CN119464231A - Stenotrophomonas maltophilia phage, composition and application thereof - Google Patents
Stenotrophomonas maltophilia phage, composition and application thereof Download PDFInfo
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- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The invention discloses a stenotrophomonas maltophilia bacteriophage, a composition and application thereof. The phage comprises phage vB_SmaSZP 9, taxonomic name Stenotrophomonas phage vB _SmaSZP 9, latin's name Stenotrophomonas phage, the phage vB_SmaSZP 9 is preserved in the Guangdong province microorganism strain preservation center, the preservation address is No. 59 building 5 of the 100 th university in Guangzhou city martyr, the preservation number is GDMCC No:64509-B1, and the preservation date is No. 2024, no. 05 and 16. The phage of the invention has wide host spectrum, strong infection capability and wide tolerance range to temperature, acid and alkali.
Description
Technical Field
The invention relates to the technical field of biology, in particular to a stenotrophomonas maltophilia bacteriophage, a composition and application thereof.
Background
The stenotrophomonas maltophilia is an obligate aerobic gram-negative non-zymobacter, is a common bacterium next to pseudomonas aeruginosa and acinetobacter, is widely distributed in water, plants and soil, and can also be lodged in respiratory tracts and intestinal tracts of people and animals. It is the only human-infected species of 17 species of the genus stenotrophomonas.
When the body resistance is reduced, the stenotrophomonas maltophilia can be secondarily infected, and the respiratory tract, urinary tract, gastrointestinal tract, skin and soft tissues, bones and joints, ophthalmic infection, meningitis, endocarditis, bacteremia and the like are caused.
The stenotrophomonas maltophilia has various drug resistance mechanisms, such as drug resistance enzyme generation, multi-drug efflux pump, SMqnr gene, biomembrane mechanism and the like, so that the stenotrophomonas maltophilia has drug resistance to most antibiotics, such as cephalosporins, carbapenems, aminoglycosides, macrolides, beta-lactams and the like, and the range of drugs selectable in clinical treatment is narrowed.
The stenotrophomonas maltophilia can form biofilms on wet surfaces (enhanced resistance to water soluble disinfectants) in direct or indirect contact with patients, such as in respiratory tubing, catheters, intravenous catheters, dialysis equipment, dental equipment and hospital plumbing systems (sinks and faucets). Bacteria within the biofilm are generally able to withstand antibiotic treatment up to 10 to 1000 times the concentration compared to planktonic bacteria, which significantly reduces the effectiveness of traditional antibiotic treatment, presenting a significant challenge for clinical treatment.
In this context, there is an urgent need to explore effective strategies to reduce the harm posed by stenotrophomonas maltophilia.
Disclosure of Invention
The main object of the present invention is to provide a stenotrophomonas maltophilia bacteriophage, a composition and an application thereof, aiming at seeking an effective strategy to reduce the harm caused by stenotrophomonas maltophilia.
To achieve the above object, the first aspect of the present invention provides a stenotrophomonas maltophilia bacteriophage comprising a bacteriophage vb_sma-SZP9, taxonomically designated Stenotrophomonas phage vB _sma-SZP9, latin's name Stenotrophomonas phage, and the bacteriophage vb_sma-SZP9 deposited at the collection center of microorganism strains in Guangdong province at the accession address of building 5, 30 th university, in Guangzhou city martyr, accession number GDMCC No:64509-B1, and the accession date of 2024, 05 months, 16 days.
The second aspect of the invention provides a stenotrophomonas maltophilia bacteriophage which comprises a bacteriophage vB_Sma-SZP25, wherein the bacteriophage vB_Sma-SZP25 is named as Stenotrophomonas phage vB _Sma-SZP25 in taxonomy, the Latin is named as Stenotrophomonas phage, the bacteriophage vB_Sma-SZP25 is preserved in the microorganism strain preservation center of Guangdong province, the preservation address is building 5 No. 59 of the university 100 in Guangzhou city martyr, the preservation number is GDMCC No:64510-B1, and the preservation date is 2024, namely 16 days 05 month.
The third aspect of the invention provides a stenotrophomonas maltophilia bacteriophage which comprises a bacteriophage vB_Sma-SZP27, wherein the bacteriophage vB_Sma-SZP27 is named as Stenotrophomonas phage vB _Sma-SZP27 in taxonomy, the Latin is named as Stenotrophomonas phage, the bacteriophage vB_Sma-SZP27 is preserved in the microorganism strain preservation center of Guangdong province, the preservation address is No. 59 building 5 of the university 100 in Guangzhou city martyr, the preservation number is GDMCC No:64511-B1, and the preservation date is No. 2024, no. 05 and 16.
The fourth aspect of the invention provides a stenotrophomonas maltophilia bacteriophage which comprises a bacteriophage vB_Sma-SZP28, wherein the bacteriophage vB_Sma-SZP28 is named as Stenotrophomonas phage vB _Sma-SZP28 in taxonomy, the Latin is named as Stenotrophomonas phage, the bacteriophage vB_Sma-SZP28 is preserved in the microorganism strain preservation center of Guangdong province, the preservation address is No. 59 building 5 of the university 100 in Guangzhou city martyr, the preservation number is GDMCC No:64624-B1, and the preservation date is No. 2024, namely No. 05 and 16.
The fifth aspect of the invention provides a stenotrophomonas maltophilia bacteriophage, which comprises a bacteriophage vB_Sma-SZP96, wherein the bacteriophage vB_Sma-SZP96 is named as Stenotrophomonas phage vB _Sma-SZP96 in taxonomy, the Latin is named as Stenotrophomonas phage, the bacteriophage vB_Sma-SZP96 is preserved in the microorganism strain preservation center of Guangdong province, the preservation address is No. 59 building 5 of the university 100 in Guangzhou city martyr, the preservation number is GDMCC No:64625-B1, and the preservation date is No. 2024, namely, no. 05 and 16 days.
In a sixth aspect, the present invention provides a stenotrophomonas maltophilia phage composition comprising: at least two of phage vB_Sma-SZP9, phage vB_Sma-SZP25, phage vB_Sma-SZP27, phage vB_Sma-SZP28 and phage vB_Sma-SZP96, said phage vB_Sma-SZP9, phage vB_Sma-SZP25, phage vB_Sma-SZP27, phage vB_Sma-SZP28 and phage vB_Sma-SZP96 are designated Stenotrophomonas phage vB_Sma-SZP9、Stenotrophomonas phage vB_Sma-SZP25、Stenotrophomonas phage vB_Sma-SZP27、Stenotrophomonas phage vB_Sma-SZP28 and Stenotrophomonas phage vB _Sma-SZP96, respectively, latin is designated Stenotrophomonas phage, said phage vB_Sma-SZP9, said phage vB_Sma-SZP25, said phage vB_Sma-SZP27, said phage vB_Sma-SZP28 and said phage vB_Sma-SZP96 are all deposited at the microorganism culture collection, and the microorganism deposit address: building 5 of road 100 college No. 59 in Guangzhou city martyr has corresponding preservation numbers of GDMCC No:64509-B1, GDMCC No:64510-B1, GDMCC No:64511-B1, GDMCC No:64624-B1 and GDMCC No:64625-B1, and preservation date of 2024, month 05 and 16 days.
In some embodiments, the titers of phage vB_Sma-SZP9, phage vB_Sma-SZP25, phage vB_Sma-SZP27, phage vB_Sma-SZP28, and phage vB_Sma-SZP96 are all 10 7-1010 pfu/mL.
In some embodiments, the phage vB_Sma-SZP9, the phage vB_Sma-SZP25, the phage vB_Sma-SZP27, the phage vB_Sma-SZP28, and the phage vB_Sma-SZP96 are all at a multiplicity of infection of 0.00001-10.
In a seventh aspect, the present invention provides a phage preparation comprising a stenotrophomonas maltophilia phage or a stenotrophomonas maltophilia phage composition.
In some embodiments, the phage preparation has a pH of 5-10.
In the phage preparation, when the phage preparation comprises phage vB_Sma-SZP9, phage vB_Sma-SZP25, phage vB_Sma-SZP27, phage vB_Sma-SZP28 and phage vB_Sma-SZP96, the ratio of the number of living bodies of phage vB_Sma-SZP9, phage vB_Sma-SZP25, phage vB_Sma-SZP27, phage vB_Sma-SZP28 and phage vB_Sma-SZP96 is (1-3): (1-3), and/or the content of phage vB_Sma-SZP9, phage vB_Sma-SZP25, phage vB_Sma-SZP27, phage vB_Sma-SZP28 and phage vB-SZP 96 is equal to or greater than 57 mL.
The phage preparation also comprises pharmaceutically acceptable auxiliary materials, wherein the pharmaceutically acceptable auxiliary materials comprise one or more of a stabilizer, a dispersing agent, a solvent and a filler.
The phage preparation comprises a liquid preparation, a freeze-dried preparation or an oral solid preparation.
In an eighth aspect, the present invention provides the use of a phage preparation according to the seventh aspect for lysing stenotrophomonas maltophilia.
In a ninth aspect, the present invention provides a pharmaceutical composition comprising a stenotrophomonas maltophilia bacteriophage or a stenotrophomonas maltophilia bacteriophage composition.
In the technical scheme of the invention, phage vB_Sma-SZP9, phage vB_Sma-SZP25, phage vB_Sma-SZP27, phage vB_Sma-SZP28 and phage vB_Sma-SZP96 have strong infection capability, can quickly kill host bacteria in a culture medium, have wide temperature and acid-base tolerance range, and have good inhibition and killing effects on proliferation of the stenotrophomonas maltophilia. The phage genome in the stenotrophomonas maltophilia phage composition provided by the invention does not carry virulence genes, drug resistance genes and lysogenic genes, does not cause the transmission of virulence genes and drug resistance genes, and is safe to use. The phage composition provided by the invention has broad spectrum and can kill more than 90% of drug-resistant stenotrophomonas maltophilia. The phage composition can be used for preparing phage preparations such as spraying agents, injection and the like, can be used for killing the stenotrophomonas maltophilia in the environment and treating the infection caused by the stenotrophomonas maltophilia of organisms, and can effectively solve the problem of difficult treatment caused by drug resistance of the stenotrophomonas maltophilia and biofilm.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings that are required in the embodiments or the description of the prior art will be briefly described, and it is obvious that the drawings in the following description are only some embodiments of the present invention, and other drawings may be obtained according to the structures shown in these drawings without inventive effort for a person skilled in the art.
FIG. 1 is a plaque morphology and electron microscopy of phage vB_Sma-SZP9, phage vB_Sma-SZP25, phage vB_Sma-SZP27, phage vB_Sma-SZP28 and phage vB_Sma-SZP 96;
FIG. 2 is a genomic analysis of phage vB_Sma-SZP 9;
FIG. 3 is a genomic analysis of phage vB_Sma-SZP 25;
FIG. 4 is a genomic analysis of phage vB_Sma-SZP 27;
FIG. 5 is a genomic analysis of phage vB_Sma-SZP 28;
FIG. 6 is a genomic analysis of phage vB_Sma-SZP 96;
FIG. 7 is a graph of one-step growth of phage vB_Sma-SZP9, phage vB_Sma-SZP25, phage vB_Sma-SZP27, phage vB_Sma-SZP28 and phage vB_Sma-SZP 96;
FIG. 8 is a graph of the optimal multiplicity of infection for phage vB_Sma-SZP9, phage vB_Sma-SZP25, phage vB_Sma-SZP27, phage vB_Sma-SZP28 and phage vB_Sma-SZP 96;
FIG. 9 is a graph of the thermal stability of phage vB_Sma-SZP9, phage vB_Sma-SZP25, phage vB_Sma-SZP27, phage vB_Sma-SZP28 and phage vB_Sma-SZP 96;
FIG. 10 is a graph of the pH stability of phage vB_Sma-SZP9, phage vB_Sma-SZP25, phage vB_Sma-SZP27, phage vB_Sma-SZP28 and phage vB_Sma-SZP 96;
FIG. 11 is a host profile of phage vB_Sma-SZP9, phage vB_Sma-SZP25, phage vB_Sma-SZP27, phage vB_Sma-SZP28, and phage vB_Sma-SZP96 and phage compositions;
FIG. 12 is a graph showing the effectiveness of a phage composition in killing drug resistant Aeromonas maltophilia;
FIG. 13 is a graph showing the effect of phage composition on the clearance of drug-resistant Z.maltophilia biofilms.
The achievement of the objects, functional features and advantages of the present invention will be further described with reference to the accompanying drawings, in conjunction with the embodiments.
Detailed Description
In order to make the objects, technical solutions and advantages of the embodiments of the present invention more clear, the technical solutions of the embodiments of the present invention will be clearly and completely described below. The specific conditions are not noted in the examples and are carried out according to conventional conditions or conditions recommended by the manufacturer. The reagents or apparatus used were conventional products commercially available without the manufacturer's attention. In addition, the meaning of "and/or" as it appears throughout includes three parallel schemes, for example "A and/or B", including the A scheme, or the B scheme, or the scheme where A and B are satisfied simultaneously. In addition, the technical solutions of the embodiments may be combined with each other, but it is necessary to base that the technical solutions can be realized by those skilled in the art, and when the technical solutions are contradictory or cannot be realized, the combination of the technical solutions should be regarded as not exist and not within the protection scope of the present invention. All other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention.
The stenotrophomonas maltophilia has various drug resistance mechanisms, such as drug resistance enzyme generation, multi-drug efflux pump, SMqnr gene, biomembrane mechanism and the like, so that the stenotrophomonas maltophilia has drug resistance to most antibiotics, such as cephalosporins, carbapenems, aminoglycosides, macrolides, beta-lactams and the like, and the selection range of the drugs selectable in clinical treatment is narrowed.
Among these, stenotrophomonas maltophilia can form biofilms on wet surfaces (enhanced resistance to water soluble disinfectants) in direct or indirect contact with patients, such as in respiratory tubes, catheters, intravenous catheters, dialysis equipment, dental equipment and hospital plumbing (sinks and faucets). Bacteria within the biofilm are generally able to withstand antibiotic treatment up to 10 to 1000 times the concentration compared to planktonic bacteria, which significantly reduces the effectiveness of traditional antibiotic treatment, presenting a significant challenge for clinical treatment.
In this context, there is an urgent need to explore new and innovative therapeutic strategies. Phage therapy has attracted considerable attention as an emerging therapeutic modality. Phages are a class of viruses specific to bacteria, with a high degree of host specificity. They eventually lead to bacterial disruption and death by infecting the bacteria and undergoing a process of replication and proliferation. Unlike traditional antibiotics, phage therapy has little effect on the normal microbiota of the body while killing bacteria, thus reducing the risk of possible side effects and drug resistance. This property makes phage therapy a great potential in the treatment of drug-resistant bacterial infections.
However, single phage therapy may suffer from a variety of problems including bacterial resistance to single phage, coexistence of multiple bacterial strains, and phage stability and adaptability challenges. Thus, there is an urgent need to explore effective strategies to reduce the harm posed by stenotrophomonas maltophilia to improve the specificity, effectiveness and adaptability of the treatment.
In view of this, a first aspect of the present invention provides a stenotrophomonas maltophilia bacteriophage comprising a bacteriophage vB_Sma-SZP9, designated as Stenotrophomonas phage vB _Sma-SZP9 in taxonomy, designated as Stenotrophomonas phage in Latin, wherein the bacteriophage vB_Sma-SZP9 is deposited at the Guangdong province microorganism strain collection at accession number: GDMCC No:64509-B1, and at accession number: 2024, 05 months and 16 days, floor 5 of the university, line 100, in Guangzhou city martyr.
The second aspect of the embodiment of the invention provides a stenotrophomonas maltophilia bacteriophage which comprises a bacteriophage vB_Sma-SZP25, wherein the bacteriophage vB_Sma-SZP25 is named as Stenotrophomonas phage vB _Sma-SZP25 in taxonomy, the Latin is named as Stenotrophomonas phage, the bacteriophage vB_Sma-SZP25 is preserved in the Guangdong province microorganism strain preservation center, the preservation address is a building 5 No. 59 of the university 100 in Guangzhou city martyr, the preservation number is GDMCC No:64510-B1, and the preservation date is 2024, 05 months and 16 days.
The third aspect of the embodiment of the invention provides a stenotrophomonas maltophilia bacteriophage which comprises a bacteriophage vB_Sma-SZP27, wherein the bacteriophage vB_Sma-SZP27 is named as Stenotrophomonas phage vB _Sma-SZP27 in taxonomy, the Latin is named as Stenotrophomonas phage, the bacteriophage vB_Sma-SZP27 is preserved in the Guangdong province microorganism strain preservation center, the preservation address is a building 5 No. 59 of the university 100 in Guangzhou city martyr, the preservation number is GDMCC No:64511-B1, and the preservation date is 2024, 05 months and 16 days.
The fourth aspect of the embodiment of the invention provides a stenotrophomonas maltophilia bacteriophage which comprises a bacteriophage vB_Sma-SZP28, wherein the bacteriophage vB_Sma-SZP28 is named as Stenotrophomonas phage vB _Sma-SZP28 in taxonomy, the Latin is named as Stenotrophomonas phage, the bacteriophage vB_Sma-SZP28 is preserved in the Guangdong province microorganism strain preservation center, the preservation address is a building 5 No. 59 of the university 100 in Guangzhou city martyr, the preservation number is GDMCC No:64624-B1, and the preservation date is 2024, 05 months and 16 days.
The fifth aspect of the embodiment of the invention provides a stenotrophomonas maltophilia bacteriophage which comprises a bacteriophage vB_Sma-SZP96, wherein the bacteriophage vB_Sma-SZP96 is named as Stenotrophomonas phage vB _Sma-SZP96 in taxonomy and Stenotrophomonas phage in Latin, and the bacteriophage vB_Sma-SZP96 is preserved in the Guangdong province microorganism strain preservation center at the preserving address of No. 59 building 5 of the university 100 in Guangzhou city martyr with the preserving number of GDMCC No:64625-B1.
In a sixth aspect, embodiments of the present invention provide a stenotrophomonas maltophilia phage composition comprising: at least two of phage vB_Sma-SZP9, phage vB_Sma-SZP25, phage vB_Sma-SZP27, phage vB_Sma-SZP28 and phage vB_Sma-SZP96, said phage vB_Sma-SZP9, phage vB_Sma-SZP25, phage vB_Sma-SZP27, phage vB_Sma-SZP28 and phage vB_Sma-SZP96 are designated Stenotrophomonas phage vB_Sma-SZP9、Stenotrophomonas phage vB_Sma-SZP25、Stenotrophomonas phage vB_Sma-SZP27、Stenotrophomonas phage vB_Sma-SZP28 and Stenotrophomonas phage vB _Sma-SZP96, respectively, latin is designated Stenotrophomonas phage, said phage vB_Sma-SZP9, said phage vB_Sma-SZP25, said phage vB_Sma-SZP27, said phage vB_Sma-SZP28 and said phage vB_Sma-SZP96 are all deposited at the microorganism culture collection, and the microorganism deposit address: building 5 of road 100 college No. 59 in Guangzhou city martyr has corresponding preservation numbers of GDMCC No:64509-B1, GDMCC No:64510-B1, GDMCC No:64511-B1, GDMCC No:64624-B1 and GDMCC No:64625-B1, and preservation date of 2024, month 05 and 16 days.
In the technical scheme of the invention, in the stenotrophomonas maltophilia phage composition, phage vB_Sma-SZP9, phage vB_Sma-SZP25, phage vB_Sma-SZP27, phage vB_Sma-SZP28 and phage vB_Sma-SZP96 have strong infection capability, can quickly kill host bacteria in a culture medium, have wide temperature and acid-base tolerance range, and have good inhibition and killing effects on proliferation of stenotrophomonas maltophilia.
In some embodiments, the titers of phage vB_Sma-SZP9, phage vB_Sma-SZP25, phage vB_Sma-SZP27, phage vB_Sma-SZP28, and phage vB_Sma-SZP96 are all 10 9-1010 pfu/mL.
In some embodiments, the phage vB_Sma-SZP9, the phage vB_Sma-SZP25, the phage vB_Sma-SZP27, the phage vB_Sma-SZP28, and the phage vB_Sma-SZP96 are all at a multiplicity of infection of 0.00001 to 10 and a titer of at least 10 7 pfu/mL. The titer of 10 9~1010 pfu/mL can be maintained after 120 minutes of treatment at 4-50 ℃ and the titer of 10 8~1010 pfu/mL can be maintained after 120 minutes of treatment at pH=4-pH=11.
In a seventh aspect, embodiments of the present invention provide a phage preparation comprising a stenotrophomonas maltophilia phage or a stenotrophomonas maltophilia phage composition. The phage mixed preparation formed by compounding the five phages has stronger cracking capability, wider cracking spectrum and wider application range, and makes up the limitation of narrow host spectrum when single phage is applied.
In some embodiments, the phage preparation has a pH of 5-10.
In the phage preparation, when the phage preparation comprises phage vB_Sma-SZP9, phage vB_Sma-SZP25, phage vB_Sma-SZP27, phage vB_Sma-SZP28 and phage vB_Sma-SZP96, the ratio of the number of living bodies of phage vB_Sma-SZP9, phage vB_Sma-SZP25, phage vB_Sma-SZP27, phage vB_Sma-SZP28 and phage vB_Sma-SZP96 is (1-3): (1-3), and the contents of phage vB_Sma-SZP9, phage vB_Sma-SZP25, phage vB_Sma-SZP27, phage vB_Sma-SZP28 and phage vB-SZP 96 are equal to or greater than 10/8 mL.
The phage preparation also comprises pharmaceutically acceptable auxiliary materials, wherein the pharmaceutically acceptable auxiliary materials comprise one or more of a stabilizer, a dispersing agent, a solvent and a filler.
The phage preparation comprises a liquid preparation, a freeze-dried preparation or an oral solid preparation. The phage formulation can be administered via aerosol inhalation, intravenous, intra-arterial, intraperitoneal, catheter, dermal, oral, topical, and rectal routes. Is used for preventing or treating infection of the stenotrophomonas maltophilia on human bodies or cultured animals by spraying, injecting or oral administration and the like. The phage preparation comprises an environmental disinfectant for improving the environment and reducing the content of the stenotrophomonas maltophilia on the surface of the medium. The phage preparation can kill the stenotrophomonas maltophilia in the infected organ through an in-vitro organ perfusion system.
The phage composition preparation provided by the invention can take the phage or phage composition as the only active ingredient. Other phages besides the phages of the invention can be contained, and the phages are compounded with the phages of the invention for use, so that the effect of preventing and controlling the stenotrophomonas maltophilia can be equivalent to or better than the phages.
It should be noted that, in the environment disinfectant provided by the invention, the concentration of each bacteriophage is not lower than 10 8 pfu/mL, and the environment disinfectant also contains other active components for inhibiting or eliminating the stenotrophomonas maltophilia in the environment.
The invention also provides application of the environment disinfectant in the environment of hospitals, social health, nursing homes, farms and object surface disinfection. The application method of the environment disinfectant comprises, but is not limited to, disinfecting and decontaminating water distribution systems, medical facilities, aquaculture facilities, public facilities or other environment surfaces in the forms of liquid soaking, spraying, combined use with an aqueous carrier and the like, and can effectively control the activity and growth of the stenotrophomonas maltophilia.
In an eighth aspect, an embodiment of the invention provides the use of a phage preparation according to the seventh aspect for lysing stenotrophomonas maltophilia.
In a ninth aspect, embodiments of the present invention provide a pharmaceutical composition comprising a stenotrophomonas maltophilia bacteriophage or a stenotrophomonas maltophilia bacteriophage composition.
Compared with the prior art, the invention has at least the following advantages and positive effects:
(1) In the technical scheme of the invention, the genomes of the phage vB_Sma-SZP9, the phage vB_Sma-SZP25, the phage vB_Sma-SZP27, the phage vB_Sma-SZP28 and the phage vB_Sma-SZ96 do not carry virulence genes, drug-resistant genes and lysogenic genes, and the transmission of the virulence genes and the drug-resistant genes is not caused, so that the use is safe.
(2) In the technical scheme of the invention, the phages vB_Sma-SZP9, the phages vB_Sma-SZP25, the phages vB_Sma-SZP27, the phages vB_Sma-SZP28 and the phages vB_Sma-SZP96 have strong infection capability, can quickly kill host bacteria in a culture medium, have wide temperature and acid-base tolerance ranges, can be used as a spraying agent and an injection, have good inhibition and killing effects on proliferation of the stenotrophomonas maltophilia, and have good treatment and protection effects on organisms (human and animals) infected with multiple drug-resistant stenotrophomonas maltophilia.
(3) In the technical scheme of the invention, the phage vB_Sma-SZP9, phage vB_Sma-SZP25, phage vB_Sma-SZP27, phage vB_Sma-SZP28 and phage vB_Sma-SZP96 composition has broad spectrum property, can kill more than 90% of drug-resistant stenotrophomonas maltophilia, and has the advantage of enhancing phage treatment effect.
(4) The phage composition provided by the invention has the potency of 10 7pfu/mL-109 pfu/mL, can effectively remove the stenotrophomonas maltophilia biofilm, and can overcome the treatment difficulty of drug resistance and biofilm of stenotrophomonas maltophilia.
In the embodiment of the application, phage vB_Sma-SZP9 is named as Stenotrophomonas phage vB _Sma-SZP9 in taxonomy and is named as Stenotrophomonas phage in Latin, and the phage vB_Sma-SZP9 is preserved in the Guangdong province microorganism strain collection, the preservation address is No. 59 building 5 of Dai 100 in Guangzhou city martyr, the preservation number is GDMCC No:64509-B1, and the preservation date is 2024, 05 and 16 days. The phage vB_Sma-SZP25 is named as Stenotrophomonas phage vB _Sma-SZP25 in taxonomy and is named as Stenotrophomonas phage in Latin, and the phage vB_Sma-SZP25 is deposited in the microorganism strain collection of Guangdong province, the deposition address is floor 5 of the university 100 in Guangzhou city martyr, the deposition number is GDMCC No:64510-B1, and the deposition date is 2024, 05 months and 16 days. The phage vB_Sma-SZP27 is named as Stenotrophomonas phage vB _Sma-SZP27 in taxonomy and Stenotrophomonas phage in Latin, and the phage vB_Sma-SZP27 is deposited in the microorganism strain collection of Guangdong province, the deposition address is floor 5 of the university 100 in Guangzhou city martyr, the deposition number is GDMCC No:64511-B1, and the deposition date is 2024, 05 months and 16 days. The phage vB_Sma-SZP28 is named as Stenotrophomonas phage vB _Sma-SZP28 in taxonomy and is named as Stenotrophomonas phage in Latin, and the phage vB_Sma-SZP28 is deposited in the microorganism strain collection of Guangdong province, the deposition address is floor 5 of the university No. 59 in Guangzhou city martyr, the deposition number is GDMCC No:64624-B1, and the deposition date is 2024, 05 months and 16 days. The phage vB_Sma-SZP96 is named as Stenotrophomonas phage vB _Sma-SZP96 in taxonomy and is named as Stenotrophomonas phage in Latin, and the phage vB_Sma-SZP96 is deposited in the microorganism strain collection of Guangdong province, the deposition address is floor 5 of the university No. 59 in Guangzhou city martyr, the deposition number is GDMCC No:64625-B1, and the deposition date is 2024, 05 months and 16 days.
The test methods for specific experimental conditions are not noted in the examples below, and are generally performed under conventional experimental conditions or under experimental conditions recommended by the manufacturer. The materials, reagents and the like used, unless otherwise specified, are those obtained commercially.
The invention will be described in detail with reference to specific examples below:
Example 1
(1) The stenotrophomonas maltophilia phage vB_Sma-SZP9, phage vB_Sma-SZP25, phage vB_Sma-SZP27, phage vB_Sma-SZP28 and phage vB_Sma-SZP96 are amplified, potency determined and concentrated.
Phage amplification, namely phage vB_Sma-SZP9, phage vB_Sma-SZP25, phage vB_Sma-SZP27, phage vB_Sma-SZP28 and phage vB_Sma-SZP96 are all obtained by separating the laboratory from the sewage of third people hospital in Shenzhen city. Taking out-80 ℃ preserved stenotrophomonas maltophilia Sma1, sma3 and Sma7, streaking LB non-resistant plates, standing and culturing overnight at 37 ℃, respectively picking single colonies into a3 mL LB culture medium, culturing overnight at 37 ℃ 200 rpm, transferring overnight bacterial liquid 1:20 into fresh 10mL LB, culturing 2h at 37 ℃ 200 rpm, adding vB_Sma-SZP9 phage solution into Sma1, respectively adding vB_Sma-SZP25, vB_Sma-SZP27 and vB_Sma-SZP28 phage solution into Sma3, adding vB_Sma-SZP96 phage solution into Sma7, culturing 2: 2h until bacterial liquid becomes clear at 37 ℃ 150 rpm with the optimal ratio of 0.001-10:1, and obtaining phage amplification liquid;
and (3) detecting phage titer by a double-layer plate method, namely performing 10-time gradient dilution on the phage suspension, taking phage diluent of each gradient, fully and uniformly mixing the phage diluent with 0.3-0.4 mL host bacterial liquid, paving double-layer agar plates, culturing overnight at a constant temperature of 37 ℃, counting plaques on each agar plate, selecting plates with about 20-200 plaques, and calculating the initial concentration of the obtained phage according to the dilution multiple to obtain phage titer (pfu/mL) =dilution multiple×number of plaques×10). The results are shown in FIG. 1:
Phage vB_Sma-SZP9 can form transparent plaques in an agar medium, has no halo around, has clear and regular edges, has a diameter of about 1.0 mm, has a titer of 1.23× 10 pfu/mL, and is a typical lytic phage;
Phage vB_Sma-SZP25 can form transparent plaques in an agar medium, has no halo around, has clear and regular edges, has a diameter of about 1.0 mm, has a titer of 9.08X10: 10 9 pfu/mL, and is a typical lytic phage;
Phage vB_Sma-SZP27 can form transparent plaques in an agar medium, has no halo around, has clear and regular edges, has a diameter of about 1.0 mm, has a titer of 1.08X10: 10 10 pfu/mL, and is a typical lytic phage;
Phage vB_Sma-SZP28 can form transparent plaques in an agar medium, has no halo around, has clear and regular edges, has a diameter of about 1.5 mm, has a titer of 7.83×10 9 pfu/mL, and is a typical lytic phage;
Phage vB_Sma-SZP96 can form transparent plaques in agar medium, has no halo around, has clear and regular edges, has a diameter of about 1.5 mm, and has a titer of 3.33X10: 10 9 pfu/mL, and is a typical lytic phage.
The phage concentration comprises concentrating amplified phage crude suspension with polyethylene glycol (PEG 8000), adding NaCl solid with final concentration of 1M into the concentrated solution, stirring for dissolving, ice bath 1h for promoting separation of phage from corresponding host bacteria fragments, 11000 g centrifuging 10min to remove cell fragments, adding 10% PEG (v/v) into supernatant, standing at 4deg.C overnight, centrifuging 11,000 g at 4deg.C for 10min, removing supernatant, gently resuspending precipitate with appropriate amount of SM buffer, and filtering with 0.22 μm filter membrane for sterilization to obtain phage concentrate.
(2) Morphological observations of phages
Transmission electron microscopy of phage were observed by TEM using phosphotungstic acid (Phosphotungstic acid, PTA) negative staining. Phage concentrate particles were dropped onto a Formvar carbon support membrane, left to stand for 10 min a to dry, and excess liquid was removed with paper. 10 μl PTA was added dropwise to dye 10 min on the membrane, deionized water was added dropwise to wash off excess PTA, excess liquid was removed with paper, and sample 10 min was dried again. The morphology of the phage was observed under TEM. The results are shown in FIG. 1.
The phage vB_Sma-SZP9 has a 24-sided head structure, a head length of about 100nm and a tail length of about 138 nm, belongs to long-tail phages, and belongs to Kyanoviridae phage families through whole genome sequence analysis according to the latest classification and naming of phages.
The phage vB_Sma-SZP25 has a 24-sided head structure, a head length of about 100 nm and a tail length of about 150 nm, belongs to long-tail phages, and belongs to Kyanoviridae phage families through whole genome sequence analysis according to the latest classification and naming of phages.
The phage vB_Sma-SZP27 has a 24-sided head structure, a head length of about 100 nm and a tail length of about 142 nm, belongs to long-tail phages, and belongs to Kyanoviridae phage families through whole genome sequence analysis according to the latest classification and naming of phages.
The phage vB_Sma-SZP28 has a 24-sided head structure, a head length of about 104 nm and a tail length of about 146 nm, belongs to long-tail phages, and belongs to Kyanoviridae phage families through whole genome sequence analysis according to the latest classification and naming of phages.
The phage vB_Sma-SZP96 has a 24-sided head structure, a head length of about 100 nm and a tail length of about 138 nm, belongs to long-tail phages, and belongs to Kyanoviridae phage families through whole genome sequence analysis according to the latest classification and naming of phages.
(3) Phage genome extraction and sequencing
1ML phage concentrate was taken, and DNase I (PROMEGA) at a final concentration of 6U/mL and RNase (TAKARA) at a final concentration of 50. Mu.g/mL were added, each 6. Mu.L, reacted at 37℃for 1h, and the exogenous nucleic acid was removed. Then adding 20 mu L of 2M ZnCl 2, uniformly mixing, reacting for 5min at 37 ℃, centrifuging for 1 min at 10g, discarding supernatant to leave a precipitate, then re-suspending the precipitate by using 500 mu L of TES buffer, reacting for 15 min at 60 ℃, adding 20 mu L of proteinase K, reacting for 90 min at 37 ℃, adding 60 mu L of 3M potassium acetate after the reaction is finished, uniformly mixing, ice-bath for 15 min, adding equal volume of phenol chloroform isoamyl alcohol (25:24:1), reversely mixing, centrifuging for 5min at 10g, taking the supernatant (the step is repeated for 3 times), adding equal volume of chloroform isoamyl alcohol (24:1), reversely mixing, centrifuging for 5min at 10g, taking the supernatant (the step is repeated for 2 times), adding equal volume of isopropyl alcohol into the supernatant, precipitating overnight, centrifuging for 5min at 10g, and removing the supernatant to leave a precipitate. The pellet was washed with 70% ethanol 2 times, centrifuged at 10 000g for 5min, the supernatant removed, the white pellet was naturally air dried, and finally the nucleic acid was dissolved with 20. Mu.L of sterile water and sent to the large gene sequencing company for whole genome sequencing. The results are shown in FIG. 2.
As can be seen from FIGS. 2 to 6, the total length of the genomes of phages vB_Sma-SZP9, vB_Sma-SZP25, vB_Sma-SZP27, vB_Sma-SZP28 and vB_Sma-SZP96 are 159670bp, 160057bp, 159891bp, 159981bp and 160628bp, respectively, and do not carry drug resistance genes, virulence genes and lysogenic genes.
The head fibrin and fiber protein sequences of phages vB_Sma-SZP9, vB_Sma-SZP25, vB_Sma-SZP27, vB_Sma-SZP28 and vB_Sma-SZP96 are shown in tables 1-5 below.
TABLE 1 vB_Sma-SZP9 lyase, tail protein and fiber protein sequences
TABLE 2 vB_Sma-SZP25 lyase, tail protein and fiber protein sequences
TABLE 3 vB_Sma-SZP27 lyase, tail protein and fiber protein sequences
TABLE 4 vB_Sma-SZP28 lyase, tail protein and fiber protein sequences
TABLE 5 vB_Sma-SZP96 lyase, tail protein and fiber protein sequences
(4) One-step growth curve determination of phage
The host bacteria cultures prepared in example 1 were inoculated into fresh medium at a ratio of 1:100 and cultured to logarithmic phase (OD 600 nm=0.5), host bacteria cultured to logarithmic phase 1: 1 mL were mixed with vB_Sma-SZP9, vB_Sma-SZP25, vB_Sma-SZP27, vB_Sma-SZP28 and vB_Sma-SZP96 phage solutions at a ratio of MOI=0.1, respectively, and after a reaction in a 37℃incubator for 5 minutes, centrifuged at 10 000rpm at room temperature for 1 min, and the pellets were left. 1 mL fresh LB medium was aspirated to blow the pellet, centrifuged at 10 rpm at room temperature for 30s, and the pellet was left to blow again (the steps of blowing, centrifuging were repeated at least 3 times). Finally, the pellet was resuspended in 10 mL fresh LB medium, and the resuspended tubes were shake-cultured in 200 rpm in a 37℃shaker for 90 min, with the first 30 min samples taken every 5 min (0, 5, 10, 15, 20, 25, 30 min), and then every 10 min (40, 50, 60, 70, 80, 90 min) samples taken every 600. Mu.L of culture mixture. A standard disposable filter membrane of 0.22 μm is selected for filtration, and the obtained filtrate is placed in a medical refrigerator. The filtrate obtained at each time was diluted in multiple ratio and the titer was measured and the experiment was repeated 3 times. And (5) counting data obtained by the experiment, and drawing a one-step growth curve. The incubation period, burst period, of phage were observed and calculated. As a result, as shown in FIG. 7, the infection host latencies of phage vB_Sma-SZP9, phage vB_Sma-SZP25, phage vB_Sma-SZP27, phage vB_Sma-SZP28 and phage vB_Sma-SZP96 were 5 min, 10 min, 0min, 5 min, 10 min, respectively, and the lysis periods were 90 min, 50min, 60 min, 50min and 30 min, respectively.
(5) Determination of the optimal multiplicity of infection of phage
Phage 1mL (titer 10 9 pfu/mL) was 10-fold diluted in LB broth, and 0.1 dilution and 0.1mL of host bacteria proliferation solution (10 8 pfu/mL) were added to LB broth of 5 mL at MOI values of 10, 1, 0.1, 0.01, 0.001, 0.0001, 0.00001, respectively, and cultured with shaking at 37℃of 200 h. The culture solution 10000 rpm is centrifuged to 5 min, the titer is measured by a double-layer plate method, and the combination with the highest titer is the optimal infection complex. The results are shown in FIG. 8.
As is clear from FIG.8, phage vB_Sma-SZP27 and phage vB_Sma-SZP28 have strong lysis capacities from MOI 10 to 0.00001, the titers of phage vB_Sma-SZP9 are 10. 9-1010 pfu/mL, the optimal multiplicity of infection of phage vB_Sma-SZP9 is 10, 1, 0.1 and 0.01, the titers of phage vB_Sma-SZP27 and phage vB_Sma-SZP25 are 1.75X10 and 5.25X10 8 pfu/mL, respectively, the optimal multiplicity of infection of phage vB_Sma-SZP25 is 10, 1, 0.1, 0.01, 0.001 and 0.0001, the titers of phage vB_Sma-SZP96 are 5.33X10 9、1.01×1011、7.83×1010、6.25×109、3.92×109 and 6.25X10. 8 pfu/mL, respectively, and the titers of phage vB_Sma-SZP96 are 10, 1, 0.1, 0.01, 0, 0.0001 and 0.0001, and 494.4980.494.
(6) Determination of phage Heat stability
Sterile 2 mL EP tubes were taken, 500 μl of phage was added, incubated at 4 ℃,25 ℃, 30 ℃, 37 ℃, 50 ℃, 60 ℃, 70 ℃ and 80 ℃ for 2 h, and phage titers after incubation at different temperatures were determined using a double-layer agar method. The results are shown in FIG. 9.
As can be seen from FIG. 9, treatment of 2h at 50℃showed that phage titers were comparable to 4℃and that phage vB_Sma-SZP9, phage vB_Sma-SZP25, phage vB_Sma-SZP27, phage vB_Sma-SZP28 and phage vB_Sma-SZP96 had excellent high temperature resistance.
(7) Determination of phage pH stability
100. Mu.L of phage were mixed with 900. Mu.L of pH buffer with pH=3-12, incubated in a37℃water bath for 2h, and titers of phage at different pH were determined using the double-layer agar method. The results are shown in FIG. 10.
As can be seen from fig. 10, phages vb_sma-SZP9, phage vb_sma-SZP25, phage vb_sma-SZP27, phage vb_sma-SZP28 and phage vb_sma-SZP96 act on 2h at ph=4-11, and still maintain high titers, indicating that the acid-base tolerance range of these five phages is wide.
(8) Determination of phage host profile
In the embodiment, 138 Shenzhen third people hospital clinical drug-resistant stenotrophomonas maltophilia is detected altogether, 150 mu L of the clinical drug-resistant stenotrophomonas maltophilia growing logarithmically is taken in a 15 mL centrifuge tube, 15 mL of 55 ℃ 0.5% agarLB is poured into the centrifuge tube, after being mixed evenly, the mixture is poured into a square plate immediately, and the square plate is uncapped in a biosafety cabinet for standing and solidification. The square plate and a 96-well plate storing Shenzhen third people hospital phage library are placed in an automatic pipetting workstation for high throughput pipetting. The plates were then incubated overnight at 37 ℃ and the screening results were recorded using a colony imager. The results are shown in FIG. 11.
As can be seen from FIG. 11, phage vB_SmaSZP 9, phage vB_SmaSZP 25, phage vB_SmaSZP 27, phage vB_SmaSZP 28 and phage vB_SmaSZP 96 form clear plaques to 47.10%, 55.80%, 51.45%, 48.55% and 47.10% of clinically resistant M.maltophilia, and form fuzzy plaques to 21.02%, 16.66%, 15.22%, 14.49% and 15.94% of clinically resistant M.maltophilia, respectively, and five phage mixtures (cocktail) form clear plaques to 81.88% of clinically resistant M.maltophilia, and form fuzzy plaques to 11.60% of clinically resistant M.maltophilia, indicating a good broad spectrum of five phage compositions.
(9) Sterilization assay in phage composition liquid medium
Thallus vB_Sma-SZP9, phage vB_Sma-SZP25, phage vB_Sma-SZP27, phage vB_Sma-SZP28 and phage vB_Sma-SZP96 are mixed according to a quantitative ratio (pfu/mL) of 1:1:1:1, so that the phage titer of the composition is 10 10 pfu/mL, and the composition is stored at 4 ℃ for standby. The log phase clinical resistant M.maltophilia was adjusted for OD 600 ≡0.2, added to 200. Mu.L phage composition (10 7pfu/mL、108 pfu/mL and 10 9 pfu/mL) in a quantitative ratio (pfu/mL) of 1:1:1:1, respectively, and cultured at rest at 37℃for OD 600 every 15 min. The results are shown in FIG. 12.
As can be seen from FIG. 12, the five M.maltophilia phage compositions rapidly killed different clinical drug-resistant M.maltophilia (numbered Sma69, sma70, sma72 and Sma79, respectively) under different infection complex conditions.
(10) Determination of the Capacity of phage compositions to clear biofilm
200. Mu.L of LB clinical resistant M.maltophilia (Sma 69, sma70, sma72 and Sma 79) suspensions (10 8 cfu/mL) were added to 96-well plates and incubated at 37℃for 24 h and 48 h to form 24 h and 48 h mature biofilms. After three washes with PBS, 200. Mu.L phage compositions (10 7pfu/mL、108 pfu/mL and 10 9 pfu/mL) were added in a quantitative ratio (pfu/mL) of 1:1:1:1:1, respectively, and left to stand at 37℃for treatment 4h. An equal volume of LB was added as a negative control. Biofilm clearance = 100% × (experimental OD 595-control OD 595)/control OD 595. The results are shown in FIG. 13.
As can be seen from FIG. 13, the five M.maltophilia phage compositions have good cleaning effects on clinical drug-resistant M.maltophilia Sma69, sma70, sma72 and Sma79 biofilms under different titers.
The foregoing is merely a preferred embodiment of the present invention and is not intended to limit the scope of the present invention, but various modifications and variations will be apparent to those skilled in the art. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present invention should be included in the scope of the present invention.
Claims (10)
1. The stenotrophomonas maltophilia phage is characterized by comprising phage vB_SmaSZP 9, wherein the taxonomic name is Stenotrophomonas phage vB _SmaSZP 9, the Latin is Stenotrophomonas phage, the phage vB_SmaSZP 9 is preserved in the Guangdong province microorganism strain preservation center, the preservation address is Guangzhou city martyr, road 100, no. 59 building 5, the preservation number is GDMCC No:64509-B1, and the preservation date is 2024, no. 05 and 16.
2. The stenotrophomonas maltophilia phage is characterized by comprising phage vB_Sma-SZP25, wherein the taxonomic name is Stenotrophomonas phage vB _Sma-SZP25, the Latin literature name is Stenotrophomonas phage, the phage vB_Sma-SZP25 is preserved in the Guangdong province microorganism strain preservation center, the preservation address is Guangzhou city martyr, road 100, no. 59 building 5, the preservation number is GDMCC No:64510-B1, and the preservation date is 2024, no. 05 and 16.
3. The stenotrophomonas maltophilia phage is characterized by comprising phage vB_Sma-SZP27, wherein the taxonomic name is Stenotrophomonas phage vB _Sma-SZP27, the Latin literature name is Stenotrophomonas phage, the phage vB_Sma-SZP27 is preserved in the Guangdong province microorganism strain preservation center, the preservation address is Guangzhou city martyr, road 100, no. 59 building 5, the preservation number is GDMCC No:64511-B1, and the preservation date is 2024, no. 05 and 16.
4. The stenotrophomonas maltophilia phage is characterized by comprising phage vB_Sma-SZP28, wherein the taxonomic name is Stenotrophomonas phage vB _Sma-SZP28, the Latin literature name is Stenotrophomonas phage, the phage vB_Sma-SZP28 is preserved in the Guangdong province microorganism strain preservation center, the preservation address is Guangzhou city martyr, road 100, no. 59 building 5, the preservation number is GDMCC No:64624-B1, and the preservation date is 2024, no. 05 and 16.
5. The stenotrophomonas maltophilia phage is characterized by comprising phage vB_Sma-SZP96, wherein the taxonomic name is Stenotrophomonas phage vB _Sma-SZP96, the Latin literature name is Stenotrophomonas phage, the phage vB_Sma-SZP96 is preserved in the Guangdong province microorganism strain preservation center, the preservation address is No. 59 building 5 of the university 100 in Guangzhou city martyr, the preservation number is GDMCC No:64625-B1, and the preservation date is No. 2024, no. 05 and 16.
6. A phage composition of stenotrophomonas maltophilia, characterized by comprising the following steps: at least two of phage vB_Sma-SZP9, phage vB_Sma-SZP25, phage vB_Sma-SZP27, phage vB_Sma-SZP28 and phage vB_Sma-SZP96, said phage vB_Sma-SZP9, phage vB_Sma-SZP25, phage vB_Sma-SZP27, phage vB_Sma-SZP28 and phage vB_Sma-SZP96 are designated Stenotrophomonas phage vB_Sma-SZP9、Stenotrophomonas phage vB_Sma-SZP25、Stenotrophomonas phage vB_Sma-SZP27、Stenotrophomonas phage vB_Sma-SZP28 and Stenotrophomonas phage vB _Sma-SZP96, respectively, latin is designated Stenotrophomonas phage, said phage vB_Sma-SZP9, said phage vB_Sma-SZP25, said phage vB_Sma-SZP27, said phage vB_Sma-SZP28 and said phage vB_Sma-SZP96 are all deposited at the microorganism culture collection, and the microorganism deposit address: building 5 of road 100 college No. 59 in Guangzhou city martyr has corresponding preservation numbers of GDMCC No:64509-B1, GDMCC No:64510-B1, GDMCC No:64511-B1, GDMCC No:64624-B1 and GDMCC No:64625-B1, and preservation date of 2024, month 05 and 16 days.
7. Phage preparation comprising a stenotrophomonas maltophilia phage according to any one of claims 1 to 5 or a stenotrophomonas maltophilia phage composition according to claim 6.
8. The phage preparation of claim 7, wherein the phage preparation has a pH of 5-10.
9. Use of a phage preparation according to claim 7 or 8 for lysing stenotrophomonas maltophilia.
10. A pharmaceutical composition comprising a stenotrophomonas maltophilia bacteriophage according to any one of claims 1 to 5 or a stenotrophomonas maltophilia bacteriophage composition according to claim 6.
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Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2004064732A2 (en) * | 2003-01-16 | 2004-08-05 | New Horizons Diagnostics Corporation | The use of bacterial phage-associated lysing proteins for preventing and treating bacterial infections in humans, animals and fowl |
| US20100322903A1 (en) * | 2008-01-10 | 2010-12-23 | Trustees Of Boston University | Engineered bacteriophages as adjuvants for antimicrobial agents and compositions and methods of use thereof |
| CN117683734A (en) * | 2023-12-18 | 2024-03-12 | 创噬纪(上海)生物技术有限公司 | Drug-resistant stenotrophomonas maltophilia phage and composition and application thereof |
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2025
- 2025-01-13 CN CN202510047135.1A patent/CN119464231A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2004064732A2 (en) * | 2003-01-16 | 2004-08-05 | New Horizons Diagnostics Corporation | The use of bacterial phage-associated lysing proteins for preventing and treating bacterial infections in humans, animals and fowl |
| US20100322903A1 (en) * | 2008-01-10 | 2010-12-23 | Trustees Of Boston University | Engineered bacteriophages as adjuvants for antimicrobial agents and compositions and methods of use thereof |
| CN117683734A (en) * | 2023-12-18 | 2024-03-12 | 创噬纪(上海)生物技术有限公司 | Drug-resistant stenotrophomonas maltophilia phage and composition and application thereof |
Non-Patent Citations (2)
| Title |
|---|
| ALISHA N MONSIBAIS等: "Enhanced Suppression of Stenotrophomonas maltophilia by a Three-Phage Cocktail: Genomic Insights and Kinetic Profiling", BIORXIV, 16 August 2024 (2024-08-16), pages 1 - 21 * |
| 田爽等: "嗜麦芽窄食单胞菌裂解性噬菌体TS-10的特性及其对细菌生物膜作用研究", 中国病原生物学杂志, vol. 18, no. 09, 14 August 2023 (2023-08-14), pages 1024 * |
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