JP6218264B2 - Degradation method of digested sludge - Google Patents

Description

  The present invention is capable of growing in a culture solution using digested sludge as a sole nutrient source, inoculated into a culture solution obtained by adding water to digested sludge, and the solid weight when cultured for 7 days under conditions of pH 5 and temperature 30 ° C. The present invention relates to a microorganism having a decrease rate (%) of 10.0% or more, and a digested sludge decomposition method for culturing such a microorganism in a culture solution containing digested sludge.

  When purifying sewage from home life or industry, generally, biological treatment such as activated sludge method is used. In biological treatment, pollutants such as organic matter, nitrogen, and phosphorus in sewage are ingested as nutrients by microorganisms that inhabit treatment tanks, thereby purifying the sewage supernatant and releasing it into natural waters such as rivers. Yes. In such a treatment tank, microorganisms grow in accordance with the amount of pollutants removed from the sewage, and a large amount of the grown microorganisms are generated as waste, which is called surplus sludge. In this way, since a large amount of sewage flows into the sewage treatment plant every day, the amount of excess sludge generated annually becomes enormous.

  Therefore, biogas (methane-containing gas) production has come to be adopted as a means for effectively using surplus sludge discharged from sewage treatment plants. Particularly well-studied is anaerobic digestion technology using anaerobic microorganisms such as methane-producing bacteria. For example, waste that undergoes methane fermentation treatment of excess sludge together with sludge, liquid organic waste, and solid organic waste A treatment method (for example, refer to Patent Document 1) and a sludge treatment apparatus provided with a methane generation tank filled with granular sludge in which anaerobic bacteria mainly composed of methanogens are self-immobilized (for example, refer to Patent Document 2) Or solubilized liquid obtained by heating the excess sludge with no alkali added, or the excess sludge with alkali concentration of 0.02N or less in an alkaline condition under anaerobic conditions by the upflow anaerobic sludge bed method A method for treating surplus sludge subjected to high-speed methane fermentation (for example, see Patent Document 3) has been reported.

  As described above, biogas production is performed as a means for effectively using surplus sludge. In biogas production, components that are easily biodegradable by anaerobic microorganisms such as methane-producing bacteria, such as water-soluble carbohydrates and proteins Although water-soluble organic substances in sewage such as lipids and intracellular components of activated sludge microorganisms are decomposed, they are derived from non-biodegradable components after degradation, such as activated sludge microorganisms and cell wall polysaccharides of methane producing bacteria. Digested sludge such as keratin derived from chitin, human hair, cellulose derived from human waste, refractory carbohydrates such as xylan is generated. The amount of digested sludge reaches about 170 million tons per year, and is the most industrial waste discharged in Japan, accounting for 44.5% of the total industrial waste (for example, Non-Patent Document 1) reference). The generated digested sludge is only partially used as a secondary material for construction materials and as a raw material for sewage compost, and most of the rest is disposed of as industrial waste without any effective use.

  Therefore, methods for reducing digested sludge have been studied. For example, a method for treating an organic waste liquid including a step of performing ozone treatment on a part of digested sludge and performing alkali treatment and anaerobic digestion after the ozone treatment (see, for example, Patent Document 4) has been reported.

JP-A-11-309438 JP-A-9-29290 Japanese Patent Laid-Open No. 09-1194 JP 2005-7339 A

2011 Business Industrial Waste Discharge / Treatment Status Report 2009 Results (Summary Version) Page 25

  So far, no technology for biologically treating digested sludge has been developed that does not involve physicochemical treatment steps. In addition, it can grow in a culture solution that uses digested sludge as its only nutrient source, and inoculates into a culture solution in which water is added to digested sludge and reduces the weight of solids when cultured for 7 days at pH 5 and temperature of 30 ° C. It has not been reported that a microorganism having a rate (%) of 10.0% or more was isolated. The subject of this invention is providing the microorganisms which decompose | disassemble the digested sludge which is the final product of a sewage treatment with microorganisms, and the digested sludge.

  The inventor has eagerly searched for microorganisms capable of decomposing digested sludge, which is the final product of sewage treatment. Although the main component of digested sludge is hardly biodegradable, it originally contained a large amount of microbial growth biomass, so we originally expected that microorganisms capable of biodegrading would be obtained relatively easily. For several years, microorganisms capable of degrading digested sludge could not be obtained. Therefore, we investigated the cause and thought that the heavy metals contained in the sludge and the aluminum flocculant used to dehydrate the digested sludge are involved in the adverse effects on the growth of microorganisms. When we tried to search for microorganisms again using digested sludge from which metal was leached, it was possible to isolate microorganisms capable of degrading digested sludge, and these microorganisms unexpectedly make digested sludge the only nutrient source. The present invention was completed after confirming that it could grow in the culture solution.

  That is, the present invention [1] can be grown in a culture solution using digested sludge as a sole nutrient source, inoculated into a culture solution obtained by adding water to digested sludge, and cultured for 7 days under conditions of pH 5 and temperature 30 ° C. A digestion sludge decomposition method for culturing a microorganism having a solid weight reduction rate (%) of 10.0% or more in a culture solution containing digested sludge, wherein the microorganism is Penicillium janthinellum A method for decomposing digested sludge characterized by being one or more microorganisms of Cunninghamella bainieri, Neosartorya fischeri, Umbelopsis isabellina, [2] The digested sludge decomposition method according to [1] above, wherein the microorganism has xylanase and chitinase activity, and [3] the microorganism is penicillin Mu Jan Cineram CedarWA2 strain (reception number: NITE AP-1523), Kaningamera bainieri CedarWA4 strain (reception number: NITE AP-1524), Neosartoria Fischeri OreWA strain (reception number: NITE AP-1522), Unveropsis Isaverina FernWA The digestion sludge decomposition method according to the above [1] or [2], which is any one or more microorganisms of a stock (reception number: NITE AP-1525), and [4] digestion sludge The method for decomposing digested sludge according to any one of the above [1] to [3], comprising a step of leaching metal in advance.

  [5] The present invention can be grown in a culture solution using [5] digested sludge as a sole nutrient source, inoculated into a culture solution in which water is added to digested sludge, and cultured for 7 days under conditions of pH 5 and temperature 30 ° C. Penicillium janthinellum, Cunninghamella bainieri, Neosartorya fischeri, Umbelopsis isabellina, whose solids weight loss (%) is more than 10.0% ), [6] the microorganism described in [5] above having xylanase and chitinase activity, and [7] Penicillium cinecineram, -1523), the micro-organism according to the above [5] or [6] And [8] Kaningamela bainieri is the Kaningamela bainieri CedarWA4 strain (reception number: NITE AP-1524), or the microorganism described in [5] or [6] above, The microorganism described in [5] or [6] above, wherein [10] Unveropsis Isaverina FernWA is characterized in that the Fischery is Neosartoria Fischeri OreWA strain (reception number: NITE AP-1522). The microorganism according to [5] or [6] above, which is a strain (reception number: NITE AP-1525).

  Furthermore, the present invention provides [11] a digested sludge degradation product obtained by inoculating the microorganism according to any one of [5] to [10] above into a culture solution containing digested sludge and culturing the microorganism. The present invention relates to a fuel containing a mixture and [12] the digested sludge decomposition product described in [11] above and a mixture of microorganisms.

  Microorganisms used in the present invention can grow in a culture solution using digested sludge as a sole nutrient source, and have a high ability to decompose digested sludge, so that digested sludge can be decomposed in large quantities at low cost, which greatly reduces industrial waste. Can be reduced. Further, the digested sludge decomposition product obtained by the present invention and a mixture of microorganisms (hereinafter also referred to as “sludge-bacteria mixture”) have a high calorific value during combustion, and thus can be used as fuel.

It is a microscope picture of the separated digested sludge decomposition microorganisms. It is a figure which shows the result of a digested sludge decomposition test. It is a figure which shows the result of the calorie | heat amount measurement in CedarWA2. It is a figure which shows the phylogenetic tree of the isolate | separated digested sludge decomposition microorganisms.

  The microorganism of the present invention can be grown in a culture solution using digested sludge as a sole nutrient source, inoculated into a culture solution obtained by adding water to digested sludge, and cultured for 7 days under conditions of pH 5 and temperature 30 ° C. Penicillium janthinellum, Cunninghamella bainieri, Neosartorya fischeri, or Umbelopsis isabellina whose solids weight loss (%) is 10.0% or more In addition, the digestion sludge decomposition method of the present invention is not particularly limited as long as it is a method of culturing the microorganism of the present invention in a culture solution containing digested sludge, By the digested sludge decomposition method of the present invention, the digested sludge can be decomposed in large quantities at low cost, and the industrial waste can be significantly reduced. It becomes ability. The present invention also relates to a mixture of digested sludge degradation product and microorganisms obtained by inoculating the microorganism of the present invention into a culture solution containing digested sludge and culturing, as well as such digested sludge degradation product and microorganisms. It relates to a fuel containing a mixture. Furthermore, the sludge-bacteria mixture obtained by the digested sludge decomposition method of the present invention is also useful as a fuel.

  In the present invention, "digested sludge" is a sludge residue composed of hardly biodegradable components that are generated when an excess of sludge is biodegraded by anaerobic microorganisms such as methane producing bacteria. In addition, chitin derived from cell wall polysaccharides of aerobic bacteria such as activated sludge microorganisms and anaerobic bacteria such as methane producing bacteria, keratin derived from human hair, cellulose derived from human waste, refractory carbohydrates such as xylan and the like are included.

  In the present invention, the “culture medium containing digested sludge as the only nutrient source” means a culture medium that does not contain nutrient sources necessary for the growth of microorganisms such as organic substances and inorganic substances other than digested sludge, and preferably suspended. Examples of the suspension culture medium include suspended digested sludge and a mixture of digested sludge and water. The pH of the culture solution is 5, and can be adjusted as appropriate. Moreover, before using a culture solution, a culture solution can be sterilized by an autoclave etc. as needed.

  In the present invention, the “decrease rate (%) of the solid weight” means (solid weight before culturing (g) −solid weight after culturing (g)) / solid weight before culturing (g) × The solid weight before culturing is the dry weight of digested sludge before culturing inoculated with microorganisms, and the solid weight after culturing is the dried sludge-bacteria mixture after culturing. Here, the dry weight is the weight (g) obtained by drying (105 ° C., 24 hours) the residue after centrifugation (3000 rpm, 10 minutes).

  In the present invention, the “culture solution containing digested sludge” is not particularly limited as long as it is a culture solution containing at least digested sludge. In addition to digested sludge and water, the culture solution may be used as necessary. Nutrient sources such as organic and inorganic substances and pH regulators can be added. Further, before using the culture solution, the culture solution may be sterilized by an autoclave or the like.

  In the present invention, the culture conditions are not particularly limited when culturing with a culture solution containing digested sludge and decomposing the digested sludge. For example, the pH of the culture solution is 4 to 8, preferably 5 to 7, and the culture temperature. Is 20 ° C. to 35 ° C., preferably 25 ° C. to 30 ° C., and the culture time is 3 to 20 days, preferably 7 to 14 days. Aeration culture, shaking culture, agitation culture, stationary culture, Or a combination of these may be used.

  The microorganisms used in the digestion sludge decomposition method of the present invention and the microorganisms of the present invention include Penicillium janthinellum, Cunninghamella bainieri having a solid weight reduction rate (%) of 10% or more, It is not particularly limited as long as it is a microorganism isolated from nature or an artificial mutant of Neosartorya fischeri or Umbelopsis isabellina, but xylanase or chitinase activity, preferably xylanase or chitinase Microorganisms having keratinase activity, microorganisms having a solid weight reduction rate (%) of 13% or more, preferably 14% or more, more preferably 18% or more, and even more preferably 25% or more, or a sludge-bacteria mixture Calorific value during combustion per unit weight (increases due to heat generation) A microorganism having a value obtained by integrating the voltage value of the temperature sensor with the combustion time (μVsec / mg) is 10,000 or more, preferably 11000 or more, more preferably 12000 or more. In addition, when using for the digestion sludge decomposition | disassembly method, you may use 2 or more types of the said microorganisms.

  The microorganisms used in the digestion sludge decomposition method of the present invention and the microorganisms of the present invention include Penicillium Jancineram CedarWA2 strain (reception number: NITE AP-1523) and Kaningamera bainieri CedarWA4 strain (reception number: NITE AP-) isolated from nature. 1524), Neosartoria Fischeri OreWA strain (reception number: NITE AP-1522), Unveropsis isaverina FernWA strain (reception number: NITE AP-1525), or mutants thereof can be particularly preferably exemplified. Such microorganisms isolated from nature were deposited on February 1, 2013, respectively, at the Patent Microorganism Depositary Center of the National Institute of Technology and Evaluation (2-5-8 Kazusa Kamashichi, Kisarazu City, Chiba Prefecture). Yes.

  The digested sludge decomposition method of the present invention preferably includes a step of leaching metal from the digested sludge in advance. The metal includes light metals such as aluminum and heavy metals such as copper, nickel, lead, zinc, and cadmium. Examples of the method for leaching the metal include a method in which digested sludge is treated with a mineral acid such as sulfuric acid and hydrochloric acid, preferably sulfuric acid, and then the acid is removed. For example, the digested sludge is treated with 2N sulfuric acid for 1 hour and then digested. A method of filtering the sludge and acid mixture and washing the filtration residue with a sufficient amount of water can be mentioned. By leaching the metal, the proliferation power of microorganisms is increased and the digestive sludge decomposition ability is improved.

  Furthermore, the mycelium or spore of the microorganism of the present invention may be used as a kit or microbial preparation for decomposing digested sludge. In that case, a method for decomposing digested sludge using the microorganism of the present invention is used. A written document can be attached.

  In the present invention, the culture conditions for inoculating the microorganism of the present invention into a culture solution containing digested sludge to obtain a mixture of digested sludge degradation product and microorganisms are the culture conditions in the digested sludge decomposition method of the present invention described above. It is the same as conditions. Such a mixture of digested sludge degradation products and microorganisms includes digested sludge degradation products, digested sludge undegraded products, proliferated microorganisms, and the like.

  As the fuel of the present invention, a solid material containing a mixture of the digested sludge decomposition product and microorganisms can be preferably exemplified. As the solid material, a culture solution after culturing is obtained by centrifuging the culture solution after culturing. Examples include a dried residue remaining after removal, and a combustible water-absorbing substance such as sawdust and rice husk added to the culture solution after culturing, and dried as necessary. The centrifugation conditions are not particularly limited, but can be appropriately adjusted depending on the amount and composition of digested sludge and the type of microorganism. For example, the number of revolutions is 1000 to 10000 rpm, preferably 2000 rpm to 5000 rpm, more preferably 2500 rpm to 3500 rpm. Yes, the time is 1 minute to 60 minutes, preferably 3 minutes to 30 minutes, more preferably 5 minutes to 15 minutes. The drying temperature and time can be appropriately adjusted depending on the amount and composition of digested sludge and the type of microorganism. For example, the temperature is 90 ° C to 120 ° C, preferably 100 ° C to 110 ° C, and the time is 12 hours. -30 hours, preferably 20 hours to 28 hours.

[Search for digested sludge-decomposing microorganisms]
(Acid treatment of digested sludge)
300 g of dehydrated digested sludge obtained from the sewage treatment plant and 600 ml of 2N sulfuric acid were added to a glass beaker and mixed, and allowed to stand at room temperature for 1 hour to leach out the metals contained in the activated sludge. The mixture of digested sludge and sulfuric acid treated with acid in this manner was filtered with gauze, and the digested sludge remaining on the gauze was washed with a sufficient amount of water to remove sulfuric acid. Each time the washing with water was repeated, the pH test paper was soaked in digested sludge, and when the pH returned to 6 or more, it was judged that the sulfuric acid was removed. Further, the digested sludge was dried at 105 ° C. for 24 hours, and then crushed with an Osterizer blender (Oster) to obtain a dry powder.

(Separation of sludge-decomposing microorganisms)
In order to obtain digested sludge-decomposing microorganisms, soil was collected from various places in Yamaguchi Prefecture (9 places such as forests), and 10 g of soil was suspended in 30 ml of sterilized water as an inoculation source. A digested sludge culture solution (hereinafter simply referred to as “digested sludge culture solution”) consisting of 200 mg of digested sludge and 10 ml of water was placed in a flask and sterilized in an autoclave (manufactured by TOMY) at 121 ° C. for 15 minutes. The pH of the digested sludge culture solution after sterilization was 5.0. It is considered that the pH decreased due to the elution of some of the components in the sludge into water by the heat treatment. Next, 0.2 ml of the inoculum was added to the sterilized digested sludge culture solution, followed by shaking culture (accumulation culture) at 30 ° C. for 7 days. 0.2 ml of a bacterial solution was collected from a flask in which microbial growth was observed by enrichment culture, inoculated into a flask containing a sterilized fresh digested sludge culture solution, and subculture was performed for 7 days. After this subculture was repeated twice, 30 μl of the bacterial solution was inoculated into YM agar medium (10 g / l glucose, 3 g / l yeast extract, 3 g / l malt extract, 5 g / l bactopeptone, 20 g / l bacto agar). Static culture was performed at 30 ° C. for 7 days. The emerged colony was picked up with a sterilized toothpick, and this was streaked on a new YM agar medium to perform pure isolation of the strain. In this way, a total of 8 isolates (FernWA, CedarWA1, CedarWA2, CedarWA3, CedarWA4, OreWA, OreYA, GalleryYA) were obtained. The obtained 8 strains are inoculated into a sterilized digested sludge culture solution, cultured for 7 days under conditions of pH 5 and temperature 30 ° C., and the results of observation with a microscope are shown in FIG.

  As is apparent from the location indicated by the arrow in FIG. 1, mycelia or spores can be observed, and all 8 strains obtained can grow in the digested sludge culture solution, that is, digested sludge is the only nutrient source. It was confirmed that it can grow in the culture solution. In addition, the black or gray particle | grains in FIG. 1 are the digested sludge intertwined with the extended mycelium.

[Digested sludge decomposition test]
(Calculation of decrease rate of solid weight)
5 mg of a thin bacterial membrane piece of each isolate was inoculated into a digested sludge culture solution that had been sterilized by autoclaving under the same conditions as described above, and cultured with shaking at pH 5 and a temperature of 30 ° C. for 7 days. Residues (digested sludge degradation product, digested sludge undegraded product) remaining after removing the culture supernatant by centrifuging (3000 rpm, 10 minutes) the microbial solution after culture with an H-19F centrifuge (manufactured by Kokusan) And a mixture of grown microorganisms) was dried (105 ° C., 24 hours) to obtain a solid. By measuring the weight of this solid matter, the reduction rate of the solid weight due to the culture was calculated as follows: solid matter weight reduction rate (%) = (solid weight before cultivation (g) −solid weight after cultivation (g )) / Obtained by solid weight (g) × 100 before culture. As a control, the digested sludge culture solution sterilized without inoculation of microorganisms was shaken for 7 days under the conditions of pH 5 and temperature 30 ° C. The average value of the reduction rate of the solid weight of each of the three isolates is shown in FIG.

(result)
As shown in FIG. 2, the decrease rate of the solid weight was 13.0% for FernWA strain, 14.0% for CedarWA1 strain and CedarWA2 strain, 14.5% for CedarWA3 strain and 18.5% for CedarWA3 strain, respectively. 18.0% and 27.0% for the OreWA strain.

[Enzyme activity test]
In order to know what components contained in activated sludge decompose and grow, the activity of degrading enzymes on digested sludge components was examined. Specifically, exocellulase, endocellulase, xylanase, chitinase, and keratinase activity were measured.

(Measuring method)
In the same manner as in Example 2, each isolate was inoculated into a sterilized digested sludge culture solution and cultured under shaking at pH 5 and a temperature of 30 ° C. for 7 days. Next, the culture solution was filtered with glass filter paper, and the obtained culture supernatant was used as an enzyme solution for various enzyme activity tests. In the measurement of exocellulase, endocellulase, xylanase and chitinase activities, 10 mg of each substrate (cellulose, carboxymethylcellulose, xylan and chitin), 0.8 ml of citrate buffer (pH 4.8) and 0.2 ml of enzyme solution were placed in a microtube. In addition, the mixture was heated at 50 ° C. for 60 minutes, and the enzyme activity was quantified by measuring the concentration of reducing sugar released during this time by the DNS method (Ghose TK, Pure and Applied Chemistry (1987) 59, 257-268). For the measurement of keratinase activity, 10 mg of keratin azul (manufactured by Sigma Aldrich), 0.8 ml of 62.5 mM Tris-HCl buffer (pH 8.5) and 0.2 ml of enzyme solution were added to a microtube and added at 50 ° C. for 60 minutes. The enzyme activity was calculated based on the method of Riffel et al. (Riffel A et al., Journal of Biotechnology (2007) 128, 693-703). The results are shown in Table 1.

(result)
From Table 1, all isolates had xylanase activity and also had chitinase activity except for CedarWA3 and GalleyYA. FernWA, CedarWA4, OreWA, and OreYA had xylanase, chitinase and keratinase activities. It is assumed that the activated sludge contains undigested plant fibers such as xylan derived from human waste. In addition, it is considered that a large amount of filamentous fungi that cannot be decomposed into methane-producing bacteria, chitin which is a cell wall component of protists, and keratin derived from human hair remain. Since these are components that are difficult to biodegrade in the aerobic microorganisms and methane producing bacteria in the activated sludge aeration tank, it is considered that the isolate grows by decomposing these components.

[Measurement of calorific value and calorific value during combustion]
(Measurement method of calorie)
As in Example 2, each isolate was inoculated into a sterilized digested sludge broth and cultured with shaking for 7 days under conditions of pH 5 and temperature 30 ° C., followed by centrifugation and drying treatment, and digested sludge degradation products and microorganisms. A solid consisting of a mixture with was obtained. The amount of heat of the solid was measured using a Thermo Plus Evo II differential type differential thermal balance (manufactured by Rigaku Corporation). The temperature inside the furnace was raised from room temperature to 800 ° C. at a rate of 20 ° C. per minute, and the amount of heat generated by combustion during this time was measured by the voltage value (μV) of the temperature sensor that increased due to heat generation.

(result)
The result of calorimetric measurement at CedarWA2 is shown in FIG. The horizontal axis is the combustion time (minutes), the right vertical axis is the amount of heat (μV), the left vertical axis is the furnace temperature (° C.), the dotted line is the furnace temperature, and the solid curve is the digestion inoculated with CedarWA2. The amount of heat of the solid after the culture of the sludge culture solution, and the dotted curve are the amount of heat of the solid after the culture of the digested sludge culture solution not inoculated with microorganisms. In the solid after the culture of the digested sludge culture medium not inoculated with microorganisms, it is about 70 μV at 530 ° C., whereas in the solid after the culture of the digested sludge culture inoculated with CedarWA2, it is more than double at 530 ° C. It was 180 μV and had a high calorific value.

(Measurement method of calorific value during combustion)
Furthermore, each isolate was inoculated into digested sludge culture broth and cultured under shaking at pH 5 and temperature 30 ° C. for 7 days. The calorific value at the time of burning per unit weight of the solid obtained from the culture broth (μVsec / mg ), And when the digested sludge culture solution is not inoculated with microorganisms and cultured with shaking for 7 days under the conditions of pH 5 and temperature 30 ° C., and the calorific value at the time of combustion per unit weight of the solid matter obtained from the culture solution is set to 1. Each isolate was inoculated into digested sludge culture broth and cultured with shaking for 7 days under the conditions of pH 5 and temperature 30 ° C., and the relative exotherm with the calorific value during combustion per unit weight of the solid matter obtained from the culture broth The amount was determined. The calorific value during combustion per unit weight was determined by integrating the voltage value of the temperature sensor, which increases due to heat generation, with the combustion time.

(result)
The results are shown in Table 2. When cultivated with any microorganism, a higher value of calorific value at the time of combustion per unit weight is obtained than when microorganisms are not inoculated. The calorific value at the time of combustion was 1.4 to 1.68 times that of fuel.

[Systematic analysis of each isolate]
(System analysis method)
The phylogenetic analysis of each obtained isolate was performed by a method of analyzing an internal transcription spacer region (Internal Transcribed Spacer: ITS) that has been proposed as the most promising DNA barcode used for fungal phylogeny ( Conrad L. Schoch et al., PNAS (2012) 109, 6241-6246). Each isolate was statically cultured in 20 ml of YM medium (10 g / l glucose, 3 g / l yeast extract, 3 g / l malt extract, 5 g / l bactopeptone) at 30 ° C. for 7 days. Bacterial membranes produced by 7 days of culture were washed with water, and then genomic DNA was extracted using Master Pure Yeast DNA Purification Kit (Epicentre). Using the extracted DNA as a template, the ITS region was PCR amplified using KOD Fx DNA polymerase (manufactured by Toyobo). ITS1 primer (5′-TCCGTAGGTGAACCTGCGG-3 ′: SEQ ID NO: 1) and ITS4 primer (5′-TCCTCCGCTTATTGATATGC-3 ′: SEQ ID NO: 2) were used as PCR reaction primers, and the PCR temperature conditions were 98 ° C. for 10 seconds. 30 cycles of 54 ° C. for 30 seconds and 68 ° C. for 60 seconds. The amplified ITS region DNA was sequenced using BigDye Terminator ver3.1 Cycle Sequencing kit (Life Technologies). The obtained sequence was input into the BLAST algorithm, and the genus species of the isolate was determined by comparing with the known DNA sequences in the GenBank, EMBL, and DDBJ gene databases.

(result)
The base sequence of the ITS region of CedarWA1 (SEQ ID NO: 3), the base sequence of the ITS region of CedarWA2 (SEQ ID NO: 4) is 99% identical to the base sequence of the ITS region of Penicillium janthinellum, and the ITS region of CedarWA3 The base sequence of SEQ ID NO: 5 and the base sequence of the ITS region of CedarWA4 (SEQ ID NO: 6) are 99% identical to the base sequence of the ITS region of Cunninghamella bainieri, and the base sequence of the ITS region of the OreWA strain (SEQ ID NO: 7) is 100% identical to the nucleotide sequence of the ITS region of Neosartorya fischeri, and the nucleotide sequence (SEQ ID NO: 8) of the ITS region of FernWA strain is the ITS of Umbelopsis isabellina. 99% identical to the base sequence of the region, The base sequence of the ITS region of the GalleryYA strain (SEQ ID NO: 9) is 95% identical to the base sequence of the ITS region of Chaetomium Globosum, and the base sequence of the ITS region of the OreYA strain (SEQ ID NO: 10) is Fusarium. It was 99% identical to the base sequence of the ITS region of Fusarium oxysporum. In addition, using the Clustal W program (Saitou & Nei, (1987) Mol. Biol. Evol. 4, 406-425, Thompson et al., Nucleic Acids Res. (1994) 22, 4673-4680), known as the neighborhood joining method. FIG. 4 shows the results of creating a phylogenetic tree based on DNA in the ITS region. As shown in FIG. 4, CedarWA1 and CedarWA2 are Penicillium janthinellum, CedarWA3 and CedarWA4 are Cunninghamella bainieri, and OreWA is Neosartoria FiseriWaN. Umbelopsis isabellina), GalleryYA strain was found to be Chaetomium Globosum, and OreYA strain was found to be Fusarium oxysporum.

  The digested sludge decomposition method of the present invention is capable of decomposing hardly biodegradable digested sludge at low cost, and has high industrial utility in the field of industrial waste treatment.

Claims (7)

The rate of decrease in the weight of solids when grown in a culture solution using digested sludge as the only nutrient source, inoculated into a culture solution in which water is added to the digested sludge and cultured at pH 5 and temperature of 30 ° C. for 7 days ( %) Is a digestion sludge decomposition method for culturing a microorganism having a digestion sludge containing a microorganism having a content of 10.0% or more, wherein the microorganism is Penicillium janthinellum, Cunninghamella bainieri A method for decomposing digested sludge, characterized in that it is one or more microorganisms of Neosartorya fischeri and Umbelopsis isabellina. 2. The digested sludge decomposition method according to claim 1, wherein the microorganism has xylanase and chitinase activities. Microorganisms are Penicillium cinecineram CedarWA2 strain (Accession number: NITE P-1523), Kaningamera bainieri CedarWA4 strain (Accession number: NITE P-1524), Neosartoria Fischeri OreWA strain (Accession number: NITE P-1522), Unveropsis The digested sludge decomposition method according to claim 1 or 2, wherein the microorganism is any one or more microorganisms of Isabela FernWA strain (Accession number: NITE P-1525). The method for decomposing digested sludge according to any one of claims 1 to 3, further comprising a step of leaching the metal from the digested sludge in advance. The rate of decrease in the weight of solids when grown in a culture solution using digested sludge as the only nutrient source, inoculated into a culture solution in which water is added to the digested sludge and cultured at pH 5 and temperature of 30 ° C. for 7 days ( %) Is 10.0% or more, Penicillium janthinellum CedarWA2 strain (Accession number: NITE P-1523), Cunninghamella bainieri CedarWA4 strain (Accession number: NITE P-1524), Neo One of the microorganisms of the Neosartorya fischeri OreWA strain (Accession number: NITE P-1522) and Umbelopsis isabellina FernWA strain (Accession number: NITE P-1525). 6. The microorganism according to claim 5, which has xylanase and chitinase activities. Method for producing a microorganism according to claim 5 or 6, was inoculated with a culture medium containing digested sludge, characterized by culturing a mixture of digested sludge decomposition products with microorganisms.