微生物分析方法

Microorganisms play an integral and often unique role in ecosystem functions, yet we know little about dominant populations that presumably play vital roles in these functions. Soil microbial communities are the most complex, diverse, and important assemblages in the biosphere.Analysis of genetic diversity in soil communities by DNA renaturation suggests that there are at least 103 or even more species per gram of soil (Torsvik et al., 1990). However, many previous studies revealed function groups with few or no known the dominant populations at the division or phylum level, perhaps because only a limited number of microorganisms have been obtained in the isolating experiments. Virtually, in many ecosystems, activity of microorganisms is limited by the availability of substrate (Belser, 1979); thus we can image how difficulty for one thousand or even more species survive in one gram of soil! Hence, most of the species cells probably are dormant in soil in without extra carbon input condition. Several experiments on organic matter decomposition support our inference, when fresh plant residues in soil, fluctuations of microbial biomass N and C could be observed (Calderόn et al., 2001; Griffiths et al., 2001; Priemé and Christensen, 2001).So, the dominant microorganism populations in soil are not only have great number of cells, but also must be active in soil. To better understand the dominant microorganism populations in soil, all activity microorganisms were assumed to take up carbon only in the form of soluble, directly consumable substrate dissolved in soil solution. Soil-water-extraction media were used (Semenov et al., 1996; Zelenev et al.,2000), two trophic groups of microorganisms (copiotrophs and oligotrophs) were selected.Semenov, A.M., Batomunkueva, B.P., Nizovtseva, D.V., Panikov, N.S., 1996. Method of determination of cellulase activity in soils and in microbial cultures, and its calibration. Journal of Microbiological Methods 24, 259–267.Zelenev, V.V., Berkelmans, R., van Bruggen, A.H.C., Bongers, T., Semenov, A.M., 2004. Daily changes in bacterial feeding nematode populations oscillate with similar periods as bacterial populations after a nutrient impulse in soil. Applied Soil Ecology 26, 93–106.Torsvik, V., Goksoyr, J., Daae, F., 1990. High diversity in DNA of soil bacteria. Applied Environmental Microbiology 56, 782–787.Belser L. W. (1979) Population ecology of nitrifying bacteria. Annual Review of Microbiology 33, 309-333.Calderόn, F.J., Jackson, L.E., Scow, K.M., Rolston, D.E., 2001. Shortterm dynamics of nitrogen, microbial activity, and phospholipids fatty acids after tillage. Soil Science Society of America Journal 65,118–126.Griffiths, B.S., Bonkowski, M., Roy, J., Ritz, K., 2001. Functional stability, substrate utilisation and biological indicators of soils following environmental impacts. Applied Soil Ecology 16, 49–61.Priemé, A., Christensen, S., 2001. Natural perturbations, drying-wetting andfreezing-thawing cycles, and the emission of nitrous oxide, carbon dioxide and methane from farmed organic soils. Soil Biology &Biochemistry 33, 2083–2091.premise received from due owing signature筛选habitat.However, none of these studies have reached the level of diversity revealed by DNA–DNA hybridization methods, Thus, despite recent expansion in divisions by several previous studies (Dojka et al., 1998; Hugenholtz et al.,1998a,b), our understanding of the extent of microbial diversity is still incomplete. More studies on a variety of soil types and habitats are needed to obtain a comprehensive view of microbial community composition and structure in soil environments.Conversion of natural ecosystems to agriculture as well as increasing intensity of tillage are known to decrease soil organic matter (SOM) levels and contribute significantly to the increase in atmospheric CO2 concentration (Lal et al., 1998). Between 1850 and 1995, SOM mineralization emitted 136 ±55 ×1015 g of CDuring several years we investigated the stages of corn residue decomposition in Mollisols and observed significant oscillations of microorganism populations: Pseudomonas is the dominant population during 40-80d decomposition period; and then at the period of 80-100d decomposition, Bacillus, Penicillium, Aspergillus, Mucor are the dominant populations; At 180d decomposition, Stachybotrys, Trichoderma, Dematium and Phoma are the dominant populations; At 240d decomposition, Polyangium, Cellvibro, Nocardia, Micromonospora and Penicillium are the dominant populations.two trophic groups of microorganisms: Zymogeneous microorganisms and Autochthonous microorganisms. The former have low substrate affinity and grow relatively fast on C rich media (1000mgCL-1), while the latter have high substrate affinity and grow more slowly, but better on C poor (10mgCl_1) than on C rich media. Copiotrophic and oligotrophic bacteria roughly correspond to r- andK-strategists, but there are many exceptions in both trophic groups.or incorporation ofCorn is the most important cash crops of China, and are mostly produced in Northeast China. This large area is covered by different type of Mollisols. These soils vary greatly in their inherent properties, like morphology, texture and mineralogy. Soils were under grassland vegetation up to about 200 years ago. Since then, they have different agricultural histories, which resulted in different degradation levels soils lost 42 and 59% of their soil organic carbon (SOC) stock upon conversion from forest to crop and from grassland to crop, respectively.In contrast, Mann (1986) calculated losses <20% of the initial stock, which wascorroborated by Kern and Johnson (1993) andConversion of natural ecosystems to agriculture as well as increasing intensity of tillage are known to decrease soil organic matter (SOM) levels and contribute significantly to the increase in atmospheric CO2 concentration (Lal et al., 1998). Between Lal, R., Follett, R.F., Kimble, J.M., Cole, C.V., 1998. The Potential of US Cropland to Sequester Carbon and Mitigate the Greenhouse Effect. Lewis Publisher, Boca Raton, FL, 128 pp.residues returned 1993).From the 1990 decade the farmers grew soybean using zero tillage (ZT) systems, which progressively replaced the more aggressive conventional tillage (CT) systemsbroth blended horizonThe main objectives of this paper are: (1) to evaluate environmental factors that regulate bacterial abundance during guano decomposition; (2) to estimate the rate of decomposition of guano with an emphasis on percentage changes in fats, protein, chitin, ash, TOC (total organic carbon), nitrogen, phosphorus, and some other elements; (3) to examine the quantitative and qualitative transformations in the bacterial population during penguin guano decomposition; (4) to isolate and identify the dominant culturable bacteria as a first step in evaluating their diversity, and to preselect strains for further phylogenetic analysis.Culturable bacteria (colony forming units, CFU) were enumerated by the spread plate method on: (a) full strength nutrient Soil Extract Agar (SEA) for the isolation of copiotrophic bacteria, and 10! diluted NA for the bacteria more oligotrophic (Ogram and Feng, 1997). Agar plates were inoculated with 100 ml from each dilution in a decimal series of the homogenate. One liter of NA includes bacto beef extract (3 g), bacto-tryptone (5 g), bacto-agar 15 g (Fenchel and Hemmingsen, 1974) in 1 l of soil extract prepared according to Klement et al. (1990). CFU growth was examined every 2–3 days through 25–30 days incubation at 10 8C. This was done to compare the kinetics of bacterial colony increment among bacterial populations, isolated in different stages of guano decomposition. At theend of incubation, CFU counts were calculated. The standard error (SE) in CFU counts was determined from four replicates. For each experimental day (0, 2, 6, 10, 15, 20, 25, 30, 42), an average of 15 colonies were randomly selected, and subcultured for purification through repeated transfers on the same medium, and physiological investigations. In total, 136 colonies were so isolated.Chitinolytic bacteria were enumerated on a solid medium containing solubledye-labeled chitin (CM-Chitin-RBV solution; LOEWE Biochemica GmbH) prepared according to Klement et al. (1990).Morphological, physiological and biochemical features of the isolated bacteria were determined on the basis of 49 tests, including growth at 4, 22 and 32 8C, Gram stain, morphology (rods or cocci), motility (det ermined in hanging drop preparations), colony pigmentation (examined macroscopically), growth in 4% NaCl, and responses in API 20NE and API ZYM systems (API bioMe´rieux; Tearle and Richard, 1987; Zdanowski and Donachie, 1993; Zdanowski and Weglenski, 2001). A dendrogram showing the hierarchical classification of isolates based on the tests described was constructed by the simple matching coefficient of Sokal and Michener (1958) in association with the weightedpair-group-average algorithm (Sneath and Sokal,1974).Ogram, A., Feng, X., 1997. Methods of soil microbial community analysis. In: Hurst, C.J., Knudsen, G.H., McInerney, M.J., Stetzenbach, L.D., Walter, M.V. (Eds.), Manual of Environmental Microbiology. ASM Press, Washington, DC, pp. 422–430.Fenchel, T., Hemmingsen, B., 1974. Manual of Microbial Ecology.Akademisk Forlag. University of Aarhus, pp. 1–235.Mollisols are the soils of grassland ecosystems. They are characterized by a thick, dark surface horizon. This fertile surface horizon, known as a mollic epipedon, results from the long-term addition of organic materials derived from plant roots.Mollisols primarily occur in the middle latitudes and are extensive in prairieregions such as the Great Plains of the US. Globally, they occupy ~7.0% of the ice-free land area. In the US, they are the most extensive soil order, accounting for ~21.5% of the land area.Mollisols are among some of the most important and productive agricultural soils in the world and are extensively used for this purpose.Mollisols are divided into 8 suborders: Albolls, Aquolls, Rendolls, Gelolls, Cryolls, Xerolls, Ustolls, and Udolls. Click here for more information about these suborders. Click here to view a map of their distribution in the US. Microbes play an important role in soil formation, ecosystem biogeochemistry,(Konopka and Turco, 1991; Hiebert and Bennett, 1992; Madsen, 1995; Richter and Markewitz, 1995), Konopka, A., Turco, R., 1991. Biodegradation of organic compounds invadose zone and aquifer sediments. Applied and EnvironmentalMicrobiology 57, 2260–2268.Hiebert, F.K., Bennett, P.C., 1992. Microbial control of silicate weatheringin organic-rich ground water. Science (Washington DC) 258, 278–281.Madsen, E., 1995. Impacts of agricultural practices on subsurface microbialecology. Advances in Agronomy 54, 1–67.Richter, D., Markewitz, D., 1995. How deep is soil? BioScience 45,600–609.Microbial community composition may be one important control on soil processes (Schimel, 1995; Cavigelli and Robertson, 2000; Balser et al., 2002).Schimel, J., 1995. Ecosystem consequences of microbial diversity andcommunity structure. In: Iii, F.C., Korner, F. (Eds.), Arctic and AlpineBiodiversity, Ecological Studies, vol. 113. Springer, New York, pp.239–254.Cavigelli, M.A., Robertson, G.P., 2000. The functional significance ofdenitrifier community composition in a terrestrial ecosystem. Ecology(Washington DC) 81, 1402–1414.Balser, T., Kinzig, A., Firestone, M., 2002. The functional consequences ofbiodiversity. In: Kinzig, A., Pacala, S., Tilman, D. (Eds.), TheFunctional Consequences of Biodiversity, Princeton University Press,Princeton, pp. 265–293.From 2003 to 2004,108 black soil samples of corn crop were taken fromYushu,Dehui,Changchun and GongzhulingoWe used three kinds of isolating methods(Thin liquid plate method, Soil particle washing technique and Soil plate method ),131 Isolates obtained were identified. The partial soil sample mycobiota and ecological characteristics were studied.Mycobiota studies were focused on soil fungi quantity, genus and species distribution of black soil corn crop .The varity of soil fungi quantity of black soil corn crop in different films were studied,the varity of genus and species in different films were studied,too. We analysed and discussed seasonal variation and veritical distribution of black soil fungi.Ecological characteristics preliminary studied the movement of black soil fungi number and the relationship with 6 kinds of environmental factors(Water contents,pH, available nitrogen, available phosphorus,available postassium and organic matter) through the timing of the corn growing.And studied the varity of black soil mycobiota in different fertilizational black soil.On the basis of morphological observation, we identified as 25 genera,55 species and 4 varieties of fungi. 14 species are the new recorded species in Jilin.25 genera, Acremonium Link, Alternaria Nees, Arthrobotrys Corda.、Aspergillus (Mich.) Link, Beauveria Vuill., Circinella sp.v.Tiegh. et Monn., Cladosporium Link, Curvularia Boed.、Epicoccum Link, Exophiala Carm., Fusarium Link, Geotrichum Link., Humicola Traaen, Kabatiella Bub.、Mucor Mich. ex Fr.、Myrothecium Tode, Neurospora Shear et Didge.、Paecilomyces Bain.、Papulaspora Preuss.、Penicillium Link, Phialophora sp.Medlar.、Pythium Pringsh.、Rhizopus Ehrenb.、Scopulariopsis Bain. and Trichoderma Pers.. 59 species刀cremonium kiliense Griitz., Acremonium strictum WGams. Alternaria alternata (Fr.) Keissl.、Arthrobotrys superba Corda.、Aspergillus sydowii (Bain.et Sart.)Thom et church.、Aspergillus carnus IV (v.Tiegh.)Thom., Aspergillus niger v.Tiegh.、Aspergillus terreus Thom var.aureus Thom&Raper.、Aspergill us如vus Link:Fr.var.flavus, Aspergillus melleus Yukawa., Aspergillus penicilloides Speg.、Aspergillus puniceus Kwon&Fenn.、Aspergillus fumigates Fres.、Aspergillus ustus (Bain)Thom&Church.、Aspergillus ochraceus Wilh.、Aspergillus versicolor(Vuill.)Tirab.vanversicolor, Aspergillus terricola March., Beauveria brongniarth (Sacc.) Petch.、Cladosporium herbarum (Pers.) Link blab Sacc, Cladosporium carpophilum Thum.、Cladosporium tenissimum Cooke.、Curvularia lunata (Walk.) Boed.、Epicoccum purpurascens Ehrenb.、Exophiala jeanselmer (Langeron) McGinnis.、Fusarium oxysporum Schlecht., Fusarium mondiforme.var.inteimedium, Fusarium merismoides Corda.、Geotrichum candidum Link.、Humicola grisea Traaen.、Humicola fuscoatra Traaen., Kabatia microsticta Bubak.、Mucor hiemalis Wehmer., Myrothecium roridum Tode, Neurospora crassa Shear et Dodge.、Paecilomyces marquandii (Mass.) Hughes., Paecilomyces varioti Bain.、Paecilomyces inflatus (Burnside) Carmichael., Paecilomyces lilacinus (Thom)Samson., Papulaspora equi Shadomy&Dixon.、Penicillium citrinum Thom.、Penicillium decumbens Thom.、Penicillium nigricans (Bain.) Thom.、Penicillium purpurogenum stoll.、Penicillium corylophilum Dierckx.、Penicillium citreomigrum Dierckx.、Penicillium rubrum Stoll.、Penicillium implicatum Biourge.、Penicillium thomii Maire.、Penicillium spinulosum Thom.、Penicillium islandicum Sopp.、Penicillium waksmanii Zal.、Penicillium restrictum Gilm.et Abbott.、Penicillium janthinellum Biourge.、Pythium acanthophoron Sid., Pythium caroliniarum Matth., Rhizopus stolonifer (Ehrenb.) Lind.、Scopulariopsis canadensis Morton&Smith.、Trichoderma atroviride P.karsten,Finl. And Trichoderma harziamum Rifai,Mycol..The results of mycobiota studies showed that Penicillium and Aspergillus is the main genus.The seasonal variation of soil fungi quantity of black soil corn crop in different films is different.There are more fungi in the topsoil(0-20cm deep) than in the subsoil layers(20-40cm deep), the composition of fungi in topsoil and subsoil horizon of all plot s is different.The results of ecological characteristics studies showed that the water contents have straight relationship with the soil fungi numbers in soil layers of normal fertilization soil, it isBacterial and fungal cell counts in the commercial formulations were determined using standard serial dilution plating on 0.1% tryptic soy agar (TSA) for bacteria and potato dextrose agar (PDA) with 50 mg chlortetracycline and 1 ml/L tergitol for fungi (Larkin, 2003). Bacterial counts of_1_109 CFU/g formulation for Bsub and fungal countsof 6.5–7_106 CFU/g for Tvir and Tharz were observed.EM (EM-1; EMRO USA, Tucson, AZ, USA), a mixLarkin, R.P., 2003. Characterization of soil microbial communities under different potato cropping systems by microbial population dynamics, substrate utilization, and fatty acid profiles. Soil Biology & Biochemistry 35, 1451–1466.The study area is located on the Snake River Plain, between4283000000 and 4383000000N and 11482000000 and 11683000000W. The sites occur across an elevation gradient ranging from 860 to1300 m. The area is classified as a temperate semi-desertActive bacteria and fungal numbers in soil were determined for each treatment using methods described by Ingham and Klein (1984). Active bacteria and fungi were estimated by taking a 1.0 ml water sample that was diluted in 9ml of a phosphate buffer (pH 6.0) and shaken at approximately 120 rpm for 5 min. For fungal biomass, a 1 ml aliquot was removed and stained with 1 ml of a 20 fg ml_1 fluorescein diacetate (FDA) solution in a 0.2 M phosphate buffer for 3min.One milliliter of 1.5% agar in a 0.1 M phosphate buffer (pH 9.5) was added to the FDA suspension. The sample was mixed and an aliquot placed on a microscope slide containing a cavity of known volume (Ingham and Klein, 1984). Immediately after preparation, slides were examined for FDA-stained hyphal length by epifluorescent microscopy.Total fungal biomasswas estimated by measuring the length and diameter ofhyphae in 3–6 fields with phase—contrast microscopy. Threeslides were evaluated from each sample and ten fields perslide were evaluated with phase contrast microscopy for totalhyphal length, and three transects were evaluated for FDAstained(active) hyphal length at 160 total magnification.Iodonitrotetrazolium (INT) stain was used for countingactive bacteria (Stadmatiadis et al., 1990). A 1 ml sample ofinitial soil suspension was diluted to a final dilution in a 0.2 mgsoil in 4ml buffer. The suspension was incubated with 4ml offiltered INT buffer for 60 min in the dark at 20 8C. Total bacteriaper ml of water were estimated from the mean number ofbacteria (fluorescent and non-fluorescent bacteria), theiraverage diameter and length per field. Three slides wereevaluated for each sample and ten fields per slide wereevaluated using epifluorescent oil-immersion microscopy todetermine numbers and size of fluorescent and total bacteria(Lodge and Ingham, 1991).Bacterial biomasswas computed fromthe numbers of activeand total bacteria and active and total fungal biomass was determined fromhyphal length. Bacterial biomass was computedfromthe number of soil bacteria per gramof soil by consideringthat the bacterial spheres were 1 mm in diameter. Active andtotal fungal biomass were computed by considering averagehyphal diameter to be 1 mm in diameter and them multiplyingIngham, E.R., Klein, D.A., 1984. Soil fungi relationships between hyphal activity and staining with fluorescein diacetate. Soil Biol. Biochem. 16, 273–278. Microorganisms exhibit an impressive diversity in their metabolic activities and in their interactions with other microbes, plants, and animals. Microbes have much shorter turnover rates than higher plants and therefore, respond more quickly to changes in land management than plants and may be the most sensitive indicator to anthropogenic activities. In the past it has been nearly impossible to discern underlying patterns within ecosystems because of the intrinsic structural and functional diversity of microbial communities. To date, only 1–5% of the world’s microorganisms have been identified (Mills et al., 2007; Nannipieri et al., 2003; Hugenholtz et al.,1998) As a result, microbial processes, the microbes are often placed in a ‘black box’, inputs and outputs measured, but most of the processes that go on inside ‘the box’ are based on inference (Mills et al., 2007; Swift et al., 2004; Brussaard et al.,1997). Recently, there has been an explosion in the identification of microorganisms in the natural world using culture independent techniques. The structure and function of these microbial communities using new molecular technologies can be determined (Mills et al., 2007; Thies, 2007; Nakatsu, 2007; Rogers et al., 2007; Suzuki et al., 1998).Mills, D.K., Entry, J.A., Gillevet, P.M., Mathee, K., 2007. Assessing microbial community diversity using amplicon length heterogeneity polymerase chain reaction. Soil Sci. Soc. Am. J. 71, 572–578.Thies, J.E., 2007. Soil microbial community analysis using terminal restriction fragment polymorphisms. Soil Sci. Soc. Am. J. 71, 579–591.Rogers, S.W., Moorman, T.B., Ong, S.K., 2007. Fluorescent in situ hybridization and micro-autoradiography applied to ecophysiology in soil. Soil Sci. Soc. Am. J.71, 620–631.Suzuki, M., Rappe, M.S., Giovannoni, S.J., 1998. Kinetic bias in estimates of Coastal Picoplankton community structure obtained by measurements ofsmall-subunit rRNA Gene PCR amplicon length heterogeneity. Appl. Environ. Microbiol. 64, 4522–4529.Thies, J.E., 2007. Soil microbial community analysis using terminal restriction fragment polymorphisms. Soil Sci. Soc. Am. J. 71, 579–591.Changes in SOM by different agricultural land uses and practices have been extensively reviewed by Mann (1986), Davidson and Ackerman (1993), Guo and Gifford (2002), and Murty et al. (2002).Guo, L.B., Gifford, R.M., 2002. Soil carbon stocks and land use change: a meta analysis. Glob. Change Biol. 8, 345–360.Murty, D., Kirschbaum, M.U.F., McMurtie, R.E., McGilvray, A., 2002. Does conversion of forest to agricultural land change soil carbon and nitrogen? A review of the literature. Glob. Change Biol. 8, 105–123.ISP 4号培养基、察氏（Czapack）培养基；添加2% 玉米秸秆species S.F (%) S.E.M species S.F (%) S.E.M Acremonium strictum 20.4%Mucor hiemalis43.2% Alternaria alternato27.2%Mucor spinescens39.6% Aspergillus carnus20.0%Neurospora crassa42.4% Aspergillus flavus18.8%Paecilomyces inflatus44.4% Aspergillus fumigatus27.2%Paecilomyces lilacinus 38.8% Aspergillus melleus21.2%Paecilomyces marquandii 34.4% Aspergillus niger84.0%Paecilomyces varioti31.2% Aspergillus ochraceus 18.8%Papulaspora equi27.6% Aspergillus penicilloides 20.8%Penicillim rubrum44.8% Aspergillus puniceus16.4%Penicillim thomii41.2% Aspergillus sydowil18.0%Penicillium citreonjgrum 23.2% Aspergillus terreus31.2%Penicillium citrinum36.8% Aspergillus terricola23.2%Penicillium corylophilum 40.8% Aspergillus ustus15.2%Penicillium decumbens 34.8% Aspergillus versicolor14.8%Penicillium implicatum39.2% Beauverja brongniartii 26.0%Penicillium janthinellum 23.6% Cladosporium berbarum 18.8%Penicillium lslandicum 23.2% Cladosporium carpophilum 24.4%Penicillium purpurogenum 23.6% Cladosporium tenissimum 20.0%Penicillium restrictum94.0% Curvularia lunata25.2%Penicillium spinulosum 27.2% Epicoccum purpurascens 23.2%Penicillium waksmanii 26.8% Exophiala jeanselmer 19.6%Penicillum nigricans21.2% Fusarium merismoides 78.4%Phoma nebulosa49.2% Fusarium moniliforme26.8%Phoma putaminum84.4% Fusarium oxysporum19.6%Rhizopus stolonifer79.2% Geotrichum candidurn 28.8%Scopulariopsis canadensis 7.6% Humicola fuscoatra24.4%Trichoderma atroviride 83.6% Humicola grisea27.2%Trichoderma harziamum 21.6% Kabatia microsticta22.4%Aspergillus niger, Fusarium merismoides, Penicillium restrictum, Phoma putaminum, Trichoderma atroviride, Arthrobacter globiformis, Bacillus subtilis, Cellulomonas flavigena, Pseudomonas synxantha, Streptomyces albus,species S.F (%) S.E.M species S.F (%) S.E.M Achromobacter cholinophagum 27.6%Flavobacterium johnsoniae21.2% Acinetobacter calcoaceticus 31.6%Micrococcus halobius 39.2% Acinetobacter johnsonii 24.8%Micrococcus luteus51.6% Agrobacterium radiobacter var.I28.4%Nocardia rubropertincta50.8% Agrobacterium radiobacter var.II30.4%Pantoea agglomerans49.6% Alcaligenes faecalis 35.6%Pantoea soilia22.4% Arthrobacter citreus 8.8%Proteus vulgaris25.6% Arthrobacter globiformis 85.6%Pseudomonas fluorescens var.Ⅰ7.6% Bacillus cerecus 20.8%Pseudomonas pseudoalaligenes11.2% Bacillus megaterium 25.2%Pseudomonas stutzeri14.8% Bacillus mycoides 18.8%Pseudomonas synxantha83.6% Bacillus subtilis 86.8%Streptomyces albus87.6% Brevibacillus brevis 24.4%Streptomyces caeruleus35.6% Brevibacterium linens 19.2%Streptomyces canescens9.2% Cellulomonas fimi 19.6%Streptomyces chromfuscus10.4% Cellulomonas flavigena 87.2%Streptomyces rseoviolaceus48.4% Corynebacterium mucifaciens 18.8%Xanthobacter flavus14.4% Erwinia amylovora 22.4%species S.F (%) S.E.M species S.F (%) S.E.M Acremonium strictum 20.4%Penicillium purpurogenum 23.6% Alternaria alternato27.2%Penicillium restrictum94.0% Aspergillus carnus20.0%Penicillium spinulosum 27.2% Aspergillus flavus18.8%Penicillium waksmanii 26.8% Aspergillus fumigatus27.2%Penicillum nigricans21.2% Aspergillus melleus21.2%Phoma nebulosa49.2% Aspergillus niger84.0%Phoma putaminum84.4% Aspergillus ochraceus 18.8%Rhizopus stolonifer79.2% Aspergillus penicilloides 20.8%Scopulariopsis canadensis 7.6% Aspergillus puniceus16.4%Trichoderma atroviride 83.6% Aspergillus sydowil18.0%Trichoderma harziamum 21.6% Aspergillus terreus31.2%Aspergillus terricola23.2%Aspergillus ustus15.2%Aspergillus versicolor14.8%Beauverja brongniartii 26.0%Cladosporium berbarum 18.8%Cladosporium carpophilum 24.4%Cladosporium tenissimum 20.0%Curvularia lunata25.2%Epicoccum purpurascens 23.2%Exophiala jeanselmer 19.6%Fusarium merismoides 43.2%Fusarium moniliforme26.8%Fusarium oxysporum19.6%Geotrichum candidurn 28.8%Humicola fuscoatra24.4%Humicola grisea27.2%Kabatia microsticta22.4%Mucor hiemalis78.4%Mucor spinescens39.6%Neurospora crassa42.4%Paecilomyces inflatus44.4%Paecilomyces lilacinus 38.8%Paecilomyces marquandii 34.4%Paecilomyces varioti31.2%Papulaspora equi27.6%Penicillim rubrum44.8% Penicillim thomii41.2% Penicillium citreonjgrum 23.2% Penicillium citrinum36.8% Penicillium corylophilum 40.8% Penicillium decumbens 34.8% Penicillium implicatum39.2% Penicillium janthinellum 23.6% Penicillium lslandicum 23.2%。