KR20210097024A - Active Treatment Agent Composition for Simultaneous Control of Olpidiopsis blight and Red-Rot Disease - Google Patents

Abstract

The present invention relates to an active treatment agent which can effectively control Olpidiopsis porphyrae var. koreanae that causes the greatest damage to laver farms, thereby preventing disease of laver, promoting the growth of laver, and improving the quality of the harvested laver. More specifically, the present invention relates to an active treatment agent composition for simultaneous control of Olpidiopsis porphyrae var. koreanae and Pythium porphyrae comprising a mixture of lactic acid, propionic acid, and hydrochloric acid.

Description

Active Treatment Agent Composition for Simultaneous Control of Olpidiopsis blight and Red-Rot Disease

The present invention can effectively control both cystic bacillus and red seaweed, which cause the greatest damage to laver farms, at the same time, thereby promoting the growth of laver and improving the quality of the harvested laver while preventing the disease of laver. It relates to an active treatment agent for the simultaneous control of

Seaweed has been used for thousands of years worldwide as food and dietary supplements, animal feed, chemicals and biofuels. In particular, in Asia, seaweed is mainly used as food, and the aquaculture industry of laver, seaweed, and kelp has developed. According to statistics in 2017, seaweed accounts for 29.6% of the total production and 71% of the total production value among laver, seaweed, and kelp, which are the main cultivated species. is showing

The types of laver cultivated in Korea include Pyropia yezoensis , Pyropia dentata , and Pyropia seriata . there is an increasing trend. The production of laver is greatly affected by natural factors such as seawater, climatic and marine conditions, as well as seaweed causing fading of laver fronds and loss of fronds from laver. In particular, in the case of seaweed, the quality of laver is reduced or production is reduced, causing great damage to fishermen. The representative fish diseases that occur in Korea are Olpidiopsis blight, Green-spot disease, and Red-rot disease. Occurs mainly in March.

Among the seaweed, red seaweed and cysticercosis caused by Pythium porphyrae and Olpidiopsis spp. belonging to the Oomycete River cause the greatest damage to seaweed farms. This is closely related to the reproductive and infection mechanism characteristics of the two pathogens, which are capable of continuously forming and releasing scavengers that directly infect seaweed. For this reason, if two types of seaweed appear at the same time in laver farms, it will cause great damage to the laver farms, and red seaweed and cystic bacillus almost always occur together in laver farms.

In Korean laver farms, inorganic acid, which is 35% hydrochloric acid, has been mainly used as a method to control seaweed. Since the use of inorganic acids was restricted after enacting the standard for use of acid treatment agents as a notification in 1994, a mixed solution mixed with organic acids and inorganic acids has been developed and used as an active treatment agent to replace inorganic acids since 1998. Currently, all active treatment agents that can be used in laver farms contain 8.0~9.5% hydrochloric acid and 10% or more organic acid. Realistically, organic acids are very expensive, so it is rarely used more than 10%, and if hydrochloric acid is not added, it is unrealistic because fishermen do not use it at all because the deposits are not removed.

The organic acid, which is the main component of the active treatment agent, is an acidic organic compound that inhibits the growth of mold and microorganisms and is widely used to prevent food spoilage. The antibacterial effect and action range of organic acids are different depending on the type and biochemical characteristics of the organic acid. As for the antibacterial effect of organic acids related to cystic larvae, the efficacy of lactic acid, malic acid, citric acid, gluconic acid, and tartaric acid has been reported for the red cyst bacillus that causes cystic larvae. In addition, it is reported that there is no control effect because it does not damage the cystic bacteria in the laver cells, which is the host, even when hydrochloric acid is treated, which was the existing method for controlling cystic bacteria.

In addition, since cystic bacillus and cystic leprosy almost always occur in seaweed farms, effective control of cystic leprosy cannot be achieved unless both diseases are caught at the same time. In the case of controlling only one disease, the possibility of spreading other diseases easily increases, but there is no treatment that can catch both diseases. As fishermen respond only by treating high concentrations of hydrochloric acid, many illegal activities and various social problems have occurred.

In addition, the active treatment agent developed to replace hydrochloric acid has a higher treatment cost and longer treatment time than hydrochloric acid.

Registered Patent No. 10-1752610 Patent Publication No. 10-2000-0060843 Japanese Registered Patent No. 3595004 Japanese Patent No. 3631466

The present invention provides an active treatment agent composition for controlling laver oomycete disease, which can effectively control cystic bacillus and red seaweed at the same time, thereby promoting the growth of laver and improving the quality of harvested laver while preventing the disease of laver. aim to

The present invention for achieving the above object relates to an active treatment agent composition for controlling Cystic disease, characterized in that it comprises a mixture of propionic acid and hydrochloric acid.

As can be seen in the following examples, hydrochloric acid, which has been conventionally used to control oomycete cyst, does not show a great effect on the control of cystic bacillus, and the antibacterial effect of organic acid on cystic bacillus has not been reported either. However, the mixture of propionic acid and hydrochloric acid of the present invention can very effectively control Cysbacteria.

The composition of the present invention also exhibits a synergistic effect on the control of cysts by mixing propionic acid and hydrochloric acid, and thus has excellent efficacy in controlling cystic bacteria as well as cystic larvae, and can be used as an active treatment composition for simultaneous control of cystic bacteria and cystic larvae. there is. According to the composition of the present invention, since it is possible to simultaneously control Cystic bacillus and red cyst, it is possible to substantially control oomycete cyst.

In order to further amplify the control effect against erythrasma using the composition of the present invention, it is more preferable to further contain lactic acid.

According to the permissible component content of the active treatment agent in Article 4 of the active treatment agent usage standards for seaweed farms, the total content of chlorine ions, sulfate ions, and nitrate ions in the active treatment agent containing organic acids as the main component is 8% or more and 9.5% or less. Accordingly, the weight ratio of hydrochloric acid in the composition of the present invention is preferably 8% to 9.5%, but the above range is not excluded. As can be seen in the Examples, even when containing 5 wt% of HCl, which is a concentration lower than the above range, an excellent control effect was exhibited, and the control efficacy is not reduced just because it is out of the above range. Hydrochloric acid in the composition of the present invention is one of the main components that produce a synergistic effect of the composition, and hydrochloric acid itself does not show the control effect of oomycete cyanobacteria at the corresponding concentration, but when mixed with propionic acid, the effect of remarkably increasing the control effect In addition, when mixed with lactic acid, it showed a synergistic effect on the control effect on red seaweed.

The concentration of propionic acid in the composition of the present invention is preferably 0.5 to 10% by weight. In addition, in the composition containing additional lactic acid, the weight ratio of lactic acid and propionic acid is preferably 1:9 to 9:1, and more preferably 7:3 to 5:5, which shows similar control effects against Cystic bacillus and cysticercosis. desirable. However, the ratio can be appropriately adjusted according to the occurrence pattern of Cystic bacillus and Rhododendron blight. That is, as the ratio of lactic acid increases, it is advantageous for the control of cystic bacilli, and as the ratio of propionic acid increases, it is advantageous for the control of cystic. The weight ratio of propionic acid may be appropriately adjusted.

The active treatment composition of the present invention can control oomycete cysts by diluting seawater and treating seaweed for 5 seconds to 1 minute. The dilution ratio may be 10 to 100 times (v/v), but is not limited thereto, and may be appropriately adjusted according to the type or frequency of occurrence of oomycete cyst.

As described above, according to the active treatment composition for controlling seaweed oomycete of the present invention, it exhibits a control effect not only on red seaweed but also on cystic bacillus, which has not been an effective active treatment agent in the prior art, and effectively treats both cystic bacilli and cystic bacillus at low concentrations. It can prevent damage to laver farms and improve the quality of harvested laver.

1 is a photomicrograph and cultured photo of radioactive laver.
Figure 2 is a microscopic photograph of the seaweed thallus infected with rhododendron bacillus and cysticercosis.
Figure 3 is a graph showing the anti-infection efficacy of red seaweed according to the treatment concentration of propionic acid and lactic acid.
Figure 4 is a graph showing the anti-infection efficacy of propionic acid or a mixed solution of lactic acid and hydrochloric acid against red seaweed.
5 is a graph showing the anti-infection efficacy for red seaweed according to the dilution ratio of propionic acid or a mixed solution of lactic acid and hydrochloric acid.
6 is a graph showing the efficacy of inhibiting infection against red seaweed according to the dilution ratio of a 9.5% aqueous hydrochloric acid solution.
7 is a graph showing the anti-infection efficacy against red seaweed according to the dilution ratio of the mixed solution of lactic acid, propionic acid and hydrochloric acid.
8 is a graph showing the efficacy of inhibiting infection against Cystic bacillus according to the dilution ratio of a 9.5% aqueous hydrochloric acid solution.
9 is a graph showing the anti-infection efficacy against Cystic bacillus according to the dilution ratio of a mixed solution of propionic acid and hydrochloric acid.
Figure 10 is a graph showing the efficacy of inhibiting infection against Cystic bacillus according to the dilution ratio of the mixed solution of lactic acid and hydrochloric acid.
11 is a graph showing the anti-infection efficacy against Cystic bacillus according to the dilution ratio of the mixed solution of lactic acid, propionic acid and hydrochloric acid.
12 is a graph showing the control effect of red cyst and cysticercosis according to the mixing ratio of propionic acid and lactic acid.

Hereinafter, the present invention will be described in more detail with reference to the accompanying examples. However, these embodiments are merely examples for easily explaining the content and scope of the technical idea of the present invention, and thereby the technical scope of the present invention is not limited or changed. It will be natural for those skilled in the art that various modifications and changes can be made within the scope of the technical spirit of the present invention based on these examples.

[Example]

Cultivation of seaweed and strains

1) Radial laver ( Pyropia yezonesis )

Radial laver was purchased from the Mokpo Seaweed Bio Research Center and cultured alone in PES (Provasoli Enriched Seawater) medium. Seawater filtered with filter paper (Whatman No. 2) was sterilized for 20 minutes at 121°C and 1.2 atm using an autoclave. Culture conditions were 10°C, 12:12 hr L:D cycle, white light (>50 μmol/m ² ·s ¹ ), and aeration was performed. The culture medium was changed every 7 days.

1A is a microscopic image (scale bar: 20 μm) of radiograss, and B is a photograph of culturing the thallus of radiopatterned laver in PES medium.

2) Red Pythium porphyrae

The rhododendron bacillus was isolated and cultured from the laver fronds infected with rhododendron collected in Hoe-dong, Jindo-gun. FIG. 2A is a microscopic photograph of a seaweed frond infected with rhododendron bacillus. Unlike normal cells in FIG. 1 , in FIG. 2A , traces of hyphae passing through laver cells like threads can be seen, and the perforated cells show red color due to contraction of laver cells than normal laver cells. Infected parts of collected seaweed were cut and subcultured until separated from cornmeal agar medium. The isolated strain was subcultured in cornmeal agar medium at 20° C., 12:12 hr L:D cycle with a cycle of 21 days. When used in the experiment, the mycelium isolated by suspension culture in Arasaki B medium for 7 to 10 days under the same light and temperature as the subculture conditions was used.

3) Cyst ( Olpidiopsis pyropiae )

Cystic bacteria were isolated and cultured by transferring the bacteria to the laver fronds infected with cystic bacilli collected in Hoe-dong, Jindo-gun. B of FIG. 2 is a micrograph of laver fronds infected with Cystic bacteria, showing that the seaweed cells have turned yellow due to Cystic bacteria infection. The culture conditions were 15° C., 12:12 hr L:D cycle, white light (>50 μmol/m ² ·s ¹ ), and new radiopatterns were added every 5 to 7 days to maintain Cyst bacteria.

Induction of cyst infection and measurement of infection rate

1) Red seaweed

The release of spores was induced by exposing the mycelium-type red sea bacillus suspended in Arasaki B medium for 7 to 10 days to sterilized seawater for 24 to 30 hr. The number of released resident cells was measured using a hemocytometer after fixing 3.7% formaldehyde in a fixative diluted 1/500. 2.5×10 ³ cell/ml zoospores were cultured in sterilized seawater together with radiopatterns to infect them. Infection was observed and photographed using an optical microscope (OLYMPUS, BX50) and cellSens program. The infection rate was measured by comparing the infected area to the total area of the photograph taken using Adobe Photoshop, an image program.

In the following examples, the statistical difference in the infection rate and the number of infected cells according to the type of treatment solution and the dilution factor was confirmed through one-way ANOVA. The difference according to the type and dilution ratio of each treatment solution was confirmed through post-hoc test.

2) Cystobacteria

After releasing the spores from the laver fronds infected with Cystic bacillus, the infected water containing the spores was filtered through a 40 μm filter to infect radiopatterns. Observation and imaging of infection was carried out in the same manner as that of the rhododendron disease.

Efficacy test of the active treatment agent against red seaweed

1) Infection analysis according to the concentration of propionic acid and lactic acid treatment

In order to analyze the minimum effective concentrations of propionic acid and lactic acid, which have excellent inhibitory effects on rhododendron infection, samples for each concentration (0.1, 0.25, 0.5, 1 wt%) were prepared and then treated on the leaflets infected with rhododendron for 15 seconds. Immediately after the treatment, the fronds were washed in sterile seawater and cultured for 3 days to confirm that the infection of the control group had progressed to 80% or more, and then the infection rate of the treatment group was compared and measured.

3 shows the results, propionic acid (PPA) was not significantly different from the infection rate of the control group until 0.25% treatment, the infection rate of the 0.5% treatment group (52.7%) was lower than the infection rate of the control group (90.9%), The 1% treatment group showed the lowest infection rate (5.63%). In addition, in FIG. 3 , it can be seen that lactic acid (LA) exhibits an effect at a lower concentration than propionic acid. The infection rate of 84.5% in the 0.1% treatment group was higher than the infection rate of 4.0% in the 0.25% treatment group, 4.2% in the 0.5% treatment group, and 3.9% in the 1% treatment group, but it was significantly lower than the infection rate of 90.9% in the control group.

The pH of propionic acid was 2.99 in 1% solution and 3.83 in 0.1% solution, and the pH of lactic acid was 2.24 in 1% solution and 3.02 in 0.1% solution. Lactic acid was stronger than propionic acid.

2) Infection analysis according to the treatment of organic acid and hydrochloric acid mixture

According to the standards for use of the active treatment agent for seaweed farms, the allowable content of each component in the active treatment agent containing organic acids as the main component is 10% or more for organic acids, and the total content of chlorine ions, sulfate ions and nitrate ions is 8% or more and 9.5% or less, sulfuric acid ions and nitrate ions are each less than 2%.

Accordingly, an active treatment agent having a composition of propionic acid (10 w%) + hydrochloric acid (9 w% or 5 w%), lactic acid (10 w%) + hydrochloric acid (9 w% or 5 w%) was prepared. After the active agent was diluted 1/30 with sterilized seawater, it was treated for 15 seconds on the thallus infected with rhododendron, and then immediately washed with sterile seawater. After culturing for 3 days, the infection rate of the treated group was compared and measured when the infection of the control group progressed more than 80%.

The infection rate of the control group was 83.7%, compared with the infection rate of 61.9% in the 10% propionic acid treatment group, 69.2% in the 5% hydrochloric acid treatment group, and 65.3% in the 9.5% hydrochloric acid treatment group, the 10% propionic acid and 5% hydrochloric acid or 9.5% hydrochloric acid mixture treatment group Infection rates were 3.4% and 3.0%, respectively, and the effect was very remarkable when treated with a mixture of propionic acid and hydrochloric acid.

On the other hand, since the infection rate of the 10% lactic acid treatment group was already very low at 1.9%, even when additional 5% hydrochloric acid or 9.5% hydrochloric acid was mixed, there was no significant difference compared to that treated with lactic acid alone.

3) Analysis of infection according to dilution ratio of organic acid and hydrochloric acid mixture

Infection was analyzed in the same manner as in 2) except that the organic acid and hydrochloric acid mixture was diluted and treated.

The infection rate of the 10% propionic acid and 9.5% hydrochloric acid mixture increased as the dilution ratio increased, and the infection rate of the 80-fold dilution solution was similar to that of the control group. For reference, when only 9.5% hydrochloric acid was diluted and treated, there was no significant difference in infection rate with the control group (see FIG. 6 ). The infection rate of the 50-fold dilution treatment group of propionic acid was 21.3%, which was significantly reduced to 25% of the infection rate of 86.1% of the control group. The concentration of propionic acid in the solution was 0.2%, suggesting that there is a synergistic effect of the mixed solution because the propionic acid solution (FIG. 3) or the hydrochloric acid solution (FIG. 6) alone at the corresponding concentration did not show the effect of controlling erythrasma.

The 10% lactic acid and 9.5% hydrochloric acid mixture showed a tendency to increase the infection rate as the dilution ratio increased, but there was no significant difference. The infection rates of the 50-fold, 60-fold, and 80-fold dilutions were 3.3, 4.2, and 5.2%, respectively, which was about 3.8 to 6.0% compared to the infection rate of 86.1% in the control group. Since lactic acid itself has a very large infection inhibitory effect, the synergistic effect by mixing with hydrochloric acid was weak compared to propionic acid, but again, when converting the concentration with reference to FIGS. 3 and 6, the synergistic effect of mixing is shown could confirm that

4) Analysis of infection by treatment with lactic acid, propionic acid and hydrochloric acid mixture

Lactic acid, propionic acid, and hydrochloric acid were mixed so as to have the concentrations shown in Table 1 below to prepare an activity inhibitor, and then the degree of inhibition of erythrasma infection was evaluated in the same manner as in 2) except that the diluent was treated. The results are shown in Table 1 and FIG. 7 . 7, A, B, and C represent the experimental groups 1, 2, and 3 of Table 1, respectively.

The pH of the solution according to the dilution ratio of 9.5% hydrochloric acid and the composition in Table 1 was measured with a pH meter (NeoMet, pH/ISE Meter pH-250L) with a glass electrode (Hemilton, Liq-Glass BNC) attached, and the results are shown in the table. 2 is described.

Efficacy test of active treatment agent against Cystic disease

1) Analysis of infection according to dilution ratio of organic acid and hydrochloric acid mixture

In the same manner as for cystic larvae, the mixed solution of organic acid and hydrochloric acid was diluted and treated on laver fronds infected with Cystic bacillus, and the number of infected cells was observed.

First, FIG. 8 is a graph showing the infection rate of Cystobacteria according to the dilution factor of 9.5% hydrochloric acid. The infection rate decreased by about 20% according to the treatment with the 9.5% hydrochloric acid dilution, but it was not significant.

9 is a graph showing the rate of infection with Cystobacteria according to the dilution treatment of a mixture of 10% propionic acid and 9.5% hydrochloric acid. The number of infected cells in the group treated with a mixture of propionic acid and hydrochloric acid diluted 30, 40, and 50 times was lower than the number of infected cells in the control group (98.6 cells). There was no significant difference between the number of infected cells in the 30-fold dilution treatment group (6.6) and the number of infected cells in the 40-fold dilution treatment group (10.5 cells), which was less than the number of infected cells in the 50-fold dilution treatment group (30.2 cells).

The 10% lactic acid and 9.5% hydrochloric acid mixture showed a tendency to increase the infection rate as the dilution ratio increased. The number of infected cells when treated with 30-fold, 40-fold, and 50-fold dilutions was 60.6, 73.8, and 89.8, respectively, which was 61 to 91% of the number of infected cells in the control group, 98.6. The number of infected cells in the 50-fold dilution treatment was not significantly different from the control group. The diluent containing lactic acid showed a strong effect against red seaweed, but the dilution of lactic acid had a lower effect than propionic acid against Cystic bacteria.

2) Analysis of infection by treatment with lactic acid, propionic acid and hydrochloric acid mixture

The degree of inhibition of Cystic bacillus infection was evaluated in the same manner as in 1), except that lactic acid, propionic acid and hydrochloric acid were mixed to the concentrations shown in Table 3 below to prepare an inhibitor, and then the diluent was treated. The results are shown in Table 3 and FIG. 11 .

Confirmation of the control effect of red cyst and cystic bacteria according to the concentration of the composition for controlling oomycete cysticercosis

In order to confirm the optimal composition of the composition for the simultaneous control of rhododendron and Cystic disease, the control effect was compared with 9.5% hydrochloric acid and a mixture of lactic acid and propionic acid by changing the ratio of lactic acid and propionic acid.

12 is a graph showing the treatment results using a 30-fold dilution, LA is lactic acid and CP is propionic acid. From FIG. 12, it is shown that when the weight ratio of propionic acid and lactic acid is 7:5 to 3:5, it is possible to simultaneously control red seaweed and Cystobacteria with excellent efficiency.

Claims (9)

An active agent composition for controlling Cystic disease, characterized in that it comprises a mixture of propionic acid and hydrochloric acid.
Active treatment composition for the simultaneous control of Cystic bacillus and cysticercosis, characterized in that it comprises a mixture of propionic acid and hydrochloric acid.
3. The method according to claim 2,
Active treatment composition for the simultaneous control of Cystic bacillus and cysticercosis, characterized in that it further contains lactic acid.
4. The method according to claim 2 or 3,
The concentration of hydrochloric acid in the composition is 8 to 9.5% by weight of the active treatment agent composition for the simultaneous control of Cystic bacillus and red cyst.
4. The method according to claim 2 or 3,
The concentration of propionic acid in the composition is 0.5 to 10% by weight of the active treatment agent composition for the simultaneous control of Cystic bacillus and erythrasma.
4. The method according to claim 3,
The active treatment composition for the simultaneous control of Cystic bacillus and red cyst, characterized in that the weight ratio of lactic acid and propionic acid in the composition is 1:9 to 9:1.
7. The method of claim 6,
The active treatment composition for the simultaneous control of Cystic bacillus and cystic bacillus, characterized in that the higher the content of propionic acid in the composition, the more severe the cystic bacillus compared to cystic bacillus in the composition.
4. The method according to claim 2 or 3,
The active treatment composition for the simultaneous control of Cystic bacillus and erythrasma, characterized in that the active treatment composition is diluted with seawater and treated for 5 seconds to 1 minute.
9. The method of claim 8,
The dilution ratio is 10 to 100 times (v / v), characterized in that the active treatment agent composition for the simultaneous control of Cystic bacillus and red cyst.

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