Phomopsis Sp Research Articles

Tracing the mass flow from glucose and phenylalanine to pinoresinol and its glycosides in Phomopsis sp. XP-8 using stable isotope assisted TOF-MS

Phomopsis sp. XP-8, an endophytic fungus from the bark of Tu-Chung (Eucommia ulmoides Oliv) showed capability to biosynthesize pinoresinol (Pin) and pinoresinol diglucoside (PDG) from glucose (glu) and phenylalanine (Phe). To verify the mass flow in the biosynthesis pathway, [13C6]-labeled glu and [13C6]-labeled Phe were separately fed to the strain as sole substrates and [13C6]-labeled products were detected by ultra-high-performance liquid chromatography-quadrupole time of flight mass spectrometry. As results, [13C6]-labeled Phe was incorporated into [13C6]-cinnamylic acid (Ca) and p-coumaric acid (p-Co), and [13C12]-labeled Pin, which revealed that the Pin benzene ring came from Phe via the phenylpropane pathway. [13C6]-Labeled Ca and p-Co, [13C12]-labeled Pin, [13C18]-labeled pinoresinol monoglucoside (PMG), and [13C18]-labeled PDG products were found when [13C6]-labeled glu was used, demonstrating that the benzene ring and glucoside of PDG originated from glu. It was also determined that PMG was not the direct precursor of PDG in the biosynthetic pathway. The study identified the occurrence of phenylalanine- lignan biosynthesis pathway in fungi at the level of mass flow.

Polyketides and nitrogenous metabolites from the endophytic fungus Phomopsis sp. D15a2a

Three new polyketides, phomopones A−C (1–3), one new cyclic tetrapeptide, 18-hydroxydihydrotentoxin (4), and a new amide, 6-hydroxyenamidin (5) together with a known derivative, enamindin (6) were obtained from the endophytic fungus Phomopsis sp. D15a2a isolated from the plant Alternanthera bettzickiana. The structures of the new compounds were elucidated by 1D, 2D NMR and HRMS data. The absolute configurations of the isolated metabolites were determined either by X-ray crystallography, Marfey’s method or by converting the compounds to Mosher esters.

Derivatization of Natural Compound β-Pinene Enhances Its In Vitro Antifungal Activity against Plant Pathogens.

Background: The development of new antifungal agents has always been a hot research topic in pesticide development. In this study, a series of derivatives of natural compound β-pinene were prepared, and the antifungal activities of these derivatives were evaluated. The purpose of this work is to develop some novel molecules as promising new fungicides. Methods: Through a variety of chemical reactions, β-pinene was transformed into a series of β-pinene-based derivatives containing amide moieties and acylthiourea moieties. The antifungal activities of these derivatives against five plant pathogens including Colletotrichum gloeosporioides, Fusarium proliferatum, Alternaria kikuchiana, Phomopsis sp. and Phytophthora capsici were tested; preliminary structure–activity relationship was discussed. Results: Some derivatives exhibited moderate or significant antifungal activity due to the fusion of the amide moiety or the acylthiourea moiety with the pinane skeleton. The structure–activity relationship analysis showed that the fluorine atom and the strong electron withdrawing nitro group, or trifluoromethyl group on the benzene ring of the derivatives had a significant effect on the improvement of the antifungal activity against Colletotrichum gloeosporioides, Fusarium proliferatum, Alternaria kikuchiana and Phomopsis sp. Meanwhile, the introduction of an ethyl group at the meta-position on the benzene ring of the derivatives could improve the antifungal activity against Phytophthora capsici. Compounds 4e, 4h, 4q, 4r exhibited broad-spectrum antifungal activity against the tested strains. Compound 4o had significant antifungal activity against Phytophthora capsici (IC50 = 0.18 μmol/L). These derivatives were expected to be used as precursor molecules for novel pesticide development in further research.

Anti-inflammatory diterpenoids from an endophytic fungus Phomopsis sp. S12

Three new pimarane-type diterpenoids, along with two known compounds, were isolated from an endophytic fungus Phomopsis sp. S12. Their structures were determined by comprehensive analyses of HRESIMS, NMR, CD, as well as single-crystal X-ray diffractions. Compound 3 exhibited anti-inflammatory activity against the production of IL-1β and IL-6 induced by lipopolysaccharide (LPS) in macrophages. A putative biosynthetic pathway for compounds 1–5 were also proposed.

Two New Naphthalene Derivatives from the Fermentation Products of an Endophytic Fungus Phomopsis sp.

Two new naphthalene derivatives, 5-methoxy-2-methyl-7-(3-methyl-2-oxobut-3-enyl)-1-naphthaldehyde (1) and 2-(hydroxymethyl)-5-methoxy-7-(3-methyl-2-oxobut-3-enyl)-1-naphthaldehyde (2), together with two known naphthalene derivatives (3 and 4), were isolated from the fermentation products of an endophytic fungus Phomopsis sp. Their structures were elucidated by spectroscopic methods, including extensive 1D and 2D NMR techniques. Compounds 1 and 2 were evaluated for their anti-methicillin-resistant Staphylococcus aureus (anti-MRSA) activity. The results showed that compounds 1 and 2 showed good inhibition with IZD of 14.5 ± 1.2 and 15.2 ± 1.3 mm, respectively.

Recovery of gold from electronic wastewater by Phomopsis sp. XP-8 and its potential application in the degradation of toxic dyes

Environmentally friendly, efficient, and economical methods of gold recovery are significant challenges for high-tech industries and environmental protection. In this study, Phomopsis sp. XP-8, a filamentous endophytic fungus, demonstrated great potential for selectively recovering gold from electronic wastewater without any pretreatment and was shown to be convenient and safe in practical application. Under optimum conditions, the gold recovery rate was more than 80% at a dilute concentration of Au3+. Model-fitting analysis indicated the adsorption of Au3+ was well described by the pseudo-second-order kinetic model and the Langmuir isotherm model. The maximum Au3+ adsorption was from 208 to 280 mg/g within the temperature range of 20–50 °C. The data from electronic wastewater revealed its great potential for selective recovery of gold from complex aqueous solutions. Additionally, the formed nanogold-bioconjugates exhibited efficient degradation of toxic dyes in wastewater, which demonstrated the potential application of these byproducts produced via the biosorption process.

Morphological Characterization and Molecular Identification of Stem-end Rot Associated Fungal Species Isolated from ‘Karutha Colomban’ Mango Fruits in Sri Lanka

Purpose: Stem-End Rot (SER) is one of the most frequently found mango postharvest diseases in many countries including Sri Lanka. Availability of a significantly detailed study on morphological and molecular characterization of the SER associated fungal species is comparatively low. The present study has been carried out considering the above fact. Research Method: SER disease associated fungi were isolated and identification was carried out using morphological characteristics. Identification was confirmed by phylogenetic analysis of newly generated ITS sequencing data using Maximum Likelihood (ML) with RAxML software (Ver.8.2.10). Findings: From the morphological studies it was observed that the fungal isolates SER1, SER2, SER3 and SER6 produced conidia on Potato Dextrose Agar. Both morphological and molecular results revealed that the isolates of SER1, SER4, SER5, and SER6 which were isolated from diseased Karutha Colomban mango fruits with SER (in Sri Lanka) belong to Lasiodiplodia sp., Nodulisporium sp., Xylaria feejeensis and Pestalotiopsis sp. respectively. Both SER2 and SER3 were confirmed as Phomopsis sp. During the present study two new fungal species (Xylaria spp. and Nodulisporium spp.) were also identified for the first time from SER of Karutha Colomban mango fruits. Research Limitations: The pathogenicity of these isolates will be tested in near future, since the study was mainly focused on the isolation and identification of the SER associated fungal species. Originality/ Value: Results of the present study will be very valuable for stakeholders in Agricultural sector, before designing control strategies at post-harvest level in order to minimize the loss and extend the storage life of mango.

Dicerandrol B: a natural xanthone dimer induces apoptosis in cervical cancer HeLa cells through the endoplasmic reticulum stress and mitochondrial damage.

BackgroundDicerandrol B is a natural antitumor agent that can be isolated from the endophytic fungus, Phomopsis sp. The present study investigated the effects of dicerandrol B on human cervical cancer HeLa cells.Materials and methodsIn this study, dicerandrol B was identified by electrospray ionization mass spectrometry and nuclear magnetic resonance spectroscopy. We used MTT to detect the cell viability. Flow cytometry was used to analyze the apoptosis and cell cycle. Western blot was used to examine the expression of related proteins.ResultsDicerandrol B was isolated from the endophytic fungus Phomopsis sp. The MTT assay and flow cytometry showed that dicerandrol B significantly inhibited HeLa cell viability and induced G2/M cell cycle arrest. Western blot analysis demonstrated that dicerandrol B increased the levels of GRP78, ubiquitin, cleaved PARP, and Bax protein, decreased the levels of PARP and Bcl-2 protein, and caused an increase in the Bax/Bcl-2 ratio in HeLa cells. Dicerandrol B increased the production of ROS in HeLa cells, which was attenuated by the antioxidant N-acetyl-l-cysteine.ConclusionThese findings suggest that dicerandrol B induces apoptosis in human HeLa cells, possibly through the endoplasmic reticulum stress and mitochondrial apoptotic pathways. This suggests that dicerandrol B possesses strong anticancer activity in cervical cancer and provides insight into the underlying mechanisms.

Biosynthesis of antibacterial compound against multidrug resistant foodborne pathogens by Phomopsis sp. XP-8

Multi-drug resistance of foodborne pathogens has been greatly highlighted in food safety and caused a significant challenge for the development of new antibiotics and safe preservatives. The ethanol extract of Phomopsis sp. XP-8 broth showed significant inhibitory effects on multi-drug resistance of foodborne pathogens. After purification with thin-layer chromatography and liquid chromatographic analysis, the antibacterial compound was identified as methyl quercetin according to FTIR spectrum analysis and ESI-Q-TOF-MS analysis. Via substrate-feeding experiments, the mass flow for the biosynthesis of methyl quercetin was defined as (starting from glucose to phenylalanine): cinnamic acid, p-coumaric acid, quercetin, and methyl quercetin. The study revealed the capability of methyl quercetin to inhibit foodborne multi-drug resistant Escherichia coli, Staphylococcus aureus, and Salmonella shubra, as well as the capability of Phomopsis sp. XP-8 to produce methyl quercetin, showing potential application in food control.

A New Benzenoid Derivative from an Endophytic Fungus in Peperomia sui

A new benzenoid, phomopsuidiol (1), was isolated from nature for the first time from the n-BuOH extract derived from cultures of the endophytic fungus 10F0011 found in the whole plant of Peperomia sui. The 10F0011 strain was identified as Phomopsis sp. The structure of the isolated compound was elucidated by one- and two-dimensional nuclear magnetic resonance and mass spectrometry and by comparison with related compounds from the literature.

Conditioned media and organic elicitors underpin the production of potent antiplasmodial metabolites by endophytic fungi from Cameroonian medicinal plants.

Plasmodial resistance to artemisinin-based combination therapies emphasizes the need for new drug development to control malaria. This paper describes the antiplasmodial activity of metabolites produced by endophytic fungi of three Cameroonian plants. Ethyl acetate extracts of fungi cultivated on three different media were tested against Plasmodium falciparum chloroquine-sensitive (Pf3D7) and chloroquine-resistant (PfINDO) strains using the SYBR green florescence assay. Selected endophytes were further grown in potato dextrose broth supplemented with small organic elicitors and their extracts tested for activity. The effect of elicitors on de novo metabolite synthesis was assessed by reverse-phase HPLC. Activity screening of 81 extracts indicated that Aspergillus niger 58 (IC50 2.25-6.69μg/mL, Pf3D7), Fusarium sp. N240 (IC50 1.62-4.38μg/mL, Pf3D7), Phomopsis sp. N114 (IC50 0.34-7.26μg/mL, Pf3D7), and Xylaria sp. N120 (IC50 2.69-6.77μg/mL, Pf3D7) produced potent extracts when grown in all three media. Further culture of these endophytes in potato dextrose broth supplemented with each of the eight small organic elicitors and subsequent extracts screening indicated the extract of Phomopsis sp. N114 grown with 1% 1-butanol to be highly selective and extremely potent (IC50 0.20-0.33μg/mL; SI > 666). RPHPLC profiles of extracts of Phomopsis sp. N114 grown with or without 1-butanol showed some peaks of enhanced intensities in the former without any qualitative change in the chromatograms. This study showed the ability of selected endophytes to produce potent and selective antiplasmodial metabolites in varied culture conditions. It also showed how the production of desired metabolites can be enhanced by use of small molecular weight elicitors.

Protoilludane, Illudalane, and Botryane Sesquiterpenoids from the Endophytic Fungus Phomopsis sp. TJ507A.

To explore the chemical diversity of metabolites from endophytic fungi, the strain Phomopsis sp. TJ507A, isolated from the medicinal plant Phyllanthus glaucus, was investigated. A 2,3- seco-protoilludane-type sesquiterpenoid (1), eight protoilludane-type sesquiterpenoids (2-9), four illudalane-type sesquiterpenoids (10a/10b, 11, and 12), and a botryane-type sesquiterpenoid (13) in addition to seven known sesquiterpenoids (14-20) were identified from the liquid culture of the fungus. Structures of the isolated compounds, including their absolute configurations, were elucidated based on extensive spectroscopic analyses, a modified Mosher analysis, electronic circular dichroism (ECD) calculations, and [Rh2(OCOCF3)4]-induced ECD spectra as well as X-ray crystallographic analyses. Compound 1 represents the first example of a naturally occurring sesquiterpenoid containing the unusual 2,3- seco-protoilludane scaffold. Compounds 1 ( p < 0.001); 2-6, 15, and 18 ( p < 0.01); and 7, 9, and 20 ( p < 0.05) displayed β-site amyloid precursor protein cleaving enzyme 1 (BACE1) inhibitory activities ranging from 19.4% to 43.8% at the concentration of 40 μM. LY2811376 was used as the positive control with an inhibitory activity of 38.6% ( p < 0.01). Furthermore, none of these compounds showed obvious hepatotoxicity at concentration of 40 μM.

Chemical interaction of endophytic fungi and actinobacteria from Lychnophora ericoides in co-cultures

Microorganisms interact chemically in natural environments; however, the compounds and mechanisms involved in this phenomenon are still poorly understood. Using the cocultivation approach, changes in metabolic profiles due to interactions between endophytic fungal and actinobacterial strains isolated from the plant Lychnophora ericoides (Asteraceae) were assessed. The production of the cytotoxic compound cytochalasin H by the fungus Phomopsis sp. FLe6 was remarkably inhibited in solid and liquid co-cultures with the actinobacteria Streptomyces albospinus RLe7. This was a consequence of the fungal growth inhibition caused by antifungal compounds produced by S. albospinus RLe7, including amphotericin B. Cytochalasin H is not toxic to S. albospinus RLe7, suggesting that this microorganism does not require a defense mechanism to prevent the potentially harmful effects of such fungal compound. By exhibiting various competitive phenotypes, these microbes can control each other’s growth when sharing an environment.

Strategies to enhance the production of pinoresinol and its glucosides by endophytic fungus (Phomopsis sp. XP-8) isolated from Tu-chung bark

To improve the production yield of (+)-pinoresinol (Pin), (+)-pinoresinol monoglucoside (PMG), and (+)-pinoresinol diglucoside (PDG), different methods were conducted, including co-culture with resveratrol-producing Alternaria sp. MG1 spores and addition of Tu-chung in a medium at the start of cultivation, ultrasound treatment (40 kHZ, 10 min) on 5-day culture, and addition of ethanol and sodium butyrate on Day 3, followed by cultivation for an additional period of 2 days. At the end of the cultivation period (5 days), the liquid phase was collected for product analysis. Cells were collected for the determination of gene expression levels and then used in bioconversion using resting cells for another period of 2 days. The liquid phase was measured to determine the output of the target products and the expression levels of the key genes related to the biosynthesis of these compounds. Consequently, co-culture with Alternaria MG1 and addition of Tu-chung bark in the medium efficiently increased Pin, PMG, and PDG production yield in the biosynthesis systems using potato dextrose broth medium and resting cells of Phomopsis sp. XP-8. The key genes related to the biosynthesis of these compounds were significantly upregulated. However, in the majority of cases, the addition of ethanol and sodium butyrate, and ultrasound treatment decreased the production yield of Pin, PMG, and PDG. The change in production yield was not consistently accompanied by a change in gene expression.

Polyoxygenated Cyclohexenoids with Promising α-Glycosidase Inhibitory Activity Produced by Phomopsis sp. YE3250, an Endophytic Fungus Derived from Paeonia delavayi.

Seven new polyoxygenated cyclohexenoids, namely, phomopoxides A-G (1-7), were isolated from the fermentation broth extract of an endophytic fungal strain Phomopsis sp. YE3250 from the medicinal plant Paeonia delavayi Franch. The structures of these compounds were established by spectroscopic interpretation. The absolute configurations of compounds 1 and 4 were confirmed by X-ray crystallographic analysis and chemical derivative approach. All isolated compounds showed weak cytotoxic activities toward three human tumor cell lines (Hela, MCF-7, and NCI-H460) and weak antifungal activities against five pathogenic fungi (Candida albicans, Aspergillus niger, Pyricularia oryzae, Fusarium avenaceum, and Hormodendrum compactum). In addition, compounds 1-7 showed a promising α-glycosidase inhibitory activity with IC50 values of 1.47, 1.55, 1.83, 2.76, 2.88, 3.16, and 2.94 mM, respectively, as compared with a positive control of acarbose (IC50 = 1.22 mM).

Heterologous expression of Oenococcus oeni sHSP20 confers temperature stress tolerance in Escherichia coli.

Small heat shock proteins (sHSPs) are heat shock proteins sized 12-43kDa that can protect proteins from denaturation, particularly under high temperature; sHSPs thus increase the heat tolerance capability of an organisms enabling survival in adverse climates. sHSP20 is overexpressed in Oenococcus oeni in response to low temperatures. However, we found that overexpression of sHSP20 in Escherichia coli BL21 increased the microbial survival ratio at 50°C by almost 2h. Adding sHSP20 to the glutamate dehydrogenase solution significantly increased the stability of the enzyme at high temperature (especially at 60-70°C), low pH values (especially below 6.0), and high concentration of metal ions of Ga2+, Zn2+, Mn2+, and Fe3+. Notably, the coexpression of sHSP20 significantly enhanced soluble expression of laccase from Phomopsis sp. XP-8 (CCTCCM209291) in E. coli without codon optimization, as well as the activity and heat stability of the expressed enzyme. In addition to the chaperone activity of sHSP20 in the gene containing host in vivo and the enzyme heat stability in vitro, our study indicated the capability of coexpression of sHSP20 to increase the efficiency of prokaryotic expression of fungal genes and the activity of expressed enzymes. Graphical abstract ᅟ.

Neofusicoccum luteum as a pathogen on Tejocote (Crataegus mexicana)

Tejocote ( Crataegus mexicana ), a small pome crab-apple-like fruit, is becoming economically important in California with increasing production, so consideration of diseases that hinder the yield is important. Diseased trees of tejocote were observed in four orchards of Riverside and San Diego Counties of California. Ten symptomatic/asymptomatic samples were studied from each of the orchards. Five most frequently isolated fungi were identified on the basis of morphological characters and sequence data of the internal transcribed spacer ITS1-5.8S-ITS2 and partial β-tubulin gene. Three isolates were identified as Neofusicoccum luteum and two as Phomopsis sp. Pathogenicity tests were conducted by inoculating detached shoots of healthy tejocote trees. Significant lesions were observed on all shoots inoculated with the three N. luteum isolates (designated UCR1190, UCR1191, and UCR1192), but not on the shoots inoculated with other isolates or the non-inoculated controls. Results indicated that all three N. luteum isolates are aggressive pathogens on tejocote. This pathosystem should be further studied with a goal of designing appropriate disease management strategies.

Antifungal property of chili, shallot and garlic extracts against pathogenic fungi, Phomopsis spp., isolated from infected leaves of para rubber (Hevea brasiliensis Muell. Arg.)

The aim of this study was to evaluate the antifungal property of extracts of chili, shallot and garlic (local varieties in Sisaket, Thailand) against pathogenic fungi, Phomopsis spp., which were isolated from infected leaves of para rubber (Hevea brasiliensis Muell. Arg.). Seven isolates of Phomopsis spp. namely Phomopsis sp. SSK1.1, SSK1.2, SSK3.1, SSK4.1, SSK5.1, SSK5.2 and SSK7.1 were identified on the basis of morphological characteristics. Fresh plants were extracted with water to obtain crude extracts and their antifungal properties were tested on potato dextrose agar (PDA) media. The study demonstrated that increasing the concentrations (20%, 40%, 60% or 80%) of the chili extract exhibited a dependent increase in the inhibitory level on mycelial growth of Phomopsis spp. SSK3.1, SSK4.1 and SSK5.2. The inhibitory level on mycelial growth of shallot extract also increased in a dose-dependent manner in all isolates of Phomopsis. The garlic extract had significant inhibition on the growth of all isolates with complete inhibition at 80% concentration. The highest levels of percentage inhibition of mycelial growth were with garlic extract followed by shallot and chili extracts, respectively. The study also showed that these plant extracts contained some polyphenols (apigenin, gallic acid, catechin, quercetin, kaempferol and tannic acid) which are well-known compounds possessing antifungal activity. Therefore, it is possible that the antifungal properties of these plant extracts were partly due to these polyphenols or unknown active compounds which could not be analyzed in this study. Collectively, these results suggest that local varieties of both shallot and garlic possess strong antifungal properties.

Metabolites Produced by an Endophytic Phomopsis sp. and Their Anti-TMV Activity.

The fermentation and isolation of metabolites produced by an endophytic fungus, which was identified as Phomopsis sp. FJBR-11, based on phylogenetic analysis, led to the identification of six compounds, including dothiorelones A–C, and H, and cytosporones C and U. Among these compounds, cytosporone U exhibited potent inhibitory activity against Tobacco mosaic virus (TMV). Moreover, the crude and a purified exopolysaccharide were proved to possess strong inhibitory effects against the virus infection.

Chaetochromones A-C, Three New Polyketides from Mangrove Plant Derived Endophytic Fungus Phomopsis sp. SCSIO 41006.

Three new polyketides, chaetochromones A-C (1-3), together with a chromone (4), were isolated from the ethyl acetate extract of mangrove-derived fungus Phomopsis sp. SCSIO 41006. Their structures were elucidated by means of spectroscopic techniques (UV, IR, MS, 1D- and 2D-NMR). The absolute configurations of the new compounds were established by CD data.