Trichoderma: A Treasure House of Structurally Diverse Secondary Metabolites With Medicinal Importance.

Fungi play an irreplaceable role in drug discovery in the course of human history, as they possess unique abilities to synthesize diverse specialized metabolites with significant medicinal potential. Trichoderma are well-studied filamentous fungi generally observed in nature, which are widely marketed as biocontrol agents. The secondary metabolites produced by Trichoderma have gained extensive attention since they possess attractive chemical structures with remarkable biological activities. A large number of metabolites have been isolated from Trichoderma species in recent years. A previous review by Reino et al. summarized 186 compounds isolated from Trichoderma as well as their biological activities up to 2008. To update the relevant list of reviews of secondary metabolites produced from Trichoderma sp., we provide a comprehensive overview in regard to the newly described metabolites of Trichoderma from the beginning of 2009 to the end of 2020, with emphasis on their chemistry and various bioactivities. A total of 203 compounds with considerable bioactivities are included in this review, which is worth expecting for the discovery of new drug leads and agrochemicals in the foreseeable future. Moreover, new strategies for discovering secondary metabolites of Trichoderma in recent years are also discussed herein.

New Enantiomers of a Nor-Bisabolane Derivative and Two New Phthalides Produced by the Marine-Derived Fungus Penicillium chrysogenum LD-201810.

Marine-derived fungi are a treasure house for the discovery of structurally novel secondary metabolites with potential pharmaceutical value. In this study, a pair of new nor-bisabolane derivative enantiomers (±)-1 and two new phthalides (4 and 5), as well as four known metabolites, were isolated from the culture filtrate of the marine algal-derived endophytic fungus Penicillium chrysogenum LD-201810. Their structures were established by detailed interpretation of spectroscopic data (1D/2D NMR and ESI-MS). The optical resolution of compound (±)-1 by chiral HPLC successfully afforded individual enantiomers (+)-1 and (-)-1, and their absolute configurations were determined by TDDFT-ECD calculations. Compound (±)-1 represents the first example of bisabolane analogs with a methylsulfinyl substituent group, which is rare in natural products. All of the isolated compounds 1-7 were evaluated for their cytotoxic activity against A549, BT-549, HeLa, HepG2, MCF-7, and THP-1 cell lines, as well as for antifungal activity against four plant pathogenetic fungi (Alternaria solani, Botrytis cinerea, Fusarium oxysporum, and Valsa mali). Compound 2, a bisabolane-type sesquiterpenoid, was shown to possess excellent activity for control of B. cinerea with half-maximal inhibitory concentration (IC50) of 13.6 µg/mL, whereas the remaining investigated compounds showed either weak or no cytotoxic/antifungal activity in this study.

Cytotoxic Secondary Metabolites Isolated from the Marine Alga-Associated Fungus Penicillium chrysogenum LD-201810.

A new pentaketide derivative, penilactonol A (1), and two new hydroxyphenylacetic acid derivatives, (2'R)-stachyline B (2) and (2'R)-westerdijkin A (3), together with five known metabolites, bisabolane-type sesquiterpenoids 4-6 and meroterpenoids 7 and 8, were isolated from the solid culture of a marine alga-associated fungus Penicillium chrysogenum LD-201810. Their structures were elucidated based on extensive spectroscopic analyses, including 1D/2D NMR and high resolution electrospray ionization mass spectra (HRESIMS). The absolute configurations of the stereogenic carbons in 1 were determined by the (Mo2(OAc)4)-induced circular dichroism (CD) and comparison of the calculated and experimental electronic circular dichroism (ECD) spectra, while the absolute configuration of the stereogenic carbon in 2 was established using single-crystal X-ray diffraction analysis. Compounds 2 and 3 adapt the 2'R-configuration as compared to known hydroxyphenylacetic acid-derived and O-prenylated natural products. The cytotoxicity of 1-8 against human carcinoma cell lines (A549, BT-549, HeLa, HepG2, MCF-7, and THP-1) was evaluated. Compound 3 exhibited cytotoxicity to the HepG2 cell line with an IC50 value of 22.0 µM. Furthermore, 5 showed considerable activities against A549 and THP-1 cell lines with IC50 values of 21.2 and 18.2 µM, respectively.

New sesquiterpenoids with COX-2 inhibitory activity from the medical plant Physalis. alkekengi L. var. franchetii.

Phytochemical investigations on Physalis. alkekengi L. var. franchetii, a widespread traditional Chinese medicine, led to the isolation and identification of three new sesquiterpenoids physalisitins A-C (1-3). Their structures were elucidated by NMR and HRESIMS analysis, and their absolute configurations were determined by quantum chemical NMR and ECD calculations, as well as by comparing their optical rotation values with those known analogues. All of the isolated compounds were evaluated for their cyclooxygenase-2 (COX-2) inhibitory activity. Compounds 1-3 dose-dependently inhibited the COX-2 enzyme with IC50 values of 3.22 ± 0.25, 6.35 ± 0.84, and 11.13 ± 1.47 µM, respectively.

Antimicrobial Secondary Metabolites from the Seawater-Derived Fungus Aspergillus sydowii SW9.

Marine-derived fungi are considered to be valuable producers of bioactive secondary metabolites used as lead compounds with medicinal importance. In this study, chemical investigation of the seawater-derived fungus Aspergillus sydowii SW9 led to the isolation and identification of one new quinazolinone alkaloid, 2-(4-hydroxybenzyl)-4-(3-acetyl)quinazolin-one (1), one new aromatic bisabolene-type sesquiterpenoid, (2) and one new chorismic acid analogue (3), as well as two known alkaloids (compounds 4 and 5). Their structures were determined by extensive 1D/2D NMR and mass spectrometric data, and the absolute configurations of 2 and 3 were assigned by the analysis of ECD spectra aided by quantum chemical computations. Compounds 1, 2, and 4 exhibited selective inhibitory activities against the human pathogenic bacteria Escherichia coli, Staphylococcus aureus, S. epidermidis, and Streptococcus pneumoniae, with MIC values ranging from 2.0 to 16 µg/mL.

Antineoplastic Constituents from the Chemical Diversified Extract of Radix puerariae.

To enhance the structural diversity of isoflavonoids and provide more derivatives for the biological screening, a semisynthetic mixture was generated by diversification of the crude extract of Radix puerariae (Pueraria montana var. lobata) through the chemical reaction with hydrazine hydrate. Eleven 3,4-diarylpyrazoles (1-11) and two 5-phenyl-6-benzyldihydropyridazinones (12 and 13) were isolated from the semisynthetic mixture, and their structures were identified by spectroscopic methods in combination with X-ray crystallographic analysis. Among them, nine compounds (5-13) were new derivatives. All the compounds were evaluated on the inhibitory activities against the prostate cancer cell lines LNCaP and PC3. Compounds 12 and 13 were found to exhibit much more potent inhibitory activities against the androgen dependent LNCaP cells than the androgen independent PC3 cells. Rapid synthesis of new 3,4-diarylpyrazoles and two 5-phenyl-6-benzyldihydropyridazinones with significant biological activity highlights the great potential of one-pot combinatorial modification for the diversification of natural products.

Complete genome sequence analysis of duck circovirus strains from Cherry Valley duck.

To investigate molecular epidemiology of DuCV in Cherry Valley ducks in China, the complete genomes of six DuCV strains, which were detected from Cherry Valley ducks in China between 2007 and 2008, were sequenced. Sequence and phylogenetic analysis were carried out to compare these six strains with another 27 DuCV strains from Mulard duck, Muscovy duck, Pekin ducks and Mule duck. The analysis showed that the six DuCV strains exhibited typical genetic features of the family of DuCV, such as a stem-loop structure, three major open reading frames (Rep, Cap and ORF3), four intergenic repeats and the conserved motifs for rolling circle replication and for the dNTP binding domain located in the Rep protein. Phylogenetic analysis of the nucleotide sequences of the complete genome and Cap gene of these strains together with those that have been previously published demonstrated two distinct DuCV genotypes. The DuCV strains with complete genomes containing 1988 and 1989 nucleotides clustered in genotype A, whereas the strains with complete genomes containing 1991, 1992, 1995 and 1996 nucleotides lay in genotype B. The six DuCV strains from Cherry Valley ducks were divided into the two groups. The results of the study provides some insight into the variation of DuCVs in Cherry Valley ducks.

Genetic diversity of parvovirus isolates from dogs and wild animals in China.

We isolated three new parvovirus variants in China. The isolate from a blue fox was related to feline parvovirus, but possessed a mutation of VP2 residue A300P. Isolates from a raccoon dog and a masked civet were antigenically similar to canine parvovirus-2a but had a substitution of VP2 residue G300S.

Development of an indirect ELISA for the detection of duck circovirus infection in duck flocks.

Infection with duck circovirus (DuCV) is associated with growth retardation and developmental problems in farmed ducks. To detect DuCV-specific antibody in duck serum, an indirect enzyme-linked immunosorbent assay (iELISA) method was developed using the recombinant capsid protein antigen prepared by cloning the cap (Cap) gene of DuCV FJ0601 strain into pET-32a (+) vector and expressed in Escherichia coli. Using the optimized iELISA method, DuCV-specific antibodies were detected in 157 (12.96%) of 1211 samples obtained from 17 (89.47%) of 19 meat duck flocks aged from 25 to 40 days and in 89 (22.08%) of 403 samples obtained from 9 (75%) of 12 breeder flocks aged from 14 to 61 weeks. These results indicated that the iELISA method is useful for serological diagnosis of DuCV infection and epidemiological investigation.

Evaluation of two experimental infection models for Avibacterium paragallinarum.

The clinical symptoms of infectious coryza are multiple and include nasal discharge, facial swelling, lacrimation, and anorexia. In general, the disease is not fatal to chicken; so, in experiments where animals are infected with Avibacterium paragallinarum, there have been debates about conducting the challenge model and evaluating the clinical signs. In this experiment, 150 chickens, aged 30 days, were randomly divided into different groups. Some groups were infected with the 'in-contact' challenge model and others with the artificial intrasinus-injection-route model. The bacterial isolates used were three field isolates of different serogroups of A. paragallinarum, including Hpg-8 (Page serovar A), CCM6075 (Page serovar B) and Hpg-668 (Page serovar C). During this study, a scoring system was used to record the clinical signs of the infected birds and evaluate the pathogenic diversity of the two models. The final results indicated that the 'in-contact' challenge model of the three isolates showed a more reliable representation of the natural infection under field conditions than the artificial intrasinus-injection-route model. Thus, on carrying out animal experiments, the effect of 'in-contact' challenge model is more accurate than the artificial intrasinus-injection-route model.

[Sequence analysis for the complete provial genome of endogenous avian leukosis virus strain SD0501].

The genomic DNA extracted from chicken embryo fibroblasts (CEF) of SPF chickens from three chicken farms was used as template to amplify the ALV proviral DNA by PCR with four pairs of primers, high positive detection rates of gag - gene (29/46), pol - gene (27/46), env - gene (24/46) and LTR fragment (31/46) were achieved. Eight continuous and overlapping fragments were amplified from one DNA sample with 8 pairs of primers according to published sequences, then cloned into the TA vector and se quenced. The complete sequence of the whole genome of ALV strain SD0501 was established and analyzed with DNAstar software. Comparisons of SD0501 sequence with that of other representative endogenous avian virus strains demonstrated that the genomes of ALV were relatively conservative, the nucleotide identity of all the strains was over 99.1%, and env - gene was over 98.5%. However, a low identity was demonstrated among the representative strains of different subgroups, especially, the env - gene showed obvious difference, the corresponding identity was as low as 56.3% - 91.5%.