Erwinia vibrio bacteriophage and application thereof

Abstract

The invention relates to the technical field of microorganisms, and provides a vibrio owneri bacteriophage and application thereof, the vibrio owneri bacteriophage is vibrio owneri bacteriophage VOP240901, and the preservation number is CCTCC (China Center For Type Culture Collection) NO: M20242007. The Erwinia vibrio bacteriophage VOP240901 is a broad-spectrum bacteriophage, can split vibrio alginolyticus, vibrio parahaemolyticus and Erwinia vibrio, can stably exist in the environment with the temperature of 4-55 DEG C and the pH value of 3-11, is suitable for killing and preventing aquatic vibrio microorganisms, especially Erwinia vibrio, and has the sterilization rate of 90%.

Description

Technical Field

[0001] The present invention relates to the technical field of microorganisms, and in particular to a Vibrio owensii phage and its application. Background Art

[0002] Vibrio owensii is widely distributed in the marine environment. Currently, there are two common Chinese names, Vibrio owensii and Vibrio owensii. Vibrio owensii is considered to be a branch of Vibrio harveyi and was identified and named by Australian microbiologist Leigh Owensii in 2010. It was isolated from diseased shrimp in a lobster farm. Recent reports have shown that Vibrio owensii is a virulent intestinal pathogen that can cause acute hepatopancreatic necrosis in the seedlings of Litopenaeus vannamei, lesions in lobster larvae, and is also associated with albinism in corals. However, there is still relatively little research on Vibrio owensii.

[0003] Phages are viruses that specifically lyse bacteria, have strong host specificity, and are widely distributed in most ecological environments on Earth. The application of phage preparations in the aquaculture field has gradually matured. For example, the phage product pipelines for aquatic pathogenic bacteria such as Vibrio, Aeromonas, and Pseudomonas aeruginosa are increasing. With the withdrawal of antibiotic restriction laws and the increasingly strict national supervision of antibiotics, phages that have been dormant have returned to people's vision. Compared with antibiotics, phages will degrade themselves after killing bacteria, will not produce pollution and drug resistance, and are more environmentally friendly. Existing patents are mostly for phages against Vibrio alginolyticus and Vibrio vulnificus, and there are no phages against Vibrio owensii. Therefore, a phage that specifically lyses Vibrio owensii is needed. Summary of the Invention

[0004] In view of this, the present invention has screened out a phage of Vibrio owensii, which can simultaneously lyse a variety of aquatic pathogenic vibrios such as Vibrio parahaemolyticus, Vibrio alginolyticus, and Vibrio harveyi, and has application value.

[0005] The technical solution of the present invention is realized as follows: In the first aspect, the present invention provides a Vibrio owensii phage, which is Vibrio owensis phage VOP240901, and the deposit number is CCTCC NO: M20242007.

[0006] On the basis of the above technical solution, preferably, the Vibrio owensii phage VOP240901 can survive under the condition of 4 - 55 °C.

[0007] On the basis of the above technical solution, preferably, the Vibrio owensii phage VOP240901 can survive under the condition of pH 3 - 11.

[0008] Based on the above technical solutions, preferably, the titer of the Vibrio anguillarum phage VOP240901 is 10 9- 10 11 PFU / mL.

[0009] Based on the above technical solutions, preferably, the multiplicity of infection (MOI) of the Vibrio anguillarum phage VOP240901 is 0.001 - 10.

[0010] In a second aspect, the present invention also provides an aquatic feed additive, comprising the Vibrio anguillarum phage VOP240901.

[0011] In a third aspect, the present invention also provides a microbial bactericide, comprising the Vibrio anguillarum phage VOP240901.

[0012] In a fourth aspect, the present invention also provides an application of a Vibrio anguillarum phage in killing Vibrio microorganisms in aquatic products, and the Vibrio microorganisms are one or more of Vibrio parahaemolyticus, Vibrio alginolyticus, Vibrio harveyi, and Vibrio anguillarum.

[0013] The Vibrio anguillarum phage and its application of the present invention have the following beneficial effects compared with the prior art:

[0014] (1) The Vibrio anguillarum phage VOP240901 of the present invention is a broad-spectrum phage, which can lyse Vibrio alginolyticus, Vibrio parahaemolyticus, and Vibrio anguillarum, and can stably exist in the environment of 4 - 55°C and pH 3 - 11. It is suitable for killing and preventing Vibrio microorganisms in aquatic products, and has the application value of industrialization and productization.

[0015] (2) The present invention has formulated a variety of culture media and screened out the optimal combination of the inoculation culture medium and the fermentation culture medium. At this time, the phage VOP240901 reaches the highest titer of 5.1×10 9 PFU / mL. When fermenting in a small volume (20 mL), the best MOI of the phage VOP240901 is 0.01, and when fermenting in a large volume (6 L), the best MOI of the phage VOP240901 is 0.04. At the same time, the best fermentation temperature is preferably 30°C and the best pH is 7.5, obtaining a phage solution with a higher titer.

[0016] (3) The Vibrio anguillarum phage VOP240901 of the present invention has been shown by the shrimp challenge experiment that the survival rate of the shrimp treated with the phage reaches 88.33%, which is much higher than the survival rate of 31.67% in the challenge group, and has the value of practical application. Description of the Drawings

[0017] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the following will briefly introduce the drawings required for the description of the embodiments or the prior art. Obviously, the drawings in the following description are only some embodiments of the present invention. For those of ordinary skill in the art, without creative efforts, other drawings can be obtained based on these drawings.

[0018] Figure 1 It is the morphology of the Vibrio owensis phage VOP240901 of the present invention;

[0019] Figure 2 It is the result graph of the temperature tolerance of the Vibrio owensis phage VOP240901 of the present invention;

[0020] Figure 3 It is the result graph of the pH tolerance of the Vibrio owensis phage VOP240901 of the present invention.

[0021] Figure 4 It is the result graph of the Vibrio owensis phage VOP240901 of the present invention in the shrimp challenge experiment. Detailed implementation manners

[0022] The following will clearly and completely describe the technical solutions in the embodiments of the present invention in combination with the embodiments of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, rather than all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative efforts belong to the scope of protection of the present invention.

[0023] The Vibrio owensis phage VOP240901 of the present invention was deposited at the China Center for Type Culture Collection (CCTCC) on September 20, 2024. The deposit address is: Wuhan University, Wuhan, China; the deposit number is CCTCC NO: M20242007; the phage was identified as being in a viable state on September 27, 2024.

[0024] Example 1 Phage isolation

[0025] The present invention screens phages with a broader host range by co-incubating various different types of vibrios with environmental water samples. Specifically, phages are isolated by the mixed culture method and the double-layer plate method. The implementation method is as follows:

[0026] (1) Collect 50 water samples from aquaculture waters. The water samples are from the waters of a certain farm in Zhangzhou, Fujian. After centrifugation at 5000 RPM, filter with a 0.22 μm filter, and take the filtrate for standby.

[0027] (2) Five glycerol bacteria of Vibrio parahaemolyticus, five of Vibrio alginolyticus, five of Vibrio harveyi, and five of Vibrio owensii were selected. The numbers of these vibrios are: FR1, FR2, FR3, VP240901, VP240902, RZ1, RZ2, RZ3, RZ4, RZ5, VH1.1601, VH240501, VH240502, VH240507, VH240508, VO240301, VO240302, VO240303, VO240304, VO240305. They were inoculated into 5 mL of 2 wt% NaCl-TSB liquid medium at an inoculation ratio of 2 wt%, and cultured in a shaker at 30 °C and 200 RPM for 1 - 2 h until the logarithmic growth phase, and used as the incubated vibrio bacterial liquid for standby. All the above strains are from the laboratory strain bank.

[0028] (3) 200 μL of the above incubated vibrio bacterial liquid was added to 20 mL of fresh 2 wt% NaCl-TSB medium respectively, mixed and incubated with 5 mL of environmental water samples, and cultured overnight in a shaker at 30 °C and 200 RPM. The enriched phage filtrate was obtained by filtering with a 0.22 μm filter head.

[0029] (4) Using NA solid medium as the bottom plate, 2 wt% NaCl-TSB semi-solid, the gradient-diluted phage filtrate, and the above 20 host vibrios were mixed and poured on the NA bottom plate to form a double-layer plate, and incubated overnight in a 30 °C incubator. At an appropriate dilution gradient, single plaques were formed on the double-layer plate of vibrio VO240305, and this vibrio was used as the host bacterium.

[0030] (5) Single plaques were picked and added to the bacterial liquid of the host bacterium in the logarithmic growth phase for co-incubation, and steps (2) and (3) were repeated. Finally, plaques with uniform morphology were formed on the double-layer plate, indicating that the purification was completed. As Figure 1 shown, the phage formed shallow and small plaques on the double-layer plate, with a plaque diameter of about 0.5 mm. It was named Vibrio owensis phage VOP240901 (hereinafter referred to as "phage VOP240901"), and was deposited in the China Center for Type Culture Collection (CCTCC) on September 20, 2024.

[0031] The components of NA medium are: 1000 mL of sterile water, 3 g of beef extract, 5 g of peptone, 2.5 g of glucose, 18 g of agar;

[0032] The components of NaCl-TSB medium are: 1000 mL of sterile water, 17 g of peptone, 3 g of soybean peptone, 25 g of sodium chloride, 2.5 g of dipotassium hydrogen phosphate, 2.5 g of glucose;

[0033] The components of the NaCl-TSB semi-solid medium are: 1000 mL of sterile water, 17 g of peptone, 3 g of soybean peptone, 25 g of sodium chloride, 2.5 g of dipotassium hydrogen phosphate, 2.5 g of glucose, and 5 g of agar.

[0034] Example 2 Determination of the host range of phage VOP240901 by the spot method

[0035] The implementation method is as follows:

[0036] (1) Select 20 strains of Vibrio parahaemolyticus, 20 strains of Vibrio alginolyticus, 14 strains of Vibrio harveyi, and 20 strains of Erwinia carotovora. These vibrios are all from the company's pathogen seed bank. After purification on TCBS plates, the 16S rDNA is determined to identify the species.

[0037] (2) Pick single colonies of the above vibrios, place them in a 2 wt% NaCl-TSB medium and culture overnight. Take the culture solution and mix it with 2 wt% NaCl-TSB semi-solid and pour it onto a NA bottom plate to form a double-layer plate containing bacterial lawns.

[0038] (3) Pipette 5 μL of phage VOP240901 lysate and titrate it onto the above bacterial lawns. Incubate overnight in an oven at 30 °C and observe whether clear lysis plaques appear to determine the lysis spectrum of the phage's host bacteria. The results are shown in the following table. "+" represents the appearance of clear lysis plaques, and "-" represents non-lysis. The results are shown in Table 1.

[0039] Table 1 Lysis spectrum of the host bacteria of phage VOP240901

[0040] Vibrio alginolyticus Lysis Vibrio parahaemolyticus Lysis Vibrio harveyi Lysis Vibrio owensii Lysis VA240714 + VP240901 + VH240601 + V0241001 - VA240715 - VP240902 + VH240602 + V0241002 + VA240716 - VP240903 - VH240603 + V0241003 + VA240717 - VP240904 - VH240604 - V0241004 + VA240718 - VP240905 - VH240605 + V0241005 - VA240719 + VP240906 - VH240606 + V0241006 + VA240720 + VP240907 - VH240607 - V0241007 + VA240721 + VP240908 - VH240608 - V0241008 + VA240722 - VP240909 - VH240609 + V0241009 + VA240723 - VP240910 - VH240610 + V0241010 + VA240724 + VP240911 + VH240611 + V0241011 + VA240725 - VP240912 - VH240612 + V0241012 + VA240726 + VP240913 - VH240613 + V0241013 + VA240727 - VP240914 - VH240614 + V0241014 + VA240728 - VP240915 - V0241015 + VA240801 + VP240916 - V0241016 + VA240802 - VP240917 - V0241017 + VA240803 - VP240918 + V0241018 + VA240804 - VP240919 - V0241019 + VA240805 - VP240920 - V0241020 +

[0041] Table 1 results show that the lysis rates of phage VOP240901 against Vibrio parahaemolyticus and Vibrio alginolyticus are 20% and 35% respectively, and the lysis rates against Vibrio harveyi and Erwinia carotovora are 78.57% and 90%, indicating that the phage is more inclined to lyse Vibrio harveyi and Erwinia carotovora.

[0042] Example 3 Detection of the temperature and pH tolerance of phage VOP240901

[0043] The implementation method is as follows:

[0044] (1) Take sterile water to dilute phage VOP240901 to a final concentration of 8×10 9 PFU / mL for standby.

[0045] (2) Take 100 μL of the above-mentioned VOP240901 phage liquid, add it to a 2 mL EP tube containing 900 μL of sterile water, and place it in environments at 4°C, 15°C, 25°C, 35°C, 45°C, 55°C, 65°C, and 75°C for 2 h respectively. Then, detect the titer of phages in each EP tube.

[0046] (3) Take 100 μL of the VOP240901 phage liquid obtained in step (1), add it to a 2 mL EP tube containing 900 μL of sterile water with a pH of 1 - 13 that has been prepared, and place it for 2 h. Then, detect the titer of phages in each EP tube. The results are shown in Figure 2 and Figure 3 。

[0047] Figure 2 As shown, for temperature stability, phage VOP240901 remains stable in the environment of 4 - 55°C, with a small fluctuation in titer. At 65°C, the titer drops significantly, dropping to 10 4 ~10 5 PFU / mL, and phage VOP240901 is completely inactivated when the environmental pH reaches 75°C.

[0048] For pH stability, Figure 3 It can be seen that phage VOP240901 can remain stable under the conditions of pH 3 - 11. When the environmental pH reaches 2 and 12, the titer of phage VOP240901 drops significantly, dropping to 10 3 ~10 4 PFU / mL, and phage VOP240901 is completely inactivated when the environmental pH reaches 1 and 13.

[0049] Example 4 Optimal medium combination for the fermentation of phage VOP240901

[0050] The implementation method is as follows:

[0051] The components of medium 1 include the following ingredients: 1000 mL of sterile water, 12 g of tryptone, 2 g of soy peptone, 1 g of yeast extract powder, 10 g of sodium chloride, 20 g of culture-grade sea salt, 0.8 g of calcium chloride, 0.7 g of magnesium chloride, 1.5 g of dipotassium hydrogen phosphate, 1.8 g of glucose. Adjust to pH = 7.8 and sterilize at 121°C for 20 minutes.

[0052] The components of medium 2 include the following ingredients: 1000 mL of sterile water, 6 g of peptone, 1 g of yeast extract powder, 1.2 g of ammonium ferric citrate, 10 g of sodium chloride, 20 g of culture-grade sea salt, 1 - 2 g of calcium chloride, 5 - 6 g of magnesium chloride, 0.008 g of disodium hydrogen phosphate. Adjust to pH = 7.8 and sterilize at 121°C for 20 minutes.

[0053] The components of Medium 3 include the following: 1000 mL of sterile water, 10 g of tryptone, 2 g of yeast extract powder, 2 g of beef extract powder, 10 g of sodium chloride, 20 g of culture-grade sea salt, 1.5 g of glucose, adjusted to pH = 7.8, and sterilized at 121 °C for 20 minutes.

[0054] Pick a single colony of the host bacterium from the TCBS plate and inoculate it into 20 mL of fresh medium to obtain the inoculated host bacterium solution. The three types of inoculation media selected are the above-mentioned ones, and they are cultured on a shaker at 30 °C and 200 RPM. Monitor the OD600 of the bacterium solution every 1 h until the OD600 of the inoculated bacterium solution reaches above 1. At this time, the state of the host bacterium is good and it can be used for inoculation into the fermentation medium.

[0055] Take the above-mentioned inoculated bacterium solution and inoculate the host bacterium solution into 20 mL of the above three types of media at an inoculation amount of 2 wt% as the fermentation bacterium solution. Culture it on a shaker at 30 °C and 200 RPM, and detect the OD600 of the bacterium solution every 10 min until the OD600 of the inoculated medium reaches 0.1. At this time, the concentration of the host bacterium is 5×10 7 CFU / mL.

[0056] Take 1 mL of the VOP240901 bacteriophage solution and add it to the fermentation bacterium solution prepared with the above different fermentation media. Incubate it in the dark for 15 min, and then culture it on a shaker at 30 °C and 200 RPM for 16 h. Detect the titer, and the results are shown in Table 2.

[0057] Table 2 Medium combinations for the fermentation of bacteriophage VOP240901

[0058] Inoculation medium Fermentation medium Titer (PFU / mL) Medium 1 Medium 1 <![CDATA[6.68×10 8 > Medium 1 Medium 2 <![CDATA[5.1×10 9 > Medium 1 Medium 3 <![CDATA[5.5×10 6 > Medium 2 Medium 1 <![CDATA[3.2×10 8 > Medium 2 Medium 2 <![CDATA[7.8×10 8 > Medium 2 Medium 3 <![CDATA[2.05×10 6 > Medium 3 Medium 1 <![CDATA[2×10 7 > Medium 3 Medium 2 <![CDATA[6.1×10 8 > Medium 3 Medium 3 <![CDATA[3.5×10 6 >

[0059] As shown in Table 2, the inoculation culture should preferably be Medium 1, and the fermentation medium should preferably be Medium 2. At this time, the highest titer of bacteriophage VOP240901 is 5.1×10 9 PFU / mL.

[0060] Example 5 Optimal multiplicity of infection (MOI) of bacteriophage VOP240901

[0061] The optimal multiplicity of infection (MOI) of a bacteriophage refers to the ratio of the number of bacteriophages to the number of host bacterium cells during infection. Different MOIs will have different effects on the fermentation effect of the bacteriophage. Therefore, the purpose of this example is to find the optimal MOI for bacteriophage fermentation, and the implementation method is as follows:

[0062] All the fermentation systems in this example are consistent with the implementation method in Example 4. The fermentation system is 20 mL, and the inoculation amount of the host bacterium is 2 wt%.

[0063] Furthermore, the inoculation medium was selected as Medium 1, and the fermentation medium was selected as Medium 2. The results are shown in Table 3.

[0064] Table 3 Titer of Bacteriophage VOP240901

[0065] MOI Titer (PFU / mL) 0.001 <![CDATA[5.3×10 9 > 0.01 <![CDATA[4.32×10 10 > 0.1 <![CDATA[6.6×10 9 > 1 <![CDATA[9.06×10 8 > 10 <![CDATA[8.38×10 7 >

[0066] As can be seen from Table 3, the optimal MOI was 0.01, and at this time, the titer of bacteriophage VOP240901 was 4.32×10 10 PFU / mL.

[0067] Example 6 Scale-up Fermentation of Bacteriophage VOP240901

[0068] In order to obtain a bacteriophage solution with a higher titer and connect the industrial fermentation production process, a 6L fermenter was used to scale up the fermentation of the bacteriophage.

[0069] (1) Further, according to the conditions of Example 5 above, the inoculation medium was selected as Medium 1, the fermentation medium was selected as Medium 2, the MOI was controlled at 0.01, and the fermentation of bacteriophage VOP240901 was scaled up to a 6L system. Gradient temperatures were set to optimize the optimal temperature for large-scale fermentation. The temperature settings were 25, 30, 35, and 40 °C respectively. The inoculation amount of the host bacteria was 2 wt%, and bacteriophage VOP240901 with a final concentration of 5×10 5 / PFU / mL was added after the OD600 of the bacterial solution in the fermentation system reached 0.1. The results are shown in Table 4.

[0070] Table 4 Effect of Temperature on the Titer of Bacteriophage VOP240901

[0071]

[0072]

[0073] As can be seen from Table 4, the optimal fermentation temperature was 30 °C, and at this time, the titer of bacteriophage VOP240901 was 7.5×10 10 PFU / mL.

[0074] (2) Further, on the basis of the optimal fermentation temperature of 30 °C, the optimal pH for fermentation was sought. A gradient pH fermentation medium was set for bacteriophage fermentation, and the fermentation steps were the same as in Example 4. The results are shown in the following table.

[0075] Table 5 Effect of pH on the Titer of Bacteriophage VOP240901

[0076] pH Titer (PFU / mL) 6.5 <![CDATA[8.06×10 9 > 7 <![CDATA[7.37×10 10 > 7.5 <![CDATA[9.2×10 10 > 8 <![CDATA[6.18×10 10 >

[0077] As can be seen from Table 5, the optimal fermentation pH is 7.5, and the titer of phage VOP240901 at this time is 9.2×10 10 PFU / mL.

[0078] Furthermore, based on the optimal MOI obtained in Example 5, the MOI feeding ratio was optimized and fine-tuned again. The selected MOIs are as follows. All other control conditions follow the above optimal fermentation conditions. The inoculation medium is Medium 1, the fermentation medium is Medium 2, the fermentation temperature is 30°C, and the pH is controlled at 7.5. The results are shown in Table 6:

[0079] Table 6 Effect of MOI on the titer of phage VOP240901

[0080] MOI Titer (PFU / mL) 0.005 <![CDATA[3.5×10 10 > 0.01 <![CDATA[8.63×10 10 > 0.02 <![CDATA[9×10 10 > 0.04 <![CDATA[1.25×10 10 > 0.06 <![CDATA[9.62×10 10 >

[0081] As shown in Table 6, when MOI = 0.04, the best titer of phage VOP240901 obtained is 1.25×10 11 PFU / mL. This shows that the optimal MOI of phage VOP240901 during large-scale fermentation is slightly different from that in the small-scale system. In this invention, the optimal MOI for 6L large-scale fermentation is slightly higher than that for 20mL small-scale fermentation.

[0082] Example 7 Effect of phage VOP240901 on the prevention and treatment of vibriosis in shrimp

[0083] To verify the control effect of phage on vibrio in actual shrimp farming, a virulence experiment using vibrio VO240305 was carried out to characterize the prevention and treatment effect of phage. The steps are as follows:

[0084] (1) Prepare 300 healthy white shrimp at 7 days old and culture them in a 20L glass water tank. Control the water temperature at about 25°C and divide them into 5 groups: G1, G2, G3, G4, G5, with 60 shrimp in each group. Among them, every 20 shrimp are used as a subgroup for parallel repetition. Among them, G1 is the blank group without adding vibrio for virulence; G2 is the phage control group, only adding phage solution; G3 is the virulence group, only adding virulent vibrio solution; G4 is the phage treatment group, adding phage solution 1 day after adding virulent vibrio solution for virulence; G5 is the antibiotic treatment group, adding antibiotic 1 day after adding virulent vibrio solution for virulence.

[0085] (2) After the shrimp adapt to the water body, prepare to pour the vibrio VO240305 used for virulence into the glass water tank for immersion virulence experiment. The used vibrio VO240305 is first cultured to a concentration of 2×10 9 CFU / mL and added to the virulence groups G3, G4, and G5 to make the final concentration of vibrio in the water tank 2×10 6CFU / mL. In Group G2, phage liquid was directly added to make the final concentration 2×10 7 PFU / mL.

[0086] (3) One day after Vibrio challenge, phage liquid was added to Group G4 to make the final concentration of phages in the water tank 2×10 7 PFU / mL, and doxycycline with a final concentration of 10 mg / L was added to Group G5. The survival status of the shrimp in all groups was observed 3 days after treatment.

[0087] (4) The results are as Figure 4 shown. The survival rates of Groups G1 and G2 were 98.33% and 96.67% respectively, indicating that phages hardly affected the survival of shrimp. The survival rate of the challenged group G3 was only 31.67%, while the survival rates of the phage treatment group G4 and the antibiotic treatment group G5 were 88.33% and 75% respectively, indicating that phage treatment improved the survival rate of Vibrio-infected shrimp, and the effect of phage treatment was better than that of the antibiotic treatment group.

[0088] The above are only the preferred embodiments of the present invention and are not intended to limit the present invention. Any modifications, equivalent replacements, improvements, etc. made within the spirit and principles of the present invention shall be included within the protection scope of the present invention.

Claims

1. A Vibrio Erwinia phage, characterized in that: The bacteriophage is Vibrio erwinia phage VOP240901, and its deposit number is CCTCC NO: M20242007.

2. The Vibrio erwinii phage according to claim 1, characterized in that: The titer of the Vibrio erwinii phage VOP240901 is 10 9- 10 11 PFU / mL.

3. The Vibrio erwinia phage according to claim 1, characterized in that: The MOI of the Vibrio erwinia phage VOP240901 is 0.001-10.

4. An aquatic feed additive, characterized in that: The invention comprises the Vibrio erwinia phage according to any one of claims 1 to 3.

5. A microbial bactericide, characterized in that: The invention comprises the Vibrio erwinia phage according to any one of claims 1 to 3.

6. Use of the Vibrio erwinii phage according to any one of claims 1 to 3 in killing microorganisms of the genus Vibrio, characterized in that: The Vibrio microorganism is one or more of Vibrio parahaemolyticus, Vibrio alginolyticus, Vibrio harveyi and Vibrio erwinia.

Images