular analyses would provide clues for the anti-C. albicans mechanism of TA and the defensive response mechanism of C. albicans. TA is thus recognized as a promising new anti-C. albicans leading compound that alleviates the hazard of C. albicans infection in human beings.

An ACE2-Based Decoy Inhibitor Effectively Neutralizes SARS-CoV-2 Omicron BA.5 Variant.

The recently circulating SARS-CoV-2 Omicron BA.5 is rampaging the world with elevated transmissibility compared to the original SARS-CoV-2 strain. Immune escape of BA.5 was observed after treatment with many monoclonal antibodies, calling for broad-spectrum, immune-escape-evading therapeutics. In retrospect, we previously reported Kansetin as an ACE2 mimetic and a protein antagonist against SARS-CoV-2, which proved potent neutralization bioactivity on the Reference, Alpha, Beta, Delta, and Omicron strains of SARS-CoV-2. Since BA.5 is expected to rely on the interaction of the Spike complex with human ACE2 for cell entry, we reasonably assumed the lasting efficacy of the ACE2-mimicking Kansetin for neutralizing the new SARS-CoV-2 variant. The investigation was accordingly performed on in vitro Kansetin-Spike binding affinity by SPR and cell infection inhibition ability with pseudovirus and live virus assays. As a result, Kansetin showed dissociation constant KD and half inhibition concentration IC50 at the nanomolar to picomolar level, featuring a competent inhibition effect against the BA.5 sublineage. Conclusively, Kansetin is expected to be a promising therapeutic option against BA.5 and future SARS-CoV-2 sublineages.

Efficacy and safety of dual antiplatelet therapy after percutaneous coronary drug-eluting stenting: A network meta-analysis.

BACKGROUND: To evaluate the efficacy and safety of dual antiplatelet regimens after coronary drug-eluting stenting by network meta-analysis (NMA). METHODS: PubMed, The Cochrane Library, Embase, and Web of Science databases were electronically searched to collect randomized controlled trials (RCTs) of the comparison of different dual antiplatelet regimens after coronary drug-eluting stenting from inception to September 1st, 2021. Two reviewers independently screened literature, extracted data, and assessed the risk bias of included studies. Stata 16.0 software was used for NMA. RESULTS: A total of 27 RCTs involving 79,880 patients were included. The results of NMA: in terms of myocardial infarction (MI), other 3 interventions were higher than the long-term dual antiplatelet therapy (L-DAPT) (the standard dual antiplatelet therapy [Std-DAPT] [odds ratio [OR] = 1.82, 95%confidence interval [CI]: 1.49-2.21), the aspirin monotherapy after short-term dual antiplatelet therapy (S-DAPT + As) (OR = 2.06, 95%CI: 1.57-2.70), the P2Y12 inhibitor monotherapy after short-term dual antiplatelet therapy (S-DAPT + P2Y12) (OR = 1.71, 95%CI: 1.29-2.28)]. In terms of stent thrombosis, other 3 interventions were higher than L-DAPT [Std-DAPT (OR = 2.18, 95%CI: 1.45-3.28), S-DAPT + As (OR = 2.32, 95%CI: 1.52-3.54), S-DAPT + P2Y12 (OR = 2.31, 95%CI: 1.22-4.36)]. There was no statistically significant difference among the 4 interventions in prevention of stroke and all-cause mortality (P > .05). In terms of cardiovascular and cerebrovascular adverse events, other 3 interventions were higher than L-DAPT (Std-DAPT [OR = 1.28, 95%CI: 1.12-1.45], S-DAPT + As [OR = 1.27, 95%CI: 1.09-1.48], S-DAPT + P2Y12 [OR = 1.24, 95%CI: 1.01-1.52]). In terms of safety, bleeding rate of other 3 interventions were lower than L-DAPT (Std-DAPT [OR = 0.67, 95%CI: 0.52-0.85], S-DAPT + As [OR = 0.51, 95%CI: 0.39-0.66], S-DAPT + P2Y12 [OR = 0.36, 95%CI: 0.26-0.49]). Two interventions were lower than L-DAPT (S-DAPT + As [OR = 0.77, 95%CI: 0.65-0.90], S-DAPT + P2Y12 [OR = 0.54, 95%CI: 0.44-0.66]). S-DAPT + As was higher than L-DAPT (OR = 1.42, 95%CI: 1.10-1.83). CONCLUSIONS: S-DAPT + P2Y12 has the lowest bleeding risk, while L-DAPT has the highest bleeding risk. In the outcome of MI, stent thrombosis, and cardiovascular and cerebrovascular adverse events, L-DAPT has the best efficacy. In the outcome of stroke and all-cause mortality, the 4 interventions were equally effective.

First Report of Postharvest Stem End Rot of mango Fruit (Mangifera indica) Caused by Diaporthe pseudomangiferae in China.

Mango (Mangifera indica L.) is considered one of the most important tropical or subtropical fruit crops (Nelson et al.2008). China is the second-largest producer of mango (Kuhn et al. 2017). In June 2021, postharvest stem-end rot disease was observed on Narcissus mango (about 20% of the fruits showed similar symptoms of infections) in local agricultural market of Guangzhou, China. Black rot symptomatic lesions were observed on the fruit surface, which initially started from the stem end and progresses into decay, turning brown. To isolate and identify the pathogen, small pieces (3-5 mm2) were excised from the lesion margins of the fruits (n=54), which were surface sterilized by 1% NaOCl (1 min), 70% ethanol (30 s) and then washed twice with sterile distilled water. After sterilization, the tissues were cultured on potato dextrose agar (PDA). Three morphologically similar isolates (SXM-1/2/3) were obtained and the representative isolate SXM-1 was analyzed. Colonies surface initially had white-gray moderate aerial mycelia, in reverse umber with patches of pale luteous to luteous. On malt extract agar (MEA) surface dirty white, reverse greyish sepia with patches of sienna. Conidiomata pycnidia, black, erumpent to superficial on PDA, globose with neck, ostiole exuding cream conidial droplets; Conidiophores hyaline, smooth, 1-3-septate, branched, densely aggregated, cylindrical, straight to sinuous, 22-42 × 2.8-3.7 µm. Alpha conidia (n = 50) aseptate, hyaline, smooth, fusiform to somewhat short cylindrical, 3.2-11 ×1.3-3.5 µm. Beta conidia (n = 30) hyaline, smooth, curved or hamate 14.8-33.6 × 1.1-2.6 µm. According to morphological characterization, the representative isolate SXM-1 was similar to Diaporthe pseudomangiferae CBS 101339 (Gomes et al. 2013). For molecular identification, the internal transcribed spacer (ITS) region, histone H3 (HIS) and ß-tubulin (TUB) genes (White et al. 1990; Carbone et al. 1999; Glass et al.1995) were amplified and sequenced, which were deposited in GenBank (ON243823, ON254656, ON254655). BLASTN analysis revealed that DNA sequences of the isolates (SXM-1/2/3) showed 99% identity with those of D. pseudomangiferae (MG576128.1, KC344149, MN329124.1), respectively. A phylogenetic tree analysis based on the concatenated sequences confirmed the isolates as D. pseudomangiferae. Pathogenicity tests were made with the representative isolate SXM-1. Healthy fruits were inoculated with 5 mm mycelial discs of the representative isolate SXM-1 after being wounded with a needle or non-wounded, control fruits were inoculated with sterilized PDA plugs. All inoculated and control fruits were incubated in the dark at 26°C for 7 days post-inoculation. Control fruits remained asymptomatic, whereas inoculated fruits were dark brown necrotic lesions with a roughly circular shape around the inoculation sites. Pathogenicity tests were performed in triplicate. The pathogenic isolates were successfully reisolated, thus confirming Koch's postulates. D. pseudomangiferae was associated with fruit peel of mango in Mexico and the Dominican Republic, and it has also been reported to cause inflorescence rot, rachis canker, and flower abortion in mango (Gomes et al. 2013; Serratodiaz et al. 2014). To our knowledge, this is the first report of D. pseudomangiferae causing postharvest stem-end rot of mango fruits in China. This finding suggests that D. pseudomangiferae is a potential problem for mango fruit production in China, and it is important to establish an adequate and effective control management of this disease. References: Nelson, S. C. 2008.Mango Anthracnose (Colletotrichum gloeosporiodes). Publication PD-48. Cooperative Extension Service, College of Tropical Agriculture and Human Resources, University of Hawai'i at Manoa, U.S.A. Kuhn, D. N., et al. 2017. Front. Plant Sci. 8:577. White, T. J., et al. 1990. Page 315 in: PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego. Carbone, I., et al. 1999. Mycologia. 91:553. Glass, N. L., et al. 1995. Appl. Environ. Gomes R. R., et al. 2013. Persoonia. 1:31. Serratodiaz, L. M., et al. 2014. Plant Dis. 98:1004. * These authors contributed equally to this work and should be considered co-first authors. The authors declare no conflict of interest. Keywords: Stem-end rot, Diaporthe pseudomangiferae, Mango, China.

Disclosure of the Molecular Mechanism of Wheat Leaf Spot Disease Caused by Bipolaris sorokiniana through Comparative Transcriptome and Metabolomics Analysis.

Wheat yield is greatly reduced because of the occurrence of leaf spot diseases. Bipolaris sorokiniana is the main pathogenic fungus in leaf spot disease. In this study, B. sorokiniana from wheat leaf (W-B. sorokiniana) showed much stronger pathogenicity toward wheat than endophytic B. sorokiniana from Pogostemon cablin (P-B. sorokiniana). The transcriptomes and metabolomics of the two B. sorokiniana strains and transcriptomes of B. sorokiniana-infected wheat leaves were comparatively analyzed. In addition, the expression levels of unigenes related to pathogenicity, toxicity, and cell wall degradation were predicted and validated by quantitative reverse transcription polymerase chain reaction (qRT-PCR) analysis. Results indicated that pathogenicity-related genes, especially the gene encoding loss-of-pathogenicity B (LopB) protein, cell wall-degrading enzymes (particularly glycosyl hydrolase-related genes), and killer and Ptr necrosis toxin-producing related unigenes in the W-B. sorokiniana played important roles in the pathogenicity of W-B. sorokiniana toward wheat. The down-regulation of cell wall protein, photosystem peptide, and rubisco protein suggested impairment of the phytosynthetic system and cell wall of B. sorokiniana-infected wheat. The up-regulation of hydrolase inhibitor, NAC (including NAM, ATAF1 and CUC2) transcriptional factor, and peroxidase in infected wheat tissues suggests their important roles in the defensive response of wheat to W-B. sorokiniana. This is the first report providing a comparison of the transcriptome and metabolome between the pathogenic and endophytic B. sorokiniana strains, thus providing a molecular clue for the pathogenic mechanism of W-B. sorokiniana toward wheat and wheat's defensive response mechanism to W-B. sorokiniana. Our study could offer molecular clues for controlling the hazard of leaf spot and root rot diseases in wheat, thus improving wheat yield in the future.

Microarray profiling analysis identifies the mechanism of miR-200b-3p/mRNA-CD36 affecting diabetic cardiomyopathy via peroxisome proliferator activated receptor-γ signaling pathway.

OBJECTIVE: Current study focused on the influence of miR-200b-3p on cardiocyte apoptosis of diabetic cardiomyopathy (DCM) by regulating CD36 and peroxisome proliferator-activated receptor γ (PPAR-γ) signaling pathway. METHODS: Bioinformatic analysis was used to analyze differentially expressed microRNA (miRNAs), messenger RNAs (mRNAs) and activated pathways in DCM. And then quantitative real-time polymerase chain reaction (qRT-PCR) was conducted to verify expression of miR-200b-3p and CD36 in DCM model rats and glucose treated H9c2 cell line. Luciferase reporter assay was used to verify the transcriptional regulation of agomiR-200b-3p and investigate the relationship between miR-200b-3p and CD36. Flow cytometry was performed to assess cardiocyte apoptosis in different interference conditions. Echocardiography was used to illustrate the ejection fraction rate and fraction shortening rate of DCM model rats. Next, hematoxylin-eosin (H&E) staining assay was carried out to reveal structures of cardiocyte tissues with transfection in different conditions. Masson trichrome staining was used to evaluate myocardial fibrosis. Western blot analysis was used to detect the expression levels of PPAR-γ signaling-related protein PPAR-γ and Bcl-2. RESULTS: miR-200b-3p was low-expressed while CD36 was overexpressed in DCM. AgomiR-200b-3p could inhibit the expression of CD36 to regulate cardiocyte apoptosis in DCM. CD36 activated PPAR-γ signaling pathway in DCM. Silencing CD36 or GW9662 treatment protect rat against DCM. CONCLUSION: miR-200b-3p targeted CD36 to regulate cardiocyte apoptosis of DCM by activating PPAR-γ signaling pathway.

Highly Substituted Benzophenone Aldehydes and Eremophilane Derivatives from the Deep-Sea Derived Fungus Phomopsis lithocarpus FS508.

Five new benzophenone derivatives named tenellones D⁻H (1⁻5), sharing a rare naturally occurring aldehyde functionality in this family, and a new eremophilane derivative named lithocarin A (7), together with two known compounds (6 and 8), were isolated from the deep marine sediment-derived fungus Phomopsis lithocarpus FS508. All of the structures for these new compounds were fully characterized and established on the basis of extensive spectroscopic interpretation and X-ray crystallographic analysis. Compound 5 exhibited cytotoxic activity against HepG-2 and A549 cell lines with IC50 values of 16.0 and 17.6 µM, respectively.

Influence of specific HSP70 domains on fibril formation of the yeast prion protein Ure2.

Ure2p is the protein determinant of the Saccharomyces cerevisiae prion state [URE3]. Constitutive overexpression of the HSP70 family member SSA1 cures cells of [URE3]. Here, we show that Ssa1p increases the lag time of Ure2p fibril formation in vitro in the presence or absence of nucleotide. The presence of the HSP40 co-chaperone Ydj1p has an additive effect on the inhibition of Ure2p fibril formation, whereas the Ydj1p H34Q mutant shows reduced inhibition alone and in combination with Ssa1p. In order to investigate the structural basis of these effects, we constructed and tested an Ssa1p mutant lacking the ATPase domain, as well as a series of C-terminal truncation mutants. The results indicate that Ssa1p can bind to Ure2p and delay fibril formation even in the absence of the ATPase domain, but interaction of Ure2p with the substrate-binding domain is strongly influenced by the C-terminal lid region. Dynamic light scattering, quartz crystal microbalance assays, pull-down assays and kinetic analysis indicate that Ssa1p interacts with both native Ure2p and fibril seeds, and reduces the rate of Ure2p fibril elongation in a concentration-dependent manner. These results provide new insights into the structural and mechanistic basis for inhibition of Ure2p fibril formation by Ssa1p and Ydj1p.

Studying the effects of chaperones on amyloid fibril formation.

The results of cell and animal model studies demonstrate that molecular chaperones play an important role in controlling the processes of protein misfolding and amyloid formation in vivo. In addition, chaperones are involved in the appearance, propagation and clearance of prion phenotypes in yeast. The effect of chaperones on amyloid formation has been studied in great detail in recent years in order to elucidate the underlying mechanisms. An important approach is the direct study of effects of chaperones on amyloid fibril formation in vitro. This review introduces the methods and techniques that are commonly used to control and monitor the time course of fibril formation, and to detect interactions between chaperones and fibril-forming proteins. The techniques we address include thioflavin T binding fluorescence and filter retardation assays, size-exclusion chromatography, dynamic light scattering, and biosensor assays. Our aim in this review is to provide guidance on how to embark on study of the effect of chaperones on amyloid fibril formation, and how to avoid common problems that may be encountered, using examples and experience from the authors' lab and from the wider literature.

Alcohol oxidase (AOX1) from Pichia pastoris is a novel inhibitor of prion propagation and a potential ATPase.

Previous results suggest that methylotrophic yeasts may contain factors that modulate prion stability. Alcohol oxidase (AOX), a key enzyme in methanol metabolism, is an abundant protein that is specific to methylotrophic yeasts. We examined the effect of Pichia pastoris AOX1 on prion phenotypes in Saccharomyces cerevisiae. The S. cerevisiae prion states [PSI(+)] and [URE3] arise from aggregation of the proteins Sup35p and Ure2p respectively, and correlate with the ability of Sup35p and Ure2p to form amyloid-like fibrils in vitro. We found that expression of P. pastoris AOX1 in S. cerevisiae had no effect on propagation of the [PSI(+)] prion, but inhibited propagation of [URE3]. Addition of AOX1 early in the time-course of fibril formation inhibits Ure2p fibril formation in vitro. AOX1 has not previously been identified as an ATPase. However, we discovered that in addition to its flavin adenine dinucleotide-dependent AOX activity, AOX1 possesses ATPase activity. This study identifies AOX1 as a novel prion inhibitory factor and a potential ATPase.