A septic tank biodegradation agent

Abstract

The present invention belongs to the technical field of microbial application, and specifically relates to a septic tank biodegrader, including Bacillus tekila YZS-C01, Bacillus saffron YZS-C03, Alcaligenes faecalis QQB-A05 and Pseudomonas putida ZTB-A29. The composite microbial preparation can effectively degrade solidified objects in the septic tank, remove malodorous pollutants, and then purify the air through the synergistic promotion of multiple microorganisms, thereby reducing the frequency of septic tank cleaning.

Description

Technical Field

[0001] The invention belongs to the technical field of microbial application, and in particular relates to a septic tank biological decomposer. Background Art

[0002] Septic tanks are equipment for treating feces and filtering and precipitating them. Solidified objects are decomposed at the bottom of the tank, and hydrated objects on the upper layer flow into the pipes, preventing pipe blockage and giving solidified objects (feces and other garbage) sufficient time to hydrolyze. Septic tanks need to be cleaned regularly, and the cleaning cycle is related to the temperature of feces and sewage, air temperature, building properties, and drainage water quality and volume. Rhizosphere-related specifications require a cleaning cycle of 3 to 12 months. Frequent cleaning work has increased labor and management costs and has caused impacts and inconveniences to the surrounding environment. Septic tank biodegraders can greatly degrade solidified objects in septic tanks, improve the environment of septic tanks and their affiliated pipes, reduce the number of cleanings, and at the same time improve the living environment of the majority of people living within the range of odor sources such as fertilizer tanks. Summary of the invention

[0003] The object of the present invention is to provide a septic tank biodegrader. The composite microbial preparation can effectively degrade solidified objects in the septic tank, remove odor pollutants, and further purify the air through the synergistic promotion of multiple microorganisms, thereby reducing the frequency of septic tank cleaning.

[0004] To achieve the above object, the present invention adopts the following technical solution:

[0005] The invention firstly provides a septic tank biodegrader, which comprises Bacillus tequila YZS-C01, Bacillus shafusae YZS-C03, Alcaligenes faecalis QQB-A05 and Pseudomonas putida ZTB-A29.

[0006] Among them, the Bacillus equilensis YZS-C01 is classified and named Bacillus equilensis (Bacillus equilensis), and was deposited in the General Microbiology Center of China Microorganism Culture Collection Administration on July 9, 2021. The deposit address is: No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing, China, and the deposit number is: CGMCC No. 22856.

[0007] Among them, the Bacillus safensis YZS-C03, whose classification name is Bacillus safensis, was deposited in the General Microbiology Center of China Microorganism Culture Collection Administration on February 1, 2021. The deposit address is: No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing, China, and the deposit number is: CGMCC No. 21798.

[0008] Among them, the faecalis QQB-A05 is classified and named Alcaligenes faecalis, and was deposited in the General Microbiology Center of China National Microbiological Culture Collection on September 2, 2021. The deposit address is: No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing, China, and the deposit number is: CGMCC No. 23341.

[0009] Among them, the Pseudomonas putida ZTB-A29 is classified and named Pseudomonas putida (Pseudomonas putida), and was deposited in the General Microbiology Center of China Microorganism Culture Collection Administration on October 16, 2020. The deposit address is: No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, China, and the deposit number is: CGMCC No. 20897.

[0010] The colony characteristics and bacterial morphology of the above-mentioned Bacillus tequila YZS-C01 are as follows:

[0011] The colonies formed by strain YZS-C01 after culturing on NA plates for 24 hours are light yellow, translucent, dull, round, and non-fluid, with a colony diameter of 4-5 mm; the bacterial body morphology is rod-shaped, Gram-positive, with spores and no capsule.

[0012] The physiological and biochemical characteristics of the above-mentioned Bacillus tekila YZS-C01 are:

[0013] Catalase reaction was negative, VP assay was negative, MR assay was negative, glucose acid production test was positive, glucose gas production test was negative, citrate test was negative, nitrate reduction was positive, starch hydrolysis was positive, indole test was negative, malonic acid assay was negative, and H2S production test was positive.

[0014] The 16S rDNA sequence (SEQ ID NO.1) of the above-mentioned Bacillus tequilensis YZS-C01 was compared with the sequence in the GenBank database. The results showed that YZS-C01 and Bacillus tequilensis were on the same branch, and the similarity of its 16S rDNA sequence with Bacillus tequilensis (KY819004.1) was 100%. Combined with the colony morphology, physiological and biochemical characteristics and 16S rDNA sequence analysis, it was preliminarily identified as Bacillus tequilensis.

[0015] The colony characteristics and bacterial morphology of the above-mentioned Bacillus sabdariffa YZS-C03 are as follows:

[0016] The colonies formed after culturing Bacillus shaforniensis YZS-C03 on NA plate medium for 24 hours are light yellow, dull, round, non-fluid, Gram-positive, with spores and no capsule; the colony diameter is 4-5 mm.

[0017] Physiological and biochemical characteristics of the above-mentioned Bacillus sabdariffa YZS-C03:

[0018] Catalase reaction was negative, VP assay was negative, MR assay was negative, glucose acid production test was negative, glucose gas production test was negative, citrate test was negative, nitrate reduction was positive, starch hydrolysis was positive, indole test was negative, malonic acid assay was negative, and H2S production test was negative.

[0019] The 16S rDNA sequence of the above Bacillus safensis was compared with the sequence in the GenBank database. The results showed that YZS-C03 and Bacillus safensis were on the same branch, and its 16S rDNA sequence was 100% similar to Bacillus safensis (MT501806.1). Combining the colony morphology, physiological and biochemical characteristics and 16S rDNA sequence analysis, it was preliminarily identified as Bacillus safensis.

[0020] The colony characteristics and bacterial morphology of the above-mentioned Alcaligenes faecalis QQB-A05 are as follows:

[0021] The colonies formed after the strain QQB-A05 was cultured on the NA plate for 24 hours were light yellow, translucent, slightly shiny, round, non-fluid, and had a colony diameter of 3-4 mm; the bacteria were rod-shaped, Gram-negative, without spores and capsules.

[0022] The physiological and biochemical characteristics of the above-mentioned Alcaligenes faecalis QQB-A05 are:

[0023] The catalase reaction was negative, the VP assay was negative, the MR assay was positive, the glucose acid production test was positive, the glucose gas production test was negative, the citrate test was positive, the nitrate reduction was positive, the starch hydrolysis was negative, the indole test was negative, the malonic acid assay was negative, and the H2S production test was positive.

[0024] The 16S rDNA sequence (SEQ ID NO.1) of the above-mentioned Alcaligenes faecalis QQB-A05 was compared with the sequence in the GenBank database. The results showed that QQB-A05 and Alcaligenes faecalis were on the same branch, and the similarity of its 16S rDNA sequence with Alcaligenes faecalis (KX548264.1) reached 98.67%. Combined with the colony morphology, physiological and biochemical characteristics and 16S rDNA sequence analysis, it was preliminarily identified as Alcaligenes faecalis.

[0025] The colony characteristics and bacterial morphology of the above-mentioned Pseudomonas putida ZTB-A29 are as follows:

[0026] The colonies formed by strain ZTB-A29 after culturing on NA plates for 24 hours are light yellow, translucent, dull, round, and non-fluid, with a colony diameter of 4-5 mm; the bacteria are rod-shaped, Gram-negative, without spores and capsules.

[0027] The physiological and biochemical characteristics of the above-mentioned Pseudomonas putida ZTB-A29 are:

[0028] The catalase reaction was negative, the VP assay was negative, the MR assay was positive, the glucose acid production test was positive, the glucose gas production test was negative, the citrate test was positive, the nitrate reduction was positive, the starch hydrolysis was negative, the indole test was negative, the malonic acid assay was positive, and the H2S production test was positive.

[0029] The 16S rDNA sequence (SEQ ID NO.1) of the above-mentioned Pseudomonas putida was compared with the sequence in the GenBank database, and the results showed that ZTB-A29 and Pseudomonas putida were on the same branch, and the similarity of its 16S rDNA sequence with Pseudomonas putida (CP026115.2) was 99.72%. Combining the colony morphology, physiological and biochemical characteristics and 16S rDNA sequence analysis, ZTB-A29 was identified as Pseudomonas putida.

[0030] Furthermore, the mass ratio of solid bacterial powder or fermentation liquid of Bacillus tergillosis YZS-C01, Bacillus shafuss YZS-C03, Bacillus faecalis QQB-A05 and Pseudomonas putida ZTB-A29 in the septic tank biodegrader is (2-5): (2-5): (1-2): (0.1-1).

[0031] Furthermore, the total live bacteria concentration in the septic tank biodegrader is (5-8)×10 8cfu / ml or (5~8)×10 8 cfu / g.

[0032] Furthermore, the septic tank biodegrader is a liquid preparation or a solid preparation.

[0033] Furthermore, the preparation method of the solid preparation of the septic tank biodegrader is as follows: solid bacterial powders of Bacillus tergrillii YZS-C01, Bacillus shafusus YZS-C03, Bacillus faecalis QQB-A05 and Pseudomonas putida ZTB-A29 are mixed in a mass ratio to prepare a solid mixed bacterial powder; a carrier is mixed with the solid mixed bacterial powder in a mass ratio of 100:2 to 5 to obtain a solid preparation of the septic tank biodegrader; the carrier is bran and rice bran mixed in any proportion.

[0034] Furthermore, the preparation method of the septic tank biodegrader liquid preparation is as follows: the fermentation broths of Bacillus tergillus YZS-C01, Bacillus shafusus YZS-C03, Bacillus faecalis QQB-A05 and Pseudomonas putida ZTB-A29 are mixed in a mass ratio of (2-5): (2-5): (1-2): (0.1-1) to obtain the obtained product.

[0035] The beneficial effects of the present invention are:

[0036] The septic tank biodegrader provided by the present invention is compounded by using the screened Bacillus tekila YZS-C01, Bacillus shafusus YZS-C03, Bacillus faecalis QQB-A05 and Pseudomonas putida ZTB-A29 with strong organic matter degradation ability and deodorization effect. The septic tank biodegrader is rich in beneficial bacteria and can effectively degrade solidified objects in the septic tank through the synergistic effect of multiple bacteria species, remove odor pollutants, and then purify the air, thereby reducing the frequency of septic tank cleaning and improving the ecological environment. DETAILED DESCRIPTION

[0037] The present invention is further described below in conjunction with specific embodiments.

[0038] Bacteria materials:

[0039] (1) Bacillus equilensis YZS-C01, whose classification name is Bacillus equilensis, was deposited in the General Microbiology Center of China National Microbiological Culture Collection Administration on July 9, 2021, with the deposit address at No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, China, with the deposit number: CGMCC No. 22856, and the deposit address: Beijing, China, which has been made public.

[0040] (2) Bacillus safensis YZS-C03, whose classification name is Bacillus safensis, was deposited in the General Microbiology Center of China National Microbiological Culture Collection Administration on February 1, 2021. The deposit address is No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing, China. The deposit number is: CGMCC No. 21798, and the deposit address is: Beijing, China, which has been made public.

[0041] (3) Alcaligenes faecalis QQB-A05, whose classification name is Alcaligenes faecalis, was deposited in the General Microbiology Center of China National Microbiological Culture Collection Administration on September 2, 2021. The deposit address is No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing, China. The deposit number is: CGMCC No. 23341, and the deposit address is: Beijing, China, which has been made public.

[0042] (4) Pseudomonas putida ZTB-A29, whose classification name is Pseudomonas putida, was deposited in the General Microbiology Center of China National Microbiological Culture Collection on October 16, 2020, with the deposit address being Beijing, China, with the deposit number being CGMCC No. 20897, and the deposit address being No. 3, Yard No. 1, Beichen West Road, Chaoyang District, Beijing, China, which has been made public.

[0043] The invention provides a septic tank biodegrader, which comprises Bacillus tekila YZS-C01, Bacillus saffron YZS-C03, Bacillus faecalis QQB-A05 and Pseudomonas putida ZTB-A29; the mass ratio of solid bacterial powder or fermentation liquid of Bacillus tekila YZS-C01, Bacillus saffron YZS-C03, Bacillus faecalis QQB-A05 and Pseudomonas putida ZTB-A29 in the septic tank biodegrader is (2-5): (2-5): (1-2): (0.1-1); the total live bacteria concentration in the septic tank biodegrader is (5-8)×10 8 cfu / ml or (5~8)×10 8 cfu / g.

[0044] In the present invention, the septic tank biodegrader is preferably a solid preparation or a liquid preparation. When the septic tank biodegrader is a solid preparation, the septic tank biodegrader includes a carrier. In the present invention, the carrier is bran or rice bran. The total mass ratio of the carrier bran and rice bran to Bacillus tekila YZS-C01, Bacillus sabifon YZS-C03, Bacillus faecalis QQB-A05 and Pseudomonas putida ZTB-A29 is preferably 100: (2 to 5). In the present invention, when the septic tank biodegrader is a solid preparation, the fermentation broth of Bacillus tekila YZS-C01, Bacillus sabifon YZS-C03, Bacillus faecalis QQB-A05 and Pseudomonas putida ZTB-A29 is prepared into solid bacterial powder by freeze drying or spray drying, and then mixed in proportion.

[0045] In the present invention, when the septic tank biodegrader is a liquid preparation, it is obtained by mixing the fermentation broths of fermented Bacillus tergillus YZS-C01, Bacillus shaforniensis YZS-C03, Bacillus faecalis QQB-A05 and Pseudomonas putida ZTB-A29 in a mass ratio of (2-5): (2-5): (1-2): (0.1-1).

[0046] In the present invention, the preparation method of the solid bacterial powder preferably includes the following steps: activating the bacterial strain and then conducting continuous two-stage seed liquid culture, inoculating the fermentation medium for fermentation and concentrating to obtain a concentrated bacterial liquid, mixing the concentrated bacterial liquid of Bacillus faecalis QQB-A05 and Pseudomonas putida ZTB-A29 with a protective agent (skim milk powder) and freeze-drying to obtain the solid bacterial powder; mixing the fermentation liquid of Bacillus tergillus YZS-C01 and Bacillus shaforensis YZS-C03 with light calcium carbonate and spray-drying to obtain the solid bacterial powder.

[0047] In the present invention, the strain is inoculated into NB culture medium and cultured for 15 to 25 hours to obtain a primary seed solution; the culture temperature is preferably 25 to 35°C, and the culture speed is preferably 150 to 350 r·min -1 .

[0048] After obtaining the primary seed solution, the present invention transfers the primary seed solution to NB liquid culture medium for 10 to 20 hours to obtain the secondary seed solution. In the present invention, the inoculation amount of the primary seed solution is preferably 0.5% to 1.0% (V / V), the culture is preferably carried out in a fermenter, sterile air is introduced during the culture, the fermenter ventilation manually adjusts the speed and air flow to adjust DO (DO40%), the culture temperature is 25°C to 35°C, and the culture speed is 200 to 350 r·min -1The culturing time is preferably 10 to 20 hours. In the present invention, the OD of the secondary seed solution is 600 It is preferably 0.6 to 1.0.

[0049] After obtaining the secondary seed liquid, the present invention connects the secondary seed liquid to the fermentation medium for fermentation culture to obtain the fermentation liquid. In the present invention, the inoculation amount of the secondary seed liquid is 0.5% to 1.0% (V / V), and the tank pressure during the fermentation culture process is maintained at 0.05MPa; sterile air is introduced during the culture process, the fermentation tank ventilation manually adjusts the speed and air flow to adjust DO (DO40%), the culture temperature is 25°C to 35°C, and the culture speed is 200 to 350r·min -1 The culturing time is preferably 15 to 35 hours. In the present invention, the OD of the secondary seed solution is 600 It is preferably 0.8 to 1.5.

[0050] After obtaining the fermented bacterial liquid, the present invention concentrates the fermented bacterial liquid to obtain a concentrated bacterial liquid. In the present invention, the concentration multiple of the concentrated bacterial liquid is 5 to 20 times; the concentration method is that the fermented bacterial liquid of Bacillus faecalis QQB-A05 and Pseudomonas putida ZTB-A29 is obtained by high-speed centrifugation; the fermented bacterial liquid of Bacillus tequila YZS-C01 and Bacillus sabdariffa YZS-C03 is obtained by ceramic membrane filtration. The present invention does not specifically limit the specific parameter settings of the concentration equipment, and conventional parameter settings can be used to achieve the above-mentioned multiple concentrations.

[0051] After obtaining the fermented bacterial liquid, the fermented bacterial liquid of Bacillus faecalis QQB-A05 and Pseudomonas putida ZTB-A29 is centrifuged in a high-speed centrifuge to obtain a concentrated bacterial liquid, a protective agent (skim milk powder) is added to the concentrated bacterial liquid, and then solid bacterial powder is obtained by vacuum freeze drying, wherein the mass ratio of the concentrated liquid to the protective agent (skim milk powder) is 10:1, and the control conditions of the vacuum freeze drying process are as follows: cooling to -40°C for 3 to 5 hours, maintaining for 3 to 5 hours, and then vacuuming, passing hot water to control the temperature at -20°C, sublimation, and removing moisture. The fermented bacterial liquid of Bacillus tekila YZS-C01 and Bacillus shafusus YZS-C03 is filtered through a ceramic membrane to obtain a concentrated bacterial liquid, and light calcium carbonate is added to the concentrated bacterial liquid and then spray-dried to obtain a solid bacterial powder, the mass ratio of the concentrated liquid to the light calcium carbonate is 100:2-5, and the control indicators of the spray drying process are that the material inlet temperature is 150-200°C, the outlet temperature is 80-85°C, and the material drying time in the tower is 15-30S.

[0052] Example 1

[0053] (1) Preparation of bacterial liquid and solid bacterial powder of Alcaligenes faecalis QQB-A05:

[0054] Preparation of primary seed solution of Alcaligenes faecalis QQB-A05:

[0055] Activate the Alcaligenes faecalis QQB-A05 strain, pick a single colony, inoculate it into 150 ml NB medium (sterilized at 121°C for 30 min), and incubate at 28°C, 180 r·min -1 The mixture was shaken and cultured for 20 h under the same conditions to obtain the primary seed solution.

[0056] Preparation of secondary seed solution of Alcaligenes faecalis QQB-A05:

[0057] The first-grade seed liquid was inoculated into a sterile seed tank with an inoculation amount of 0.5% to 1.0% (V / V). The culture conditions were: ventilation was manually adjusted to adjust the speed and air flow to adjust DO (DO40%), the culture temperature was 25℃ to 35℃, and the speed was 200 to 350 r·min -1 , tank pressure 0.05MPa; culture time 10-15h. The OD of the secondary seed solution 600 The value is between 0.6 and 0.8, at which time the strain has the strongest activity.

[0058] Preparation of Alcaligenes faecalis QQB-A05 fermentation broth:

[0059] The secondary seed liquid of Alcaligenes faecalis QQB-A05 was inoculated into a sterile fermentation tank at an inoculation rate of 0.5% to 1.0% (V / V). The composition of the fermentation medium was as follows (per liter): 25-50 grams of yeast extract and 12-20 grams of maltose syrup. The culture conditions were: ventilation manually adjusted the speed and air flow to adjust DO (DO40%), the culture temperature was 25℃ to 35℃, and the speed was 200 to 350 r·min -1 , tank pressure 0.05MPa; culture time 15-20h, pH rises to 7.5-8.0, OD 600 When the value is 0.8-1.0, the fermentation is terminated and the mature fermentation liquid is obtained.

[0060] Preparation of Alcaligenes faecalis QQB-A05 powder:

[0061] The fermented mature bacterial liquid obtained above was centrifuged in a high-speed centrifuge, and then a protective agent (skim milk powder) was added and mixed evenly. The mass ratio of the concentrated bacterial liquid to the protective agent (skim milk powder) was 10:1, and a solid-liquid mixture was obtained, which was vacuum-freeze-dried. The vacuum-freeze-drying conditions were as follows: cooling to -40°C for 3 to 5 hours, maintaining for 3 to 5 hours, then vacuumizing, passing hot water to control the temperature at -20°C, sublimating, and removing moisture. The solid bacterial powder of Alcaligenes faecalis QQB-A05 was obtained, and the water content of the bacterial powder was less than 5% (W / W), and the number of viable bacteria was 2 to 5×10 11cfu / g.

[0062] (2) Preparation of Pseudomonas putida ZTB-A29 bacterial liquid and solid bacterial powder:

[0063] Activate the ZTB-A29 strain of Pseudomonas putida, pick a single colony, inoculate it into 150 ml NB medium (sterilized at 121°C for 30 min), and incubate at 28°C, 180 r·min -1 The mixture was shaken and cultured for 20 h under the same conditions to obtain the primary seed solution.

[0064] Preparation of secondary seed solution of Pseudomonas putida ZTB-A29:

[0065] The first-grade seed liquid was inoculated into a sterile seed tank with an inoculation amount of 0.5% to 1.0% (V / V). The culture conditions were: ventilation was manually adjusted to adjust the speed and air flow to adjust DO (DO40%), the culture temperature was 25℃ to 35℃, and the speed was 200 to 350 r·min -1 , tank pressure 0.05MPa; culture time 10-15h. The OD of the secondary seed solution 600 The value is between 0.6 and 0.8, at which time the strain has the strongest activity.

[0066] Preparation of Pseudomonas putida ZTB-A29 fermentation broth:

[0067] The secondary seed liquid of Pseudomonas putida ZTB-A29 was inoculated into a sterile fermentation tank at an inoculation rate of 0.5% to 1.0% (V / V). The composition of the fermentation medium was as follows (per liter): 25-50 grams of yeast extract and 12-20 grams of maltose syrup. The culture conditions were: ventilation manually adjusted the speed and air flow to adjust DO (DO40%), the culture temperature was 25℃ to 35℃, and the speed was 200 to 350 r·min -1 , tank pressure 0.05MPa; culture time 15-20h, pH rises to 7.5-8.0, OD 600 When the value is 0.8-1.0, the fermentation is terminated and the mature fermentation liquid is obtained.

[0068] Preparation of Pseudomonas putida ZTB-A29 bacterial powder:

[0069] The fermented mature bacterial liquid obtained above was centrifuged in a high-speed centrifuge, and then a protective agent (skim milk powder) was added and mixed evenly. The mass ratio of the concentrated bacterial liquid to the protective agent (skim milk powder) was 10:1, and a solid-liquid mixture was obtained, which was then freeze-dried in vacuum. The conditions for vacuum freeze-drying were: cooling to -40°C for 3 to 5 hours, maintaining for 3 to 5 hours, then vacuumizing, passing hot water to control the temperature at -20°C, sublimating, and removing moisture. The solid bacterial powder of Pseudomonas putida ZTB-A29 was obtained, and the moisture content of the bacterial powder was less than 5% (W / W), and the number of viable bacteria was 2 to 5×10 11 cfu / g.

[0070] (3) Preparation of Bacillus tergillosis YZS-C01 bacterial liquid and solid bacterial powder:

[0071] Activate the YZS-C01 strain of Bacillus tekila, pick a single colony, inoculate it into 150 ml NB medium (sterilized at 121°C for 30 min), and incubate at 28°C, 180 r·min -1 The mixture was shaken and cultured for 20 h under the same conditions to obtain the primary seed solution.

[0072] Preparation of secondary seed solution of Bacillus tekila YZS-C01:

[0073] The first-grade seed liquid was inoculated into a sterile seed tank with an inoculation amount of 0.5% to 1.0% (V / V). The culture conditions were: ventilation was manually adjusted to adjust the speed and air flow to adjust DO (DO40%), the culture temperature was 30℃ to 35℃, and the speed was 200 to 350 r·min -1 , tank pressure 0.05MPa; culture time 15-20h. The OD of the secondary seed solution 600 The value is between 0.6 and 0.8, at which time the strain has the strongest activity.

[0074] Preparation of fermentation broth of Bacillus tequila YZS-C01:

[0075] The secondary seed liquid of Bacillus tekila YZS-C01 is inoculated into a sterile fermentation tank at an inoculation rate of 0.5% to 1.0% (V / V). The composition of the fermentation medium is as follows (per liter): yeast extract: 10-20 grams, maltose syrup: 25-35 grams, magnesium chloride: 0.2-0.5 grams, calcium chloride: 0.1-0.4 grams, manganese chloride: 0.1-0.4 grams, dipotassium hydrogen phosphate: 1.0-2.5 grams. The culture conditions are: ventilation manually adjusts the speed and air flow to adjust DO (DO40%), the culture temperature is 30℃ to 35℃, and the speed is 200 to 350r·min -1 , tank pressure 0.05MPa; culture time 30-35h, pH rises to 7.5-8.0, OD 600 The value is 1.2-1.5. When the spores are more than 90% mature after sampling and microscopic examination, the fermentation is terminated to obtain the mature fermentation liquid.

[0076] Preparation of Bacillus tequila YZS-C01 powder:

[0077] The fermented mature bacterial liquid obtained above was filtered through a ceramic membrane, and then light calcium carbonate was added and mixed evenly. The mass ratio of the concentrated bacterial liquid to the light calcium carbonate was 100:2-5, and a solid-liquid mixture was obtained, which was spray-dried. The spray-drying conditions were: the material inlet temperature was 150-200°C, the outlet temperature was 80-85°C, and the material was dried in the tower for 15-30 seconds. The solid bacterial powder of Bacillus tequila YZS-C01 was obtained, and the moisture content of the bacterial powder was less than 5% (W / W), and the number of viable bacteria was 2-5×10 11 cfu / g.

[0078] (4) Preparation of Bacillus shafusus YZS-C03 bacterial liquid and solid bacterial powder:

[0079] Activate the Bacillus sabdariffa YZS-C03 strain, pick a single colony, inoculate it into 150 ml NB medium (sterilized at 121°C for 30 min), and incubate at 28°C, 180 r·min -1 The mixture was shaken and cultured for 20 h under the same conditions to obtain the primary seed solution.

[0080] Preparation of secondary seed solution of Bacillus sabdariffa YZS-C03:

[0081] The first-grade seed liquid was inoculated into a sterile seed tank with an inoculation amount of 0.5% to 1.0% (V / V). The culture conditions were: ventilation was manually adjusted to adjust the speed and air flow to adjust DO (DO 40%), the culture temperature was 30°C to 35°C, and the speed was 200 to 350 r·min. -1 , tank pressure 0.05MPa; culture time 15-20h. The OD of the secondary seed solution 600 The value is between 0.6 and 0.8, at which time the strain has the strongest activity.

[0082] Preparation of Bacillus saffron YZS-C03 fermentation broth:

[0083] The secondary seed liquid of Bacillus sabdariffa YZS-C03 is inoculated into a sterile fermentation tank at an inoculation rate of 0.5% to 1.0% (V / V). The composition of the fermentation medium is as follows (per liter): yeast extract: 10-20 grams, maltose syrup: 25-35 grams, magnesium chloride: 0.2-0.5 grams, calcium chloride: 0.1-0.4 grams, manganese chloride: 0.1-0.4 grams, dipotassium hydrogen phosphate: 1.0-2.5 grams. The culture conditions are: ventilation manually adjusts the speed and air flow to adjust DO (DO40%), the culture temperature is 30℃ to 35℃, and the speed is 200 to 350r·min -1 , tank pressure 0.05MPa; culture time 30-35h, pH rises to 7.5-8.0, OD 600 The value is 1.2-1.5. When the spores are more than 90% mature after sampling and microscopic examination, the fermentation is terminated to obtain the mature fermentation liquid.

[0084] Preparation of Bacillus sabdariffa YZS-C03 powder:

[0085] The fermented mature bacterial liquid obtained above was filtered through a ceramic membrane, and then light calcium carbonate was added and mixed evenly. The mass ratio of the concentrated bacterial liquid to light calcium carbonate was 100:2-5, and a solid-liquid mixture was obtained, which was spray-dried. The spray-drying conditions were: the material inlet temperature was 150-200°C, the outlet temperature was 80-85°C, and the material was dried in the tower for 15-30 seconds. The solid bacterial powder of Bacillus shafusus YZS-C03 was obtained, and the moisture content of the bacterial powder was less than 5% (W / W), and the number of viable bacteria was 2-5×10 11 cfu / g.

[0086] Example 2

[0087] The solid bacterial powders of Bacillus teteguila YZS-C01, Bacillus shafusus YZS-C03, Bacillus faecalis QQB-A05 and Pseudomonas putida ZTB-A29 obtained in the above Example 1 were mixed in a mass ratio of 5:5:2:1 to obtain a solid mixed bacterial powder. Then, the carrier bran and rice bran (the bran and rice bran were mixed in any mass ratio) were mixed with the above solid mixed bacterial powder in a mass ratio of 100:5 to obtain a solid preparation 1 of a septic tank biodegrader, wherein the total live bacteria concentration in the septic tank biodegrader was 8×10 8 cfu / g.

[0088] The solid bacterial powders of Bacillus teteguila YZS-C01, Bacillus shafusus YZS-C03, Bacillus faecalis QQB-A05 and Pseudomonas putida ZTB-A29 obtained in the above Example 1 were mixed in a mass ratio of 2:2:1:0.1 to obtain a solid mixed bacterial powder. Then, the carrier bran and rice bran (the bran and rice bran were mixed in any mass ratio) were mixed with the above solid mixed bacterial powder in a mass ratio of 100:2 to obtain a solid preparation 2 of a septic tank biodegrader, wherein the total live bacteria concentration in the septic tank biodegrader was 5×10 8 cfu / g.

[0089] The solid bacterial powders of Bacillus teteguila YZS-C01, Bacillus shafusus YZS-C03, Bacillus faecalis QQB-A05 and Pseudomonas putida ZTB-A29 obtained in the above Example 1 were mixed in a mass ratio of 3:3:2:0.5 to obtain a solid mixed bacterial powder. Then, the carrier bran and rice bran (the bran and rice bran were mixed in any mass ratio) were mixed with the above solid mixed bacterial powder in a mass ratio of 100:3 to obtain a solid preparation 3 of a septic tank biodegrader, wherein the total live bacteria concentration in the septic tank biodegrader was 6×10 8 cfu / g.

[0090] Example 3

[0091] The fermented mature bacterial liquids of Bacillus teteguila YZS-C01, Bacillus shafusus YZS-C03, Bacillus faecalis QQB-A05 and Pseudomonas putida ZTB-A29 obtained in the above Example 1 were mixed in a mass ratio of 5:5:2:1 to obtain a septic tank biodegrader liquid preparation 1.

[0092] The fermented mature bacterial liquids of the above-mentioned Bacillus teteguila YZS-C01, Bacillus shafusus YZS-C03, Bacillus faecalis QQB-A05 and Pseudomonas putida ZTB-A29 were mixed in a mass ratio of 2:2:1:0.1 to obtain a septic tank biodegrader liquid preparation 2.

[0093] The fermented mature bacterial liquids of Bacillus tertequila YZS-C01, Bacillus shafusus YZS-C03, Bacillus faecalis QQB-A05 and Pseudomonas putida ZTB-A29 obtained in the above Example 1 were mixed in a mass ratio of 3:3:2:0.5 to obtain a septic tank biodegrader liquid preparation 3.

[0094] Example 4

[0095] Take a fecal sample from a septic tank, weigh 300 grams and put it into a transparent plastic box, and add 200 mL of sterile water. After the fecal sample in the plastic box floats up, record the thickness of the scum, and add 5% of the weight of the feces to obtain a septic tank biodegradation agent solid preparation 1 in Example 2 as a treatment group; a control group without any degrading agent is used, and three replicates are set for each group. Regularly observe the thickness of the scum in the plastic box, color changes, etc.

[0096] Table 1 Determination of biodegradation effect of septic tank biodegraders

[0097]

[0098] As can be seen from Table 1, during the implementation of this embodiment, the decomposition of feces in the plastic box showed a change characteristic of first floating up and then settling. After adding the septic tank biodegrader liquid preparation 1 obtained in Example 2, the feces scum was accelerated to degrade and settle, and the color of the feces liquid gradually turned black. On the 10th day, the scum in the treatment group was 100% settled, the bottom residue degradation rate was 69.82%, and the feces liquid was black; while the average scum sedimentation rate of the control group was 32.50%, and the natural degradation rate of the bottom residue was only 8.25%, indicating that the septic tank biodegrader liquid preparation 1 of the present invention has a significant biodegradation effect.

[0099] Example 5

[0100] The deodorization effect of a septic tank in a certain community was measured on site. After odor analysis and olfactory perception, a septic tank with a very obvious odor was selected. The septic tank biodegradation agent liquid preparation 2 obtained in Example 3 of the present invention was added at 0.5 L / m 2 Spray evenly on the surface. After spraying, use an air sampling bag to collect samples every 2 hours to measure the concentrations of NH3 and H2S. The detection of NH3 uses Nessler's reagent colorimetry, and the detection of H2S uses methylene blue colorimetry. Regularly observe the thickness and color changes of septic tank scum.

[0101] Table 2 Determination of the removal effect of NH3 and H2S by septic tank biodegraders

[0102]

[0103] Table 3 Determination of biodegradation effect of septic tank biodegraders

[0104]

[0105] As can be seen from Table 2, after the septic tank biodegradation agent liquid preparation 2 of the present invention is sprayed to treat feces, the removal rate of ammonia and hydrogen sulfide is significant. After 4 hours of application, the removal rate of ammonia and hydrogen sulfide is still above 60%, and the deodorization effect is significant. As can be seen from Table 3, during the treatment of the septic tank biodegradation agent liquid preparation 2 of the present invention, the scum and bottom feces are accelerated to degrade, the scum thickness first increases and then decreases, the scum hardness gradually softens and turns black, and the scum degradation rate reaches 60% after 10 days of treatment, and the biodegradation effect is significant.

[0106] The above description is only a preferred embodiment of the present invention. All equivalent changes and modifications made according to the scope of the patent application of the present invention should fall within the scope of the present invention.

Claims

1. A septic tank biodegrader, characterized in that: Including Bacillus tequila YZS-C01 ( Bacillus tequilensis )、Bacillus saffron YZS-C03( Bacillus safensis ), Alcaligenes faecalis QQB-A05 ( Alcaligenes faecalis ) and Pseudomonas putida ZTB-A29 ( Pseudomonas putida ); the deposit number of the Bacillus tequila YZS-C01 is: CGMCC No.22856; the deposit number of the Bacillus shafusus YZS-C03 is: CGMCC No.21798; the deposit number of the Alcaligenes faecalis QQB-A05 is: CGMCC No.23341; the deposit number of the Pseudomonas putida ZTB-A29 is: CGMCC No.20897.

2. The septic tank biodegrader according to claim 1, characterized in that: The mass ratio of the solid bacterial powder or fermentation liquid of Bacillus tequila YZS-C01, Bacillus shafusus YZS-C03, Bacillus faecalis QQB-A05 and Pseudomonas putida ZTB-A29 is (2-5): (2-5): (1-2): (0.1-1).

3. The septic tank biodegrader according to claim 1, characterized in that: The total live bacteria concentration is (5~8)×10 8 cfu / ml or (5~8)×10 8 cfu / g.

4. The septic tank biodegrader according to claim 1, characterized in that: The septic tank biodegrader is a liquid preparation or a solid preparation.

5. The septic tank biodegrader according to claim 1, characterized in that: The preparation method of the septic tank biodegrader solid preparation is as follows: solid bacterial powders of Bacillus tequila YZS-C01, Bacillus shafusus YZS-C03, Bacillus faecalis QQB-A05 and Pseudomonas putida ZTB-A29 are mixed in a mass ratio to prepare a solid mixed bacterial powder; a carrier is mixed with the solid mixed bacterial powder in a mass ratio of 100:2-5 to obtain the septic tank biodegrader solid preparation; the carrier is bran and rice bran mixed in any proportion.

6. The septic tank biodegrader according to claim 5, characterized in that: The fermentation broth of Bacillus tequila YZS-C01, Bacillus shafusus YZS-C03, Bacillus faecalis QQB-A05 and Pseudomonas putida ZTB-A29 is mixed in a mass ratio of (2-5): (2-5): (1-2): (0.1-1) to obtain the fermentation broth.

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