Isolation and Characterization of Antibiotics from Soil FUNGI for the Development of Drugs

Abstract

During the present study, 680 root samples with adhering soil of 65 wild and cultivated plan species belonging to 59 genera and 19 families viz, Amaranthaceae (Amaranthus virides L, Aerva javancia (Burm.F.) Merrill, Digera muricata (L.) Mart.); Asclepiadaceae (Calotropis procera (Wild) R.BR.); Boraginace (Hellotropium europeam L.); Cannaceae (canna indica L.); Caricaceae (carica papaya L.); Chenopodiaceae (Beta vulgaris L., Chenopodium album L., Spinacea oleracea L.); Compositae (Conyza bonariensis (L.) Cronq., Hekuanthus annuus L., Lactuca sativa L., Launea mudicaulis Hook. F.); Convolvulaceae (Convolvulus arvensis L.); Cruciferae (Brassica juncea (L.) Czem & Coss, B.oleracea L., Var.capitata L., B.rapa L., Raphanus sativus L.); Cucurbitacea (Citrullus lanatus (Thumb.) Mastsum & Nakai, Cucumis sativus L., Cucurbita moschata L., Lagenaria siceraria (Mol.) standl., Luffa aegyptiaca Mill., Momordica charantia L.); Cyperarceae (Cyperus rotundus L.); Euphorbiaceae (Euphorbia hirta L.); Gramineae (Avena sativa L., Cenchrno setigerus Vahl, Cynodon dactylon (l.) Pers, Oryza sativa L., Pennisetum americanum (L.) Leeke, Satsria verticilata L., Sorghum bicolor L., Triticum aestivum L., Zea mays L.); Papilionaceae (Alhaji murorum Medik, Arachis hypogaea L., Cicer arietinum L., Cyamopsis tetragonoloba (L.) Taub., Glycine max (L.) Merr., Lens culinaris Medik., Leucaena leucocephala Lam., Medicago sativa L., Melilotus alba Medik., Phaseolus vulfaris L., Pisum sativum L., Sesbania sesban (L.) Merr., Trifolium alexandrunum (L.) Spergue ex. Turrill, Trigonella foenum-graecum L., Vigna munga (L.) Hepper, V.radiata (L.) Wilczek); Malvaceae (Abelmoschus esculentus (L.) Moench., Abutilon indicum (L.) Sweet, Gossypium arboretum L.); Pedaliaceae (Sesamum indicum L.); Piperaceae (Piper betel L.); Solanum melongena L., S.nigrum L., S.surrenttense Burm.F); Umbelliferae (Coriandrum sativum L., Daucus carota L.) were collected from Karachi University Campus, Malir, Darsano, Chano, Khator, ShahFaisal colony from Karachi Division; Gharo and Ghuji from Thatta District; and Hub, Baluchistan. Fungi associated with rhizosphere and rhizoplane were isolated and identified. Fungi isolated from rhizoplane were identified as Alternaria alternata (Fr.) Keissler (34 hosts), Aspergillus candidua Link ex link (1 host). A flavus kink ex gray (25 hosts). A fumigatus Fres. (5 hosts). A niger Van Tieghem (32 hosts), A nidulans (Eidam) Winter (13 hosts), A terreus Thom (19 hosts), Chaetomium flavum omvik (1 host), C.globosum kunze ex staud (13 hosts), C.indicum Corda (5 hosts), Cladosporium sp., (5 hosts), Cunningharmella echinulata (Thaxt) Thaxt., (10 hosts). Curvularia clavata jain (3 hosts). C.lunata (Wakker) Boedjin (19 hosts), Drechsler australiensis (Bugni) Subram. & Jain ex M.B Ellis (40 hosts), D.halodes (Drechsler) Subram & Jain (8 hosts), D.hawiiensis (Bugni) Subram. & Jain ex M.B Ellis (2 hosts), Fusarium culmurum (W.G Sm.) Sacc. (2 hosts), F.moniliforme sheld (6 hosts), F.oxysporum Schlecht emend. Snyed. & Hans (19 hosts), F.semitectum Berk and Rav. (3 hosts), F.solani (Mart.) Appel & Wollenw. Emend snyd. & Hans (62 hosts),Helicocephalum sp. (1 host), Humicola grisea Traaen (1 host), Macrophomina Phaseolina (Tassi) Goid. (44 hosts), Memnoniella echinata (Riv.) Galloway (1 host), Myrothecium roridum Tode ex Steudel (1 host), Niprospora oryzae (Berk. & Br.) Petch (2 hosts), Paecilomyces lilacinus (Thom.) Samson (3 hosts), Penicillium waksmanni Zaleski (1 host), Phialophora fastigiata (Lagerb. & Mekin) Conant (1 host), Rhizoctonia solani Kuhn (40 hosts), Rhizopus stolonifera (Ehrenb. Ex Link) Lind. (12 hosts), Trichoderma viride pers ex. Gray (4 hosts) and Verticillium chlamydosporium Goddard (1 host). Fungi isolated from rhizosphere were identified as Alternaria alternata (Fr.) Keissler (36 hosts), Articulospora sp. (1 host), Aspergillus canadidus Link ex link (5 hosts), A flavus link ex Gray (57 hosts), A fumigatusfres. (6 hosts), Aglaucus Link (2 hosts), Anidulans (Eidam) Winter (21 hosts), Aniger Van Tieghem (62 hosts), Asulphureus (Fres.) Thom. & Church (10 hosts), Aterreus Thom (48 hosts), Aversicolor (Vuill) Tiraboschi (1 host), Cephalosporium sp. (8 hosts), Chaetomium flavum Omvik (4 hosts), C.globosum kunze ex staud (11 hosts), C,indicum corda (8 hosts), Cladosporium sp. (9 hosts), Cunninghamella echinulate (Thaxt) Thaxt. (6 hosts), Curvularia clavata Jain (3 hosts), C.lunata (wakker) Boedjin (3 hosts), Drechslera austrakiensis (Bugni) Subram. & Jain ex M.B. Ellis (32 hosts), D.hawaiiensis (Bungi) subram. & Jain ex M.B.Ellis (1 host), Fusarium culmuuuurum (W.G.Sm.) Sacc, (2 hosts),F.moniliforme Sheld (8 hosts), F.oxysporum Schlet emend Snyd & Hans (12 hosts), F.proliferatum (Matsushima) Nirenberg (1 host), F.semitectum Berk & Rav, (7 hosts), F,solani (Mart.) Appel & Wallenw. Emend Synd & Hans (26 hosts), Gliomastix murorum Grey (1 host), Macrophomina Phaseolina (Tassi) Goid (22 hosts), Memnoniella echinata (Riv.) Galloway (2 hosts), Monodictys puterdinis (Wallr.) Hughes (1 host), Myrothecium cinctum (Corda) Sacc. (3 hosts), M. roridum Tode ex steudel (5 hosts), Nigrospora oryzae (Berk. & Br.) Petch (2 hosts), Pacilomyces lilacinus (Thom) Samson (9 hosts),Penicillium aspermum (Shear) n.Comb (1 host), P.citrinum Thom (1 host), P.chrysogenum Thom (1 host), P.funiculosum Thom (3 hosts), P.javanicum Van Beyma (1 host), P.luteum Zukal (7 hosts), P.purpurenscens (Sopp) n.Comb (1 host), P.purpurogenum Stoll (5 hosts), P.raistrickii Smith (1 host), P.regulosum Thom (6 hosts), P.wasksmanni Zaleski (2 hosts), Philophora fastigiata ( Lundberg & Melin) Conant. (1 host), Rhizoctonia solani Kuhn (59 hosts), Rhizopus Stolonifer (Ehrenb. Ex. Link) Lind (3 hosts), Scopulariopsis brumptii Salvanet-Duval (2 hosts), Stachybotrys atra Corda (1 host), S.parvispora Hughes (1 host), Trichoderma harzianum Rifai (4 hosts), T.koningii Oudem (1 host) and T.viride Pers ex. Gray (7 hosts). Fifty Four species of fungi/isolates belonging to 18 genera were tested inti vitro for their antigungal activity against Macrophhomina phaseolina (Tassi) Goid, Rhizoctonia solani Kuhn Fusarium solani (Mart.) Appel & Wallenw. Emend Snyd & Hans, F.oxysporum Schlecht emend Snyd & Hans (Plant pathogens), Aspergillus fumigatus Fres. And Candida albicans. Trichoderma viride (S-2), and Paecilomyces lilacinus(S-1) inhibited all the six test pathogens. P.lilacinus (S-4), Chaetomium flavum, Aspergillus candidus and Satchybotrys atra were found active against five pathogens except C.albicans. Whereas P.lilacinus (S-2) and (S-3) were active against five pathogens except R.solani and Afumigatus respectively. Memnomiella echinata, Penicillium raistrickii and Verticillium chlamydosporium showed activity against four test pathogens. Fifty Five species/isolates of fungi belonging to 18 genera were tested in Vitro against 5 bacterial species Viz, Bacillus subtilis (Ehrenberg) Cohn, Staphylococcus aureus Rosenbach (Gram+ve) and Escherichia coil (Migula) Castellani and Chalmers,Pseudomonas aeruginosa (Schraeter) Migula and Salmonella typhimurium (Loeffler) Castellani and Chalmers(Gram-ve). Aspergillus flavus, Curvularia clavata, Fusarium Oxysproum, F.solani (S-2), Penicillium citrinum, P.raistrickii, stachybotrys atra (S-1 & S-2) and Verticillium chlamydosporium were found to inhibit all the test bacteria and produced zone of inhibitions. Aspergillus candidus, Cephalosporium sp., Memnoniella echinata, Myrothecium roridum, P.liliacinus (S-1, S-2 &S-3), S.parvispora inhibited all the test bacteria except S.aureus whereas Aspergillus Sulphureus, Drechslera hawaiiensis, F.solani(S-2) and Trichoderna viride (S-2) also inhibited all the test bacteria except P.aeruginosa. Culture filtrates of most of the test fungi showed better antibacterial activity than antifungal activity. Lyophilization of culture filtrates was found to enhance antibacterial activity of culture filtrates. Where antibiotic production of test fungi were tested in vitro against test fungal pathogens using cellophane paper method; all the test fungi A candidus, A flavus, A fumigatus, C.flavum, C.globosum (S-1), C.indicum, F.solani, F.oxysporum, G.murorum, M.echinata, M.cinctum, M.roridum, P.lilacinus (S-1, S-2,S-3 & S-4), P.raistrickii, S.atra(S-1 & S-2), S.parvispora, Trichoderma Spp, and Talaromyces Flavaus were found to produce fungistatic metabolites against M.phaseolina R.solani, F.solani, F.oxysporum, Afumigatus and C. albicans. However metabolites of Acandidus and S.parvispora showed fungicidal activity against M.phaseolina. Ethyl-acetate fractions of culture filtrates f F.solani, Trichoderma spp., C.globosum, C.flavum, S.atra and P.lilacinus were found more active against test bacteria than n-hexane fractions, indicating intermediary polar nature of most of the antimicrobial compound produced by these fungi. It is interesting to note that n-hexane fraction of some fungi were found active against some bacteria where ethyl-acetate fraction of these fungi were found inactive. In the present study, mutagenesis with UV or acridine orange has induced or enhanced antimicrobial activity in some fungi, although some strains has lost or reduced their activity. Less antimicrobial activity was found in solvent fractions of mycelium as compared to culture filtrates. However methanol and chloroform soluble fractions of Mycelium of Acandidus, chaetomium spp, P.lilacinus, M.echninata, Trichoderma spp. and s.atra were also found active against some test bacteria. Five compunds were isolated and purified from ethyl-acetate fractions of culture filtrate of Memniniella echinata and some of them characterized on the basis of spectroscopic techniques. Two compounds named ME-2 and ME-5 showed significant antibacterial activity. Whereas four compounds from S.parvispora and five from C.flavum were isolated and purified their culture filtrates and spectral data were obtained. Two compounds named CE-2 and CE-3 from C.flavum showed significant antibacterial activity against B.subtilis, S,typhimurium, S.aureus and P.aeruginosa. This is first report of antibacterial activity of pure compounds of S.parvispora and C.flavum.