Pantoea_yangzhengvirus_transmission_biological_control_bacterium_yn60_and_applications_thereof

Abstract

The application discloses a positive flavivirus transmission biocontrol bacterium Pantoea_YN60 and application thereof. The application provides a bacterial strain with a preservation number of CGMCC No. 39790. A new strain Pantoea_YN60 of the Pantoea ananatis species has a good prevention and control effect on Zika virus and dengue virus. The bacterial strain has the advantages of stable, efficient and environment-friendly inhibition on mosquito-borne flavivirus. Meanwhile, the application also provides a method for placing the bacteria-containing plant to prevent and control the mosquito-borne virus, and has a good application prospect in preventing and controlling mosquito-borne flavivirus infectious diseases and the like.

Description

Technical Field

[0001] This invention relates to a biocontrol bacterium for the transmission of flavivirus, Pantoea_YN60, and its application, particularly to Pantoea_YN60 and its application in the prevention and control of flavivirus, belonging to the field of microbial technology. Background Technology

[0002] Mosquito-borne flaviviruses are major pathogens causing serious human diseases, including hemorrhagic fever, biphasic fever, arthritis, and neurotropic disorders, infecting hundreds of millions of people and causing numerous deaths annually. Dengue virus is the most prevalent mosquito-borne flavivirus, having appeared or re-emerged in over 100 countries worldwide, potentially infecting approximately 390 million people annually. This disease is primarily distributed in subtropical and tropical regions, including Yunnan and Guangdong provinces in my country. Between 2015 and 2017, Zika virus experienced a large-scale epidemic in the Pacific Islands and the Americas, with over 223,000 confirmed cases reported; this outbreak was declared a Public Health Emergency of International Concern in 2016. Zika virus infection is associated with severe neurological complications, including Guillain-Barré syndrome in adults and microcephaly in newborns. The increasing frequency of mosquito-borne flavivirus outbreaks has become a major global public health problem. These flaviviruses maintain their life cycle between vertebrate hosts and mosquitoes. Mosquitoes acquire viral particles by sucking the blood of an infected host. The virus invades the mosquito's intestinal epithelial cells, subsequently infecting the hemocoel tissue and migrating to the salivary glands, enabling the infected mosquito to transmit the virus upon subsequent bites. Therefore, regulating the interaction between the mosquito's gut and the virus may be an effective strategy for intervening in and protecting mosquitoes from flavivirus infection.

[0003] The gut of mosquitoes is home to a rich microbial community. This symbiotic flora not only maintains homeostasis within mosquitoes but also plays a complex role in determining their ability to transmit viruses. For example, *Serratia marcescens* in the gut secretes bacterial effector factors that deglycosylate mucin bound to the intestinal epithelial cell membrane, thereby promoting dengue and Zika virus infection. Conversely, *Aedes albopictus* YN46 in the gut can inhibit the transmission of mosquito-borne flavivirus by secreting glucose dehydrogenase to inactivate the envelope protein. *Chlorobacterium* strains (Csp_P and Csp_BJ) inhibit arbovirus infection by secreting enzymes. Csp_P secretes a protease that degrades virus particles, while Csp_BJ produces a specific lipase that destroys the viral envelope. *Enterobacter holmieae* B17 confers resistance to Zika and dengue viruses by secreting sphingosine. Furthermore, the endosymbiotic microorganism Wolbachia can inhibit viral replication by regulating the immune or physiological responses of mosquitoes. In recent years, the interactions between mosquitoes, gut microbiota, and viruses have attracted widespread attention because the regulation of gut microbiota can affect the ability of mosquitoes to transmit pathogenic viruses. Therefore, regulating the mosquito gut microbiota may provide a new strategy for controlling the spread of flavivirus diseases. Summary of the Invention

[0004] The problem the invention aims to solve

[0005] The technical problem to be solved by this invention is to provide a new, efficient, stable and environmentally friendly microbial resource and its application method to overcome the shortcomings of existing mosquito-borne flavivirus (especially Zika virus and dengue virus) control methods, so as to overcome the environmental pollution and mosquito resistance of traditional chemical control.

[0006] Solution for solving the problem

[0007] [1]. A strain with accession number CGMCC No.39790.

[0008] [2]. A composition comprising the strains as described in [1] and / or their extracts;

[0009] Optionally, the composition is a microbial preparation;

[0010] Optionally, the composition is used for the prevention and control of arboviruses;

[0011] Optionally, the strain and / or its extract are used as the active ingredient of the composition.

[0012] [3]. Any of the following (i) to (iii) applications of Pantoea ananatis and / or its extracts:

[0013] (i) Application in the preparation of products for controlling the spread of arboviruses;

[0014] (ii) Application in the preparation of products that inhibit the susceptibility of vectors to arboviruses;

[0015] (iii) Application in the prevention and control of vector-borne viruses.

[0016] [4]. According to the application described in [3], wherein,

[0017] The Pantoea ananatis strain includes strains with a 16S rRNA gene sequence that are at least 97% homologous to the sequence shown in SEQ ID NO: 1;

[0018] Preferably, the Pantoea ananatis comprises strains whose 16S rRNA gene sequence includes the sequence shown in SEQ ID NO: 1;

[0019] More preferably, the Pantoea ananatis includes the Pantoea_YN60 strain, whose accession number is CGMCC No.39790.

[0020] [5]. According to the application described in [3] or [4], wherein the extract of Pantoea ananatis contains endotoxins produced by Pantoea ananatis;

[0021] Preferably, the extract of Pantoea ananatis contains O-specific polysaccharides of Pantoea ananatis.

[0022] [6]. According to any one of [3] to [5], wherein the arbovirus includes one or more of Bunyaviridae, Flaviviridae, Reoviridae and Togaviridae;

[0023] Preferably, the arbovirus includes one or more of the genera Flavivirus, Alphavirus, and Orthobunyavirus.

[0024] [7]. The application according to any one of [3] to [6], wherein the arbovirus is a flavivirus;

[0025] Optionally, the vector-borne viruses include dengue virus and / or Zika virus.

[0026] [8]. The application according to any one of [3] to [7], wherein the vector of the arbovirus includes one or more of mosquitoes, ticks or sandflies;

[0027] Preferably, the vector for the vector-borne virus is a mosquito;

[0028] More preferably, the mosquitoes include one or more of Culex, Culiseta, Anopheles, and Aedes;

[0029] Optionally, the mosquito is an Aedes mosquito, including one or more of Aedes aegypti, Aedes albopictus, Aedes polynesiensis, Aedes australis, Aedes cinereus, Aedes rusticus, and Aedes vexans.

[0030] [9]. The application according to any one of [3] to [8], wherein the product comprises one or more of a reagent or a kit; optionally, the reagent is a microbial preparation.

[0031]

[10] . Any of the following methods (a) to (c):

[0032] (a) A method for controlling the spread of arboviruses, the method comprising applying to a vector Pantoea ananatis and / or an extract thereof, or a composition as described in [2];

[0033] (b) A method for inhibiting the susceptibility of a vector to an insect-borne virus, the method comprising applying to the vector Pantoea ananatis and / or its extracts, or a composition as described in [2];

[0034] (c) A method for controlling arboviruses, the method comprising spraying Pantoea ananatis and / or its extracts onto the area to be controlled, or spraying the composition as described in [2] onto the area to be controlled.

[0035] The effects of the invention

[0036] This invention reports for the first time a novel strain of the species *Pantoea ananatis*, *Pantoea_YN60*, which exhibits excellent control effects against Zika virus and dengue virus. The discovery of this strain enriches the resources of beneficial microorganisms, and it possesses the advantages of stable, efficient, and environmentally friendly inhibition of mosquito-borne flaviviruses. Furthermore, this invention proposes a method for controlling insect-borne viruses by placing plants containing bacteria, which shows great promise for applications in the prevention and control of mosquito-borne flavivirus infectious diseases. Attached Figure Description

[0037] Figure 1 The growth curve of Pantoea_YN60 in brain heart infusion liquid medium.

[0038] Figure 2 The inhibitory effect of Pantoea_YN60 on dengue virus-infected Aedes albopictus mosquitoes.

[0039] Figure 3 The inhibitory effect of Pantoea_YN60 on Zika virus-infected Aedes albopictus mosquitoes.

[0040] Figure 4 The inhibitory effect of Pantoea_YN60 on dengue virus infection in Aedes aegypti mosquitoes.

[0041] Figure 5 The inhibitory effect of Pantoea_YN60 on Zika virus-infected Aedes aegypti mosquitoes is shown.

[0042] Figure 6 The inhibitory effect of the O antigen of Pantoea_YN60 on dengue virus infection in Aedes albopictus mosquitoes.

[0043] Figure 7 The inhibitory effect of the O antigen of Pantoea_YN60 on Zika virus-infected Aedes albopictus mosquitoes.

[0044] Figure 8 The inhibitory effect of the O antigen of Pantoea_YN60 on dengue virus infection of Aedes aegypti mosquitoes.

[0045] Figure 9 The inhibitory effect of the O antigen of Pantoea_YN60 on Zika virus-infected Aedes aegypti mosquitoes was investigated.

[0046] Figure 10 To investigate the inhibitory effect of placing Pantoea_YN60-containing daisies on dengue virus-infected Aedes albopictus mosquitoes.

[0047] Figure 11 To investigate the inhibitory effect of placing Pantoea_YN60-containing daisies on Zika virus-infected Aedes albopictus mosquitoes.

[0048] Figure 12To investigate the inhibitory effect of placing Pantoea_YN60-containing daisies on dengue virus-infected Aedes aegypti mosquitoes.

[0049] Figure 13 To investigate the inhibitory effect of placing Pantoea_YN60-containing daisies on Zika virus-infected Aedes aegypti mosquitoes. Detailed Implementation

[0050] Various exemplary embodiments, features, and aspects of the present invention will be described in detail below. The term "exemplary" as used herein means "serving as an example, embodiment, or illustration." Any embodiment described herein as "exemplary" is not necessarily to be construed as superior to or better than other embodiments.

[0051] Furthermore, to better illustrate the present invention, numerous specific details are set forth in the following detailed embodiments. Those skilled in the art should understand that the present invention can be practiced without certain specific details. In other instances, methods, means, apparatus, and steps well known to those skilled in the art have not been described in detail in order to highlight the spirit of the present invention.

[0052] Unless otherwise stated, all units used in this specification are international standard units, and all numerical values ​​and ranges appearing in this invention should be understood to include systematic errors that are unavoidable in industrial production.

[0053] In this specification, the word "may" has two meanings: to perform a certain process and not to perform a certain process.

[0054] In this specification, references to "some specific / preferred embodiments," "other specific / preferred embodiments," "implementation," etc., refer to specific elements (e.g., features, structures, properties, and / or characteristics) related to that embodiment, which are included in at least one of the embodiments described herein and may or may not be present in other embodiments. Furthermore, it should be understood that these elements may be combined in any suitable manner in various embodiments.

[0055] In this specification, the range of values ​​referred to as "value A to value B" refers to the range including the endpoint values ​​A and B.

[0056] In this specification, "optional" or "optionally" means that the event or situation described below may or may not occur, and the description includes both the scenario in which the event occurs and the scenario in which the event does not occur.

[0057] The terms "comprising" and "having," and any variations thereof, are intended to cover non-exclusive inclusion. For example, a process, method, apparatus, product, or device that includes a series of steps is not limited to the steps or modules listed, but may optionally include steps not listed, or may optionally include other steps inherent to such process, method, product, or device.

[0058] In this invention, "multiple" refers to two or more. "And / or" describes the relationship between related objects, indicating that three relationships can exist. For example, A and / or B can represent: A alone, A and B simultaneously, or B alone. The character " / " generally indicates that the preceding and following related objects have an "or" relationship.

[0059] In this specification, the interchangeable terms “polynucleotide” and “nucleic acid” refer to polymeric forms of nucleotides (ribonucleotides or deoxyribonucleotides) of any length. Therefore, this term includes, but is not limited to, single-stranded, double-stranded, or multi-stranded DNA or RNA, genomic DNA, cDNA, DNA-RNA hybrids, or polymers containing purine and pyrimidine bases or other natural, chemically or biochemically modified, non-natural, or derived nucleotide bases.

[0060] In this invention, the terms “peptide,” “polypeptide,” and “protein” are used interchangeably herein and refer to a polymeric form of amino acids of any length, which may include encoded and non-coded amino acids, chemically or biochemically modified or derived amino acids, and polypeptides having a modified peptide backbone.

[0061] In this invention, the term "conservative amino acid substitution" refers to the interchangeability of amino acid residues with similar side chains in a protein. For example, a group of amino acids with aliphatic side chains consists of glycine, alanine, valine, leucine, and isoleucine; a group of amino acids with aliphatic-hydroxyl side chains consists of serine and threonine; a group of amino acids with amide-containing side chains consists of asparagine and glutamine; a group of amino acids with aromatic side chains consists of phenylalanine, tyrosine, and tryptophan; a group of amino acids with basic side chains consists of lysine, arginine, and histidine; a group of amino acids with acidic side chains consists of glutamic acid and aspartic acid; and a group of amino acids with sulfur-containing side chains consists of cysteine ​​and methionine. Exemplary conserved amino acid substituents are: valine-leucine-isoleucine, phenylalanine-tyrosine, lysine-arginine, alanine-valine-glycine, and asparagine-glutamine.

[0062] In this invention, a polynucleotide or polypeptide has a certain percentage of “sequence identity” with another polynucleotide or polypeptide, meaning that the percentages of bases or amino acids are the same when aligned, and that the bases or amino acids are in the same relative positions when the two sequences are compared. Sequence identity can be determined in many different ways. To determine sequence identity, various convenient methods and computer programs (e.g., BLAST, T-COFFEE, MUSCLE, MAFF T, etc.) can be used to align sequences, which are available on the World Wide Web at websites including ncbi.nlm.nili.gov / BLAST, ebi.ac.uk / Tools / msa / tcoffee / , ebi.ac.uk / Tools / msa / muscle / , and mafft.cbrc.jp / alignment / software / . See, for example, Alts Chul et al. (1990), J. Mol. Bioi. 215:403-10.

[0063] The technical solution of the present invention will be described in detail below.

[0064] (Pantoea ananatis)

[0065] In some aspects of the present invention, a novel strain of the genus *Pantoea*, specifically *Pantoea ananatis*, named Pantoea_YN60, is provided. The morphological characteristics of Pantoea_YN60 are as follows: Gram-negative, single colonies are round, pale yellow, smooth, and opaque.

[0066] The YN60 strain from *Pantoea ananatis* was deposited on July 22, 2026, at the China General Microbiological Culture Collection Center (CGMCC), classified as *Pantoea ananatis*; accession number: CGMCC No. 39790; address of the depository: No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing; Institute of Biology, Chinese Academy of Sciences; postal code: 100101. In this invention, the YN60 strain is also referred to as the *Pantoea_YN6* strain or *Pantoea ananatis* YN60, or simply *Pantoea_YN60*.

[0067] In some alternative embodiments, *Pantoea ananatis* is not limited to the above-described preserved strains, but may also be strains substantially similar in nature to the above-described preserved strains. A strain substantially similar in nature refers to a strain classified as *Pantoea ananatis*, and specifically, a strain that has at least the same degree of biological function as the strain preserved in this invention.

[0068] In some optional embodiments, *Pantoea ananatis* may comprise a strain having a 16S rRNA gene sequence having at least 97% homology, preferably at least 98% homology, more preferably at least 98.5%, at least 98.7%, at least 99%, at least 99.5%, or at least 99.9% homology to the 16S rRNA gene sequence of the strain deposited in this invention. The 16S rRNA gene sequence of the strain deposited in this invention is as follows (SEQ ID NO: 1):

[0069]

[0070] In some preferred embodiments, Pantoea ananatis contains the Pantoea_YN60 strain.

[0071] <Composition>

[0072] In some aspects of the invention, a composition is provided comprising Pantoea ananatis and / or its extracts as described above.

[0073] In some embodiments, the composition is a microbial preparation.

[0074] In some embodiments, the composition is used to control arboviruses.

[0075] In some embodiments, the Pantoea ananatis and / or its extracts are used as the active ingredients of the composition.

[0076] In some more specific implementations, examples of arboviruses include African Swine Fever virus, Tick-borne Encephalitis virus, Rift Valley Fever virus, Colorado Tick Fever virus, Equine Encephalosis virus, Chikungunya virus, Dengue virus (DV), Zika virus (ZV), and West Nile virus.

[0077] In some preferred embodiments, the arbovirus originates from the Flaviviridae family and the Flavivir genus.

[0078] In some exemplary embodiments, the present invention provides a microbial preparation for controlling orthoflavin virus, containing the above-mentioned Pantoea ananatis and / or its extract as active ingredients.

[0079] In some preferred embodiments, the vector virus is dengue virus (such as dengue virus type 2, hereinafter referred to as DENV2) and / or Zika virus.

[0080] In some exemplary embodiments, the present invention provides a microbial preparation for the prevention and control of dengue virus, containing Pantoea ananatis and / or its extracts as active ingredients.

[0081] In some exemplary embodiments, the present invention provides a microbial preparation for the prevention and control of Zika virus, containing the above-mentioned Pantoea ananatis and / or its extract as active ingredients.

[0082] <Application and Methods of Pantoea ananatis and / or its Extracts>

[0083] In some aspects of the invention, any of the following (i) to (iii) applications of Pantoea ananatis and / or its extracts are provided:

[0084] (i) Application in the preparation of products for controlling the spread of arboviruses;

[0085] (ii) Application in the preparation of products that inhibit the susceptibility of vectors to arboviruses;

[0086] (iii) Application in the prevention and control of vector-borne viruses.

[0087] Pantoea ananatis and its compositions are described in the preceding sections on Pantoea ananatis and Compositions.

[0088] In some implementation schemes, the control of vector-borne virus transmission, the inhibition of the susceptibility of the transmission vector to vector-borne viruses, and the prevention and control of vector-borne viruses are achieved by inhibiting the infection of the transmission vector by vector-borne viruses.

[0089] In some implementations, the inhibition of vector-borne virus infection includes reducing the viral load and infection rate of the vector-borne virus in the vector when the vector comes into contact with the vector-borne virus.

[0090] In some exemplary embodiments, the inhibition of vector-borne virus infection of the transmission vector includes reducing the viral load and infection rate of the vector-borne virus (e.g., dengue virus (such as dengue fever type 2 virus) and / or Zika virus) carried by the transmission vector when the transmission vector comes into contact with the vector-borne virus.

[0091] In some exemplary embodiments, an environment unfavorable to virus transmission is pre-established in the vector by using Pantoea ananatis and / or its extracts before or simultaneously with the vector's contact with the vector-borne virus.

[0092] In some exemplary embodiments, the infection rate of the virus to the vector is further reduced by using Pantoea ananatis and / or its extracts to colonize the midgut.

[0093] In some exemplary embodiments, compared to vectors not carrying the strain, vectors treated with Pantoea ananatis and / or its extracts exhibited significantly reduced viral load and infection rate upon contact with the arbovirus.

[0094] In some embodiments, the extract of Pantoea ananatis contains endotoxins extracted from Pantoea ananatis.

[0095] In this invention, "endotoxin" refers to the main component of the outer membrane of Gram-negative bacteria, usually referring to lipopolysaccharide (LPS) or its biologically active fragments, derivatives or modifications.

[0096] In some embodiments, the method for extracting LPS according to the present invention can be any known means capable of isolating lipopolysaccharide or its lipid A-containing fragment from Gram-negative bacteria, including but not limited to the phenol-water method, butanol method, Triton X-114 phase separation method, guanidine salt lysis method, or the use of commercial endotoxin extraction kits.

[0097] In some exemplary embodiments, the present invention uses an extraction kit to extract LPS, which yields LPS with high purity and structural integrity.

[0098] In some preferred embodiments, the Pantoea ananatis extract contains O-specific polysaccharides from Pantoea ananatis.

[0099] In this invention, "O-specific polysaccharide," also referred to herein as O antigen, refers to a polysaccharide chain at the distal end of a lipopolysaccharide (LPS) molecule, composed of multiple repeating oligosaccharide units. These repeating units typically contain 2–8 monosaccharide residues, which can be linearly linked by glycosidic bonds or form branched structures. Differences in the sugar composition, sequence, and linkage mode of O-specific polysaccharides determine the O serotype of Gram-negative bacteria.

[0100] In this invention, the method for extracting O-specific polysaccharide (O antigen) can be any known means capable of isolating O-specific polysaccharide chains, immunogenic fragments thereof, or molecular fragments containing one or more repeating oligosaccharide units from Gram-negative bacteria or their lipopolysaccharides, including but not limited to: releasing O antigen from purified LPS by chemical cleavage, such as acid hydrolysis, hydrazine hydrolysis, or alkaline degradation; directly extracting from bacterial cells, such as isolating and releasing polysaccharides after treating bacterial cells with trichloroacetic acid, acetic acid, or heat; specifically cleaving O antigen chains using bacteriophages or enzymes; or expressing O antigen gene clusters in a heterologous host and recovering the secreted polysaccharides through genetic engineering.

[0101] In some embodiments, the present invention hydrolyzes purified LPS with acetic acid solution at a temperature not exceeding 100°C to obtain O antigen in the supernatant of the hydrolysate.

[0102] In some exemplary embodiments, the present invention uses purified LPS hydrolyzed with 1% acetic acid at 100°C to obtain O antigen (supernatant).

[0103] In the above aspects of the invention, a fundamental characteristic of arboviruses is their replication within mammalian and insect cells. Many arboviruses are transmitted to mammals via mosquitoes, ticks, or sandflies. In some embodiments, arboviruses include one or more of the families Bunyaviridae, Flaviviridae, Reoviridae, and Togaviridae. In some specific embodiments, arboviruses include one or more of the genera Flavivirus, Alphavirus, and Orthobunyavirus.

[0104] In some more specific implementations, examples of arboviruses include African Swine Fever virus, Tick-borne Encephalitis virus, Rift Valley Fever virus, Colorado Tick Fever virus, Equine Encephalosis virus, Chikungunya virus, Dengue virus (DV), Zika virus (ZV), and West Nile virus.

[0105] In some preferred embodiments, the arbovirus originates from the Flaviviridae family and the Flavivirus genus. In some more preferred embodiments, the arbovirus is dengue virus (such as dengue type 2 virus) and / or Zika virus.

[0106] In the above aspects of the present invention, the vector for the vector-borne virus can be a mosquito, tick, or sandfly. In some embodiments, the vector for the vector-borne virus is a mosquito (a mosquito-like insect), and correspondingly, the vector-borne virus can be a mosquito-borne virus.

[0107] In some embodiments of the invention, "mosquito" or "mosquito" refers to insects of the family Culicidae. The mosquitoes of the invention may include adult mosquitoes, mosquito larvae, pupae, or their eggs. Typically, the mosquito life cycle comprises four distinct stages: egg, larva, pupa, and adult. Thus, the mosquito life cycle begins when eggs are placed on the surface of water (e.g., Culex, Culiseta, and Anopheles) or on damp, flooded soil (e.g., Aedes). Most eggs hatch into larvae within 48 hours. The larvae live in water, feeding on microorganisms and organic matter, and then surface to breathe. They molt four times, growing larger after each molt, and on the fourth molt, the larva becomes a pupa. The pupal stage is a resting stage, a non-feeding period of approximately two days. During this time, the mosquito reaches adulthood. When development is complete, the pupal skin ruptures and the adult mosquito emerges.

[0108] In some embodiments of the present invention, the mosquitoes belong to one or more of the subfamilies Anophelinae and Culicinae. In some specific embodiments of the present invention, the mosquitoes include one or more of Culex, Culiseta, Anopheles, and Aedes.

[0109] In some more specific embodiments of the present invention, the mosquitoes include, but are not limited to, Aedes, such as one or more of Aedes aegypti, Aedes albopictus, Aedes polynesiensis, Aedes australis, Aedes cinereus, Aedes rusticus, and Aedes vexans.

[0110] In some preferred embodiments of the invention, the mosquitoes are Aedes aegypti and / or Aedes albopictus.

[0111] In the foregoing aspects of the invention, the term “control” refers to reducing, inhibiting, regulating and / or blocking the progression and / or spread of pathogens / diseases, especially arboviruses / diseases caused by arbovirus infection.

[0112] In the foregoing aspects of the invention, "host" refers to a single-celled or multicellular organism in which an arbovirus can replicate, including cell lines and animals. In some embodiments of the invention, the host can be any mammal capable of serving as a host for an arbovirus, such as a primate, preferably a human. In other embodiments, the host can also be a rodent, such as a mouse or rat.

[0113] In the above aspects of the invention, the product may be a reagent or kit, as well as any other suitable form of product.

[0114] In some specific implementations, the reagent is a microbial preparation.

[0115] In some embodiments, the viable count of Pantoea ananatis and / or its extracts in the product (e.g., a microbial preparation, reagent, or kit) is not less than 1 × 10⁻⁶. 6 CFU (Colony-Forming Unitsper Gram).

[0116] In some aspects of the invention, any one of the following methods (a) to (c) is also provided:

[0117] (a) A method for controlling the spread of arboviruses, the method comprising applying Pantoea ananatis and / or its extracts, or a combination as described above, to a vector;

[0118] (b) A method for inhibiting the susceptibility of a vector to an arbovirus, the method comprising applying to the vector Pantoea ananatis and / or its extracts, or a composition as described above;

[0119] (c) A method for controlling vector-borne viruses, the method comprising spraying Pantoea ananatis and / or its extracts onto an area to be controlled, or spraying a composition as described above onto an area to be controlled, preferably, bringing the vector in the controlled area into contact with Pantoea ananatis and / or its extracts, or the composition as described above.

[0120] In some embodiments, the method for controlling the spread of vector-borne viruses, the method for inhibiting the susceptibility of vectors to vector-borne viruses, and / or the method for preventing and controlling vector-borne viruses includes colonizing the strain in the vector's intestines by feeding.

[0121] In some embodiments, the viable count of Pantoea ananatis and / or its extracts is 1 × 10⁻⁶ during feeding. 5 CFU -1×10 8 CFU.

[0122] In some exemplary embodiments, the method includes soaking sterile cotton balls in a 10% sucrose solution containing the strain, wherein the sucrose solution contains 1 × 10⁻⁶ strains. 6 CFU. Soaked cotton balls were placed in the container of the vector and fed as its sole food source for 7 days. This allowed the vector to ingest the bacterial strain by continuously consuming the bacterial sucrose solution, thereby achieving colonization of Pantoea_YN60 in the vector's midgut.

[0123] In some embodiments, the method for controlling the spread of vector-borne viruses, the method for inhibiting the susceptibility of vectors to vector-borne viruses, and / or the method for preventing and controlling vector-borne viruses includes introducing Pantoea ananatis and / or its extracts into plants, and placing the plants containing Pantoea ananatis and / or its extracts in the area to be controlled, so that vectors in the area to be controlled ingest Pantoea ananatis and / or its extracts by feeding on, probing, contacting, or dwelling on the plants; Pantoea ananatis and / or its extracts reduce the susceptibility of the vectors to vector-borne viruses within the vectors, thereby blocking or reducing the spread and diffusion of vector-borne viruses within the vector population.

[0124] In some preferred embodiments, the plants are common plants that naturally exist in the area to be controlled or are easily obtained and cultivated. These plants include, but are not limited to, common crops (such as rice, wheat, corn, cotton, vegetables, etc.), horticultural plants (such as roses, chrysanthemums, pothos, etc.), turfgrass, pasture grasses, common greening trees or shrubs, and naturally growing weeds in fields. These common plants are used by vectors to visit flowers, probe, feed on sap, or serve as habitats.

[0125] In some specific embodiments, Pantoea ananatis and / or its extracts are sprayed onto the leaves, stems, flowers, or fruits of the plant in the form of a live bacterial suspension or wettable powder. During normal foraging or resting, the vector ingests Pantoea ananatis and / or its extracts orally or through skin contact with the sprayed plant surface. Pantoea ananatis and / or its extracts then colonize the midgut of the vector.

[0126] In some embodiments, the Pantoea ananatis and / or its extracts can also be absorbed into the plant tissue through root irrigation or seed treatment, so that the plant containing the strain carries the active strain in whole or in part.

[0127] In some exemplary embodiments, the plant is Leucanthemum vulgare, and more specifically, Pantoea_YN60 is sprayed onto the flowers of the Leucanthemum vulgare using a spray bottle.

[0128] Example

[0129] The embodiments of the present invention will be described in detail below with reference to examples. However, those skilled in the art will understand that the following examples are for illustrative purposes only and should not be considered as limiting the scope of the invention. Unless otherwise specified in the examples, conventional conditions or conditions recommended by the manufacturer are followed. Reagents or instruments whose manufacturers are not specified are all commercially available conventional products.

[0130] Example 1: Isolation and Identification of Pantoea_YN60

[0131] Adult female Aedes albopictus mosquitoes were collected in Yiliang County, Yunnan Province using a mosquito trapping method. The mosquitoes were disinfected with 75% ethanol, rinsed twice with sterile phosphate buffered saline solution, and their midguts were dissected. The slurries were then homogenized using a grinder in 300 μL of sterile phosphate buffered saline solution. The homogenized solution was then diluted 10... -1 Up to 10 -4100 μL of the culture was then spread onto blood agar plates and incubated at 30°C for 24 h. After 24 h, newly grown bacteria were picked from the plates and transferred to brain and heart infusion liquid medium, which was then incubated at 30°C and 220 rpm in a shaker until turbidity was reached. Subsequently, the isolated bacteria were classified by whole-genome sequencing, average nucleotide identity (ANI), and digital DNA-DNA hybridization (dDDH) comparisons (Table 1), thus obtaining strain Pantoea_YN60. The 16S rRNA sequence of this strain is shown in SEQ ID NO: 1.

[0132] Table 1. Genomic relationships between Pantoea_YN60 and type strains of closely related species in the genus Pantoea.

[0133]

[0134] As shown in Table 1, the ANI and dDDH of the Pantoea_YN60 strain of the present invention are both greater than 90% compared with the known reference strain Pantoea ananatis, thus confirming that the Pantoea_YN60 strain of the present invention belongs to the new strain of the Pantoea ananatis species.

[0135] Pantoea_YN60 is Gram-negative. Single colonies of this bacterium on blood agar or brain and heart infusion agar are round, pale yellow, smooth, and opaque.

[0136] Example 2: Growth performance identification of Pantoea_YN60

[0137] The preserved strain Pantoea_YN60 was activated by streak plating onto blood agar plates and incubated at 30°C for 24 hours. After 24 hours, newly grown single colonies were picked from the plates and transferred to brain and heart extract liquid medium, which was then incubated at 30°C and 220 rpm on a shaker. 1 mL of bacterial culture was collected at 0, 6, 12, 24, 48, 72, and 96 hours after the start of incubation. The culture was then diluted 10⁻⁶ times. -1 Up to 10 -20 100 μL of the brain heart extract was then spread onto a plate and incubated at 30°C for 24 hours. Colony forming units (CFU) were then calculated based on the bacterial count on the plate to quantify the growth performance of Pantoea_YN60. Figure 1 As shown.

[0138] Example 3: Identification of the inhibitory effect of Pantoea_YN60 on dengue virus-infected Aedes albopictus (Jiangsu strain)

[0139] Soak sterile cotton balls in a 10% sucrose solution containing or without the bacteria Pantoea_YN60 (bacterial concentration of 1 × 10⁻⁶). 6 CFU. Then, using this as food for Aedes albopictus mosquitoes, they were fed for 7 days to colonize the midgut of Pantoea_YN60. Next, extracorporeal membrane feeding was performed. One day in advance, Aedes albopictus mosquitoes were placed in paper cups without food, and the cups were covered with damp paper towels to maintain moisture. Dengue virus type 2 (DENV2; New Guinea C strain; virus concentration 1 × 10⁻⁶) was added. 6 Plaque-forming units (PFU / ml) were mixed with human blood in an equal volume. The mixture was then added to the storage container of the extracorporeal membrane feeding system and placed on a paper cup. The Aedes albopictus mosquitoes in the cup began to feed. After 30 minutes, the storage container was removed and soaked in 10% 84 disinfectant. The Aedes albopictus mosquitoes were anesthetized at 4°C for 15 minutes. Then, the blood-feeding mosquitoes were transferred to a new mosquito cup and continued to be fed for 8 days. Finally, the mosquitoes were euthanized, and the DENV2 infection rate was detected by real-time quantitative PCR.

[0140] Real-time quantitative PCR detection method: Total RNA was extracted from homogenized mosquitoes using the Multisource RNA Miniprep Kit (AP-MN-MS-RNA-250; Axygen) and reverse transcribed into complementary DNA (cDNA) using the iScript cDNA Synthesis Kit (170-8890; Bio-Rad). Absolute quantification of the genome copies of DENV2 (DENV2-F: 5'-CATTCCAAGTGAGAATCTCTTTGTCA-3(SEQ ID NO: 2)'; DENV2-R: 5'-CAGATCTCTGATGAATAACCAACG-3(SEQ ID NO: 3)'; DENV2-probe: 5' FAM-ATGCTGAAACGCGAGAGAAACCGC-3' TAMRA(SEQ ID NO: 4)) and ZIKV (ZIKV-F: 5'-CATTCCAAGTGAGAATCTCTTTGTCA-3'(SEQ ID NO: 5); ZIKV-R: 5'-CCACTAACGTTCTTTTGCAGACAT-3'(SEQ ID NO: 6); ZIKV-probe: 5' FAM-AGCCTACCTTGACAAGCARTCAGACACTCAA3' TAMRA(SEQ ID NO: 7)) was performed by qPCR using primers and probes. The cycling conditions were as follows: 95°C for 2 minutes, followed by 40 cycles of 95°C for 10 seconds and 60°C for 30 seconds. To generate a standard curve, in vitro transcribed RNA was prepared using the MegaScript T7 kit (AM1333; Ambion) and purified using the MegaClear transcription cleanup kit (AMS908; Ambion). The purified RNA was quantified on a NanoDrop 2000 spectrophotometer (Thermo Fisher Scientific) to calculate the copy number. The viral genome copy number per mosquito was calculated according to the standard curve. The limit of detection (LOD) for this assay was determined to be 10³ genome copies. Samples below this threshold were assigned a baseline value of 1 copy to facilitate downstream logarithmic transformation.

[0141] The results showed that, compared with Aedes albopictus mosquitoes not colonized with Pantoea_YN60, the viral load and infection rate of DENV2 in Aedes albopictus mosquitoes colonized with Pantoea_YN60 were significantly reduced. Figure 2 As shown. Therefore, Pantoea_YN60 can inhibit dengue virus infection in Aedes albopictus mosquitoes.

[0142] Example 4: Identification of the inhibitory effect of Pantoea_YN60 on Zika virus-infected Aedes albopictus mosquitoes

[0143] Soak sterile cotton balls in a 10% sucrose solution containing or without the bacteria Pantoea_YN60 (bacterial concentration of 1 × 10⁻⁶). 6 CFU. Then, using this as food for Aedes albopictus mosquitoes, they were fed for 7 days to colonize the Pantoea_YN60 bacteria in the midgut. Next, extracorporeal membrane feeding was performed. One day in advance, Aedes albopictus mosquitoes were placed in paper cups without food, and the cups were covered with damp paper towels to maintain moisture. Zika virus (ZIKV; GZ01 strain; virus concentration 1 × 10⁻⁶) was added. 6 PFU / ml) was mixed with human blood in an equal volume and added to the storage container of the extracorporeal membrane feeding system. The container was then placed on a paper cup, where Aedes albopictus mosquitoes began to feed. After 30 minutes, the storage container was removed and soaked in 10% 84 disinfectant. The Aedes albopictus mosquitoes were anesthetized at 4°C for 15 minutes. Then, the blood-feeding mosquitoes were transferred to a new mosquito cup and continued to be fed for 8 days. Finally, the mosquitoes were euthanized, and the infection rate of ZIKV was detected by real-time quantitative PCR.

[0144] The results showed that, compared with Aedes albopictus mosquitoes not colonized with Pantoea_YN60, the viral load and infection rate of ZIKV in Aedes albopictus mosquitoes colonized with Pantoea_YN60 were significantly reduced. Figure 3 As shown. Therefore, Pantoea_YN60 can inhibit Zika virus infection in Aedes albopictus mosquitoes.

[0145] Example 5: Identification of the inhibitory effect of Pantoea_YN60 on dengue virus infection of Aedes aegypti mosquitoes (Rockefeller strain)

[0146] Soak sterile cotton balls in a 10% sucrose solution containing or without the bacteria Pantoea_YN60 (bacterial concentration of 1 × 10⁻⁶). 6 CFU. Then, this was used as food for Aedes aegypti mosquitoes, and they were fed for 7 days to colonize the Pantoea_YN60 bacteria in the midgut of Aedes aegypti. Next, extracorporeal membrane feeding (ECM) was performed. One day in advance, Aedes aegypti mosquitoes were placed in paper cups without food, and the cups were covered with damp paper towels to maintain moisture. DENV2 and human blood were mixed in equal volumes and added to the reservoir of the ECM feeding system, which was then placed on the paper cups. The Aedes aegypti mosquitoes in the paper cups began to feed. After 30 minutes, the reservoirs were removed and soaked in 10% 84 disinfectant. The Aedes aegypti mosquitoes were anesthetized at 4°C for 15 minutes, and then the blood-feeding mosquitoes were transferred to new mosquito cups for continued feeding for 8 days. Finally, the mosquitoes were euthanized, and the DENV2 infection rate was detected by real-time quantitative PCR.

[0147] The results showed that, compared with Aedes aegypti mosquitoes not colonized with Pantoea_YN60, the viral load and infection rate of DENV2 in Aedes aegypti mosquitoes colonized with Pantoea_YN60 were significantly reduced. Figure 4 As shown. Therefore, Pantoea_YN60 can inhibit dengue virus infection in Aedes aegypti mosquitoes.

[0148] Example 6: Identification of the inhibitory effect of Pantoea_YN60 on Zika virus-infected Aedes aegypti mosquitoes

[0149] Soak sterile cotton balls in a 10% sucrose solution containing or without the bacteria Pantoea_YN60 (bacterial concentration of 1 × 10⁻⁶). 6 The bacteria (Pantoea_YN60) were then used as food for Aedes aegypti mosquitoes for 7 days to colonize their midguts. Next, extracorporeal membrane feeding (ECM) was performed. One day prior, Aedes aegypti mosquitoes were placed in paper cups without food, covered with a damp paper towel for moisture retention. ZIKV and human blood were mixed in equal volumes and added to the storage container of the ECM feeding system, which was then placed on the paper cups. The mosquitoes in the cups began to feed. After 30 minutes, the storage container was removed and soaked in 10% 84 disinfectant. The mosquitoes were anesthetized at 4°C for 15 minutes, then the blood-feeding mosquitoes were transferred to new mosquito cups for another 8 days of feeding. Finally, the mosquitoes were euthanized, and the ZIKV infection rate was detected by real-time quantitative PCR.

[0150] The results showed that, compared with Aedes aegypti mosquitoes not colonized with Pantoea_YN60, the viral load and infection rate of ZIKV in Aedes aegypti mosquitoes colonized with Pantoea_YN60 were significantly reduced. Figure 5 As shown. Therefore, Pantoea_YN60 can inhibit Zika virus infection in Aedes aegypti mosquitoes.

[0151] Example 7: Identification of the inhibitory effect of Pantoea_YN60 O antigen on dengue virus infection in Aedes albopictus mosquitoes

[0152] LPS was extracted from Pantoea_YN60 using an LPS extraction kit (17141; iNtRON). The purified LPS was hydrolyzed with 1% acetic acid at 100°C to obtain the O antigen (supernatant). Residual solvent and acetic acid were removed by rotary evaporation, and the O antigen was resuspended in sterile water. One day in advance, Aedes albopictus mosquitoes were placed in paper cups without food, and the cups were covered with a damp paper towel for moisture retention. DENV2 was mixed with human blood in an equal volume, with or without the addition of 20 μg / mL purified O antigen. The mixture was then added to the reservoir of an in vitro feeding system and placed on the paper cups. The Aedes albopictus mosquitoes in the cups began to feed. After 30 minutes, the reservoirs were removed and immersed in 10% 84 disinfectant. The Aedes albopictus mosquitoes were anesthetized at 4°C for 15 minutes, and then the blood-feeding mosquitoes were transferred to new mosquito cups for continued feeding for 8 days. Finally, the mosquitoes were euthanized, and the DENV2 infection rate was detected by real-time quantitative PCR.

[0153] The results showed that, compared with Aedes albopictus mosquitoes not supplemented with O antigen, the viral load and infection rate of DENV2 in Aedes albopictus mosquitoes supplemented with O antigen were significantly reduced. Figure 6 As shown. Therefore, the O antigen of Pantoea_YN60 can inhibit dengue virus infection of Aedes albopictus mosquitoes.

[0154] Example 8: Identification of the inhibitory effect of Pantoea_YN60 O antigen on Zika virus-infected Aedes albopictus mosquitoes

[0155] LPS was extracted from Pantoea_YN60 using an LPS extraction kit (17141; iNtRON). The purified LPS was hydrolyzed with 1% acetic acid at 100°C to obtain the O antigen (supernatant). Residual solvent and acetic acid were removed by rotary evaporation, and the O antigen was resuspended in sterile water. One day prior to the experiment, Aedes albopictus mosquitoes were placed in paper cups without food, and the cups were covered with a damp paper towel for moisture retention. ZIKV was mixed with an equal volume of human blood, with or without the addition of 20 μg / mL of purified O antigen. The mixture was then added to the reservoir of an in vitro feeding system and placed on the paper cups. The Aedes albopictus mosquitoes in the cups began to feed. After 30 minutes, the reservoirs were removed and immersed in 10% 84 disinfectant. The Aedes albopictus mosquitoes were anesthetized at 4°C for 15 minutes, and then the blood-feeding mosquitoes were transferred to new mosquito cups for continued feeding for 8 days. Finally, the mosquitoes were euthanized, and the ZIKV infection rate was detected by real-time quantitative PCR.

[0156] The results showed that, compared with Aedes albopictus mosquitoes not supplemented with O antigen, the viral load and infection rate of ZIKV in Aedes albopictus mosquitoes supplemented with O antigen were significantly reduced. Figure 7 As shown. Therefore, the O antigen of Pantoea_YN60 can inhibit Zika virus infection of Aedes albopictus mosquitoes.

[0157] Example 9: Identification of the inhibitory effect of Pantoea_YN60 O antigen on dengue virus infection in Aedes aegypti mosquitoes

[0158] LPS was extracted from Pantoea_YN60 using an LPS extraction kit (17141; iNtRON). The purified LPS was hydrolyzed with 1% acetic acid at 100°C to obtain the O antigen (supernatant). Residual solvent and acetic acid were removed by rotary evaporation, and the O antigen was resuspended in sterile water. One day prior to the experiment, Aedes aegypti mosquitoes were placed in paper cups without food, and the cups were covered with a damp paper towel for moisture retention. DENV2 was mixed with an equal volume of human blood, with or without the addition of 20 μg / mL of purified O antigen. The mixture was then added to the reservoir of an in vitro feeding system and placed on the paper cups. The Aedes aegypti mosquitoes in the cups began feeding. After 30 minutes, the reservoirs were removed and immersed in 10% 84 disinfectant. The Aedes aegypti mosquitoes were anesthetized at 4°C for 15 minutes. Mosquitoes that had fed on blood were then transferred to new mosquito cups for further rearing for 8 days. Finally, the mosquitoes were euthanized, and the DENV2 infection rate was detected by real-time quantitative PCR.

[0159] The results showed that, compared with Aedes aegypti mosquitoes not supplemented with O antigen, the viral load and infection rate of DENV2 in Aedes aegypti mosquitoes supplemented with O antigen were significantly reduced. Figure 8 As shown. Therefore, the O antigen of Pantoea_YN60 can inhibit dengue virus infection of Aedes aegypti mosquitoes.

[0160] Example 10: Identification of the inhibitory effect of Pantoea_YN60 O antigen on Zika virus-infected Aedes aegypti mosquitoes

[0161] LPS was extracted from Pantoea_YN60 using an LPS extraction kit (17141; iNtRON). The purified LPS was hydrolyzed with 1% acetic acid at 100°C to obtain the O antigen (supernatant). Residual solvent and acetic acid were removed by rotary evaporation, and the O antigen was resuspended in sterile water. One day prior to the experiment, Aedes aegypti mosquitoes were placed in paper cups without food, and the cups were covered with a damp paper towel for moisture retention. ZIKV was mixed with an equal volume of human blood, with or without the addition of 20 μg / mL purified O antigen. The mixture was then added to the reservoir of an in vitro feeding system and placed on a paper cup. The Aedes aegypti mosquitoes in the cups began feeding. After 30 minutes, the reservoir was removed and immersed in 10% 84 disinfectant. The Aedes aegypti mosquitoes were anesthetized at 4°C for 15 minutes. Mosquitoes that had fed on blood were then transferred to new mosquito cups for further rearing for 8 days. Finally, the mosquitoes were euthanized, and the ZIKV infection rate was detected by real-time quantitative PCR.

[0162] The results showed that, compared with Aedes aegypti mosquitoes not supplemented with O antigen, the viral load and infection rate of ZIKV in Aedes aegypti mosquitoes supplemented with O antigen were significantly reduced. Figure 9 As shown. Therefore, the O antigen of Pantoea_YN60 can inhibit Aedes aegypti virus infection in mosquitoes.

[0163] Example 11: Identification of the inhibitory effect of placing Pantoea_YN60-containing daisies on dengue virus-infected Aedes albopictus mosquitoes.

[0164] Use a spray bottle to apply Pantoea_YN60 (at a concentration of 1 × 10⁻⁶). 7 (CFU / mL) was sprayed onto the flowers of *Panax chinensis*, and then the *Panax chinensis* were placed in mosquito cages containing *Aedes albopictus*, allowing the mosquitoes to feed on the nectar containing *Panax chinensis*_YN60 and obtain the bacteria into their intestines. Five days later, *Aedes albopictus* were collected into paper cups for external membrane feeding. DENV2 was mixed with human blood in equal volumes, and the mixture was added to the storage container of the external membrane feeding system and placed on a paper cup. The *Aedes albopictus* in the paper cup began to feed. After 30 minutes, the storage container was removed and soaked in 10% 84 disinfectant. The *Aedes albopictus* were anesthetized at 4°C for 15 minutes, and then the blood-feeding mosquitoes were transferred to new mosquito cups for continued feeding for 8 days. Finally, the mosquitoes were euthanized, and the DENV2 infection rate was detected by real-time quantitative PCR.

[0165] The results showed that, compared with Aedes albopictus mosquitoes that fed on daisies without Pantoea_YN60, the viral load and infection rate of DENV2 in Aedes albopictus mosquitoes that fed on daisies containing Pantoea_YN60 were significantly lower. Figure 11 As shown. Therefore, placing sea daisies containing Pantoea_YN60 can inhibit dengue virus infection of Aedes albopictus mosquitoes.

[0166] Example 12: Identification of the inhibitory effect of placing Pantoea_YN60-containing daisies on Zika virus-infected Aedes albopictus mosquitoes

[0167] Pantoea_YN60 was sprayed onto the flowers of *Aedes albopictus* using a spray bottle. The *Aedes albopictus* were then placed in a mosquito cage containing *Aedes albopictus* mosquitoes, allowing the mosquitoes to feed on the nectar containing Pantoea_YN60 and acquire the bacteria for intestinal feeding. Five days later, *Aedes albopictus* mosquitoes were collected into paper cups for external membrane feeding. ZIKV was mixed with human blood in equal volumes and added to the storage container of the external membrane feeding system. The container was then placed on a paper cup, where *Aedes albopictus* mosquitoes began feeding. After 30 minutes, the storage container was removed and soaked in 10% 84 disinfectant. *Aedes albopictus* mosquitoes were anesthetized at 4°C for 15 minutes. Mosquitoes that had fed on blood were then transferred to new mosquito cups for continued feeding for 8 days. Finally, the mosquitoes were euthanized, and the ZIKV infection rate was detected by real-time quantitative PCR.

[0168] The results showed that, compared with Aedes albopictus mosquitoes that fed on daisies without Pantoea_YN60, the viral load and infection rate of ZIKV in Aedes albopictus mosquitoes that fed on daisies containing Pantoea_YN60 were significantly lower. Figure 11 As shown. Therefore, placing sea daisies containing Pantoea_YN60 can inhibit Zika virus infection of Aedes albopictus mosquitoes.

[0169] Example 13: Identification of the inhibitory effect of placing Pantoea_YN60-containing daisies on dengue virus-infected Aedes aegypti mosquitoes.

[0170] Pantoea_YN60 was sprayed onto the flowers of *Aedes aegypti* using a spray bottle. The *Aedes aegypti* were then placed in a mosquito cage containing *Aedes aegypti* mosquitoes, allowing the mosquitoes to feed on the nectar containing Pantoea_YN60 and acquire the bacteria for intestinal feeding. Five days later, *Aedes aegypti* mosquitoes were collected into paper cups for extracorporeal membrane feeding (ECMF) blood feeding. DENV2 was mixed with an equal volume of human blood and added to the storage container of the ECMF system, which was then placed on a paper cup. The *Aedes aegypti* mosquitoes in the cup began feeding. After 30 minutes, the storage container was removed and soaked in 10% 84 disinfectant. The *Aedes aegypti* mosquitoes were anesthetized at 4°C for 15 minutes. Then, the blood-feeding mosquitoes were transferred to new mosquito cups for continued feeding for 8 days. Finally, the mosquitoes were euthanized, and the DENV2 infection rate was detected by real-time quantitative PCR.

[0171] The results showed that, compared with Aedes aegypti mosquitoes that fed on daisies without Pantoea_YN60, the viral load and infection rate of DENV2 in Aedes aegypti mosquitoes that fed on daisies containing Pantoea_YN60 were significantly lower. Figure 12 As shown. Therefore, placing sea daisies containing Pantoea_YN60 can inhibit dengue virus infection of Aedes aegypti mosquitoes.

[0172] Example 14: Identification of the inhibitory effect of placing Pantoea_YN60-containing daisies on Zika virus-infected Aedes aegypti mosquitoes.

[0173] Pantoea_YN60 was sprayed onto the flowers of *Aedes aegypti* using a spray bottle. The *Aedes aegypti* were then placed in a mosquito cage containing *Aedes aegypti* mosquitoes, allowing the mosquitoes to feed on the nectar containing Pantoea_YN60 and acquire the bacteria into their intestines. Five days later, *Aedes aegypti* mosquitoes were collected into paper cups for external membrane feeding. ZIKV was mixed with human blood in equal volumes and added to the storage container of the external membrane feeding system. The container was then placed on a paper cup, where *Aedes aegypti* mosquitoes began feeding. After 30 minutes, the storage container was removed and soaked in 10% 84 disinfectant. The *Aedes aegypti* mosquitoes were anesthetized at 4°C for 15 minutes. Mosquitoes that had fed on blood were then transferred to new mosquito cups and fed for another 8 days. Finally, the mosquitoes were euthanized, and the ZIKV infection rate was detected by real-time quantitative PCR.

[0174] The results showed that, compared with Aedes aegypti mosquitoes that fed on daisies without Pantoea_YN60, the viral load and infection rate of ZIKV in Aedes aegypti mosquitoes that fed on daisies containing Pantoea_YN60 were significantly reduced. Figure 13 As shown. Therefore, placing sea daisies containing Pantoea_YN60 can inhibit Zika virus infection of Aedes aegypti mosquitoes.

[0175] The above results indicate that Pantoea_YN60 has a significant ability to inhibit mosquito-borne flavivirus, Zika virus, and dengue virus infections, making it a broad-spectrum and highly effective biocontrol strain. Pantoea_YN60 can be used for the control of mosquito-borne flavivirus, Zika virus, and dengue virus by direct spraying onto plants.

[0176] It should be noted that although the technical solution of the present invention has been described with specific examples, those skilled in the art will understand that the present invention should not be limited thereto.

[0177] The various embodiments of the present invention have been described above. These descriptions are exemplary and not exhaustive, nor are they limited to the disclosed embodiments. Many modifications and variations will be apparent to those skilled in the art without departing from the scope and spirit of the described embodiments. The terminology used herein is chosen to best explain the principles, practical application, or technical improvements to the embodiments in the market, or to enable others skilled in the art to understand the embodiments disclosed herein.

Claims

1. A strain of bacteria, with accession number CGMCC No. 39790.

2. A composition comprising the strain as described in claim 1 and / or its extract; Optionally, the composition is a microbial preparation; Optionally, the composition is used for the prevention and control of arboviruses; Optionally, the strain and / or its extract are used as the active ingredient of the composition.

3. Any of the following applications (i) to (iii) of Pantoea ananatis and / or its extracts: (i) Application in the preparation of products for controlling the spread of arboviruses; (ii) Application in the preparation of products that inhibit the susceptibility of vectors to arboviruses; (iii) Application in the prevention and control of vector-borne viruses.

4. The application according to claim 3, wherein, The Pantoea ananatis strain includes strains with a 16S rRNA gene sequence that are at least 97% homologous to the sequence shown in SEQ ID NO: 1; Preferably, the Pantoea ananatis comprises a strain whose 16S rRNA gene sequence includes the sequence shown in SEQ ID NO: 1; More preferably, the Pantoea ananatis includes the Pantoea_YN60 strain, whose accession number is CGMCC No.39790.

5. The application according to claim 3 or 4, wherein, The extract of Pantoea ananatis contains endotoxins produced by Pantoea ananatis; Preferably, the extract of Pantoea ananatis contains O-specific polysaccharides of Pantoea ananatis.

6. The application according to any one of claims 3 to 5, wherein, The arboviruses include one or more of the following families: Bunyaviridae, Flaviviridae, Reoviridae, and Togaviridae; Preferably, the arbovirus includes one or more of the genera Flavivirus, Alphavirus, and Orthobunyavirus.

7. The application according to any one of claims 3 to 6, wherein, The arbovirus is a flavivirus; Optionally, the vector-borne viruses include dengue virus and / or Zika virus.

8. The application according to any one of claims 3 to 7, wherein, The vectors of the arboviruses include one or more of mosquitoes, ticks, or sandflies; Preferably, the vector for the vector-borne virus is a mosquito; More preferably, the mosquitoes include one or more of Culex, Culiseta, Anopheles, and Aedes; Optionally, the mosquito is an Aedes mosquito, including one or more of Aedes aegypti, Aedes albopictus, Aedes polynesiensis, Aedes australis, Aedes cinereus, Aedes rusticus, and Aedes vexans.

9. The application according to any one of claims 3 to 8, wherein, The product includes one or more reagents or kits; optionally, the reagent is a microbial preparation.

10. Any of the methods described in (a) to (c) below: (a) A method for controlling the spread of arboviruses, the method comprising applying Pantoea ananatis and / or its extracts, or the composition as described in claim 2, to a vector; (b) A method for inhibiting the susceptibility of a vector to an arbovirus, the method comprising applying to the vector Pantoea ananatis and / or an extract thereof, or the composition as described in claim 2; (c) A method for controlling arboviruses, the method comprising spraying Pantoea ananatis and / or its extracts onto an area to be controlled, or spraying the composition of claim 2 onto an area to be controlled.

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