Application of cellulase in degrading bacterial biofilm

Abstract

The invention discloses an application of cellulase in degrading a bacterial biofilm. The invention researches and develops a new application of cellulase, namely a new application in degrading a bacterial biofilm; based on research of the invention, the cellulase can effectively degrade bacterial biofilms on instruments such as medical catheters and prevent and treat infection caused by the bacterial biofilms on the instruments such as the medical catheters; the invention provides a novel scheme and approach for preventing and treating diseases caused by bacterial biofilm infection on instruments such as medical catheters and the like.

Description

Technical Field

[0001] The present application relates to the technical field of cellulase, and in particular to the application of cellulase in degrading bacterial biofilm. Background Art

[0002] Cellulase is a general term for a group of enzymes that can degrade cellulose and generate glucose. It is widely present in organisms in nature, such as bacteria, fungi, and animals. Cellulase is a group of complex enzymes, mainly composed of exo-β-glucanase, endo-β-glucanase and β-glucosidase, and can also contain highly active xylanase. Cellulase can hydrolyze cellulose and can be used as an additive in the fields of medicine, textiles, daily chemicals, papermaking, food fermentation, industrial washing, wastewater treatment and feed.

[0003] Although cellulase has a wide range of cellulose degradation effects, the amino acid sequence homology of cellulase produced by different bacteria is very low, and the specific applications are also completely different. Therefore, the development of new cellulases or the development of new applications of cellulases has always been an important research topic in this field. Summary of the invention

[0004] The purpose of the present application is to provide a new application of cellulase in degrading bacterial biofilm.

[0005] This application adopts the following technical solutions:

[0006] The first aspect of the present application discloses the use of a cellulase in degrading bacterial biofilms. The cellulase has a sequence as shown in SEQ ID NO.1.

[0007] It should be noted that the creative research of the present application found that the structural domain of amino acids 89-380 at the C-terminus of the Mycobacterium tuberculosis Rv0062 protein can act alone and can effectively degrade bacterial biofilms on medical catheters, thereby preventing or treating diseases related to bacterial biofilm infections of medical catheters; therefore, the present application cloned, expressed, and purified the structural domain using a prokaryotic expression system to obtain a new cellulase that can effectively degrade bacterial biofilms on medical catheters and prevent and treat medical catheter-related infections, namely, the cellulase of the sequence shown in SEQ ID NO.1, and proposed a new application of the cellulase, namely, application in degrading bacterial biofilms.

[0008] The second aspect of the present application discloses the use of a cellulase in preparing an agent for degrading bacterial biofilms. The cellulase has a sequence as shown in SEQ ID NO.1.

[0009] It can be understood that the present application has developed a new application of the cellulase with the sequence shown in SEQ ID NO.1, namely, its application in degrading bacterial biofilms; based on the research findings, the cellulase with the sequence shown in SEQ ID NO.1 can be used to prepare reagents for degrading bacterial biofilms.

[0010] The third aspect of the present application discloses the use of a cellulase in preparing an agent for treating or preventing bacterial biofilm infection. The cellulase has the sequence shown in SEQ ID NO.1.

[0011] It should be noted that the present application has found that the cellulase with the sequence shown in SEQ ID NO.1 can degrade bacterial biofilms, and therefore, it can also be used to prepare reagents for treating or preventing bacterial biofilm infections.

[0012] The fourth aspect of the present application discloses the use of a cellulase in the preparation of a device for treating or preventing bacterial biofilm infection. The cellulase has the sequence shown in SEQ ID NO.1.

[0013] It can be understood that, since the cellulase of the present application can degrade bacterial biofilms, it can be used to clean or soak various instruments, such as medical instruments, thereby playing a role in treating or preventing the formation of bacterial biofilms on these instruments.

[0014] The medical device of the present application particularly includes a medical catheter. The bacterial biofilm of the present application particularly includes a biofilm formed by Escherichia coli and / or Pseudomonas aeruginosa.

[0015] The fifth aspect of the present application discloses a reagent for degrading bacterial biofilm, wherein the reagent contains a cellulase with a sequence shown in SEQ ID NO.1.

[0016] It should be noted that the present application has found that the cellulase with the sequence shown in SEQ ID NO.1 has the function of degrading bacterial biofilms. Therefore, the cellulase can be used alone or added to existing bacterial biofilm degradation reagents to improve the degradation efficiency and quality of bacterial biofilms.

[0017] The sixth aspect of the present application discloses a reagent for cleaning instruments, wherein the cleaning reagent contains a cellulase having a sequence shown in SEQ ID NO.1.

[0018] It should be noted that the instrument cleaning reagent of the present application can add the cellulase of the present application on the basis of the existing cleaning reagent, so that the cleaning reagent has the effect of degrading bacterial biofilm, or by using the cleaning reagent of the present application for cleaning or soaking, the instrument can have the effect of preventing the formation of bacterial biofilm.

[0019] The beneficial effects of this application are:

[0020] The present application has developed a new application of cellulase, namely, the degradation of bacterial biofilm. Based on the research findings of the present application, the cellulase of the present application can effectively degrade bacterial biofilm on medical catheters and other devices, prevent and treat infections caused by bacterial biofilm on medical catheters and other devices, and provide a new solution and approach for preventing and treating diseases caused by bacterial biofilm infection on medical catheters and other devices. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] Figure 1 This is a diagram showing the PCR amplification results of the cellulase Rv0062p gene in the examples of the present application;

[0022] Figure 2 This is a diagram showing the construction result of the recombinant cloning plasmid of the cellulase Rv0062p gene in the examples of the present application;

[0023] Figure 3 It is a comparison result diagram of the sequencing results of the cellulase Rv0062p gene in the examples of the present application;

[0024] Figure 4 This is a diagram showing the construction result of the recombinant expression plasmid of the cellulase Rv0062p gene in the examples of the present application;

[0025] Figure 5 This is a diagram showing the expression and purification results of the cellulase Rv0062p protein in the examples of this application;

[0026] Figure 6 This is a graph showing the results of observation of the degradation effect of the cellulase Rv0062p protein on the bacterial biofilm on the medical catheter in the examples of the present application;

[0027] Figure 7 It is the result of the ELISA instrument detection of suspended bacteria after the degradation of bacterial biofilm in the example of the present application. DETAILED DESCRIPTION

[0028] The present application is described in detail below by means of specific embodiments in combination with the accompanying drawings. The following embodiments are only used to illustrate the present application and should not be construed as limiting the present application.

[0029] Example

[0030] 1. Main Materials and Instruments

[0031] Main instruments: clean bench (Suzhou Antai Air Technology Co., Ltd., model: SW-CJ-2); constant temperature incubator (Shanghai Fuma Experimental Equipment Co., Ltd., model: DDX-9505213-1); constant temperature culture shaker (Shanghai Fuma Experimental Equipment Co., Ltd., model: KYC 100B); microplate reader (Thermo Scientific, model: Multiscan FC); low temperature chromatography cabinet (Haier).

[0032] Main materials: Ec medium, NA medium (Qingdao High-tech Industrial Park Haibo Biotechnology Co., Ltd.); LB solid medium (1% Tryptone, 0.5% Yeast Extract, 1% NaCl, 1.5% Agar); TransStart FastPfuFly DNA polymerase, dNTPs, restriction endonuclease, T4 DNA ligase (TaKaRa); DNA gel recovery kit, pJET1.2 blunt vector (Thermo); FastPure Plasmid Mini Kit, BCA protein quantification kit (Nanjing Nova Biotechnology Co., Ltd.); pET16b vector (Novagen); E. coli NovaBlue, E. coli BL21 (DE3) strain (Novagen); IPTG, PMSF (Aladdin Biochemical Technology Co., Ltd.), Ni-NTA Superflow (QIAGEN); (anti)-polyhisti-dine clone HIS-1antibody (Sigma); 3K Cut-off centrifugal concentrator (PALL); double-control latex catheter (Guangzhou Weili Medical Instrument Co., Ltd.); 0.22 μm filter membrane, 48-well plate, 96-well plate (Biofil).

[0033] 2. Methods

[0034] 1. Amplification of the gene encoding the protein domain of Rv0062

[0035] The newly developed cellulase in this example is taken from the C-terminal domain of the Rv0062 protein of Mycobacterium tuberculosis, that is, the C-terminal 291 amino acids (labeled as Rv0062p protein). Therefore, the PCR amplification primers for designing and synthesizing the coding gene fragment corresponding to the 291 amino acids of the C-terminal domain of the Rv0062 protein (labeled as the Rv0062p coding gene) are as follows:

[0036] Upstream primer: 5'-TACATATG CCG CTG GCC GGAAAG-3' (SEQ ID NO. 3), wherein the "CATATG" at the 5' end is the introduced NdeI site.

[0037] Downstream primer: 5’-TA GGA TCC CTA CTG GCC GGC GTT GTG-3’ (SEQ ID NO.4), where the “GGATCC” at the 5’ end is the introduced BamHI site.

[0038] The nucleotide sequence of the Rv0062p-encoding gene is as follows:

[0039] Aacccgctggccggaaagcccttctacgtcgatcccgcctcggcggccatggtcgccgcgcgcaacgccaacc

[0040] cgccgaacgccgagctgacctccgtcgccaacaccccgcagtcctactggctcgaccaggcattcccgccggcgaccg

[0041] tcggcggcacggttgccaggtacaccggagcggcgcaggcggccggcgccatgccggttctgacgctgtatggaatcc

[0042] cccatcgcgactgcggtagctacgcatccggtgggttcgcgacgggcactgattaccgcgggtggatcgacgctgtcgc

[0043] atccggcctgggctcatcgccggcgacgatcatcgtcgaacccgatgcgctggccatggccgactgcctgtcgcctgac

[0044] cagcgccaggaacgtttcgacttggtgcgctacgccgtcgacacgctgacccgcgacccggccgctgccgtgtacgtcg

[0045] atgcggggcattcgcgctggctgagcgccgaggcaatggccgccaggctcaacgatgtcggtgtgggccgcgcgcgc

[0046] gggtttagcctcaacgtctcgaacttctacaccaccgatgaggaaatcggctatggcgaggcgatttcggggctcacgaa

[0047] cggttcgcattacgtgatcgacacgtcgcgcaacggcgccggacccgcgcccgacgccccgctcaactggtgtaaccc

[0048] cagcggccgcgccctgggcgcaccgcccaccacggcgaccgcgggcgcgcacgccgacgcttacctgtggatcaaa

[0049] cgtcccggggaatcggacggaacctgcggtcgcggggagcctcaggcgggtcggttcgttagccagtacgccatcgat

[0050] ctggcccacaacgccggccagtag(SEQ ID NO.2)

[0051] The amino acid sequence of Rv0062p is:

[0052] NPLAGKPFYVDPASAAMVAARNANPPNAELTSVANTPQSYWLDQAFPPATVGGTVARYTGAAQAAGAMPVLTLYGIPHRDCGSYASGGFATGTDYRGWIDAVASGLGSSPATIIVEPDALAMADCLSPDQRQERFDLVRYAVDTLTRDP AAAVYVDAGHSRWLSAEAMAARLNDVGVGRARGFSLNVSNFYTTDEEIGYGEAISGLTNGSHYVIDTSRNGAGPAPDAPLNWCNPSGRALGAPPTTATAGAHADAYLWIKRPGESDGTCGGEPQAGRFVSQYAIDLAHNAGQ(SEQID NO.1)

[0053] The Rv0062p coding gene was amplified by PCR using the genomic DNA of Mycobacterium tuberculosis H37Rv as a template. The PCR reaction system was: 5×PCR buffer 10μL, dNTPs 4μL, DMSO 5μL, upstream primer 1μL, downstream primer 1μL, H37Rv Genomic DNA 1μL, TransStart FastPfu Fly DNA polymerase 1μL, ddH 2 O 27 μL.

[0054] The PCR amplification conditions were: 95°C for 2 min, followed by 30 cycles of: 95°C for 20 s, 56°C for 20 s, and 72°C for 20 s. After the cycle, 72°C for 5 min and standby at 4°C.

[0055] After the PCR reaction was completed, the PCR products were separated by 1% agarose gel electrophoresis and recovered using a DNA agarose gel recovery kit.

[0056] 2. Construction of recombinant cloning plasmid pJET-Rv0062p

[0057] Ligation and transformation: Ligate the Rv0062p PCR product recovered above with the pJET1.2 blunt vector. The 10μL reaction system includes 5μL 2×Ligation buffer, 3μL recovered Rv0062p PCR product, 1μL pJET1.2 blunt vector and 1μL T4 ligase. The ligation reaction is carried out at room temperature for 20 minutes. Take 5μL of the ligation product and transform it into E.coli Novablue competent cells, spread it on LB solid medium containing 50μg / mL ampicillin, and culture it at 37℃ for 16-18 hours.

[0058] Identification: Pick a single bacterial colony grown on the solid culture medium, inoculate it into 3mL of LB liquid culture medium containing 50μg / mL ampicillin, and shake and culture it at 37℃ for 16-18 hours. Use FastPure Plasmid Mini Kit to extract the plasmid, and use restriction endonucleases BamHI&NdeI, NotI for digestion and identification, and detect the digestion product by 1% agarose gel electrophoresis. The plasmid with correct digestion identification was sent to Beijing Liuhe BGI Gene Technology Co., Ltd. for nucleotide sequence determination of the Rv0062p gene.

[0059] 3. Construction of recombinant expression plasmid pET16b-Rv0062p

[0060] The pJET-Rv0062p plasmid and pET16b vector that were sequenced correctly were double-digested with BamHI and NdeI, and the Rv0062p gene fragment and the linear pET16b vector were recovered and connected by ligase. Take 5 μL of the ligation product and transform it into E. coli Novablue competent cells. Pick a single bacterial colony grown on the solid culture medium, inoculate it into 3 mL of LB liquid culture medium containing 100 μg / mL ampicillin, shake and culture at 37°C for 16-18 hours, and extract the plasmid using FastPure PlasmidMini Kit. Double digestion identification was performed using BamHI and NdeI, and the digestion product was detected by 1% agarose gel electrophoresis.

[0061] 4. Expression, purification and concentration of Rv0062p protein

[0062] Expression: The correctly identified pET16b-Rv0062p recombinant plasmid was transformed into E. coli BL21 (DE3) to construct the E. coli recombinant strain Ec0062p overexpressing the Rv0062p protein. 1 mL of freshly cultured Ec0062p bacterial solution was inoculated into 100 ml of LB liquid medium containing 100 μg / mL ampicillin. After shaking culture at 37°C for 2.5 hours, the inducer IPTG was added to a final concentration of 1 mM, and continued to shake culture at 37°C for 3 hours to induce the expression of Rv0062p protein. After the induction expression was completed, the bacteria were collected by centrifugation at 4000g, 8 mL of lysis buffer (20mM Tris-HCl, pH 8.0, 500mM NaCl, 20% glycerol, 1mM PMSF) was added, and the bacteria were ultrasonically broken on ice. Centrifuged at 12000g for 20 minutes to separate the supernatant and the precipitate. The expression of the protein was detected by SDS-PAGE. The correctness of protein expression was confirmed by Western blot using (anti)-polyhistidine clone HIS-1 antibody.

[0063] Purification and concentration: The Rv0062p protein in the supernatant was purified using Ni-NTA Superflow affinity chromatography. The purification process is as follows: The sample was loaded onto the filler and the column was washed with 20 mL of washing buffer (lysis buffer + 40 mM imidazole). The Rv0062p protein was eluted with 10 mL of elution buffer (lysis buffer + 200 mM imidazole), and 1.0 mL of the eluate was collected per tube. The purification effect was identified by SDS-PAGE electrophoresis, and the Rv0062p protein was concentrated using a 3K cut-off centrifugal concentrator tube, and the replacement buffer was 20 mM Tris-HCl. After sterilization by filtration with a 0.22 μm filter membrane, the Rv0062p protein was quantified using a BCA protein quantification kit.

[0064] 5. Degradation of bacterial biofilm on medical catheters by Rv0062p protein

[0065] Growth of bacterial biofilm on urinary catheters and degradation of Rv0062p protein: Clinically isolated Escherichia coli and Pseudomonas aeruginosa were cultured in Ec broth and NA medium, respectively. Take 20 μL of freshly cultured bacterial culture (OD600 is about 0.4) and add it to 1 mL of fresh culture medium and mix well. Take 80 μL of this bacterial dilution and add it to a 96-well plate, and add a 0.5 cm sterile latex urinary catheter at the same time. After sealing with sealing film, place it at 37℃ and culture it for 60 hours. Discard the planktonic bacterial solution, wash it once with sterile PBS, and add 80 μL of fresh culture medium. Finally, add 20 μL of 20mM Tris-HCl buffer or 20 μL of 0.5mg / mL Rv0062p fresh culture medium and incubate it at 37℃ for 24 hours.

[0066] Crystal violet staining of biofilm on urinary catheter: Carefully remove the urinary catheter and use crystal violet to detect bacterial biofilm on the catheter. The specific operation is as follows: Place the urinary catheter in a 48-well plate and wash it twice with sterile PBS. Add 200μL of 0.1% crystal violet dye and leave it at room temperature for 15 minutes. Wash it once with sterile PBS and take a picture.

[0067] Detection of floating bacteria: After the catheter is removed, take a photo, take 100 μL of floating bacteria and add it to a 96-well plate, and use an ELISA reader to read the OD600 of the floating bacteria.

[0068] 3. Results and Discussion

[0069] 1. Amplification of the gene encoding Rv0062p

[0070] In this case, the gene encoding the functional domain Rv0062p of the Rv0062 protein of Mycobacterium tuberculosis H37Rv strain was amplified by PCR. Figure 1 As shown, Figure 1 In the figure, lanes 1 and 2 are Rv0062p PCR amplification products, and lane M is GeneRuler1kbplus DNALadder.

[0071] Figure 1 The results showed that a PCR amplification product with the expected size of about 895 bp was obtained.

[0072] 2. Construction of recombinant cloning plasmid pJET-Rv0062p

[0073] In this case, the PCR amplification product of the Rv0062p gene was connected to the cloning vector pJET1.2 blunt to construct the recombinant cloning plasmid pJET-Rv0062p, and restriction endonucleases were used for digestion identification. The results are as follows: Figure 2 shown. Figure 2It is a diagram showing the result of enzyme digestion identification of the pJET-Rv0062p recombinant cloning plasmid, wherein lanes 1-4 are the results of BamHI&NdeI digestion of pJET-Rv0062p cloning plasmid numbers 1, 2, 5 and 7, lanes 5-8 are the results of NotI digestion of pJET-Rv0062p cloning plasmid numbers 1, 2, 5 and 7, and lane M is GeneRuler 1kbplus DNALadder.

[0074] Figure 2 The results showed that after double digestion with BamHI & NdeI and single digestion with NotI, pJET-Rv0062p recombinant cloning plasmids #1, 2, and 5 (corresponding to Figure 2 DNA fragments of the expected size appeared in lanes 1, 2, 3 and lanes 5, 6, and 7 in the Figure 1, indicating the correctly constructed recombinant cloning plasmid pJET-Rv0062p.

[0075] The nucleotide sequence of Rv0062p in pJET-Rv0062p#1 was performed, and the sequencing result was compared with the Rv0062p gene sequence. Figure 3 As shown, Figure 3 In the figure, 1 is the sequencing result of Rv0062p in the pJET-Rv0062p recombinant cloning plasmid, and 2 is the nucleotide sequence of the Rv0062p gene.

[0076] Figure 3 The results showed that the sequencing results were 100% similar to the Rv0062p gene sequence, indicating that the Rv0062p gene sequence amplified by PCR and connected to the cloning vector did not have any mutations.

[0077] 3. Construction of recombinant expression plasmid pET16b-Rv0062p

[0078] In this example, the Rv0062p gene in pJET-Rv0062p#1 that was correctly identified by sequencing was subcloned into the pET16b vector to construct the recombinant expression plasmid pET16b-Rv0062p, and restriction enzyme digestion was performed for identification. The results are as follows: Figure 4 As shown, Figure 4 The results are enzyme digestion identification results of the pET16b-Rv0062p recombinant expression plasmid, wherein lanes 1 to 4 are the results of four selected recombinant clone plasmids, and lane M is GeneRuler 1kb plus DNALadder.

[0079] Figure 4 The results showed that after double digestion with BamHI and NdeI, pET16b-Rv0062p#1, 2, 3, 4 (corresponding to Figure 4Lanes 1, 2, 3, and 4 in the figure are all correctly constructed recombinant cloning plasmids pJET-Rv0062p.

[0080] 4. Expression, purification and concentration of Rv0062p protein

[0081] In this example, the correctly identified pET16b-Rv0062p#1 plasmid was transformed into the E. coli BL21 (DE3) strain to construct the Rv0062p high-expression strain Ec0062p. The expression of Rv0062p was induced by IPTG, and the bacteria were collected and ultrasonically disrupted to separate the supernatant and precipitate. The results were detected by SDS-PAGE and Western blot. Figure 5 As shown in Figure A, in Figure A, 1 is the supernatant after ultrasonic disruption and centrifugation of Ec0062p bacteria, 2 is the precipitate after ultrasonic disruption and centrifugation of Ec0062p bacteria, and M is a protein marker.

[0082] Figure 5 The results in Figure A show that Rv0062p is well expressed in the recombinant strain Ec0062p, and soluble Rv0062p protein is expressed in the supernatant.

[0083] Because the expressed Rv0062p has a histidine tag, we also performed Western blot identification using an anti-histidine antibody. The results are shown in Figure 5 As shown in Figure B, in Figure B, 1 is the supernatant after ultrasonic disruption and centrifugation of Ec0062p bacteria, 2 is the precipitate after ultrasonic disruption and centrifugation of Ec0062p bacteria, and M is a protein marker.

[0084] Figure 5 The results in Figure B show that the protein highly expressed in the Ec0062p strain is indeed the Rv0062p protein with a histidine tag.

[0085] We purified the Rv0062p protein using Ni-NTA affinity chromatography and concentrated it using a centrifugal concentrator. At the same time, we replaced the buffer with 20mM Tris-HCl. We used SDS-PAGE to detect the purified and concentrated protein. The results are as follows: Figure 5 As shown in Figure C, in Figure C, 1 to 5 are the results of tubes numbered 1 to 5, M is the protein marker, and the arrow in the figure points to the Rv0062p protein.

[0086] Figure 5 The results in Figure C show that we obtained Rv0062p protein with good purity and correct size.

[0087] 5. Degradation of bacterial biofilm on medical catheters by Rv0062p

[0088] Sterile latex urine catheters were placed in 96-well plates, and freshly cultured Escherichia coli or Pseudomonas aeruginosa were added. After biofilm formation, 0.1 mg / mL Rv0062p protein was added. After 24 hours, the biofilm on the catheter was stained with crystal violet and the number of planktonic bacteria generated in the 96-well plate was used to identify the degradation effect of Rv0062p on the biofilm on the catheter. The results are as follows: Figure 6 Shown and Figure 7 shown. Figure 6 Figure A is an experimental model of Rv0062 degradation of bacterial biofilm on latex urine catheter; Figure B is a crystal violet staining showing the degradation effect of Rv0062p on bacterial biofilm formed on the catheter; Figure C is the observation result of suspended bacteria after degradation of bacterial biofilm; in the figure, E. coli is Escherichia coli, P. aeruginosa is Pseudomonas aeruginosa, CN is the control group, i.e., 20mM Tris-HCl was added, and Rv0062p protein treatment group, i.e., 0.1mg / mLRv0062p was added. Figure 7 The results of the ELISA test of suspended bacteria after degradation of bacterial biofilm; in the figure, E. coli is Escherichia coli, P. aeruginosa is Pseudomonas aeruginosa, CN is the control group, i.e., 20mM Tris-HCl was added, and Rv0062p protein treatment group, i.e., 0.1mg / mLRv0062p was added.

[0089] Figure 6 and Figure 7 The results showed that after treatment with Rv0062p protein, the biofilms of Escherichia coli and Pseudomonas aeruginosa on the catheter were significantly reduced, while the planktonic bacteria generated in the 96 wells increased significantly, indicating that Rv0062p protein effectively degraded the bacterial biofilm formed on the catheter.

[0090] The above results indicate that the cellulase Rv0062p produced by the prokaryotic expression system in this example can effectively degrade bacterial biofilms on medical catheters and can be used to prevent and treat diseases caused by bacterial biofilm infections.

[0091] The above contents are further detailed descriptions of the present application in combination with specific implementation methods, and it cannot be determined that the specific implementation of the present application is limited to these descriptions. For ordinary technicians in the technical field to which the present application belongs, several simple deductions or substitutions can be made without departing from the concept of the present application.

Claims

1. A use of cellulase in degrading bacterial biofilm, wherein the cellulase is the sequence shown in SEQ ID NO.

1.

2. Use of a cellulase in preparing an agent for degrading bacterial biofilm, wherein the cellulase has the sequence shown in SEQ ID NO.

1.

3. Use of a cellulase in the preparation of an agent for treating or preventing bacterial biofilm infection, wherein the cellulase has the sequence shown in SEQ ID NO.

1.

4. The use according to any one of claims 1 to 3, characterized in that: The bacterial biofilm is a biofilm formed by Escherichia coli and / or Pseudomonas aeruginosa.

5. Use of a cellulase in the preparation of a device for treating or preventing bacterial biofilm infection, wherein the cellulase has the sequence shown in SEQ ID NO.

1.

6. The use according to claim 5, characterized in that: The device includes a medical catheter.

7. The use according to claim 5 or 6, characterized in that: The bacterial biofilm includes a biofilm formed by Escherichia coli and / or Pseudomonas aeruginosa.

8. A reagent for degradation of bacterial biofilm, characterized in that: A cellulase containing the sequence shown in SEQ ID NO.

1.

9. A reagent for cleaning equipment, characterized in that: A cellulase containing the sequence shown in SEQ ID NO.

1.

10. The reagent according to claim 9, characterized in that: The device includes a medical catheter.

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