CN112159767A - Method for improving biological control efficiency of acid pathogenic fungi antagonistic yeast after fruit and vegetable harvest and application - Google Patents
Method for improving biological control efficiency of acid pathogenic fungi antagonistic yeast after fruit and vegetable harvest and application Download PDFInfo
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Abstract
The invention relates to a method for improving biological control efficiency of acid pathogenic fungi antagonistic yeast after fruit and vegetable harvest, which comprises the following steps: activating bio-control yeast; performing liquid culture on the activated biocontrol yeast; the biocontrol yeast is marine yeast Rhodosporidium paludigenum Fell and Tallman, the strain preservation number is IMI 394084, organic acid is added into a culture medium, and the biocontrol yeast is applied to improving the capability of antagonistic yeast in preventing and controlling acid pathogenic fungi of picked fruits and vegetables, wherein the acid pathogenic fungi comprise: the penicillium digitatum, the penicillium italicum, the penicillium expansum and the botrytis cinerea and the sclerotinia sclerotiorum can effectively improve the survival rate of antagonistic yeast in different organic acid stresses and improve the capability of the antagonistic yeast in preventing and treating postharvest diseases, are safe, efficient, green and pollution-free, accord with the natural and environment-friendly idea, and are easy to implement large-scale equipment due to the high growth and propagation speed of microorganisms.
Description
Technical Field
The invention belongs to the field of disease control of picked fruits and vegetables, relates to an antagonistic yeast biological control technology, and particularly relates to a method for improving biological control efficiency of acid pathogenic fungi antagonistic yeast of picked fruits and vegetables and application thereof.
Background
The postharvest diseases of fruits and vegetables are mainly caused by pathogenic fungi, the losses of fruit and vegetable products in China reach hundreds of billions of yuan RMB every year, and the resource and economic losses are serious. Not only are the pathogenic fungi responsible for the enormous losses in fruit numbers, but the large amount of secondary metabolites secreted by many pathogenic fungi may also cause serious food safety problems, such as patulin produced by penicillium expansum, and the like.
The pathogenic fungi causing postharvest diseases of fruits and vegetables mainly comprise penicillium, rhizopus, alternaria, botrytis, sclerotinia and the like, and are divided into 2 types, namely acid pathogenic fungi and alkaline pathogenic fungi. Among them, the acidic pathogenic fungi, which are the main body of postharvest fungal diseases, include penicillium digitatum, penicillium italicum, penicillium expansum, etc., botrytis cinerea, etc., and sclerotinia, etc., which can infect various hosts, causing the most serious loss. The acidic pathogenic fungi can secrete organic acids such as oxalic acid, citric acid, glucuronic acid and the like in the growth and infection processes of the acidic pathogenic fungi, so that the environment and host tissues are acidified, the toxic factors of the acidic pathogenic fungi are activated, the generation of resistance signal molecules such as hydrogen peroxide and the like in peel tissues is inhibited, the plant defense reaction is hijacked, and the pathogenicity of the acidic pathogenic fungi is further enhanced.
Among the many studied possible alternatives to chemical fungicides, biological control techniques that utilize antagonistic microorganisms, particularly antagonistic yeasts, to inhibit postharvest fruit disease are one of the most interesting novel postharvest disease control methods at home and abroad. The yeast has better tolerance to environmental conditions such as low temperature, high temperature, dryness, low oxygen level, pH value fluctuation, ultraviolet radiation and the like. However, low concentrations of organic acids have been shown to still stress yeast growth and the like, inhibiting their growth, resulting in prolonged lag phases.
At present, researches on how to enhance the stable biological control effect of antagonistic yeast on acid fruit wounds are few, and the development of technologies for directionally improving the activity of fruit biocontrol yeast is severely limited. At present, the method for improving the acid resistance and the biological activity of the yeast is mainly focused on the modification of genetically engineered bacteria, and researches for effectively enhancing the biological control efficiency of antagonistic yeast by combining acidic pathogenic fungal virulence factors (oxalic acid, citric acid, glucuronic acid and the like) through a fermentation process optimization method are not reported.
Through a search of existing published patent documents, the following several related published patent documents were found:
1. a method for inhibiting fruit postharvest diseases by inducing resistance and a preparation (publication No. CN106085890B) used by the method disclose a preparation for inhibiting fruit postharvest diseases by inducing resistance, the preparation comprises marine red wintergreen yeast cell walls and water, and each 1L of the preparation contains 0.1-10 g of the marine red wintergreen yeast cell walls. The invention also provides a method for inhibiting postharvest diseases of fruits by inducing resistance by using the preparation, wherein the method comprises the following steps of pretreating the fruits in any mode before boxing and storing the fruits: firstly, soaking fruits in the preparation, draining, and then putting the soaked fruits into a container to keep the fruits in a sealed state for 23-25 hours; secondly, in the pretreatment mode, before picking the fruits, spraying the preparation on the surfaces of the fruits, and then putting the fruits into a container to keep the fruits in a sealed state for 23-25 hours; and taking the pretreated fruits out of the container and then boxing the fruits. The invention can effectively control the postharvest diseases of the fruits on the premise of not using chemical bactericides.
2. A method for regulating and controlling acid stress resistance of Torulopsis glabrata by using a transcription factor Crz1p (publication number: CN105400770B) discloses a method for regulating and controlling acid stress resistance of Torulopsis glabrata by using a transcription factor Crz1p, and belongs to the field of bioengineering. The invention correspondingly reduces or improves the acid stress resistance of the strain by deleting or over-expressing the Cgcrz1 gene of the Torulopsis glabrata. The invention also compares the cell membrane fatty acid and sterol components and the proportion and permeability of the deletion mutant strain Cgcrz1 delta under the acid stress, and the result shows that Crz1p is an essential transcription factor of the Torulopsis glabrata for resisting the acid stress, and the overexpression Crz1p can improve the acid stress resistance of the Torulopsis glabrata.
3. A co-culture method (publication number: CN109370933A) for improving acid resistance of yeast discloses a co-culture method for improving acid resistance of yeast, belonging to the technical field of bioengineering. The invention is characterized in that cultured lactic acid bacteria are cultured for 4 hours in a culture medium, then yeast is inoculated for co-culture for 4 hours, the obtained thalli are suspended in normal saline (pH is 1.9 and 30 ℃) containing lactic acid and stressed for 90 minutes, the thalli after acid stress are centrifugally washed and are dibbled on a yeast extract peptone glucose agar culture medium (containing 0.01 percent of chloramphenicol), and are statically cultured for 48 hours at 30 ℃, and the biomass of the yeast is respectively improved by 7.16 times compared with the number of cells which are not co-cultured. The method is simple and convenient to operate, and can be used for improving the stress resistance of the saccharomycetes.
Disclosure of Invention
The invention aims to overcome the defects of the prior art, provides a method for improving the control efficiency of the acid pathogenic fungi antagonistic yeast organisms after fruit and vegetable picking based on the microecology principle, including competition, parasitism and other comprehensive effects, controlling the fruit and vegetable picking diseases, is safe and efficient to use, green and pollution-free, accords with the natural and environment-friendly concept, and is easy for large-scale equipment due to the high growth and propagation speed of microorganisms.
The technical problem to be solved by the invention is realized by adopting the following technical scheme:
a method for improving biological control efficiency of acid pathogenic fungi antagonistic yeast after fruit and vegetable picking comprises the following steps:
activating bio-control yeast;
performing liquid culture on the activated biocontrol yeast;
the biocontrol yeast is marine yeast Rhodosporidium paludigenum Fell & Tallman, and the strain preservation number is IMI 394084;
adding 1-500mmol L into culture medium-1An organic acid.
Further, the organic acids include: oxalic acid, fumaric acid, citric acid, glucuronic acid, malic acid or ascorbic acid.
Further, the addition concentrations of the respective organic acids were as follows: oxalic acid 1-10mmol L-11-100mmol L of fumaric acid-11-10mmol L of citric acid-11-500mmol L of glucuronic acid-11-100mmol L of malic acid-1Or ascorbic acid 1-100mmol L-1
The activation medium in the step comprises:
the weight of the potato is 200 g,
0 to 20 grams of glucose or fructose,
1-500mmol L of glucuronic acid-1，
20 g of agarose,
1 liter of tap water, and sterilizing to obtain the activated culture medium.
And, the liquid medium in step does:
the weight of the potato is 200 g,
0 to 20 grams of glucose or fructose,
1-500mmol L of glucuronic acid-1，
Adding tap water to a constant volume of 1L, and sterilizing to obtain liquid culture.
Moreover, preferred activation media and liquid media are: the addition amount of glucose or fructose is 1-20 g, and the addition concentration of glucuronic acid is 10-200mmol L-1。
Moreover, the optimum culture time in the liquid medium was 48 hours.
An application for improving biological control efficiency of acid pathogenic fungi antagonistic yeast after fruit and vegetable picking is characterized in that: the biocontrol yeast Rhodosporidium paludigenum Fell and Tallman IMI 394084 is applied to improve the capability of antagonistic yeast in preventing and controlling acid pathogenic fungi of picked fruits and vegetables.
Moreover, said acid pathogenic fungi include: penicillium digitatum, Penicillium italicum, Penicillium expansum, Botrytis cinerea, Sclerotinia sclerotiorum.
Moreover, the method is applied to the biological control of the acid pathogenic fungi antagonistic yeast after fruit and vegetable harvest in any one of the following modes:
mode A: to a concentration of 107Spraying cell/mL antagonistic yeast on the surface of the fruit before picking to wet the surface of the fruit;
mode B: soaking picked fruit in 10% solution7Soaking in antagonistic yeast suspension of cells/mL for 1-30 minutes, and standing and airing at 25 ℃ and at a relative humidity of 90%.
The invention has the advantages and positive effects that:
(1) the fruit disease control method based on antagonistic yeast is one of the most concerned novel postharvest disease control methods at home and abroad at present, controls fruit and vegetable diseases based on the micro-ecological principle, including competition, parasitism and other comprehensive effects, is safe, green and pollution-free, accords with the natural and environment-friendly concepts, and is easy for large-scale equipment due to the high growth and propagation speed of microorganisms.
(2) The D-glucuronic acid is a chemical preparation, is a hexose widely existing in a body, is easy to dissolve in water, can be widely applied to the fermentation culture process of different types of antagonistic yeast, can reduce the damage of organic acid in the environment to the antagonistic yeast, enhances the capability of the antagonistic yeast in biologically preventing and treating postharvest diseases, and has the advantages of low cost, easy production and transportation and convenient use.
(3) The method for improving the effectiveness of the antagonistic yeast in preventing and treating the postharvest acidic pathogenic fungi can effectively improve the survival rate of the antagonistic yeast under different organic acid stresses and improve the capability of the antagonistic yeast in preventing and treating postharvest diseases.
Drawings
FIG. 1 shows the effect of different organic acids and sugar sources on the growth of marine yeast in the experiment of the present invention, wherein (a) is organic acid and (b) is different sugar source;
FIG. 2 shows the effect of different optimal carbon source concentrations and application concentrations on the incidence of marine yeast in preventing and treating citrus fruit green mold in the experiment of the present invention, wherein (a) shows the incidence of fruit wounds after treatment with different optimal carbon source concentrations, and (b) shows the incidence of fruit wounds after treatment with yeast with different concentrations;
FIG. 3 shows the effect of marine yeast cultured with different nitrogen sources on the incidence rate of green mold of citrus fruits in the experiment of the present invention, wherein (a) is the incidence rate of wound treatment and (b) is the lesion diameter of wound treatment;
FIG. 4 shows the effect of marine yeast cultured with different nitrogen sources on the incidence rate of gray mold of tomato fruits in the experiment of the present invention, wherein (a) is the incidence rate of wound treatment and (b) is the lesion diameter of wound treatment;
FIG. 5 is a graph showing the number of viable yeast cells in the plate obtained after glucuronic acid-induced culture in the experiment of the present invention;
FIG. 6 shows the result of staining yeast PI in the solution obtained after glucuronic acid induced culture in the present invention.
Detailed Description
The present invention is further illustrated by the following specific examples, which are intended to be illustrative, not limiting and are not intended to limit the scope of the invention.
Example 1
A method for improving biological control efficiency of acid pathogenic fungi antagonistic yeast after fruit and vegetable picking comprises the following steps:
activating bio-control yeast;
performing liquid culture on the activated biocontrol yeast;
the biocontrol yeast is marine yeast Rhodosporidium paludigenum Fell & Tallman, and the strain preservation number is IMI 394084.
The activation medium in the step comprises:
the weight of the potato is 200 g,
0g of glucose or fructose,
fumaric acid 1-100mmol L-1，
20 g of agarose,
1 liter of tap water, and sterilizing to obtain the activated culture medium.
And, the liquid medium in step does:
the weight of the potato is 200 g,
0g of glucose or fructose,
fumaric acid 1-100mmol L-1，
Adding tap water to a constant volume of 1L, and sterilizing to obtain liquid culture.
The liquid culture time was 24 hours.
The following method is applied to biological control of the acid pathogenic fungi antagonistic yeast after fruit and vegetable harvest: to a concentration of 107cells/mL antagonistic yeast was sprayed onto the fruit surface before picking to wet the fruit surface.
Example 2
A method for improving biological control efficiency of acid pathogenic fungi antagonistic yeast after fruit and vegetable picking comprises the following steps:
activating bio-control yeast;
performing liquid culture on the activated biocontrol yeast;
the biocontrol yeast is marine yeast Rhodosporidium paludigenum Fell & Tallman, and the strain preservation number is IMI 394084.
The activation medium in the step comprises:
the weight of the potato is 200 g,
20 g of glucose or fructose,
malic acid 1-100mmol L-1，
20 g of agarose,
1 liter of tap water, and sterilizing to obtain the activated culture medium.
And, the liquid medium in step does:
the weight of the potato is 200 g,
20 g of glucose or fructose,
malic acid 1-100mmol L-1，
Adding tap water to a constant volume of 1L, and sterilizing to obtain liquid culture.
The liquid culture time was 24 hours.
The following formula is applied to biological control of the acid pathogenic fungi antagonistic yeast after fruit and vegetable harvest: to a concentration of 107cells/mL antagonistic yeast was sprayed onto the fruit surface before picking to wet the fruit surface.
Example 3
A method for improving biological control efficiency of acid pathogenic fungi antagonistic yeast after fruit and vegetable picking comprises the following steps:
activating bio-control yeast;
performing liquid culture on the activated biocontrol yeast;
the biocontrol yeast is marine yeast Rhodosporidium paludigenum Fell & Tallman, and the strain preservation number is IMI 394084.
The activation medium in the step comprises:
the weight of the potato is 200 g,
15 g of glucose or fructose,
glucuronic acid 200mmol L-1，
20 g of agarose,
1 liter of tap water, and sterilizing to obtain the activated culture medium.
And, the liquid medium in step does:
the weight of the potato is 200 g,
15 g of glucose or fructose,
glucuronic acid 200mmol L-1，
Adding tap water to a constant volume of 1L, and sterilizing to obtain liquid culture.
The liquid medium was cultured for 48 hours.
The following method is applied to biological control of the acid pathogenic fungi antagonistic yeast after fruit and vegetable harvest: soaking picked fruit in 10% solution7Soaking in antagonistic yeast suspension of cells/mL for 15 min, and standing and air drying at 25 deg.C and relative humidity of 90%.
Experiment 1 influence of different organic acids and carbon sources on growth of marine yeast
1. Experimental materials:
antagonistic yeast: rhodosporidium toruloides PallidigenumFell & Tallman (IMI 394084)
Organic acid: d-glucuronic acid, malic acid, citric acid, oxalic acid and ascorbic acid
2. Test method
L-malic acid, oxalic acid, fumaric acid, citric acid, glucuronic acid, ascorbic acid and glucose, yeast suspension and carbon-starved yeast glucose medium (NYCB: 8g nutrient solution and 5g yeast extract in 1L tap water) were added to a 100-well honeycomb plate under aerobic conditions to give a final concentration of acid and glucose of 500, 200, 100 or 10mmol L-1. Growth was monitored by bioscreen C (Oy growth cultures Ab Ltd). The optical density (OD600nm) was measured automatically every 30 minutes using a 600nm filter, and the detection time was more than 72 hours from bioscreen C.
3. Test results
At a value of 10mmolL-1At all concentrations tested except, the addition of different carbon sources (organic acids) and their concentrations significantly affected yeast growth. The yeast grows best in a medium containing glucose, and secondly, glucuronic acid and malic acid. Fumaric acid inhibits yeast growth most strongly or is used least by yeast. Wherein the glucuronic acid is 100mmmolL-1The growth effect was the best, similar to the number of yeasts in the group with glucose as sole carbon source, and the optimal cultivation time was 48h (FIG. 1).
1. Experimental materials:
the fruit is citrus and the variety is ponkan orange. Antagonistic yeast: rhodosporidium toruloides R.paludigenumFell & Tallman (IMI 394084). Culture medium: potato dextrose deficient medium (200 g cooked potatoes and 20 g agar to volume in 1l tap water, sterilized at 121 ℃ for 20 min). Pathogenic bacteria: penicillium digitatum (Penicillium digitatum), activated for 7 days for use.
2. And (3) treatment:
(1) the fruits used in the test are slightly greenish in appearance color, free of mechanical wounds and uniform in size, and are soaked in a 0.1% sodium hypochlorite solution for disinfection for 1-2min, washed with tap water and dried at room temperature for later use.
(2) 2 wounds of uniform size (5mm) and of the same depth (2mm) as possible were made on the top of each fruit with a sterile punch. Normal antagonistic yeast and acid-adapted yeast cell suspensions were added at different concentrations in equal amounts (50. mu.l) to each wound, and equal amounts of sterile water were added as controls. Then the original wound is accessed to 1X 104cells/ml P.aeruginosa, 30 ul. Sealing with PE plastic film for moisture keeping, and storing in constant temperature and humidity storage chamber (25 deg.C, relative humidity higher than 95%). Disease incidence and lesion diameter were recorded at regular intervals every day, and the results were expressed as incidence (%) and mean lesion diameter (mm) and compared for direct bacteriostatic efficacy of different yeasts. 12 fruits were selected as one set/replicate, 3 replicates.
The effect of glucuronic acid induced culture on marine yeast control of citrus green mold disease is shown in fig. 2 (detection time is day 2).
3. Results of the experiment
In potato glucose deficient medium, treated with glucuronic acid (500mmol L)-1) After the adaptive culture, the antagonistic yeast has the best biological control effect on the morbidity of the green mold, and the biological control capability of the antagonistic yeast is respectively improved by 43.7 percent and 31.3 percent. The optimal concentration of yeast is 107cells/mL (FIG. 2).
Experiment 3 influence of different optimal carbon sources on marine yeast control of citrus green mold disease
1. Experimental materials:
the fruit is citrus and the variety is ponkan orange. Antagonistic yeast: rhodosporidium toruloides R.paludigenum Fell & Tallman (IMI 394084). Culture medium: nutrient yeast glucose liquid culture medium (8 g of beef extract and 5g of yeast powder, constant volume is set to 1L of tap water, and sterilization is carried out for 20min at 121 ℃). Carbon source of culture medium: glucose, glucuronic acid. Pathogenic bacteria: penicillium digitatum (Penicillium digitatum) was activated at 25 ℃ for 7 days for use.
2. And (3) treatment:
(1) the fruits used in the test are slightly greenish in appearance color, free of mechanical wounds and uniform in size, and are soaked in a 0.1% sodium hypochlorite solution for disinfection for 1-2min, washed with tap water and dried at room temperature for later use.
(2) 2 wounds of uniform size (5mm) and of the same depth (2mm) as possible were made on the top of each fruit with a sterile punch. Equal amounts (50. mu.l) of 1X 10 were added to each wound7cells/ml of normal antagonistic yeast and acid-adapted yeast cell suspensions, and an equal amount of sterile water was added as a control. Then the original wound is accessed to 1X 104cells/ml P.digitatum30 ul of P.aeruginosa. Sealing with PE plastic film for moisture keeping, and storing in constant temperature and humidity storage chamber (25 deg.C, relative humidity higher than 95%). Disease incidence and lesion diameter were recorded at regular intervals every day, and the results were expressed as incidence (%) and mean lesion diameter (mm) and compared for direct bacteriostatic efficacy of different yeasts. 12 fruits were selected as one set/replicate, 3 replicates.
The effect of glucuronic acid induced culture on marine yeast control of citrus green mold disease is shown in fig. 3 (detection time is day 2).
3. Results of the experiment
Glucuronic acid can significantly reduce the severity of citrus fruit green mold in a nutrient yeast glucose liquid medium. Glucuronic acid (500mmolL L-1) The treatment group was 22.1% more resistant to the yeast organism to control the citrus green mold disease than the NYDB medium treatment group (incidence), and the lesion diameter was also significantly larger than in the NYDB medium treatment group (figure 3).
Experiment 4 influence of different optimal carbon sources on prevention and treatment of tomato fruit botrytis by marine yeast
1. Experimental materials:
the fruit is tomato. Antagonistic yeast: rhodosporidium toruloides R.paludigenumFell & Tallman (IMI 394084). Culture medium: potato dextrose deficient medium (200 g cooked potatoes and 20 g agar to volume in 1l tap water, sterilized at 121 ℃ for 20 min). Carbon source of culture medium: glucose, glucuronic acid. Pathogenic bacteria: botrytis cinerea (Botrytis cinerea), activated at 25 ℃ for 7 days for use.
2. And (3) treatment:
(1) the fruits used in the test are fruits in red ripe period, have no mechanical wound and are uniform in size, and the fruits are soaked and sterilized by 0.1% sodium hypochlorite solution for 1-2min, washed by tap water and dried at room temperature for later use.
(2) 2 wounds of uniform size (5mm) and of the same depth (2mm) as possible were made on the top of each fruit with a sterile punch. Equal amounts (50. mu.l) of 1X 10 were added to each wound7cells/ml of normal antagonistic yeast and acid-adapted yeast cell suspensions, and an equal amount of sterile water was added as a control. Then the original wound is accessed to 1X 104cells/ml Botrytis cinerea B.cinerea30 ul. Sealing with PE plastic film for moisture keeping, and storing in constant temperature and humidity storage chamber (25 deg.C, relative humidity higher than 95%). Disease incidence and lesion diameter were recorded at regular intervals every day, and the results were expressed as incidence (%) and mean lesion diameter (mm) and compared for direct bacteriostatic efficacy of different yeasts. 12 fruits were selected as one set/replicate, 3 replicates.
The effect of glucuronic acid induced culture on marine yeast control of citrus green mold disease is shown in fig. 4 (detection time is day 2).
3. Results of the experiment
As shown in figure 4, after the compound culture of glucuronic acid and glucose in the potato glucose-deficient culture medium, the antagonistic yeast has better biological control effect on the incidence rate of gray mold, and the biological control capability of the antagonistic yeast is respectively improved by 29.2 percent.
1. Experimental materials:
antagonistic yeast: rhodosporidium toruloides R. paludigenum Fell & Tallman (IMI 394084)
Organic acid: d-glucuronic acid, malic acid, citric acid, oxalic acid and ascorbic acid
2. Test method
(1) Respectively taking 1mL of acid-adaptive antagonistic yeast and normal antagonistic yeast as seed liquid, inoculatingInoculating in 500mM PDB medium containing different organic acids in an amount of 1 × 106cells/mL, shaking culture at 28 ℃ and 200rpm for 48h, sampling and diluting with equal gradient 104Then, the cells were plated on PDA plates and cultured at 28 ℃ for 2 days, followed by counting.
(2) Acid-adapted antagonistic yeast and normal antagonistic yeast were inoculated into PDB medium supplemented with 500mM organic acid and not treated with organic acid, respectively, and cultured at 200rpm at 28 ℃ for 48h to reach logarithmic phase. Yeast cells were collected in the middle of log phase by centrifugation at 12000rpm for 5 min. Yeast cells were collected and cultured without addition of acid as a control. After washing twice with PBS, the yeast cells were resuspended in a solution containing 10. mu.g/mL Propidium Iodide (PI) and allowed to stand in the dark for 20 min. The cells were then washed twice with PBS and then resuspended in an equal volume of PBS and observed under a fluorescent microscope. Cells stained with PI were excited at 488 nm. Yeast cells positive for PI binding (red) are dead cells with damaged plasma membrane.
3. Test results
As shown in fig. 5 and fig. 6, after 500mM glucuronic acid adaptive induction treatment, the survival rate of rhodosporidium marinum r.paludigenum in solid plates containing different organic acids (glucuronic acid, malic acid, citric acid, ascorbic acid and oxalic acid) is obviously improved by 21.2%, 33.6%, 19.4%, 33.3% and 39.4% respectively compared with control treatment; the decline rate in the PDB liquid culture medium containing high concentration (500mM) of glucuronic acid, malic acid, citric acid, ascorbic acid and oxalic acid is obviously reduced by 4.57%, 5.41%, 4.53%, 7.66% and 13.01%, and the damage to the cytoplasmic membrane of the yeast is effectively reduced.
Although the embodiments of the present invention have been disclosed for illustrative purposes, those skilled in the art will appreciate that: various substitutions, changes and modifications are possible without departing from the spirit and scope of the invention and the appended claims, and therefore the scope of the invention is not limited to the embodiments disclosed.
Claims (10)
1. A method for improving the biological control efficiency of acid pathogenic fungi antagonistic yeast after fruit and vegetable harvest is characterized by comprising the following steps: the method comprises the following steps:
activating bio-control yeast;
performing liquid culture on the activated biocontrol yeast;
the biocontrol yeast is marine yeast Rhodosporidium paludigenum Fell & Tallman, and the strain preservation number is IMI 394084;
adding 1-500mmol L into culture medium-1An organic acid.
2. The method for improving the biological control efficacy of the picked fruit and vegetable on the antagonistic yeast of the acid pathogenic fungi, which is disclosed by claim 1, is characterized in that: the organic acid comprises: oxalic acid, fumaric acid, citric acid, glucuronic acid, malic acid or ascorbic acid.
3. The method for improving the biological control efficiency of the fruit and vegetable post-harvest acid pathogenic fungi antagonistic yeast according to claim 2, which is characterized in that: the addition concentrations of the respective organic acids were as follows: oxalic acid 1-10mmol L-11-100mmol L of fumaric acid-11-10mmol L of citric acid-11-500mmol L of glucuronic acid-11-100mmol L of malic acid-1Or ascorbic acid 1-100mmol L-1。
4. The method for improving the biological control efficacy of the picked fruit and vegetable on the antagonistic yeast of the acid pathogenic fungi, which is disclosed by claim 1, is characterized in that: the activation culture medium in the steps comprises:
the weight of the potato is 200 g,
0 to 20 grams of glucose or fructose,
1-500mmol L of glucuronic acid-1，
20 g of agarose,
1 liter of tap water, and sterilizing to obtain the activated culture medium.
5. The method for improving the biological control efficacy of the picked fruit and vegetable on the antagonistic yeast of the acid pathogenic fungi, which is disclosed by claim 1, is characterized in that: the liquid culture medium comprises the following steps:
the weight of the potato is 200 g,
0 to 20 grams of glucose or fructose,
1-500mmol L of glucuronic acid-1，
Adding tap water to a constant volume of 1L, and sterilizing to obtain liquid culture.
6. The method for improving the biological control efficacy of the picked fruit and vegetable on the antagonistic yeast of the acid pathogenic fungi, which is disclosed by claim 1, is characterized in that: preferred activation media and liquid media: the addition amount of glucose or fructose is 1-20 g, and the addition concentration of glucuronic acid is 10-200mmol L-1。
7. The method for improving the biological control efficacy of the picked fruit and vegetable on the antagonistic yeast of the acid pathogenic fungi, which is disclosed by claim 1, is characterized in that: the optimal culture time in the liquid culture medium is 48 h.
8. An application for improving biological control efficiency of acid pathogenic fungi antagonistic yeast after fruit and vegetable picking is characterized in that: the biocontrol yeast Rhodosporidium paludigenum Fell and Tallman IMI 394084 is applied to improve the capability of antagonistic yeast in preventing and controlling acid pathogenic fungi of picked fruits and vegetables.
9. The application of the biological control efficacy of the acid pathogenic fungi antagonistic yeast for improving the postharvest disease of fruits and vegetables according to claim 6, is characterized in that: the acid pathogenic fungi comprise: penicillium digitatum, Penicillium italicum, Penicillium expansum, Botrytis cinerea, Sclerotinia sclerotiorum.
10. The application of the biological control efficacy of the acid pathogenic fungi antagonistic yeast for improving the postharvest disease of fruits and vegetables according to claim 6, is characterized in that: the method is applied to biological control of the acid pathogenic fungi antagonistic yeast after fruit and vegetable harvest in any one of the following modes:
mode A: to a concentration of 107Antagonistic yeast cells/mL were sprayed onto fruit surfaces prior to pickingWetting the fruit surface;
mode B: soaking picked fruit in 10% solution7Soaking in antagonistic yeast suspension of cells/mL for 1-30 minutes, and standing and airing at 25 ℃ and at a relative humidity of 90%.
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