Fungi associated with Seeds of some economically important plants

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1 Mycopth (27) 5(1): 35-4 Fungi ssocited with Seeds of some economiclly importnt plnts Nureen Akhtr, J.H. Mirz, Rukhsn Bjw & Amn Jvid Deprtment of Mycology & Plnt Pthology, University of the Punj, Lhore Astrct Sixteen different species were isolted using Agr Plte method nd Blotter method. Isoltions were mde from the seeds of twelve plnts viz., Ze mys L., Aven stiv L., Nigell stiv L., Crum copticum (L.) Clrke, Aelmoscus esculentus L., Glycine mx (L.) Merrill, Luff cylindric (L.) Roem. Pennisetum typhoides (Burm.) Stpf., Brssic cmpestris (L.) Czern., Cicer rietinum L., Cuminum cyminum L., nd Hordeum vulgre L. Gener Isolted from seeds were Aspergillus, Penicillium, Monili, Drechsler, Mucor, Alternri, Cldosporium, Fusrium, Acremonium, Rhizopus, Tuerculri, Phom, nd Trichoderm. Among ll the tested plnts Z. mys seeds were found to e hevily colonized y fungi., A. flvus Link, A. fumigtus Fresenius nd A. niger Vn Tieghem were the most prominent fungi isolted in present study. C. cyminum, H. vulgre nd C. copticum were found to e infected only y Aspergillus. Key Words: Aspergillus, gr plte, isoltion, seeds. Introduction Post-hrvest spoilge y filmentous fungi is one of the most importnt threts ssocited with processed nd stored food products worldwide. Discolortion, qulity deteriortion, reduction in commercil vlue nd mycotoxin production hs een linked to moldy contminted foods (Moreu, 1968, Christensen nd Kufmn, 1969). This sitution is mde worse in the tropics where the wrm nd humid climtes provide these micro orgnisms with fvorle conditions for their spred nd susequent estlishment in numerous sustrtes. Three gener viz. Fusrium, Penicillium nd Aspergillus, ll potentil mycotoxin producers, could e considered the most significnt toxigenic fungi growing in processed nd stored foods. Due to their cpility to develop in wide rnge of environmentl conditions, fungi in the genus Aspergillus re comprtively more widespred thn others (Chelkowski, 1991). Aspergilli re economiclly, ecologiclly, nd mediclly importnt nd constitute lrge genus. Consequently, specil cre is to e devoted to them, especilly s they could ply n importnt role in food decy nd mycotoxin formtion under certin storge conditions. There re mny ccounts of Aspergillus occurring in processed nd stored griculturl commodities (Jvid nd Anjum, 26). Adel-Gwd nd Zohri (1993) & Mzen et l., (199) documented the spectrum nd levels of Aspergillus growing on nuts nd cotton seedssed products. The presence nd incidence of Aspergillus spp. in cerel-sed foods, milks, oils nd penuts re lso well known (Mne nd Tsurut, 1991; Adejo, 1993). Crcinogenic fltoxins produced y Aspergillus spp. re common contminnts of seeds (Klich, 22). In Pkistn, the phenomenon could e of gret concern, especilly in res where food shortges hve compelled people to consume low grde food mteril, even if moulds re visile s contminnts. The present study is imed t documenting the spectrum of Aspergillus spp. growing on seed of selected plnts. Mterils nd Methods Seeds of selected plnts i.e. Ze mys, Aven stiv, Nigell stiv, Crum copticum, Aelmoscus esculentus, Glycine mx, Luff cylindric, Pennisetum typhoides, Brssic cmpestris, Cicer rietinum, Cuminum cyminum, nd Hordeum vulgre were collected from Lhore nd vicinity. Agr plte nd Blotter method were used for isoltion of fungi (Mthur et l., 1975). In Agr plte method surfce seeds were plted septiclly on PDA (Potto Dextrose Agr) nd in lotter method on lyer of three sterilized moisten filter 1 seeds per plte. These petripltes were incuted t 25 o C. Ech smple ws replicted thrice for ech method. Fungi growing on different seeds were isolted, purified nd identified fter reference to Brron (1968), Ellis (1971), Brnett (1972) nd Rper nd Fennel (1965). Percentge occurrence of ech genus ws clculted y using the formul.

2 36 N. Akhtr et l. % occurrence of genus= No of colonies of genus X 1 Totl No of Colonies Pure cultures of fungi were mintined on PDA nd deposited to First Fungl Culture Bnk of Pkistn. All dt ws nlyzed y nlysis of vrince followed y Duncn s Multiple Rnge Test for men seprtion. (Steel nd Torrie, 198) Results nd Discussion Twenty seven Aspergillus isoltes elonging to 16 species were isolted from the 12 different plnt seeds smples. The Aspergillus species isolted from seed smples were A. flvus, A. fumigtus, A. prsiticus, A. niger, A. sydowi, A. jponicus, A. prsiticus, A. lliceus, A. terreus, A. versicolor, A. sprsus, A. clvto-flvus, A. cronrius, A. urtus, A. pulvinus nd Emericell nidulns. Among tested seeds, mximum infection ws recorded in Ze mys seeds s 2 isoltes of fungi elonging to 1 different gener viz., Aspergillus Penicillium, Monili, Drechsler, Mucor, Alternri, Cldosporium, Fusrium, Acremonium nd Rhizopus were screened. (Fig. 1). Six different species of Aspergilli i.e., A. flvus, A. niger, A. lliceus, A terreus, A. versicolor nd A. glucus cused 3% infection in Z. mys seeds. In cse of seeds of G. mx, 6 fungl gener nmely Aspergillus, Rhizopus, Mucor, Penicillium, Tuerculri nd Phom with lmost equl percentge frequency of occurrence were recorded (Fig. 2). However only one species of Aspergillus, A. flvus ws recovered in this cse. Five fungl gener i. e., Aspergillus, Trichoderm, Acremonium, Fusrium nd Monili were identified from the seeds of A. stiv (Fig. 3). In this cse Aspergillus nd Trichoderm were eqully distriuted with 18% infection in A stiv seeds. From the seeds of L. cylindric, 4 different fungl gener i.e., Aspergillus, Mucor, Monili nd Fusrium were isolted (Fig. 4). In L. cylindric, spergilli showed 6% infection in seeds. In cse of B. cmpestris, (Fig. 5) Aspergillus, Rhizopus, Monili nd Alternri were reported. In this cse Aspergillus did not pper s the prominent genus which is the only exception in present study. Species of Aspergillus, Acremonium nd Drechsler were identified from the seeds of Pennisetum typhoides (Fig. 6). Two species of Aspergillus i.e. A. sydowi nd A. jponicus constituted 5 % infection of seeds. Seeds of oth Cicer rietinum (Fig. 7) nd Aelmoscus esculentus (Fig. 8) were colonized with only with one genus i.e., Fusrium nd Monili, respectively long with Aspergillus which is in higher percentge frequency of occurrence. It ws interesting to note tht C. cyminum, H. vulgre nd C. copticum were found to e 1 percent infected only y Aspergillus (Fig. 9). The 16 different species of Aspergillus mentioned in this pper hve een reported in other food commodities elsewhere. In generl, ll the Aspergillus spp. referred to in this study re common nd distriuted in nture worldwide, nd hve een isolted in wide rry of sustrtes (Kozkiewicz, 1994). Mzen et l. (199) isolted 12 species of Aspergillus from cotton seeds nd cotton seeds products in Egypt. Mne nd Tsurut (1991) isolted 17 Aspergillus spp. from stored rice grins. Similrly, in other study, Essono et l., 27 reported different species of Aspergilli, some of which re sme s reported in present investigtion. According to them, stored food products re suitle sustrtes for the growth nd development of Aspergillus spp. However, presence of other fungl species in smple nlyzed my e due to the differences in methods of isoltion which hd greter influence (Essono et l., 27). Presently surfce steriliztion with NClO of seeds llowed the coloniztion of vriety of fungl flor, which is in line with Msikit (1995), who ttriuted this to the lck of protective shell in foods such s mize, groundnuts nd rice. Suer nd Burroughs (1986) rgued tht such method (sence of steriliztion) is likely to ring out some overestimtion in the ctul internl composition of the microflor popultion when portions of studied smples re to e plted onto culture medi for nlyses. Among ll tested seeds, Z. mys showed mximum coloniztion nd B. cmpestris lest colonized y Aspergilli memers, which support erlier reserch showing the role of the seed cot s rrier for invsion (Crter, 1973). Tnnins, wxes, mino compounds nd structurl fetures in seed cot hve een implicted in resistnce to invsion y Aspergillus species (Horn et l., 25). In contrst to mycoflor exmined on other plnt seeds viz., C. copticum, C. cyminum nd H. vulgre were found to e 1 percent infected only y Aspergillus ttriuted to the fct the source of infection y other fungi in inoculted seeds nd grin ws primrily soil. Moreover, competitive ility in fungi is complex nd depends on environmentl vriles such s sustrte composition, wter ctivity, temperture nd inoculum density (Ryner nd Weer, 1984). Aspergillus species hve een reported from diverse sustrtes ut re found most frequently in oil-rich seeds nd grins (Pyne, 1998) where they often hve competitive dvntge over other Mycopth (27) 5(1): 35-4

3 Percentge occurrence of fungl gener Percentge occurrence of fungl gener Percentge occurrence of fungl gener Study of fungi ssocited with seeds 37 fungi t wter ctivities of <.96 nd temperture of pproximtely 3 C (Mrín et l.,1998). In ddition to environmentl prmeters, physiologicl chrcters such s rte of spore germintion, rte of mycelil growth nd enzymtic cpility gretly influence competitive sprophytic ility in fungi (Grrett 197) cd c 5 d d d d d d Aspergillus Penicillium Monili Drechsler Mucor Alternri Cldosporium Fusrium Acremonium Rhizopus Fig. 1 Fungl gener isolted from Ze mys seeds Fig 1: Fungl gener isolted from Ze mys seeds Aspergillus Rhizopus Mucor Penicillium Tuerculri Phom Fig 2: Fungl gener Fig. 2 isolted Fungl gener from Glycine isolted mx from seeds. Glycine mx seeds Aspergillus Trichoderm Acremonium Fusrium Monili Fig 3: Fungl Fig.3 gener Fungl isolted gener from isolted Aven stiv from seeds. Aven stiv seeds Mycopth (27) 5(1): 35-4

4 Percentge occurrence of fungl gener Percentge occurrence of fungl gener Percentge occurrence of fungl gener 38 N. Akhtr et l c Aspergillus Mucor Monili Fusrium Fig. 4: 4 Fungl gener isolted isolted from Luff from cylindric Luff cylindric seeds. seeds Aspergillus Rhizopus Monili Alternri Fig 5: Fig. Fungl 5 Fungl gener gener isolted isolted from Brssic from cmpestris Brssic cmpestris seeds. seeds Aspergillus Acremonium Drechsrler Fig 6: Fig. Fungl 6 Fungl gener gener isolted isolted from Pennisetum from Pennisetum typhoides seeds. typhoides seeds Mycopth (27) 5(1): 35-4

5 Percentge occurrence of Aspergillus Percentge occurrence of fungl gener Percentge occurrence of fungl gener Study of fungi ssocited with seeds Aspergillus Fusrium Fig 7: Fungl Fig. 7 Fungl gener isolted gener isolted from Cicer from rietinum Cicer rietinum seeds. seeds Aspergillus Monili Fig. 8: 8 Fungl gener isolted from from Aelmoscus Aelmoscus esculentus esculentus seeds. seeds 12 1 c d Cuminum cyminum Hordeum vulgre Crum copticum Nigell stiv Fig 9: Aspergilli isolted from Fig. 9 seeds. Isolted Aspergilli from seeds Note: Vlues with different letter show significnt difference (P.5) s determined y Duncn s Multiple Rnge Test. Mycopth (27) 5(1): 35-4

6 4 N. Akhtr et l. References Adel GK, Zohri M, Fungl flor nd mycotoxins of six kinds of nuts seeds for humn consumption in Sudi Ari. Mycopthologi, 124: Adejo LO, Survey of Afltoxins nd Ochrtoxin A in stored tuers of Cyperus esculentus L. Mycopthologi, 124: Brnett HL, Hunter BB, Illustrted gener of Imperfect Fungi. Burgess Co., Miennepolis, Minnesot. Brron GL, The Gener of Hyphomyceres from Soil. Bltimre, Willims nd Wilkins. Co. Crter JBH The influence of the test, dmge nd seed dressing on the emergence of groundnut (Archis hypoge). Ann. Appl. Biol., 74: Chelkowski J, Cerel grin mycotoxins, fungi nd qulity in drying nd storge. In developments in food science: Cerel grin mycotoxins, fungi nd qulity in drying nd storge Elsevier, Amsterdm, The Netherlnds, pp Christensen CM, Kufmn HH, Grins storge. The role of fungi in qulity loss. University of Minnesot Press, Minnepolis. Ellis MB, Demticeous Hyphomycetes. CMI, Kew, Surrey, Englnd. Essono G, Ayodele M, Ako A, Foko J, Olemo S, Gockowski J, 27. Aspergillus species on cssv chips in storge in rurl res of southern Cmeroon: their reltionship with storge durtion, moisture content nd processing methods. Afri. Jour. Microiol. Res pp. 1-8 Grrett SD Pthogenic root-infecting fungi. Cmridge, Englnd: Cmridge University Press. 294 p. Horn B, Greene WRL, Dorner JW, 25. Effect of corn nd penut cultivtion on soil popultions of Aspergillus flvus nd A. prsiticus in southwestern Georgi. Ntionl Penut Reserch Lortory, U.S. Deprtment of Agriculture, Dwson, Georgi. Appl. Eviron. Microiol., 61: Jvid A, Anjum T, 26. Fungi ssocited with seeds of economiclly importnt crops in Pkistn. Pk. J. Seed Technol., 1(8 & 9):55-61 Klich MA, 22. Biogeogrphy of Aspergillus species in soil nd litter. Mycol., 94(1): Kozkiewicz, Z., Txonomy: the key to mycotoxin identifiction in food nd feedstuffs. In: Proceedings of the 6th Interntionl Working Conference on stored-product protection. Ntionl convention centre, Cnerr. Highley E, Wright EJ, Bnks HJ, Chmp BR (eds.). CAB. Mne M, Tsurut O, Mycoflor nd Mycotoxin in stored rice grin. In: Developments in food science: Cerel grin mycotoxins, fungi nd qulity in drying nd storge J Chelkowski (ed.). Elsevier, Amsterdm, The Netherlnds, pp Mrín S, Snchis V, Rmos AJ, Vins I, Mgn N Environmentl fctors, in vitro interctions, nd niche overlp etween Fusrium moniliforme, F. prolifertum, nd F. grminerum, Aspergillus nd Penicillium species from mize grin. Mycol Res., 12: Mthur SA, Mtur SB, Neergrd P, Detection of seed orne fungi in sorghum nd loction of Fusrium moniliforme in seed. Seed Sci. Technol., 3: Mzen MB, Kdey IA, Ser SM, 199. Survey of the mycoflor nd mycotoxins of cotton seeds nd cotton seed products in Egypt. Mycopthologi, 118: Moreu C, Moisissures toxiques dns l limenttion. P Lechevlier (ed.), Pris. Frnce, p. 371 Msikit W Preliminry results ssocited with the contmintion of cssv chips y fungl flor in Nigeri nd Benin. Internl reserch report 6, IITA, Idn, Nigeri Pyne GA Process of contmintion y fltoxin-producing fungi nd their impct on crops. In: Sinh KK, Bhtngr D, eds. Mycotoxins in griculture nd food sfety. New York: Mrcel Dekker. p Rper KB, Fennell DI, The genus of Aspergillus. The Willims & Wilkins Co., Bltimore. Ryner ADM, Weer JF Interspecific mycelil interctions n overview. In: Jennings DH, Ryner ADM, eds. The ecology nd physiology of the fungl mycelium. Cmridge, Englnd: Cmridge University Press. p Suer DB, Burroughs R, Disinfection of seed surfces with sodium hypochlorite. Phytopthology, 34: Steel RGD, Torrie JH, 198. Principles nd Procedures of Sttistics: A Biometricl Approch (2nd Ed.). McGrw-Hill Inc., New York. Mycopth (27) 5(1): 35-4

7 Study of fungi ssocited with seeds 41 Mycopth (27) 5(1): 35-4