WO2019027234A1 - Markers for diagnosing acinetobacter baumannii infection, and use thereof - Google Patents

Abstract

The present invention relates to markers for diagnosing Acinetobacter baumannii infection, and a use thereof. Markers of six proteins or genes coding for the proteins, provided by the present invention, enable preferential detection of a multidrug-resistant Acinetobacter baumannii infection and provide faster and more accurate test results than those from a conventional blood culture diagnosis method (bacterial culture diagnosis method) and the like, thereby being highly advantageous for rapid response in the early stages of infectious diseases, of multidrug-resistant Acinetobacter baumannii, such as septicemia.

Description

 【Specification】

Title: Marker for diagnosis of infection with Aspergillus baumannii and its use

The present application claims priority from Korean Patent Application No. 10-2017-0097726 filed on August 1, 2017, the entire contents of which are incorporated herein by reference.

The present invention relates to a marker for diagnosis of an Acinetobacter baumannii infection and a use thereof, and more particularly, to a marker for the diagnosis of an Acinetobacter baumannii infection and more particularly to a marker for beta-lactamase class D OXA-23, a carbapenem-associated resistance protein precursor Carbapenem- resistance associated protein precursor), TonB_dependent siderophore (fore receptor protein ^"in TonB side receptor protein), ompE (Outer membrane protein E), ompW (0uter membrane protein W) and beta-lactase Maze class C 73-Adc (beta- (A: //? eii? ce / ^" baumannii) infection of a protein of the lactamase class C Adc-73 or a gene encoding the protein.

Background of the Invention [0002] The bacteria in Ashitto Bacteria are gram-negative bacilli and are non-fermenting bacteria of glucose, and they are distributed as supercritical bacteria in natural environment including soil, water and sewage, human skin, oral cavity and respiratory mucosa. These bacteria are a major cause of endogenous opportunistic infections in immunocompromised high-risk groups such as neonates, spleen-treated patients, long-term steroid-treated patients, cancer patients undergoing chemotherapy, organ transplant patients, and intensive care unit patients (Patrick PR, et al. < RTI ID = 0.0 > 5. < / RTI > Acinetobacter baumanni C4c // 7e o acer baumannii is a major nosocomial pathogen, which is known to cause a variety of infections, mainly pneumonia and bacteremia. (Bergogne-Berezin and Towner, CI in Microbiol Rev. 1996: 148-65; Kolenti and Rello, Expert Op. Pharmacol., 2006 Aug; 7 (12): 1555-69). They not only represent a variety of antibiotic resistance During the treatment period, new resistance to antibiotics develops. Infection by bacteria with this resistance has a higher mortality rate because there are very few or no classes of antibiotics to choose from for treatment.

At present, the diagnosis of AcoBactor Vaumani depends on the diagnosis of bacterial culture. However, since this method takes a long time to produce the results, it is necessary to determine the therapeutic drugs and methods depending on the experience of the medical staff There is a problem in that the treatment failure rate due to this is considerable. Therefore, in order to promptly respond to the early stage of infection and appropriate treatment, it is necessary to develop the rapid diagnosis technology of Asnate thoracic buumani. In order to develop such a rapid diagnosis technology, it is necessary to closely examine the characteristics of the organism to be detected, and to select the organism which is specific to the target organism and easy to be detected. In addition, since the screening of the above-mentioned indicator material can be practically effective in view of the clinical characteristics of the patient infected with the infectious agent, setting the indicator material capable of accurately and quickly obtaining the diagnosis result is substantially time- Cost, and effort.

DETAILED DESCRIPTION OF THE INVENTION

 SUMMARY OF THE INVENTION Accordingly, the present inventors have made intensive studies to develop a rapid diagnostic technique for Asnatech Bauterni, particularly, a multidrug-resistant Acinetobacter baumanni. Among the six proteins provided by the present invention and a gene encoding the protein, The present inventors confirmed that they are useful as markers because they are highly specific to the desired detection characteristics when they are used as markers and are closely related to the clinical characteristics of infection of the bacteria.

baumannii) 감염 진단용 조성물을 제공하는 것이다. Accordingly, it is an object of the present invention to provide a method for the treatment and / or prophylaxis of a disease or condition selected from the group consisting of Beta-lactamase class D OXA-23 (Beta-lactamase class D OXA-23), Carbapenem-associated resistance protein precursor, Protein is selected from the group consisting of TonB_dependent siderophore receptor protein, ompE (Outer membrane protein E), omp (0uter membrane protein W) and beta-lactamase class C Adc-73 (Beta-lactamase class C Adc-73) One or more proteins; Or A gene encoding one or more genes encoding a protein family,

baumannii) infection.

It is another object of the present invention to provide a composition for diagnosing an Acquired Tomato Vaumani infection comprising an agent for detecting the protein or gene and an Ashkenobarbaumani infection diagnostic kit comprising the composition.

It is still another object of the present invention to provide a method for diagnosing an Ashtonobacter baumannii infection comprising the step of detecting the protein or gene from a biological sample separated from a specimen.

baumannii) 감염 진단용 제제를 제조하기 위한， 베타 -락타메이즈 클래스 D 0XA-23(Beta- lact mase class D OXA-23), 카바페넴 -연관 저항성 단백질 전구체 (Carbapenem- associated resistance protein precursor) , TonB 사이드로포어 수용체 단백질 (TonB_dependent siderophore receptor protein) , ompE (Outer membrane protein E) , ompW(0uter membrane protein W) 및 베타 -락타메이즈 클래스 C Adc-73 (Bet a- lactamase class C Adc-73)로 이루어지는 단백질군에서 선택되는 하나 이상의 단백질; 또는 상기 단백질군을 코딩하는 유전자로 이루어진 군에서 선택되는 하나 이상의 유전자의 용도를 제공하는 것이다. It is another object of the present invention to provide a method for controlling an object,

(Beta-lactase class D OXA-23), carbapenem-associated resistance protein precursor, TonB side-by-side A protein group consisting of TonB_dependent siderophore receptor protein, ompE (Outer membrane protein E), ompW (0uter membrane protein W) and Beta-lactamase class C Adc-73 (Beta-lactamase class C Adc-73) ≪ / RTI > Or a gene coding for the above-mentioned protein group.

A further object of the present invention is to provide a method of detecting a) beta-lactamase class D OXA-23, a carbapenem-associated resistance protein precursor ), TonB-dependent siderophore receptor protein, ompE (Outer membrane protein E), ompW (Outer membrane protein W) and Beta lactamase class C Adc-73 (Bet a-lactamase class C Adc -73); < / RTI &gt; Or a gene coding for the protein group; b) comparing the level of the detected protein or gene with a control; And c) providing Acinetobacter ^{^^ v \ ^ <j 'netobacter} baumaimii) infection comprising the step of the level of the detected protein or gene is determined to be infected with the control level is higher than Acinetobacter baumannii will be.

[Technical Solution] In order to achieve the above object, the present invention provides a method for producing a carbapenem-associated resistance protein (D-XA-23), a beta-lactamase class D OXA- precursor, TonB side-dependent siderophore receptor protein, ompE (Outer membrane protein E), ompW (Outer membrane protein W) and Beta lactamase class C Adc-73 Adc-73); Or a gene coding for the protein group. The present invention also provides a composition for diagnosing infection of Aspergillus baumannii.

In order to accomplish still another object of the present invention, the present invention provides a composition for diagnosing infection with Aspergillus baumannii comprising the protein or gene, and an assay kit for Aspergillus baumannii infection comprising the composition.

In order to accomplish still another object of the present invention, the present invention provides a method for detecting the protein or gene from a biological sample separated from a specimen, in order to provide information necessary for diagnosis of Ashton burberani.

In order to accomplish still another object of the present invention, the present invention provides a method for producing an agent for diagnosing infection with Aspergillus baumannii (Beta-lactamase) class D OXA-23), carbapenem-associated resistance protein precursor, TonB-dependent siderophore receptor protein, ompE (Outer membrane protein E), ompW (Outer membrane protein W) and beta- One or more proteins selected from the group consisting of Lactamase class C Adc-73 (Beta-lactamase class C Adc-73); Or a gene coding for the above-mentioned protein group.

In order to accomplish still another object of the present invention, the present invention provides a kit for detecting a) a beta-lactamase class D OXA-23 from a biological sample isolated from a specimen, a carbapenem-associated resistance protein (OmpE), ompW (Outer membrane protein W), and beta-lactamase class C Adc-73 &lt; RTI ID = 0.0 &gt; (Beta-lactamase class C Adc-73); Or a gene coding for the protein group; b) comparing the level of the detected protein or gene with a control; And c) determining that the level of the detected protein or gene is higher than the level of the control, determining that the virus is infected with Asnithobacter baumannii.

Hereinafter, the present invention will be described in detail.

In the present invention, 'expression' means that a protein or a nucleic acid is produced in a cell. A "protein" is a "polypeptide" or

Is used interchangeably with a 'peptide', for example, a polymer of amino acid residues as commonly found in proteins in nature. A "polynucleotide" or "nucleic acid" refers to deoxyribonucleotides (DNA) or ribonucleotides (RNA) in the form of single- or double-stranded. Unless otherwise constrained, also includes known analogs of natural nucleotides that are naturally encoded in the nucleic acid in a manner similar to naturally occurring nucleotides. 'mRNA' is RNA that transfers genetic information (gene-specific nucleotide sequence) to a ribosome in which amino acid sequence is specified from a specific gene during protein synthesis. The term " marker " or " biomarker " in the present invention means a substance capable of detecting Acinetobacter baumannii from other bacteria and detecting the polypeptide or a polypeptide which is specifically present only in Ashtonobacter baumannii Such as nucleic acids (e.g., mRNA), lipids, glycolipids, glycoproteins or sugars (monosaccharides, disaccharides, oligosaccharides, etc.). Also, preferably, the role of the marker in the present invention can be understood as a substance that can be diagnosed by distinguishing antibiotic-resistant Asnithobacter baumanni from antibiotic-susceptible Asnithobacter baumanni, and antibiotic resistant bacteria And include organic biomolecules such as polypeptides or nucleic acids (such as mRNA), lipids, glycolipids, glycoproteins or sugars (monosaccharides, disaccharides, sugars and the like) which show an increase or decrease in bacteria.

In the present invention, the term " diagnosis " refers to identifying the presence or characteristic of a pathological condition. In the present invention, the diagnosis may be to ascertain the presence of an Acinetobacter baumanni infection, preferably an antibiotic resistant Acinetobacter baumanni, especially a multidrug-resistant Acinetobacter baumannii, MRAB) infection. Also most preferably, in the present invention, the diagnosis can be understood as ascertaining whether Ashinobacterium mania has acquired resistance to certain antibiotics.

The inventors of the present invention have found that the detection of the six genes described below and the proteins expressed therefrom, such as Asnithobacter baumannii (in particular, Escherichia coli bacterium having antibiotic resistance), or the use of the bacteria as an infection diagnostic marker Respectively.

Accordingly, the present invention is directed to a pharmaceutical composition comprising a beta-lactamase class D OXA-23, a carbapenem-associated resistance protein precursor, (OTR), ompW (Outer membrane protein W), and beta-lactamase class C Adc-73 (Beta-actamase clad C Ad-73) One or more proteins selected from the group consisting of proteins; Or a gene coding for the protein group. The present invention also provides a composition for diagnosing infection with Aspergillus baumannii C4c. Ec. Bac er baumannii.

The beta-lactamase class D OXA-23 protein is not particularly limited as long as it is known in the art that A. baumannii expresses the protein, The Beta-lactamase class D 0XA-23 of the present invention comprises SEQ ID NO: Genbank accession no. A0X75888), or may be one comprising the amino acid sequence of SEQ ID NO: 1. The gene coding for the beta-lactamase class D OXA-23 protein is not particularly limited as long as it is known to code the protein, 1 may be a gene sequence encoding the amino acid sequence of SEQ ID NO: 1, and more preferably, the gene encoding the amino acid sequence of SEQ ID NO: 1 may be a nucleotide sequence of SEQ ID NO:

The carbapenem-associated resistance protein precursor is not particularly limited as long as it is known to express A. baumannii i7 in the art, but preferably the carbapenem-associated resistance protein precursor The penem-associated resistance protein precursor may comprise the amino acid sequence of SEQ ID NO: 3 (Genbank accession no. A0X78535), or may comprise the amino acid sequence of SEQ ID NO: 3. The gene coding for the carbapenem-associated resistance protein precursor is not particularly limited as long as it is known that the protein encodes the protein. Preferably, the gene coding for the carbapenem-associated resistance protein precursor The gene encoding the amino acid sequence of SEQ ID NO: 3 may be the gene sequence encoding the amino acid sequence, and more preferably, the gene encoding the amino acid sequence of SEQ ID NO: 3 may be the nucleotide sequence of SEQ ID NO: The TonB side-by-side receptor protein (TonB-dependent siderophore receptor protein) is described in the art.

The specific amino acid sequence is not particularly limited as long as it is known to be expressed, but preferably the TonB side-by-side receptor protein of the present invention comprises the amino acid sequence of SEQ ID NO: 5 (Genbank accession no. A0X77361) And may be composed of the amino acid sequence of SEQ ID NO: 5. The gene coding for the TonB-dependent siderophore receptor protein is not particularly limited as long as it is known to encode the protein, preferably the amino acid sequence of SEQ ID NO: 5 And more preferably the gene encoding the amino acid sequence of SEQ ID NO: 5 may be a nucleotide sequence of SEQ ID NO:

Although the specific amino acid sequence of ompE (Outer membrane protein E) is known to express A baumanni P in the art, ompE of the present invention is preferably an amino acid sequence of SEQ ID NO: 7 (Genbank accession no. A0X78734) Or may comprise the amino acid sequence of SEQ ID NO: 7. . The gene coding for the ompE (Outer membrane, protein E) may be a gene sequence encoding the amino acid sequence of SEQ ID NO: 7 without particular limitation on the specific gene sequence as long as it is known to encode the protein, More preferably, the gene encoding the amino acid sequence of SEQ ID NO: 7 may be a nucleotide sequence of SEQ ID NO: 8.

Although the specific amino acid sequence of ompW (Outer membrane protein W) is known to express A. baumannii in the art, ompW of the present invention is preferably an amino acid sequence of SEQ ID NO: 9 (Genbank accession No. AB010762) Or may consist of the amino acid sequence of SEQ ID NO: 9. The gene coding for ompW (Outer membrane protein W) may be a gene sequence coding for the amino acid sequence of SEQ ID NO: 9, and the specific gene sequence is not particularly limited as long as it is known to code the protein. More preferably, the gene encoding the amino acid sequence of SEQ ID NO: 9 may be a nucleotide sequence of SEQ ID NO: 10.

The specific amino acid sequence of the beta-lactamase class C Adc-73 (Beta-lactamase class C Adc-73) known to express A b mianniA in the art is not particularly limited. Preferably, the beta-lactamase class C Adc- Maize Class C Adc-73 is SEQ ID NO: lKGenbank accession no. A0X78355), or may be one comprising the amino acid sequence of SEQ ID NO: 11.

 The gene encoding Beta-lactamase class C Adc-73 (Beta-lactamase class C Adc-73) is not particularly limited as long as it is known to code the protein, 11, and more preferably, the gene encoding the amino acid sequence of SEQ ID NO: 11 may be a nucleotide sequence of SEQ ID NO: 12.

The Ashtonobacter baumanni may preferably be an antibiotic resistant Ashton Burton Vaumani. The antibiotics are not particularly limited as long as they are known in the art as antibiotics. However, the antibiotics are not limited to the antibiotics, and examples thereof include beta-lactam, Cephalosporin, Carbapenem, Aminoglycoside, But is not limited to, one or more selected from the group consisting of fluoroquinolone series, tetracycline series, polymyxin series, and sulfonamide series antibiotics. The β-Lactam antibiotic is not particularly limited as long as it is known in the art, but it is preferable that the antibiotic is selected from the group consisting of Ampicillin, Sulbactam, Ticarcillin, Clavui lanic acid, Piperacillin, and Tazobactam. In one embodiment, have. Such cephalosporin antibiotics are not particularly limited as long as they are known in the art, but they are not limited to the types of cells such as Cefotaxime, Ceftazidime, Cefepime and Aztreonam And the like. The carbapenem antibiotic may be one or more selected from the group consisting of Imipenem and Meropenem, although the type of the antibiotic is not particularly limited as long as it is known in the art. The aminoglycoside-based antibiotic may be one or more selected from the group consisting of amikacin and gentamicin, although the type of the antibiotic is not particularly limited as long as it is known in the art. The type of the fluoroquinolone antibiotic is not particularly limited as long as it is known in the art, but it may be Ciprofloxacin. The tetracycline antibiotic may be one or more selected from the group consisting of minocycline and tigecycline, although the type of the antibiotic is not particularly limited as long as it is known in the art. The polymyxin antibiotic is not particularly limited as long as it is known in the art, but may be a colistin. The sulphonamide antibiotic is not particularly limited as long as it is known in the art, but may be one or more selected from the group consisting of trimesferram (Tr imethopyr im) and Sul famethoxazole.

The term &quot; multidrug-resistant &quot; in the present invention means that the organism exhibits resistance to two or more drugs having different pharmacological actions at the same time. The above-mentioned Ashtonobacter baumanni of the present invention is more preferably a multidrug- It can be Bertha Baumanni. The Ashtonobacter baumanni may be selected from the group consisting of β-lactam, Cephalosporin, Carbapenem, Aminoglycoside, Fluoroquinolone, Tetracyclin ine, polymyxin, and sulphonamide antibiotics. The antimicrobial agent of the present invention can be used in a variety of applications.

The six types of the present invention. Genes and proteins expressed therefrom are used as diagnostic markers for the infection with Acinetobacter baumannii, Examination of the expression status shows not only the presence or absence of the Acinetobacter baumannii but also the antibiotic resistance or multidrug resistance of the bacterium. This is well illustrated in the embodiments of the present invention.

In an embodiment of the present invention, the six markers of the present invention are specifically resistant to multidrug resistance through comparison with A. baumannii susceptible to antibiotics. baumannii could be detected.

Based on the novel diagnostic uses of the six genes and the proteins expressed therefrom, the present invention provides a novel diagnostic utility for beta-lactamase class D 0XA-23, carbapenem-associated resistance protein A carrier protein, a precursor (Carbapenem-assisted resident protein precursor), a TonB-dependent siderophore receptor protein, an ompE (Outer membrane protein E), an ompW (Outer membrane protein W) One or more proteins selected from the group consisting of Maize Class C Adc-73 (Beta-actamase clad C Ad-733); Or a gene coding for the protein group, and a kit for diagnosing an Ashtonobacter baumannii infection comprising the composition, which comprises the composition .

The kind of the ligand that specifically binds to the six proteins is not particularly limited, and may be, for example, a peptide, an antibody, or an aptamer having a binding domain specific to the protein But is not limited thereto.

As used herein, the term " antibody " means a specific immunoglobulin as indicated in the art and directed against an antigenic site. Any of those prepared through the above-mentioned one or more protein injections or commercially available can be used. In addition, the antibody includes a polyclonal antibody, a monoclonal antibody, a chimeric antibody, and a fragment capable of binding to an epitope. The term " peptide " refers to a polypeptide that does not have the structure of the intact antibody, but has a specific antigen binding site (binding domain) directed against the antigenic site. The peptide comprises a functional fragment of an antibody molecule that is not a complete form of the antibody having two light and two heavy chains. A functional fragment of an antibody molecule means a fragment having at least an antigen-binding function. The length of the peptide is not particularly limited, but may be, for example, 2 to 100 amino acids, preferably 5 to 50 amino acids.

The " aptamer " has binding activity for a given target molecule. Refers to oligonucleotide molecules. The aptamer may be RNA, DNA, modified nucleic acid or a fused product thereof, and may be in a linear or cyclic form.

The agent for detecting the gene encoding the six proteins may preferably be an agent for detecting the mRNA expressed from the gene, but is not limited thereto. Thus, the agent for detecting the gene encoding the six proteins of the present invention is not particularly limited as long as it is a ligand that specifically binds or hybridizes to the mRNA expressed from the gene, For example, a primer pair or a probe.

The " primer " is a nucleic acid sequence having a short free 3 'hydroxy 1 group, a short nucleic acid sequence capable of base pairing with a complementary template and serving as a starting point for template strand radiation . The primer can initiate DNA synthesis in the presence of reagents (i. E., DNA polymerase or reverse transcriptase) and four different nucleoside triphosphates for polymerization reactions at the appropriate complete solution and temperature. The PCR conditions, the lengths of the sense and antisense primers can be appropriately selected according to techniques known in the art.

The present invention relates to the aforementioned six proteins. A primer pair detecting the gene (mRNA) encoding the marker is provided as follows. The gene encoding Beta-lactamase class D OXA-23 (Beta-lactamase class D OXA-23) can be detected as a primer pair represented by the nucleotide sequences of SEQ ID NOS: 13 and 14. The gene encoding the carbapenem-associated resistance protein precursor can be detected as a pair of primers represented by the nucleotide sequences of SEQ ID NOS: 29 and 30,

The gene encoding the TonB one dependent siderophore receptor protein can be detected as a pair of primers represented by the nucleotide sequences of SEQ ID NOS: 25 and 26. The gene coding for ompE (0uter membrane protein E) can be detected as a pair of primers represented by the nucleotide sequences of SEQ ID NOs: 37 and 38. The gene encoding ompW (Outer membrane protein W) can be detected as a pair of primers represented by the nucleotide sequences of SEQ ID NOS: 47 and 48. The gene encoding Beta-lactamase class C Adc-73 (Bet a-lactamase class C Adc-73) can be detected as a primer pair represented by the nucleotide sequences of SEQ ID NOS: 57 and 58.

The term " probe " refers to a nucleic acid fragment such as RNA or DNA corresponding to a few nucleotides or several hundreds of nucleotides that can specifically bind to the mRNA. The presence or absence of the specific mRNA and the amount of expression . The probe can be produced in the form of a single strand DNA probe, a double strand DNA probe, an RNA probe, or a peptide nucleic acid (PNA). The selection and modulation conditions of an appropriate probe may be appropriately selected according to techniques known in the art.

The kit is not particularly limited as long as it is known as an assay kit that provides a peptide, an antibody or an aptamer having a specific binding domain in the art as a component, but the assay kit may be any one known in the art such as Western blotting, ELISA, radioimmunoassay, Ouchteroni immunodiffusion method, Locate immunoelectrophoresis, tissue immuno staining, immunoprecipitation assay, complement fixation assay, kit for FACS or protein chip, and the like. In one embodiment, the kit comprises an antibody specific for the six marker protein. Antibodies are monoclonal antibodies, polyclonal antibodies or recombinant antibodies (chimeric antibodies) with high specificity and affinity for the marker protein and little cross-reactivity to other proteins. The kit may also comprise an antibody specific for the control protein. The kit may further comprise a reagent capable of detecting bound antibody, such as a labeled secondary antibody, chromophore ((± 1 ^" 01110 110 ^ 3), enzyme (in conjugated form with antibody) Or other substances capable of binding to the antibody, etc. The kit of the present invention may further comprise a washing solution or eluent capable of removing excess chromogenic substrate and unbound protein and retaining only the protein marker bound to the antibody . &Lt; / RTI &gt;

The kit is not particularly limited as long as it is known in the art as an assay kit for providing a primer (primer pair) or a probe as a component. For example, the kit may include PCR (polymerase chain reaction), RNase protection Analysis, Northern blotting, Southern blotting or kits for DNA microarray chips, and the like. For example, the diagnostic kit may be a diagnostic kit characterized by comprising essential elements necessary for performing a polymerase reaction. The Polymerase Enzyme Kit contains a respective pair of primers specific for the marker gene (mRNA). The primer is a nucleotide having a nucleotide sequence specific to the nucleotide sequence of each marker gene (mRNA), and has a length of about 7 bp to 50 bp, more preferably about 10 bp to 30 bp. It may also include primers specific to a nucleic acid ^"sequence of a control gene. Other Polymerase Enzyme Kit can be used in a test tube or other suitable container, a buffer solution (pH and magnesium concentration varies), deoxynucleotides (dNTPs), a DNA polymerase (such as Taq polymerase) DNAse, RNAse inhibitor DEPC-water, sterile water, and the like.

In order to provide information necessary for diagnosis of an Acinetobacter baumannii infection, the present invention provides a method for diagnosing infection with Aspergillus baumannii, comprising the steps of extracting from a biological sample isolated from a sample a beta-lactamase class D OXA-23, Dependent receptor protein (TPB), ompE (outer membrane protein E), ompWCOuter membrane protein W, and beta 1 lactamase class C Adc -73 &lt; / RTI &gt; (Beta-lactamase class C Adc-73) One or more proteins; Or a gene coding for the above-mentioned protein group,

The 'specimen' means the subject to be infected according to the method of the present invention because there is no information as to whether or not the infected person is infected with Ashinotobacter baumannii. Preferably means an analyte that does not have information about whether it has been infected with an antibiotic resistance (particularly multidrug resistant) strain of Acinetobacter baumannii.

The term " biological sample " or " sample &quot; as used herein includes blood and other solid samples of biologic origin, biopsy specimens, solid tissue samples such as tissue culture, or cells derived therefrom. But are not limited to, tissue, extract, cell lysate, whole blood (blood), plasma, serum, saliva, ocular fluid, cerebrospinal fluid, sweat, urine , Alveolar lavage fluid, etc. Preferably, the sample may include at least one selected from the group consisting of saliva, sputum, alveolar lavage fluid, urine, blood, serum and plasma.

The sample can be obtained from an animal, preferably a mammal, and most preferably from a human. The sample may be pretreated prior to use for detection or diagnosis. For example, it may include homogenization, filtration distillation, extraction, concentration, inactivation of interfering components, addition of reagents, and the like.

The method for diagnosing Ashkenobarbaumani infection can be understood by those skilled in the art as a method for providing information on the diagnosis of Ashkenobarbaumani infection through detection of a marker protein or its genes.

The term &quot; detection &quot; in the present invention means measurement and confirmation of the presence or absence of a desired substance, or measurement and confirmation of changes in the level of presence (level of expression) of a desired substance. Can be performed without limitation including all of the quantitative methods. ^"Wherein the detecting step (step 1) may be to detect a gene which may be to detect the six kinds of protein, or encoding the six kinds of protein.

The detection of the protein includes the detection of the presence or absence of an increased or decreased amount of the protein expressed by the six proteins provided by the marker in the present invention. The measurement method is not particularly limited as long as it is a protein expression assay method known in the art, but it may be preferably by an antigen-antibody binding reaction method. Specifically, protein detection in the present invention can be carried out by enzyme immunoassay (ELISA), radioimmunoassay (RIA), radial immunodiffusion, sandwich ELISA, Western blot, immunoprecipitation, , Immunoelectrophoresis, Ouchter lony immunodiffusion, immunofluorescence, enzyme substrate staining, antigen-antibody aggregation, surface plasmon resonance (SPR), and protein chips. .

The detection of the gene includes the detection of the presence or absence of the mRNA transcribed from each gene encoding the six proteins provided as markers in the present invention, or confirming the increase or decrease in the expression level of the mRNA. The measurement method is not particularly limited as long as it is a method of measuring mRNA expression known in the art, but it is preferable to use PCR, RNase protection assay, northern blotting, southern blotting, May be one selected from the group.

The method for diagnosing infection with Aspergillus puberty may further include a step (step 2) of comparing the protein expression level or mRNA expression level of the gene detected in the step of detecting the protein or gene (step 1) with the control group have.

The term &quot; control group &quot; may refer to a normal or non-infected sample from the normal subject. Preferably those not infected with the antibiotic-resistant Acinetobacter baumannii An individual infected only with a normal subject not infected with itself or with an antibiotic-susceptible organism) or a sample from the subject.

In the step of comparing the detected level with the control group of the step 2, the beta-lactamase class D OXA-23, the carbapenem-associated resistance protein precursor , the TonB-dependent siderophore receptor protein, the ompE (Outer membrane protein E), the ompW (Outer membrane protein W) and the beta-lactamase class C Adc-73 (Beta-lactamase class C Adc-73); Or a gene coding for the protein group is higher than the control level (that is, when the expression level is regulated higher than that of the control), it can be judged that the virus is infected with Ashinobacterium var. Can be judged to be infected with antibiotic resistance (especially, multidrug resistant).

Up-regulating expression of the six marker proteins or genes encoding the six marker proteins provided by the present invention is obtained when the bacterium is resistant to antibiotics in patients who have been infected with antibiotic-sensitive strains of Aspergillus puberty As shown in FIG.

The present invention relates to the use of a beta-lactamase class D OXA-23, a carbapenem-associated antagonist for the preparation of a diagnostic agent for the diagnosis of infection with Aspergillus baumannii (c / o c er bau annii) A TonB-dependent siderophore receptor protein, ompE (outer membrane protein E), ompW (Outer membrane protein W), and a beta-lactamase class C Adc -73 (Beta-lactamase class C Adc-73); Or a gene coding for the above-mentioned protein group. The present invention also relates to a method for the preparation of a pharmaceutical composition, which comprises the steps of: a) obtaining from a biological sample isolated from a sample a Beta-lactamase class D OXA-23, a carbapenemassociated resistance protein precursor, (NB-dependent siderophore receptor protein), ompE (Outer membrane protein E), ompW (Outer membrane protein W) and Beta-lactamase class C Adc-73 One or more proteins selected from the group of proteins; Or a gene coding for the protein group; b) comparing the level of the detected protein or gene with a control; And c) an Acinetobacter baumannii ^{(Ar / 'je obac e /} "baumannii) infection comprising the step of the level of the detected protein or gene is determined to be infected with Acinetobacter baumannii is higher than the control level, to provide.

The 'effective amount' of the present invention refers to an amount which, when administered to an individual, indicates an improvement, treatment, prevention, detection, diagnosis, or inhibition or reduction effect of an infection with Ashkenobacter baumanni, May be an animal, preferably a mammal, especially an animal, including a human, and may be an animal derived cell, tissue, organ, or the like. The subject may be a patient requiring the effect. The term &quot; treatment &quot; of the present invention broadly refers to ameliorating the symptoms of an Acinetobacter baumanni infection or a degenerative Ashkenobacter baumanni infection, which heals, substantially prevents, or improves the condition , And includes, but is not limited to, relieving, curing or preventing one or most of the symptoms resulting from an Acinetobacter baumannii infection.

The term "comprising" of the present invention is used synonymously with "containing" or "characterized" and does not exclude additional component elements or method steps not mentioned in the composition or method . The term &quot; consisting of &quot; means an additional element, step or Components and the like. The term "essence i al ly consisting of" means that, in the context of a composition or method, a component element or step, as well as component elements or steps, which do not materially affect its underlying properties, It means to include.

Effect of the Invention By using the six kinds of proteins provided in the present invention or the gene markers coding for the proteins, it is possible not only to detect differentiation of the multidrug-resistant Acinetobacter baumanni differentially, but also to the existing blood culture method The method of the present invention is advantageous in the early onset of the multidrug-resistant Acinetobacter baumannii infectious disease such as sepsis.

BRIEF DESCRIPTION OF THE DRAWINGS FIG. 1 is a schematic representation of the multi-drug resistant A baumannii strains KAB01 to KAB08

Show.

FIG. 2 is a schematic diagram showing a method of separating membrane proteins (MEM) and secretory proteins (including proteins from SEC and fresh) in KAB strains.

Fig. 3A shows the results of the present invention in the presence of the whole strain of the strain and the complex antibiotic in the condition (KAB03- S) without the antibiotic and in the condition of adding the compound (KAB03-R) to the versatile resistant strain KAB03 (a strain resistant to three types of combined antibiotics) (KAB03-MEM) and the number of fractions (KAB03-SEC, a concept including proteins from 0MV) in the presence of the enzyme.

FIG. 3B shows the results of the detection of the presence of the entire strain of the strain and the presence of the combined antibiotic (KAB03-S) in the absence of the antibiotic (KAB03-S) (KAB03-MEM) and the fraction (KAB03-SEC) in the cell under the same conditions. Fig. 4 is a graph showing the effect of 0 mV (Outer Membrane Vehicle) isolated from the multi-resistant A bai / roa / // (DU202) cultured in each of the conditions with Imipenem antibiotic and no antibiotic (Normal, LB medium) TEM images. ^'

Fig. 5 shows the results of the multi-agent resistance cultured in the presence of Imipenem (IM) antibiotic and in the absence of antibiotics (Normal, LB medium). The disrupted 0MV (Outer Membrane Vehicle les) dislocations were developed by SDS-PAGE and the protein was separated by molecular weight.

Figure 6a shows the results of a multi-agent resistance assay in the presence of Imipenem antibiotics and in the absence of antibiotics (Normal, LB medium). As a result of the identification of the 0MV protein species isolated, 332 proteins were identified under LB (control) conditions and 290 proteins under IM (Imipenem) conditions. Among these proteins, proteins expressed in common in both conditions Respectively.

Figure 6b shows the results of multiple cultures cultured in the presence of Imipenem antibiotic and in the absence of antibiotics (Normal, LB medium). The intracellular distribution of 0MV proteins isolated from wu / ra / // was confirmed using CELLO v2.5 software.

FIG. 7a shows the result of immunoactivity analysis of membrane protein bodies obtained with multidrug-resistant A baumannii cultured under the condition of Imipenem antibiotics by two-dimensional gel electrophoresis (left) and Western blot (right) method.

FIG. 7b shows the result of immunological activity analysis of secretory protein bodies obtained with the multidrug-resistant baumannii cultured under the condition of Imipenem antibiotic through two-dimensional gel electrophoresis (left) and Western blotting (right). FIG. 8 shows the status of acquisition of the patients to be collected on a monthly basis (true infection: colonization: resistant colonization, sensitive: colonization, negative control)

Fig. baumannii true infectivity group and colonization of the resistant bacteria.

Fig. 10 shows the result of investigation of the co-infection style of the multidrug resistant A baumanni-other microorganisms in the true infection group and the colonizat ion group.

Fig. 11 shows the result of comparative evaluation of the specificity of A baumannii-specific resistance against 23 kinds of final candidate markers. Using the primers specifically designed for each marker, various species (KAB03 (Resistant AB), PCR was performed on the genomes of AC19606 (Sensitive AB), E. coli J / (Gram negative), S. pneumonia (Gram positive) and Burkholderia sp. 24 (Gram negative) (The 23 kinds of candidate markers, which are not shown in Fig. 11, are not displayed because PCR is not performed).

BEST MODE FOR CARRYING OUT THE INVENTION Hereinafter, the present invention will be described in detail. However, the following examples are illustrative of the present invention, and the present invention is not limited to the following examples.

Example 1: Diagnosis of multidrug-resistant Acinetobacter baumannii through analysis of main characteristics and infected patients "

1-1. Acinetobacter baumannii strain collection Among 36 A baumannii strains isolated from pneumonia patients in five intensive ICU hospitals in Seoul, Gyeonggi, Daejeon / Gyeongbuk, Daegu / Gyeongbuk, Busan / Gyeongnam, Jeonju and Jeonju, eight imipenem resistant strains . These strains were named KAB (Korean Acinetobacter baumannii) ^. Respectively. (see Table 1), using the Pasteur MLST method using gene sequencing such as glutA, gyrB, gdhB, recA, cpn60, GPI and rpoD. In Table 1, A baumannii DU202 strain was collected from Busan (Dong-A University Hospital) in 2006.

【Table 11 g! T gyrB gdhB recA cpn60 6Pi rpoD ST region

DU202 46 3 2 2 142 3 423 Busan AB01 3 3 2 2 142 3 451 Daejeon

KAB02 3 3 2 2 106 3 369 Daejeon

KAB03 3 3 2 2 142 3 451 Daejeon

KAB04 3 3 2 2 94 3 191 Busan

KAB05 3 3 2 2 106 3 369 Daegu

KAB06 3 3 2 2 106 3 369 Daegu

KAB07 3 3 2 2 94 3 191 Jeonju

KAB08 1 3 3 2 2 97 3 208 games

1-2. Antibiotic Resistance Identification (MIC test) of Collected Strain The collected strains were collected in Example <1-1>. The antibiotic resistance of baumannii strains was confirmed. Specifically, as shown in Table 2, Mi crobiologi cal ly Influenced Corrosion (MIC) test was performed using a total of 17 antibiotics in the eight antibiotic clasps. The MIC test was conducted on the antibiotics listed in Table 2 below in accordance with the standard diagnosis method of the medical-related disinfection bacteria test (National Institute of Health Research and the Korean Society of Clinical Microbiology (Dist diffusion method). Among the antibiotics listed in Table 2, there are three types of antibiotics, namely, Imipenem (tolerance of 16 g / ml or more), Amikacin (aminoglycoside system, more than 16 ug / ml), Col istin For the antibiotics, the concentration of Imipenem, Amikacin, and Col istin was increased from 0.25 ug / ml to liquid culture for protein testing. In Table 2, the strains growing above the reference concentration of each antibiotic resistance are denoted by R as Resistance. The strains which can not grow even at the initial concentration are Sensitiva strain S and the strains which grow at the concentration therebetween (weak resistant strain) and I as intermediated resistance.

As a result, as shown in Table 2, all of the collected bacteria were resistant to the reference antibiotic carbapenem, and KAB01, KAB03, KAB04, and KAB08 were resistant to col ist, the last used antibiotic in the hospital Respectively.

[Table 2]

 Susceptibility

 Drug class Antimicrobial

 ABO1 KAB02 AB03 KBA04 KAB05 KAB06 AB07 AB08 202

 Ampicillin &

 R R R R I I S I R

 Sulbactam

 Ticarcillin &

 R R R R R R R R R

 (3- Lactams Clavulanic acid

 Piperacillin R R R R R R R R

Piperacillin &

 R R R R R R R R R

 Tazobactam

 Cefotaxime R R R R R R R R

Ceftazidime R R R R R R R R

Cephalosporins

 Cefepime R R R R R R R R

Aztreonam RRRRRRRRR Imipenem RRRRRRRRR

Carbapenems

 eH R R R R R R R R

Amicacin R R S R S S S S R

Aminoglycosides

 Gentamicin R R R R R R R R

Fluoroquinolone Ciprofloxacin R R R R R R R R

 Minocycline I I S S S S I

Tetracyclines

 Tigecycline S S S S S S S S

Polymyxin Colistin s R S R R s S s R

 Trimethoprim &

 Sulphonamide R S R R R R R R R

 Sulfamethoxazole

1-3. Dielectric Detection of Multidrug Resistant Acinetobacter baumannii Pathogen. Full-length genomic information for the baumannii strains was obtained from the PacBi o RSI I system according to the protocol of the manufacturer. As shown in Table 3, eight KAB strain genomes were 3.96-4.00 Mb , Total 3, 919 ~ 4, 026 (gene + pl asmid) genes, and each KAB strain has a plasmid of 70.8 ~ 121.4 Kb, and these plasmids encode 94 ~ 152 genes . The phylogenetic tree of the KAB strains collected in Example 1-1 was analyzed and the results are shown in FIG.

[Table 3]

Chromoso

 mm m KMS03 w mm KA807

^ 178,9S8 3 63 17 3 ^ 67,717

 393 39Λ 39.1 39.0 39.1 39.1 39.1 39.1 39

4140 382S 3830 »7 3893 3 ^ 2 3854 3843

404S 3733 378 $ 3738 3782 3801 37 $ 4 3761 3751 纖 18 18 18 18 18 18 18 18 18 tRNA 7β 73 n n 73 73 73 74 73 tmrWA 1 1 1 1 1 1 1 1 1

 OU202 KA801 ΚΑ8Θ2 KA803 XA80 KA80S KABOE KM07 KA808

• 7W75 73 ^ 91 mm 70884 70JBU njn 10 06

GCH - &gt; S 33.9 31.7 s 33. $ 33,5 Ϊ3.4 346

Cene - 94 113 101 1S2 94 96 95 117 cos 94 113 101 1S2 94 96 95 11?

1-4. Acinetobacter bawnannii 5 ^"· ^ ¾: separating body the use of KAB strains to identify multi-drug resistance marker membrane protein (MEM) and secreted protein (SEC, concepts that include proteins from 0MV) performing each proteomic analysis The separation technology of A b nni membrane protein and secretory protein according to the presence of antibiotics was secured and the underlying technology for analysis of proteins involved in antibiotic resistance was secured. KAB02, KAB05, KAB05, KAB05, KAB05, KAB05, KAB05, KAB05, KAB05, KAB05, KAB05, KAB05, KAB05, KAB05, KAB05, KAB05, KAB05, KAB05, and KAB06 were obtained from each strain of KAB strain by immunization with Imipenem (16 ug / ml), Amikacin , KAB07 is Imipenem alone, KAB04 is Imipenem and with Colistin, KAB01, KAB03, KAB08 is the three antibiotics 0D both added LB medium inoculated with the (LPS solution, LB-05) and 37 ^° C, 180rpm ₆₀₀ Up to 1.0 and incubated. The centrifugal separation at 8000rpm and then incubated with the cells to give a ^"medium removed (media cell). After the medium to take advantage of the quick-stand system pass through a 0.22um filter, by using ammonium sulfate Badge Protein was precipitated and centrifuged at 8000 rpm for 30 minutes. Then, ammonium sulfate was removed using a dialysis method, and secreted proteins (including proteins from SEC and 0 MV) were separated. The cell fractions were obtained by centrifuging the cells using the separated cells, and the obtained cell fractions were treated with sodium carbonate and centrifuged at an ultra-high speed (150,000 x g, 1 hour) to obtain membrane proteins MEM). Separated proteins were separated by protein-protein molecular weight using SDS-PAGE, and analyzed by LC-MS / MS using trypsin. Protein identification was performed using Mascot version 2.2 software.

[Table 4] AB03-S AB03- AB03-MEM KAB03-SEC

Significance threshoM 0.01 0.01 0.01

FDR 0,43 0.51 0.42 0.2!

Protest 118? 1120 629 776

SwnofemPAI 2905.81 2427.84 163S.87 2224.61 cello 2.S IOVB03-S AB03-R KAB03-MEM A803-SEC

Cytoplasmic 884 (74, 47) 833 (7438) 288 (45.79) 478 (61.60)

Periplasmic 143 (12.05) 133 (11.88) 126 (20.03) 147 (18.94)

Extracellular 31 (2.61) 33 (2.9S) 24 (3.82) 44 (5.67)

OuterMembrane 64iS, 39) 59.5 5.27 79 (12.56) 68 (8.76)

Inner embrane 65 (5.48) 62iS.S4) 112 (17.81) 39 (5.03)

Total 1187 1120 629 776

Proteins of membrane proteins of KAB01 ~ KAB08 were identified as 440-791 proteins and 418-799 proteins were expressed by secretory protein. Specifically, the above Tables 4, 3A, and 3B show the conditions (KAB03- R) and the absence of antibiotics (KAB03-S) with the combination of the antibiotic (three antibiotics) (KAB03-SEC) and membrane protein (KAB03-MEM) under the condition that the whole strain of the strain and the complex antibiotic are present in the culture medium. Among the proteins identified in the above experiment, the proteins having the highest expression level of 20% in each strain and the proteins that are commonly expressed were classified and set as potential candidates for marker digestion.

1-5. Separation of OMV (Outer Membrane Vehicle clones) from baiiffla // multi-resistant Acinetobacter baumannii OMV isolates was carried out by the following method. The LB medium (LPS solution, LB-05) and Imipenem (16 ug / ml) supplemented with antibiotics were added to the DU202 strain The cells were inoculated and cultured at 37 ^° C and 180 rpm from 0D ₆₀₀ to 1.0, followed by centrifugation at 8000 rpm to obtain a medium from which the cells were removed. The medium was passed through a 0.22-μm filter using a quick-stand system, and then the 0MV fraction was concentrated using a 5000 d I on filter and centrifuged for 3 hours using a 150000 × g ultracentrifuge (See Fig. 2).

The image was analyzed using a Bio-TEM apparatus to confirm the separation of 0 MV. As a result of the image analysis, it was confirmed that the separation was successful without other substances (impurities) other than 0 MV as shown in FIG. 4, It was confirmed that many 0 MV was expressed. Density analysis (Otsuka electronics, ELSZ-10000) was performed to confirm the physical differences such as the size of OMVs under the two conditions. The 0MVs secreted at both conditions were found to have similar sizes of 150nm.

1-6. 0MV analysis according to antibiotic conditions The 0MV expression pattern according to the multidrug - resistant baumannii antibiotic condition was analyzed. As in Example <1-5>, OMVs were added to OMVs isolated from conditions with and without Imipenem antibiotic (Normal, LB medium), and the proteins were separated by molecular weight using SDS-PAGE After tryptic inoculation, the cells were analyzed by LC-MS / MS, and protein identification was performed using Mascot vers ion 2.2 software. The results of analyzing 0MV obtained are as follows.

First, for the LC-MS / MS analysis, the same amount of 0 MV protein was developed using SDS-PAGE for each condition, and as a result, the overall pattern was similar in both conditions as shown in FIG. 5 , But only in some bands. As shown in FIG. 5, MS / MS analysis was performed on proteins from 8 fractions after 8 fractions were separated and in-gel digestion was performed. MS / MS analysis showed that 332 proteins were detected under LB (control) conditions and 292 proteins were detected under IM (Imipenem) condition as shown in FIG. 6A. Among these proteins, 191 proteins were commonly expressed in both conditions . Imi enem It was confirmed that only about 30% of the proteins were expressed in each condition only in the presence of antibiotics or in the absence of antibiotics (Normal, LB medium). The distribution of identified proteins was analyzed using CELLO v2.5 software. As shown in FIG. 6B, more than 70% of the proteins were related to the membrane and 30% were proteins related to the outer membrane.

1-7. Detection of A. baumannii diagnostic markers through immuno-proteomic analysis Antibodies to multidrug resistant A baumannii membrane protein and secretory protein were used to digest antigen proteins. In order to increase the accuracy of the assay, the membrane protein and secretory protein of the multidrug-resistant A baumannii KAB03 obtained under the antibiotic conditions of Example 1-4 were injected into the mouse twice at intervals of two weeks to prepare ant i -serum Only IgG was isolated and purified from each ant i -serum.

The analysis of the two-dimensional gel electrophoresis and membrane proteomic immune activity through a Western blot method is the same as Figure 7a, to therefrom in Table 5 ^'Spot NO. Antigenic proteins corresponding to M_l_01 to M_l_10 were identified. Using LC-MS analysis, Identification was performed.

In the same manner, the immunoactivity of secretory protein bodies was analyzed as shown in FIG. 7b. From these, the antigenic proteins of Spot N0.S-1_01 to S_l_21 were identified in the following Table 5 and analyzed by LC-MS analysis. Identification was performed. The resulting antigenic proteins are shown in Table 5.

[Table 5]

 Accessions

 Spot NO (Genbank Description Name MW PI mol%

 Accession no.)

CAB03.03220

 M_1 _01 Outer membrane protein A precursor 38427 5.32 92.591 5

 (AOX78807)

 KAB03_03220

 M_1 _02 Outer membrane protein A precursor 38427 5.32 92.4557

 (AOX78807)

 KAB03_03220

 M_1_03 Outer membrane protein A precursor 38427 5.32 94.3989

 (AOX78807)

 CAB03.03220

 _1 _04 Outer membrane protein A precursor 38427 5.32 80.6061

 (AOX78807)

 KAB03_03220

 M_1_05 Outer membrane protein A precursor 38427 5.32 85.535

 (AOX78807)

 KAB03_03220

 _1_06 Outer membrane protein A precursor 38427 5.32 85.51 38

 (AOX78807)

 KAB03_03220

 M_1 _07 Outer membrane protein A precursor 38427 5.32 89.41 18

 (AOX78807)

 KAB03_03035

 Phosphoenolpyruvatecarboxykinase [GTP] pKG 67377 5.18 23.401 6 (AOX78632)

 M_1 _08

 KAB03_03255

 Putative TonB-dependent receptor yncD 6861 7 5.39 38.3607 (AOX78842)

 CAB03.02937

 hypothetical protein 26489 4.8 19.3049 (AOX78535)

 _1 _09

 KAB03.03667

hypothetical protein 27633 4.59 34.8605 (AOX79231) KAB03_00249

 beta-lactamase class D OXA-23 blaOXA-23 30959 7.71 43.1952 (AOX75888)

 M 丄 10

 KAB03_02937

 hypothetical protein 26489 4.8 32.2048 (AOX78535)

 KAB03_03220

 S_1 _01 Outer membrane protein A precursor 38427 5.32 96.381

 (AOX78807)

 KAB03_03220

 S_1 _02 Outer membrane protein A precursor 38427 5.32 89.21 9

 (AOX78807)

 KAB03_03220

 S_1 _03 Outer membrane protein A precursor 38427 5.32 83.7313

 (AOX78807)

 KAB03_02755

 S_1 _04 beta-lactamase class C ADC-25 blaADC-73 43156 9.41 99.9505

 (AOX78355)

 CAB03-01862

 putative protein 20999 9.2 55.6249 (AOX77475)

 S 丄 05

 KAB03_02755

 beta-lactamase class C ADC-25 blaADC-73 431 56 9.41 31 .631 7 (AOX78355)

 KAB03_01862

 putative protein 20999 9.2 53.7909 (AOX77475)

 S_1 _06

 KAB03.02755

 beta-lactamase class C ADC-25 blaADC-73 431 56 9.41 25.21 95 AOX78355

 KAB03_00924

 C jter membrane lipoprotein omp16 precursor 22472 9.3 25.3788 (AOX76538)

 S_1 _07

 KAB03_01862

 putative protein 20999 9.2 20.7912 (AOX77475)

 KAB03_01 1 34

 Aldose 1-epimerase 41 546 9.01 1 7.5247 (AOX76747)

 KAB03_02755

 S? 08 beta-lactamase class C ADC-25 blaADC-73 431 56 9.41 49.581 5

 (AOX78355)

 KAB03_02959

 Protein to Old 46544 9.1 9 18.6914 (AOX78557)

 KAB03_01 134

 Aldose 1-epimerase 41546 9.01 25 (AOX76747)

 S 丄 09

 KAB03_02755

 beta-lactamase class C ADC-25 blaADC-73 431 56 9.41 51 .7366 (AOX78355)

 KAB03_02755

 S_1 _10 beta-lactamase class C ADC-25 blaADC-73 43156 9.41 91 .1987

 (AOX78355)

S_1 _1 1 KAB03_00939 putative protein 25544 5.47 1 3.2086 (AOX76553)

KAB03_03220

 Outer membrane protein A precursor 38427 5.32 28.777 (AOX78807)

 KAB03_00249

 oeta-lactamase class D OXA-23 blaOXA-23 30959 7J1 1 1.4325 (AOX75888)

 S_1_12

 KAB03_03220

 Outer membrane protein A precursor 38427 5.32 21 .0848 (AOX78807)

 KAB03_03359

 S 丄 1 3 Malatedehydrogenase 35301 5.2 76.4256

 (AOX78935)

 KAB03_03071

 Succinyl-CoA synthetase subunit alpha 30674 5.6 35.4554 (AOX78665)

 S_1 _14

 KAB03_03359

 Malatedehydrogenase 35301 5.2 37.0552 (AOX78935)

 KAB03_00647

 S_1_16 Cysteinesynthase B 331 35 5.55 61 .1 1 1 1

 (AOX76276)

 KAB03_01487

 S_1 _1 7 Chlorideperoxidase 1 cPO 3071 1 5.54 56.0444

 (AOX77100)

 KAB03_00924

 0 knee membrane lipoprotein omp1 6 precorrsor 22472 9.3 27.127 (AOX76538)

 S_1 _18

 KAB03_02755

 beta-lactamase class C ADC-25 blaADC-73 431 56 9.41 26.6034 (AOX78355)

 CAB03.00924

 Outer membrane lipoprotein omp16 precursor 22472 9.3 45.46 (AOX76538)

 S_1 _1 9

 KAB03_03220

 Outer membrane protein A precursor 38427 5.32 1 2.6344 (AOX78807)

 KAB03_00249

 S 丄 20 beta-lactamase class D OXA-23 blaOXA-23 30959 53.6972

 (AOX75888)

 KAB03_03425

 S 丄 21 putative protein 28814 4.91 32.3268

 (AOX78997)

1-8. Clinical characterization of patients with multiresistant A baiimannii infection

 Clinical information and specimens of Acinetobacter bamannii infected patients were collected based on IRB approval (IRB No. 2015-02-027) of Lonnam University Hospital.

환자 (내성균 진성 감염， true infect ion) , 기관지내 다제내성 Acinetobacter baumannii 집락화된 환자 (내성균 집락화， colonization) 및 약제 감수성 Acinetobacter baumannii 집락화된 환자 (감수성균 집락화， sensitive)를 대상으로 하였다 (도 8 참조). 검체로서는 상기 환자들의 폐포세척액， 소변， 혈액을 수집하였다. 균체는 60명으로부터, 폐포 세척액， 소변 및 혈액은 52명의 환자로부터 수득할 수 있었다. 또한 다제 내성 A. baumannii^] 약제 내성 여부 및 MIC (minimum inhibitory concentration) 수집하는 동시에 임상 data (나이 , 성별， A. baumannii 약제 내성 여부， colonization/true infection 여부를 조사하였고, co-infection 여부， Duration of ICU stay, mortality, APACHE II score, co-morbidity 등을 정리하였다 (데이터 미도시). Patients to be collected for the sample include patients with acute respiratory distress syndrome

Patient (true infection, true infectious), endotracheal multidrug resistance Acinetobacter baumannii colonized patients (colonization) and drug susceptibility Acinetobacter baumannii colonized patients (susceptible bacteria colonization, sensitive) (see FIG. 8). As a specimen, alveolar lavage fluid, urine, and blood of the patients were collected. From 60 patients, alveolar lavage fluid, urine and blood were obtained from 52 patients. We also investigated the clinical and clinical data (age, sex, A. baumannii drug resistance, colonization / true infection, co-infection, Duration of ICU stay, mortality, APACHE II score, and co-morbidity (data not shown).

As shown in Fig. 9, the antibiotic susceptibility between the multidrug resistant A baumannii true infectious group and the colonizat ion was compared with the antibiotic susceptibility of Colistin, amikacin, minocyl in, and tigecycl ine Immunity was found to be high in crabs, but antibiotics of Ampicillin and sulbactam, cotrimoxazole, Piperacillin and Tazobactam, Aztreonam, Cefepime and Cefotaxime showed low resistance. Also, as shown in Fig. 10, the co-infection type () -infection style of the multidrug-resistant A baumannn was examined. Patients with truly infected baumanni showed more patterns of co-infection with other strains, as shown in Table 6 below

rotal 19 patterns 24 24 Total 16 patterns 36 36

1-9. A. baumannii diagnostic marker for multi-drug resistance

In the above examples, A. baumannii genome and proteomic analysis results, And the result of analysis of the specimen,

7 were selected as final marker candidates.

[Table 7]

 Accession # log ratio

 Standard of Administration *

 Localization (Genbank

 (mol%)

 Accession no.)

Beta-lactamase class D KAB03 KAB03_00249

1 1 Extracellular

 OXA-23 (1.44) (AOX75888)

Ketol-acid KAB03_00620

2 1 cytoplasmic 1.068

 reductoisomerase (AOX76249)

3 Putative protein 1 No ID

 KAB03_01808

4 Integration host factor 1 cytoplasmic 1.038

 subunit beta (AOX77421)

 KAB03_00342

5 DNA binding protein 1 cytoplasmic 1.463

 (AOX75981)

CAB03 CAB03- 02303

6 beta-lactamase class A 1 cytoplasmic

 TEM-1D (0.489) (AOX77913)

 Outer KAB03_01748

7 TonB ᅳ dependent 1 0.987

 siderophore receptor Membrane (AOX77361)

 KAB03_03636

8 Single-stranded DNA ᅳ 1 Extracellular 1.003

 binding protein (AOX79200)

Carbapenem ᅳ associated Outer KAB03 KAB03_02937

9 resistance 1

 Membrane (0.355) (AOX78535) protein precusor

 KAB03_02013

10 Putative 17 kDa surface 1 cytoplasmic 0.980

 antigen (AOX77625)

 KAB03_01180

11 copper homeostasis protein 1 Extracellular 1.278

 (AOX76793) AB03_03492

12 D-3-phosphoglycerate 1 cytoplasmic 1.468

 dehydrogenase (AOX79064)

13 ompE (1 Outer AB03

Outer membrane KAB03 ᅳ 03141 protein E) Membrane (0.128) (AOX78734)

KAB03_00333

Argininosuccinate lyase 1 cytoplasmic 1.261

 (AOX75972)

Adel family multidrug

 Outer KAB03 KAB03_03104 efflux RND 1

transporter periplasmic Membrane (0.055) (AOX78698) adapter subunit

 Outer A1S_1296

Hep 4 - Membrane (ABO 11724)

Outer A1S_1655

Ferric siderophore 4 - receptor protein Membrane (ABO12082)

 Outer A1S_0292

OmpW (Outer membrane 4 - protein W) Membrane (ABO10762)

 Outer A1S_0255

RND efflux transporter 4 - Membrane (ABO10730)

Outer A1S_1751

AdeA 4 - Membrane (AB012178)

Outer A1S_2737

Adek 4 - Membrane (ABO13150)

Outer A1S_1749

ABC transporter-like 4-protein Membrane (AB012176) beta ᅳ lactamase class C KAB03 (2D KAB03_02755

 3 Extracellular

 ADC-73 MS / MS) (AOX78355)

* 1: Proteomic cs results

 2: Genomic cs results

 3: Immuno-proteomic cs results

4: Reference Example 2: Multidrug resistance of selection markers A. baumannii infection diagnosis ability For the markers selected in Table 7 above, the ability to diagnose these multidrug-resistant A baumannii infections was confirmed by PCR. Specifically, PCR was performed using the PCR primers shown in Table 8 for each marker protein. The PCR amplification conditions were as follows: 1 at 94 ^° C, 1 minute at 55 ^° C, and 30 minutes at 72 ^° C. Solg &Lt; / RTI &gt; 2X Taq polymerase (Solgent). For comparison, the genomes of KAB03 (Resistant AB), AC19606 (Sensitive AB), E. coli (Gram negative), 5. pneumoniae (Gram positive) and sp.KT24 (Gram positive) were electrophoresed using 2X Agalose gel Respectively. As a result of the experiment, the log ratio of the expression level of KAB03 (Resistant AB) to the expression level of AC19606 (Sensitive AB) is shown in Table 7 (add the mol% value in the expression level in parentheses). The expression of KAB03 in Table 7 indicates that the expression level was measured only in KAB03.

[Table 8] SEQ ID NO: PCR primer sequence

13 5 'F- ATG AAT TAA ATA ΤΓΤ TAC TTG CTA TGT GGT TGC

 Beta-lactamase class D

 0XA-23

14 ^' 5' R 1 TTA AAT AAT ATT CAG CTG TTT TAA TGA TTT CAT C

15 5 'F- ATG CM ATT ΤΓΤ TAC GAT AAA GAC TGT GAC

 Ketol-acid

 재ductoisomerase

 16 5 'R-TTA GTT TTT CTC GTC TAC GAT CTT G

17 5 'F- ATG GGT CTT ΊΤΤ GAT TTT GTA AAA GGT ATT G

 Putative protein

 18 5 'R - TTA TTG AGG AAT ACG TAA AAC TTG TCC T

19 5 'F-ATG ACT ACT GAA GCT CTT AAT AAG TCT

 Integration host factor

 subunit beta

 20 5 'R- TTA TTT TCC AGA TTC ATT CAC TGC ATA

 21 5 'F- ATG AAA CCG GAT ATT AGT GAA TTA TCT G

 DNA binding protein

 22 5 'R - TTA GAT TAA GAA ATC TTC AAG TTT TGC ACC

23 5 'F- ATG AGT ΑΠ CM CAT TTC CGT GTC G

 Bet-lactamase class A

 TEM-1D

 24 5 'R - TTA CCA ATG CTT AAT CAG TGA GGCA

TonB-dependent 25 5 'F- ATG AAT GTA AAC AAT AAG Απ GGG AAA G siderophore receptor

 26 5 'R - TTA ATA GTT AAA AGT ATA ACT TAA ACC ATA AGT ACG

27 5 'F- ATG CGT GGT GTA MT AAG GTT ATT TTG

 Single-stranded DNA-binding protein

 28 3 'R- TTA GAA CGG TAA ATC GTC GTC TAA ATC

Carbapenem-associ ated 29 5 'F- ATG AAA GTA TTA CGT GTT TTA GTG ACA ACTA

 res i stance

 prote in precusor 30 5 'R-TTA CCA GAA GAA GTT CAC ACC MC TTT

31 5 'F-ATG ACA GAT CM AAT ACA GAT CAT GCT

 Putat ive 17 kDa sur face

 antigen

 32 5 'R- TTA AAT ATC ATC CCA CGC ATC ΉΤ AAT C

33 5 'F-ATG AAA AAA TCA TTA TTA GCA ATT GCA CTT ATGA copper homeostas i s

 protein in

 34 5 'R-TTA ATC TTC TTT TTT CAA MC ATA ATG TTC AGC

35 5 'F- ATG AGC CAA CAT CTT TCA CTA CCT AAA

 D-3-phosphog lycate

 dehydrogenase

 36 5 'R- TTA GAA CAA TAC GCG TAC GCG MT T

37 5 'F- ATG AM TTA AAA CAT CTG AGC ACT GCA AT

 Outer membrane protein

 E

 38 5 'R-TTA GAA TTT ATA ACC AAG TTT TAA ACC GTA AG

39 5 'F- ATG TGG GGT GGT CGT TTT TCT G

 Argininosucc inate lyase

 40 5 'R- TTA AGC GTA TAA CTG TTC TAG ACG TTT AT

AdeA / Adel fami ly

 41 5 'F- ATG ATG TCG GCT AAG CTT TGG G

 mu l t i drug ef f l ux RND

transporter per ipl asmi c

 42 5 'R- TTA TGC ATT TGA AGC TGC ΊΤΤ CTG TTC

 adapter subunit

 43 F- ATC CTC GAG ATA TGA AAG ATA TAT ACG TTG AGT TTC GC

Hep

 44 R- TGA GGT ACC CGC TGC CTA AGC AGC TGT ATT

45 F- ATC CTC GAG ATA TGA CAC CTT GTT GTC TTG CCA TAT

Ferr i c s iderophore

 R-TGA GGT ACC ATA GTT AAA AGT ATA ACG TAA ACC ATA AGT receptor protein

 46

 ACG

OmpW 47 F- ATC CTC GAG ATA TGG GGG TGT CTT CAT TTA CTT TTG 48 R- TGA GGT ACC GAA TTT ATA GCT ATA GCC TAA AGT GTA MC

49 F- ATC CTC GAG ATA TGT TGG CM CAG GCG CTT TAC TA

 RND ef f lux transporter

 50 R- TGA GGT ACC CAG ATA CGT TAA CCA GCT GTT TAA TTC

51 F- ATC CTC GAG ATA TGT CAT GGT TGC GCC CC

 AdeA

 52 R- TGA GGT ACC TAG TTG TTT GAA GAT ATT AAT GTT GAC CC

53 F- ATC CTC GAG ATA TGC AAA AAG TAT GGT CTA TTT CAG G

Ade

 54 R-TGA GGT ACC TTG CTT TTT AAG TTC AGC ACT AGA TG

55 F- ATC CTC GAG ATA TGC AAA AAG TAT GGT CTA TTT CAG G

ABC transporter l lke

 protein

56 R- TGA GGT ACC TTG CTT TTT AAG TTC AGC ACT AGA TG ^'

57 F- ATG CGA TTT AAA AAA ATT TCT TGT CTA CTT TTA TC bet a-lactamase class C

ADC- 73

 58 R-TTA TTT CTT TAT TGC ATT CAG CAC AGC ACA GCA TAA

11 and Fig _. As shown in Fig. 12, Beta-lactamase class D OXA-23, Carbapenem-associa ed residence protein precursor, T iB 1 dependent siderophore receptor, Outer membrane protein E (OmpE), Outer membrane protein W (OmpW) , and bet a-lactamase class C ADC-73 were used as markers, it was confirmed that the differential detection ability of antibiotic-resistant bacteria and multidrug-resistant bacteria was excellent.

INDUSTRIAL APPLICABILITY As described above, the use of the six kinds of proteins provided in the present invention or the gene markers coding for the protein not only makes it possible to differentially detect the infection of the multidrug-resistant Acinetobacter baumannii, Since it provides faster and more accurate test results than conventional blood culture methods (diagnosis of bacterial culture), it has a very advantageous advantage in the early response to the early stage of multidrug-resistant Ashtonobacter baumannii infection such as sepsis. Therefore, It is very likely to be used in the field.

Claims

Claims:  The present invention relates to a beta-lactamase class D OXA-23, a carbapenem-associated associative resistance protein precursor, a TonB sidropore receptor protein (S) selected from the group consisting of TonB-dependent siderophore receptor protein, ompE (Outer membrane protein E), ompW (Outer membrane protein W) and Beta-lactamase class C Adc-73 (Beta-lactamase class C Adc-73) One or more proteins; Or a gene coding for said protein group. 2. The composition for diagnosing infection with Aspergillus oryzae baumannii according to claim 1, 2. The composition of claim 1, wherein the Ashtonobacter baumanni is an antibiotic resistant Ashtonobacter baumanni. 3. The composition of claim 2, wherein the resistance is multidrug resistant. [Claim 4] beta-lactase Maze class D 0XA-23 (Beta-lactamase class D OXA-23), carbazole penem - more resistance protein precursor (Carbapenem- associ ated resi stance protein precursor), the pore receptor proteins TonB ^■ side ( One or more proteins selected from the group consisting of TonB_dependent siderophore receptor protein, ompE (outer membrane protein E), ompWCOuter membrane protein W and beta-lactamase class C Adc-73 (Beta-lactamase class C Adc-73); Or a gene coding for said protein group, wherein said composition comprises at least one gene selected from the group consisting of a gene encoding said protein group and a gene encoding said protein group. 5. The composition according to claim 4, wherein the agent for detecting proteinol is a peptide, an antibody, or a umbrella having a binding domain specific for the protein. 6. The method according to claim 1, wherein the agent for detecting the gene is selected from the group consisting of a single stranded DNA probe, a double stranded DNA probe, an RNA probe, and a PNMPeptide nucleic acid Probe; Or a primer pair. 7. A kit for the diagnosis of an Ashtonobacter baumannii infection comprising the composition of claim 1. 8. The method of claim 1, wherein the biological sample isolated from the sample is selected from the group consisting of Beta-lactamase class D OXA-23 (Beta-lactamase class D OXA-23), carbapenem Carbapenem-associated associative protein precursor, TonB-dependent siderophore receptor protein, ompE (outer membrane protein E), ompW (outer membrane protein W), and beta-lactamase One or more proteins selected from the group consisting of class C Adc-73 (Bet a-lactamase class C Adc-73); Or a gene encoding said protein group. 9. The method according to claim 8, wherein the sample comprises any one or more selected from the group consisting of acupuncture, sputum, alveolar lavage fluid, urine, blood, serum and plasma How to . [Claim 10] The method of claim 18, wherein the protein is detected by an antigen- Lt; / RTI &gt; Claim 111  11. The method of claim 10, wherein the protein detection is selected from the group consisting of enzyme immunoassay (ELISA), radioimmunoassay (RIA), radial immunodiffusion, sandwich ELISA, Western blot, immunoprecipitation, Selected from the group consisting of immunohistochemical staining, immunoelectrophoresis, Ouchter lony immunofluorescence, immunofluorescence, enzyme substrate staining, antigen-antibody aggregation, surface plasmon resonance (SPR) and protein chip &Lt; / RTI &gt; 12. The method according to claim 8, wherein the gene detection is performed by any one selected from the group consisting of PCR, RNase protection assay, northern blotting, southern blotting and DNA chip Lt; / RTI &gt; 13. The method of claim 8, wherein the method further comprises comparing the level of the detected protein or gene to a control. [14] The method of claim 13, wherein if the level of the detected protein or gene is higher than the control level, it is determined that the virus is infected with Asnithobacter baumannii. 15. The method of claim 8, wherein the Ashtonobacter baumanni is an antibiotic resistant Ashton Burton Vaumani. 16. The method of claim 1,

(Beta-l actamase c lass D OXA-23), a carbapenem-associated resistance protein precursor for the preparation of a drug for the diagnosis of infectious diseases, , TonB one dependent siderophore receptor protein, ompE (Outer membrane protein E), ompW (Outer membrane protein W) and Beta-lactamase class C Adc-73 C Adc-73); Or a gene encoding said protein group. 17. The method of claim 1, further comprising the steps of: a) obtaining from the biological sample isolated from the sample a beta-lactamase class D OXA-23, a carbapenem-associated resistance protein precursor protein precursor, TonB-dependent siderophore receptor protein, ompE (Outer membrane protein E), ompW (0uter membrane protein W) and Beta 1lactamase class C Adc-73 actamase clad C Adc-73); Or a gene coding for the protein group; b) comparing the level of the detected protein or gene with a control; And c) determining that the level of the detected protein or gene is higher than the control level, determining that the virus is infected with Asnithobacter baumannii.