CN110295118A - A kind of degraded cellulose, the Tarlaromyces flavus bacterial strain for adsorbing cadmium and its application - Google Patents
A kind of degraded cellulose, the Tarlaromyces flavus bacterial strain for adsorbing cadmium and its application Download PDFInfo
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- B—PERFORMING OPERATIONS; TRANSPORTING
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Abstract
The present invention provides a kind of degraded cellulose, the Tarlaromyces flavus bacterial strain of absorption cadmium, the classification naming of the bacterial strain is Tarlaromyces flavus (Talaromycesveerkampii) CAF2, deposit number are as follows: ACCC 32732.The concentration of cadmium ions tolerance range of the Tarlaromyces flavus bacterial strain is 0-100mg/L；Its energy cellulase-producing, and have good degradation effect to rice straw, it is applied in the degradation of rice straw, in the 15th day half that is degraded；Meanwhile the bacterial strain have adsorbing heavy metal cadmium ability, applied cadmium pollution soil improvement in, the 30th day Tarlaromyces flavus bacterial strain to every adsorption rate be more than 74%.The bacterial strain is suitable for the biological prosthetic of cadmium pollution rice field, while degrading rice straw, can also achieve the effect that rehabilitating soil metal cadmium pollution.
Description
Technical field
The present invention relates to technical field of environmental management, in particular to a kind of degraded cellulose, the Tarlaromyces flavus bacterium for adsorbing cadmium
Strain and its application.
Background technique
With the rapid development of industry, heavy metal pollution of soil phenomenon is also on the rise.Common soil pollution heavy metal
Mainly there are Hg, Cd, Pb, As etc., as such heavy metal migrates in ecological environment, converts, and constantly accumulates, reach a fixed limit
When angle value, the life and health for endangering animals and plants, the mankind will be recycled by food chain.
Chinese patent CN 102286383A discloses a kind of application of yellow indigo plant shape bacterium in terms of phytopathogen prevention and treatment,
The bacterial strain is able to suppress a variety of pathogens, as Peanut Web Blotch Disease bacterium, tomato early blight bacterium, Rhizoctonia solani Kuhn, Rhizoctonia solani Kahn,
Valsa mali, tobacco brown spot pathogen etc.；In addition, the hemicellulose that the research report in relation to Tarlaromyces flavus includes the strain drops
(1. Yin are small, Tan Jiajin, mono- plant of Tarlaromyces flavus (Talaromycesflavus) of Fang Aiqin for solution characteristic and its phosphorus decomposing characteristic
Measurement [J] Nanjing Forestry University's journal (natural science edition) of SH16 phosphorus decomposing characteristic, 2017 (5)；2. Gu Wenjie, Xu Youquan, Xu
Form-intelligence waits the screening of efficient hemicellulose degradation bacterium and identification [J] microorganism journal, 2012,52 (10) in acid soil).
Currently, temporarily having no the report in relation to the Tarlaromyces flavus with degraded cellulose simultaneously, adsorbing heavy metal cadmium ability.
Summary of the invention
The present invention provides a kind of degraded cellulose, adsorb cadmium Tarlaromyces flavus bacterial strain and its application, its purpose is to
So that Tarlaromyces flavus bacterial strain is suitable for the biological prosthetic of cadmium pollution rice field, while degrading rice straw, can also reach a reparation huge sum of money
Belong to the effect of contaminated soil.
In order to achieve the above object, the invention provides the following technical scheme:
A kind of degraded cellulose, the Tarlaromyces flavus bacterial strain for adsorbing cadmium, the classification naming of the bacterial strain are Tarlaromyces flavus
(Talaromycesveerkampii) CAF2, deposit number are as follows: ACCC 32732, biological characteristics are as follows: bacterium colony quality is cotton-shaped
And velvet-like, edge white to faint yellow, middle part light green, produce yellow green spore；Most suitable culture pH is 6.5；Most suitable cultivation temperature
It is 28 DEG C；Spore can be produced by cultivating 3~5 days under the conditions of 25~28 DEG C of solid fermentation；Can be with glucose, sucrose, wheat bran or stalk
Microbe carbon source is grown；It can be grown by microorganism nitrogen source of peptone, yeast extract or corn flour.
Preferably, the concentration of cadmium ions tolerance range of the Tarlaromyces flavus bacterial strain is 0~100mg/L.
The present invention also provides a kind of application of above-mentioned Tarlaromyces flavus bacterial strain in rice straw degradation.
Preferably, above-mentioned application includes the following steps:
(1) rice straw fluid nutrient medium is prepared；
(2) the Tarlaromyces flavus bacterial strain is prepared into seed liquor, is connected to rice straw Liquid Culture by 10% inoculum concentration
In base, shaken cultivation 5~30 days under 28 DEG C, the revolving speed of 120r/min.
It is highly preferred that the rice straw fluid nutrient medium is by CMC-Na, KH2PO4、MgSO4·7H2O, NaCl, albumen
Peptone, yeast extract, distilled water and rice straw composition.
It is highly preferred that the rice straw fluid nutrient medium includes following parts by weight of component: 10 parts of CMC-Na, KH2PO4 1
Part, MgSO4·7H2After 0.5 part of O, 0.5 part of NaCl, 2 parts of peptone, 0.5 part of yeast extract, 1000 parts of distilled water, and shearing
10 parts of rice straw.
Preferably, evaluation Tarlaromyces flavus bacterial strain includes by the rice after step (2) are cultivated to rice straw degradation effect
Straw solution is centrifuged, is cleaned and drying and processing, and rice straw weight-loss ratio is calculated.
The present invention also provides a kind of application of Tarlaromyces flavus bacterial strain in cadmium pollution soil improvement.
Above-mentioned application includes the following steps:
(1) the Tarlaromyces flavus bacterial strain is obtained into conidia powder after solid fermentation, drying and crushing；
(2) spare after drying, clean by cadmium pollution soil, pulverizing, being sieved；
(3) conidia powder obtained by step (1) is added step (2) treated in cadmium pollution soil, contain soil miospore
Amount is 1 × 106~1 × 107CFU/g is cultivated 10~50 days naturally at room temperature after adding water and stirring.
Preferably, after step (3) is cultivated 30 days naturally Tarlaromyces flavus bacterial strain to every adsorption rate be more than 74%.
Above scheme of the invention have it is following the utility model has the advantages that
Tarlaromyces flavus bacterial strain CAF2 provided by the invention, classification naming are Tarlaromyces flavus
(Talaromycesveerkampii) WAF6 is preserved in Chinese agriculture Microbiological Culture Collection administrative center, deposit number
Are as follows: ACCC 32732.It can cellulase-producing, and have good degradation effect to rice straw；It is applied in rice straw
In the degradation of stalk, rice straw was in the 15th day half that is degraded；Meanwhile the bacterial strain has the ability of adsorbing heavy metal cadmium, it can
It reduces the biological effectiveness of the heavy metal cadmium in contaminated soil and there is good colonization ability in the soil, applied in cadmium
Contaminated soil administer in, after the 30th day, Tarlaromyces flavus bacterial strain to every adsorption rate be more than 74%.The bacterial strain is suitable for cadmium pollution
Rice field it is biological prosthetic, while degrading rice straw, can also achieve the effect that repairing heavy metal in soil pollutes.
Detailed description of the invention
Fig. 1 is the strain culturing 25 days disintegration effects to rice straw of the invention；
Fig. 2 is the degradation effect of different strains of the invention to rice straw；
Fig. 3 is the adsorption effect of bacterial strain CAF2 of the invention in cadmium pollution soil.
Specific embodiment
To keep the technical problem to be solved in the present invention, technical solution and advantage clearer, below in conjunction with specific implementation
Example is described in detail.
Embodiment 1: this example demonstrates that the present invention uses culture medium
(1) isolation medium: CMC culture medium: sodium carboxymethylcellulose (CMC-Na) 10.0g, KH2PO4 0.25g, potato
Juice 100mL, agar powder 18.0g, distilled water 1000mL.
(2) purifying, Storaged media: PDA culture medium: potato 200g, glucose 20.0g, agar powder 18.0g, distillation
Water 1000mL.
(3) culture medium containing cadmium ion: potato 200g, glucose 20.0g, agar powder 18.0g, distilled water 1000mL, root
Various concentration concentration of cadmium ions solid medium is configured to according to needing to add mother liquor containing cadmium ion (20mg/mL).
(4) screening and culturing medium:
A. Congo red culture medium: CMC-Na 2.0g, KH2PO40.5g, MgSO4·7H2O 0.25g, (NH4)2SO4
1.0g, Congo red 0.1g, agar powder 18.0g, distilled water 1000mL；
B. secondary screening culture medium: CMC-Na 5.0g, KH2PO41.0g, MgSO4·7H2O 0.5g, NaNO33.0g, KCl
0.5g, FeCl3·6H2O 0.01g, distilled water 1000mL；
(5) rice straw fluid nutrient medium (by weight): 10 parts of CMC-Na, KH2PO41 part, MgSO4·7H2O 0.5
10 parts of rice straw after part, 0.5 part of NaCl, 2 parts of peptone, 0.5 part of yeast extract, 1000 parts of distilled water, and shearing.
Embodiment 2: strains tested isolates and purifies
Strains tested isolates and purifies in the present invention method particularly includes: dilution plate rubbing method is used, by pedotheque
10 are made with sterile water by gradient dilution-3-10-5Bacteria suspension, and be coated on isolation medium, it is placed in 26-28 DEG C of constant temperature training
It supports and is cultivated 2-3 days in case, the different fungal bacterial strains being separated to are subjected to scribing line purifying and preservation in PDA culture medium, will be purified
Good strain is put in 4 DEG C of Storage in refrigerator.
The classification naming of the bacterial strain is Tarlaromyces flavus (Talaromycesveerkampii) CAF2, deposit number are as follows:
ACCC 32732, biological characteristics are as follows: bacterium colony quality is cotton-shaped and velvet-like, edge white to faint yellow, middle part light green, production are yellowish green
Color spore；Most suitable culture pH is 6.5；Most suitable cultivation temperature is 28 DEG C；3~5 days are cultivated under the conditions of 25~28 DEG C of solid fermentation i.e.
Spore can be produced；It can be grown using glucose, sucrose, wheat bran or stalk as microbe carbon source；It can be with peptone, yeast extract or corn
Powder is that microorganism nitrogen source is grown.
Embodiment 3: the cadmium ion tolerance evaluation of bacterial strain CAF2
The cadmium ion tolerance evaluation method of bacterial strain CAF2 of the present invention, specifically includes the following steps:
(1) prepare the PDA culture medium containing different concentration of cadmium ions, concentration is set as 0,10,50,80,100,150,
200mg/L；
(2) bacteria cake is made with 5mm punch in bacterial strain, is aseptically inoculated in step (1) described plating medium,
It is cultivated 3-5 days in 26-28 DEG C of constant incubator, observes the growing state of bacterial strain.
Bacterial strain CAF2 growing state on the plate for the concentration of cadmium ions that concentration of cadmium ions is 0~100mg/L is good, and
It is grown on the plate that concentration of cadmium ions is 150mg/L and receives inhibition completely, therefore the concentration of cadmium ions tolerance range of the bacterial strain is 0-
100mg/L。
Embodiment 4: the degraded cellulose merit rating of bacterial strain CAF2
Specifically includes the following steps:
(1) primary dcreening operation of bacterial strain: the fungal bacterial strain spore suspension isolated and purified is coated on PDA, in 28 DEG C of perseverances
It is cultivated 2-3 days in warm incubator, uniformly to the growth of media surface bacterium colony, bacteria cake is made with 5mm punch and is inoculated in the Congo
It on red culture medium, is placed in 28 DEG C of constant incubators and cultivates 3-5 days, whether have hydrolysis, and measure if observing around bacteria cake
Colony diameter (d) and hydrolytic circle (D)；
(2) measurement of carboxymethylcelluloenzyme enzyme (CMCase) living:
1) Specification Curve of Increasing: drying to constant weight under the conditions of 80 DEG C for DEXTROSE ANHYDROUS, and being prepared into concentration is 1.0mg/
The glucose standard of mL is separately added into glucose standard 0,0.2,0.4,0.6,0.8,1.0mL in 6 clean tubes, mends
Distilled water is added boiling water bath 5min after 1.5mLDNS reagent, is settled to 25mL after cooling, measures OD under wavelength 540nm to 2.0mL
Value draws standard curve using Excel.
2) crude enzyme liquid preparation and enzyme activity determination: seed liquor is prepared using the fungal bacterial strain that primary dcreening operation obtains, by 10% inoculation
Amount is connected in secondary screening culture medium, and liquid amount is 90mL/250mL triangular flask, 28 DEG C of constant-temperature shaking cultures, respectively 3,5,7,9,11
It takes fermentation liquid 1.5mL in centrifuge tube, and 10000r/min centrifugation 10min obtains crude enzyme liquid, measures bacterial strain in different fermentations
Between under carboxymethylcelluloenzyme enzyme (CMCase) vigor." Li Jing, Zhang Hanneng, Zhao's Chong wait to enzyme activity determination method reference literature
Highly effective cellulose degradation bacteria separation screening, composite microbial system building and straw degradative effect analysis apply with environmental organism journal,
2016,22 (4): the method being distributed in 0689-0696.." measure the time of occurrence that post analysis records highest enzyme activity.
As shown in table 1, in order to illustrate the effect of bacterial strain more of the present invention, table 1 provides the degraded cellulose of different strains
Indexes of capability evaluation.The ratio size of hydrolytic circle (D) and colony diameter (d) can only qualitative performance's bacterial strain there is CMC-Na
Capacity of decomposition, and enzyme activity size is then the quantitative target for determining strains for degrading cellulose ability；As shown in Table, bacterial strain CAF1,
The highest enzyme activity time of occurrence of CAF2 is identical but the highest enzyme activity of CAF2 is slightly larger than CAF1.
Each strains for degrading cellulose merit rating of table 1.
Embodiment 5: application of the bacterial strain CAF2 in rice straw degradation
Specifically includes the following steps:
(1) disintegration effect of the bacterial strain to rice straw: rice straw is cut into about 0.5cm*3cm strip with scissors, by water
Rice straw substitutes carbon source (CMC-Na) in secondary screening culture medium, and seed liquor is connected to by 0.5g rice straw/bottle by 10% inoculum concentration
Liquid amount is in 45mL/250mL triangular flask, and 28 DEG C, constant-temperature shaking culture under the conditions of 120r/min, routine observation rice straw collapse
Solution situation simultaneously records.
(2) degradation effect of the bacterial strain to rice straw: crossing 40 meshes after rice straw is crushed, rice straw is substituted multiple
Carbon source (CMC-Na) in culture medium is sieved, 2.0g rice straw/bottle, seed liquor, which is connected to liquid amount, by 10% inoculum concentration is
In 90mL/250mL triangular flask, to access 10mL distilled water as control, 28 DEG C, constant-temperature shaking culture under the conditions of 120r/min, 25
After it, culture 5000r/min is centrifuged 10min, supernatant is abandoned, is cleaned 3-5 times repeatedly with distilled water, 80 DEG C are dried to perseverance
Weight calculates weight-loss ratio.
If strains tested is disintegrated effect as shown in table 2. to rice straw, strains tested is inoculated in the training of rice straw liquid
After supporting base culture 25 days, bacterial strain CAF1 and CF3 are respectively at first 5 days, 15 days, and rice straw is without apparent metamorphosis；And from
Starting within 5th day, rice straw is gradually degraded under bacterial strain CAF2 effect, in 15 days, the rice straw for the half that can degrade.
For different strains to the disintegration effect of rice straw as shown in Figure 1, under bacterial strain CAF2 effect, rice straw is degraded one
Half.
2. rice straw of table is disintegrated degree
Note :+: stalk is degraded less；++: stalk is degraded half；+++: stalk is degraded substantially；++++: stalk is complete
Degradable
As shown in Fig. 2, each bacterial strain has certain degradation to rice straw, using the present embodiment bacterial strain to rice straw
Calculation method in the degradation effect of stalk about weight-loss ratio obtains: the degradation effect of bacterial strain CAF2 is preferable, and degradation rate is
The degradation rate of 33.97%, bacterial strain CAF1 are 25.22%, and the degradation effect of bacterial strain CF3 is worst, only 2.94%.
Embodiment 6: application of the bacterial strain CAF2 in cadmium pollution soil improvement
Specifically includes the following steps:
1) by different strains testeds by solid fermentation, drying, crushing and etc. obtain spore pulvis, and detection level；
2) it will be air-dried for examination soil natural, remove the sundries such as plant roots and stems, it is spare to pulverize 80 meshes of sieving；
3) it is dispensed for examination soil, dress 100g cadmium pollution soil in 250mL beaker adds conidia powder by different disposal, and adjust
Inoculum concentration makes each fungal spore content 1 × 10 in soil6-1×107CFU/g, while control group is set；
4) soil incubation, each processing addition 100mL water are simultaneously stirred evenly with glass bar, put at room temperature cultivation 10 naturally~
50 days；
5) the 0th, 10,20,30d be measured by sampling effective cadmium content of soil, after every sub-sampling, be added equivalent amount of water with
Ensure soil moisture regime；
As shown in figure 3, with the extension of soil incubation time, effective cadmium content in soil was gradually reduced, at the 30th day
Afterwards, compared to initial effective cadmium content, the adsorption rate of bacterial strain has reached 74.27%, illustrates bacterial strain in the present invention in the soil
With good colonization ability, and to provide available microbial resources in heavy metal pollution repairing and treating.
The above is a preferred embodiment of the present invention, it is noted that for those skilled in the art
For, without departing from the principles of the present invention, it can also make several improvements and retouch, these improvements and modifications
It should be regarded as protection scope of the present invention.
Claims (10)
1. a kind of degraded cellulose, the Tarlaromyces flavus bacterial strain for adsorbing cadmium, which is characterized in that the classification naming of the bacterial strain is champac
Shape bacterium (Talaromycesveerkampii) CAF2, deposit number are as follows: ACCC 32732, biological characteristics are as follows: bacterium colony quality
Cotton-shaped and velvet-like, edge white to faint yellow, middle part light green produces yellow green spore；Most suitable culture pH is 6.5；Most suitable culture
Temperature is 28 DEG C；Spore can be produced by cultivating 3~5 days under the conditions of 25~28 DEG C of solid fermentation；It can be with glucose, sucrose, wheat bran or straw
Stalk is that microbe carbon source is grown；It can be grown by microorganism nitrogen source of peptone, yeast extract or corn flour.
2. Tarlaromyces flavus bacterial strain according to claim 1, which is characterized in that the concentration of cadmium ions of the Tarlaromyces flavus bacterial strain is resistance to
It is 0~100mg/L by range.
3. a kind of application of Tarlaromyces flavus bacterial strain as claimed in claim 1 or 2 in rice straw degradation.
4. application according to claim 3, which comprises the steps of:
(1) rice straw fluid nutrient medium is prepared；
(2) the Tarlaromyces flavus bacterial strain is prepared into seed liquor, is connected in rice straw fluid nutrient medium by 10% inoculum concentration,
Shaken cultivation 5~30 days under 28 DEG C, the revolving speed of 120r/min.
5. application according to claim 4, which is characterized in that the rice straw fluid nutrient medium by CMC-Na,
KH2PO4、MgSO4·7H2O, NaCl, peptone, yeast extract, distilled water and rice straw composition.
6. application according to claim 4, which is characterized in that the rice straw fluid nutrient medium includes following parts by weight
Component: CMC-Na10 parts, KH2PO41 part, MgSO4·7H2O0.5 parts, NaCl0.5 parts, 2 parts of peptone, 0.5 part of yeast extract, steaming
10 parts of rice straw after 1000 parts of distilled water, and shearing.
7. application according to claim 3, which is characterized in that evaluation Tarlaromyces flavus bacterial strain is to rice straw degradation effect packet
It includes and the rice straw solution after step (2) culture is centrifuged, is cleaned and drying and processing, calculate rice straw weight-loss ratio.
8. a kind of application of Tarlaromyces flavus bacterial strain as claimed in claim 1 or 2 in cadmium pollution soil improvement.
9. application according to claim 8, which comprises the steps of:
(1) the Tarlaromyces flavus bacterial strain is obtained into conidia powder after solid fermentation, drying and crushing；
(2) spare after drying, clean by cadmium pollution soil, pulverizing, being sieved；
(3) conidia powder obtained by step (1) is added step (2) treated in cadmium pollution soil, make soil miospore content 1
×106~1 × 107CFU/g is cultivated 10~50 days naturally at room temperature after adding water and stirring.
10. application according to claim 9, which is characterized in that the Tarlaromyces flavus bacterial strain after step (3) is cultivated 30 days naturally
To every adsorption rate be more than 74%.
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| CN113717860B (en) * | 2021-07-07 | 2023-05-16 | 昆明理工大学 | Application of Huang Lanzhuang bacteria in conversion of total saponins of Notoginseng radix into small-polarity ginsenoside |
| CN113373069A (en) * | 2021-08-05 | 2021-09-10 | 中国科学院东北地理与农业生态研究所 | Straw saprophytic fungus and application thereof |
| CN113373069B (en) * | 2021-08-05 | 2022-12-06 | 中国科学院东北地理与农业生态研究所 | Straw saprophytic fungus and application thereof |
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