Preliminary investigation of the chemo diversity of bioactive molecules produced by endophytic fungi isolated from Manihot utilisima leaf

Endophytic fungi have demonstrated a harmless relationship living within the internal tissues of several plant hosts and at the same time produce diverse important bioactive compounds having a wide range of pharmaceutical applicability. In this study, nine endophytic fungi, eight from Cassava leaf: Clg1, Clg2, Clr4, Clr5, Clr6, Clr7, Clr9 and one from Cassava mid-rib: CRs3 with distinct cultural features were isolated from healthy leaves of Manihot utilisima and axenic cultures were fermented on sterile rice medium for 21 days. The antimicrobial and antioxidant activities of the fungal crude extracts were evaluated. Chemical analyses of the metabolic profiles of each fungal extract revealed the presence of nine known compounds with established biological activities. Each fungal extract exhibited antimicrobial activities against at least one Gram positive and Gram negative bacteria with an inhibition zone that ranged from 2 – 6 mm. Clg2 and CRs3 fungal extracts demonstrated moderate potential to scavenge free radical with an inhibition of 58 and 60% respectively. Septicine, cyclo(prolylvalyl), pentenedioc acid, neurolenin B, rubrofusarin, p-Hydroxybenzoic acid, protocatechuic acid, citreoisocoumarin, palitantin and pestalio pyrone were the compounds detected in the fungal fermentation products. Our findings reveal that M. utilisima leaves harbor endophytic fungi with unique chemodiversity of bioactive secondary metabolites needed for development of new drugs.

Fungal Extract of Lasiodiplodia pseudotheobromae IBRL OS-64 Inhibits the Growth of Skin Pathogenic Bacterium and Attenuates Biofilms of Methicillin-Resistant Staphylococcus aureus

Background: The emergence of multidrug-resistant pathogens associated with biofilm formation can cause life-threatening infections to humans. Therefore, the present study aims to evaluate the effects of the fungal extract of Lasiodiplodia pseudotheobromae (L. pseudotheobromae) Industrial Biotechnology Research Laboratory (IBRL) OS-64 on bacterial cells and the biofilm formation of methicillin-resistant Staphylococcus aureus (MRSA). Methods: Broth microdilution and semi-quantitative adherence assays were conducted to determine the anti-biofilm activity of the fungal extract. Light and scanning electron microscopy (SEM) analyses were performed to observe the effect of the fungal extract on biofilm formation by MRSA. Results: The transmission electron microscopy (TEM) microphotographs showed that the bacterial cells were severely damaged upon 24 h exposure to the extract and displayed several symptoms such as cell shrinkage and breakage. Meanwhile, results from the antibiofilm study indicated the extract attenuated the initial and preformed biofilms of MRSA by 80.82% and 61.39%, respectively. The initial biofilm was more sensitive to the extract compared to the pre-formed biofilm, as evidenced by the light microscopy and SEM observations that demonstrated more severe bacterial cell damage on the initial biofilms compared to pre-formed biofilms. Conclusion: The ethyl acetate extract of L. pseudotheobromae IBRL OS-64 significantly inhibited bacterial cells growth and eliminated biofilm formation by MRSA.

Effect of Culture Medium Incorporated with Ocimum sanctum Extract in Enhancing Anti-MRSA activity of Endophytic Fungus, Lasiodiplodia pseudotheobromae

The effect of incorporating plant extract in the culture medium on anti-MRSA activity of Lasiodiplodia pseudotheobromae IBRL OS-64, was investigated in the present study. On disk diffusion assay, the ethyl acetate fungal extract from culture medium supplemented with host plant extract (HPE) of Ocimum sanctum leaves demonstrated good anti-MRSA activity with a diameter inhibition zone of 22.6±0.6 mm. Meanwhile, the minimal inhibition concentration (MIC) values of the extract from YES broth and YES broth incorporated with HPE were 1000 µg/mL and 250 µg/mL, respectively. The MBC values were 8000 µg/mL and 500 µg/mL, respectively. The YES + HPE extracts exerted bactericidal effect against the test bacteria since the MBC/MIC ratio was less than or equal to 4. The time-kill study revealed a 90% of growth reduction of MRSA ATCC 33591 after 16 h exposure to the fungal extract cultured in YES + HPE. Ironically, for fungal extract grown in YES broth, time-kill curve showed a regrowth pattern of bacterial cells after 24 h exposed to the extract. Therefore, the present study suggested that the addition of HPE in the culture medium could enhance the anti-MRSA activity of endophytic fungus, L. pseudotheobromae IBRL OS-64 against MRSA ATCC 33591.

“Enhancement of Vincristine Under In Vitro Culture of Catharanthus Roseus Supplemented With Alternaria Sesami Endophytic Fungal Extract As Biotic Elicitor”

Vincristine, one of the major vinca alkaloid of Catharanthus roseus(L.) G. Don. (Apocynaceae) was enhanced under in vitro culture of C.roseus using fungal extract of an endophyte Alternaria sesami isolated from the surface-sterilized root cuttings of C.roseus. Vindoline, a precursor molecule of Vincristine was detected for the first time from the fungal endophyte A.sesami which was used as biotic elicitor to enhance Vincristine content in the C.roseus callus.It was identified using high performance liquid chromatography and mass spectroscopy techniques by matching retention time and mass data with reference molecule. Supplementing heat sterilized A.sesami endophytic fungal culture extract into callus culture medium of C. roseus enhanced the Vincristine content in C. roseus callus by 21.717% after 105 day culture.

Comparative efficacy of commercially available deoxynivalenol (DON) detoxifying feed additives on growth performance, total tract digestibility of components and physiological responses in nursery pigs fed diets formulated with naturally contaminated corn

Comparative efficacy of deoxynivalenol (DON) detoxifying feed additives (FA) was evaluated in growth performance (Exp. 1) and apparent total tract digestibility (ATTD) (Exp. 2) nursery pig studies. Six corn-soybean-meal based diets were used: 1) positive control (PC, formulated with <1.5 ppm DON corn), negative control (NC, formulated with 5.5 ppm DON corn), NC+FA1 (clay plus yeast cell wall extract), NC+FA2 (aluminosilicate), NC+FA3 (aluminosilicate plus fungal extract), and NC+FA4 (Sodium metabisulfite, SMB). In Exp. 1, 144 pigs [(Body weight (BW), 10.2±0.1kg)] were housed (4 pigs/pen), allocated to diets (n=6) based on BW, and fed for 4-wk. The BW and feed intake were monitored weekly. On d 7, one pig/pen was bled for plasma and euthanized for organ weight and tissue samples. Assayed DON concentration in PC, NC, NC+FA4 was 0.29, 2.86 and 1.21 ppm, respectively. In wk-1, the ADG of pigs fed NC+FA4 was not different (P>0.05) to that of pigs fed PC diet but greater (P=0.01) than for pigs fed NC without or with other FA. Pigs fed NC and NC+FA2 had lower (P=0.026) ADFI than pigs fed PC and NC+FA3. Pigs fed NC+FA4 had greater (P=0.003) G:F than pigs fed the other diets. Diets had no effect (P>0.05) on ADG, ADFI, and G: F after first week, plasma concentration of urea and creatinine or liver and spleen weight. Pigs fed NC diets had greater (P=0.01) jejunal mRNA expression of superoxide dismutase 1 relative to pigs fed PC or NC plus FA. Jejunal histomorphology and mRNA expression of nutrient transporters, inflammatory cytokines and tight junction proteins and ceca digesta concentration of short chain fatty acids were not affected (P>0.05) by the diet. In Exp. 2, 24 barrows (BW 10.2 ± 0.3 kg) were individually placed in metabolism crates and allocated to four diets: PC, NC, NC+FA3 and NC+FA4 (n=6) containing TiO2 as digestibility marker. Pigs were adjusted to diets for 5 d, followed by a 2-d grab fecal sample collection. Pigs fed PC and NC+FA4 diets had higher ATTD of dry matter, gross energy, and crude protein than NC fed pigs. The FA3 was intermediate in digestibility response. In conclusion, feed additives containing sequestering component plus fungal extract or SMB in DON-contaminated feed resulted in commensurate nursery pig performance to PC. The tested feed additives mitigated intestinal oxidative stress through decreased expression of genes for superoxide dismutase.

A diverse global fungal library for drug discovery

Background Secondary fungal metabolites are important sources for new drugs against infectious diseases and cancers. Methods To obtain a library with enough diversity, we collected about 2,395 soil samples and 2,324 plant samples from 36 regions in Africa, Asia, and North America. The collection areas covered various climate zones in the world. We examined the usability of the global fungal extract library (GFEL) against parasitic malaria transmission, Gram-positive and negative bacterial pathogens, and leukemia cells. Results Nearly ten thousand fungal strains were isolated. Sequences of nuclear ribosomal internal transcribed spacer (ITS) from 40 randomly selected strains showed that over 80% were unique. Screening GFEL, we found that the fungal extract from Penicillium thomii was able to block Plasmodium falciparum transmission to Anopheles gambiae, and the fungal extract from Tolypocladium album was able to kill myelogenous leukemia cell line K562. We also identified a set of candidate fungal extracts against bacterial pathogens.

Bioactive potential of endophytic fungus Chaetomium globosum and GC–MS analysis of its responsible components

The recent exploration of various medicinal plants for bioactive potential has led to the growing interest to explore their endophytes for such bioactive potential which may turn out to be better option than the plants. In the present study, Chaetomium globosum, an endophytic fungus isolated from Moringa oleifera Lam has been explored for its various biological activities. The chloroformic extract of C. globosum showed good antimutagenicity against the reactive carcinogenic mutagen, 2-aminofluorene (2-AF) in Ames test. The antiproliferative activity against various cell lines such as HCT-15, HeLa and U87-MG was found to be dose dependent and the viability reduced to 9.26%, 15.7% and 16.3%, respectively. Further, the chloroformic fungal extract was investigated for free radical scavenging activity using 2, 2-diphenyl-1-picrylhydrazyl (DPPH) and 2,2′-azino-bis(3-ethyl-benzthiazolin-6-sulfonic acid) assay which showed the IC50 value of 45.16 µg/ml and 50.55 µg/ml, respectively. The fungal extract also showed good ferric reducing power. Total phenolic and flavonoid content was found to be in linear relationship with the antioxidant potential of the fungal extract. High performance liquid chromatography showed the presence of phenolics which may help to combat the free radicals. The presence of various bioactive compounds was analysed by GC–MS which endorsed Chaetomium globosum to be a promising candidate for drug development.