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CN-122012417-A - Acinetobacter baumannii phage, depolymerizing enzyme, and preparation methods and applications thereof

CN122012417ACN 122012417 ACN122012417 ACN 122012417ACN-122012417-A

Abstract

The invention relates to the technical field of phages, and particularly discloses a Acinetobacter baumannii phage, a depolymerizing enzyme, a preparation method and application thereof. The Acinetobacter baumannii bacteriophage P919 provided by the invention has strong specific cracking capacity on the Acinetobacter baumannii of the KL3 type, and the encoded depolymerizing enzyme has excellent temperature stability below 70 ℃, so that the capsular polysaccharide of the Acinetobacter baumannii of the KL3 type can be effectively degraded.

Inventors

  • ZHANG WEI
  • YAN XIAONA
  • XU SIXIANG
  • Zheng Xiangkuan
  • TAN ZHONGMING
  • DU HONG
  • ZHENG YISHAN
  • CHEN LONG
  • LI CHENXI
  • LI PEI
  • Fang Huanxin

Assignees

  • 南京农业大学三亚研究院

Dates

Publication Date
20260512
Application Date
20260414

Claims (10)

  1. 1. The Acinetobacter baumannii phage (Acinetobacter baumannii phage) P919 is characterized in that the Acinetobacter baumannii phage P919 is preserved in China Center for Type Culture Collection (CCTCC) No. M20252192 at the 10 th month 11 of 2025.
  2. 2. A phage depolymerizing enzyme gene, which is isolated from Acinetobacter baumannii phage P919 as set forth in claim 1, and has a sequence shown in SEQ ID No. 1.
  3. 3. A phage depolymerizing enzyme P919DPO, which is encoded by the phage depolymerizing enzyme gene of claim 2, and has an amino acid sequence shown in SEQ ID NO. 2.
  4. 4. The method for producing phage depolymerizing enzyme P919DPO according to claim 3, comprising the steps of: Cloning a phage depolymerizing enzyme P919DPO gene fragment shown in SEQ ID No.1, connecting the P919DPO gene fragment to a pET28a plasmid vector to obtain a recombinant plasmid, transferring the recombinant plasmid into a host cell, screening to obtain a recombinant host cell containing the recombinant plasmid, culturing the recombinant host cell to obtain a culture solution, and collecting liquid after solid-liquid separation of the culture solution to obtain the phage depolymerizing enzyme P919DPO.
  5. 5. The method for producing phage depolymerizing enzyme P919DPO according to claim 4, wherein said host cell is E.coli DE3.
  6. 6. The method for producing phage depolymerizing enzyme P919DPO according to claim 4, wherein the culture conditions are 37-38 ℃, 180-200 rpm.
  7. 7. The method for preparing phage depolymerizing enzyme P919DPO according to claim 4, wherein phage depolymerizing enzyme P919DPO gene fragment shown in SEQ ID NO.1 is amplified by using phage P919 genome DNA as a template and SEQ ID NO. 3-SEQ ID NO.4 as a primer.
  8. 8. The use of acinetobacter baumannii bacteriophage P919 according to claim 1 or phage depolymerase P919DPO according to claim 3 for preparing KL 3-type acinetobacter baumannii disinfection products, wherein said acinetobacter baumannii bacteriophage P919 or phage depolymerase P919DPO is used for specifically degrading KL 3-type acinetobacter baumannii capsular polysaccharide.
  9. 9. The use according to claim 8, wherein said acinetobacter baumannii bacteriophage P919 or said bacteriophage depolymerase P919DPO promotes phagocytosis thereof by macrophages by degrading the KL3 acinetobacter baumannii capsular polysaccharide.
  10. 10. The use according to claim 9, wherein the biocidal product is a phage depolymerase P919DPO solution at a concentration of 1.5 μg/mL to 150 μg/mL.

Description

Acinetobacter baumannii phage, depolymerizing enzyme, and preparation methods and applications thereof Technical Field The invention relates to the technical field of phages, in particular to a Acinetobacter baumannii phage, a depolymerizing enzyme, a preparation method and application thereof. Background Acinetobacter baumannii is widely existed in natural environment, is an opportunistic pathogen, can infect people, various aquatic animals, domestic fowls and wild animals, and seriously threatens public health safety. This pathogen is the leading cause of nosocomial infections in humans and can cause pneumonia, bacteremia, endocarditis, skin and soft tissue infections, urinary tract infections, meningitis, etc. The main difficulties in treating acinetobacter baumannii infections are their multiple antimicrobial resistance and numerous capsule types. In recent years, such pathogens have developed a broad range of resistance to most first-line antibiotics used in clinical practice. Of these 70% are carbapenem-resistant Acinetobacter baumannii CRAB, while carbapenem antibiotics are considered the last line of defense against gram-negative bacteria. Mortality from hospital acquired pneumonia and blood stream infections caused by CRAB has been reported to be nearly 70%. Antibacterial drugs (such as polymyxin and tigecycline) currently used to treat CRAB are only the last therapeutic means due to their pharmacokinetic properties and toxicity. One of the key virulence factors of acinetobacter baumannii is capsular polysaccharide, which plays a key role in evading the host immune system, resisting desiccation and enhancing drug resistance. Therefore, CPS is also a key target for the treatment of Acinetobacter baumannii infection. At least 237 k-site reference sequences have been identified and assigned more than 65 k-forms. Among them, KL3, KL2, KL9, KL13 and KL49 capsule types are most common. Carbapenem-resistant acinetobacter baumannii lacks therapeutic means due to severe antibiotic resistance. It is necessary to develop a product that specifically degrades the capsule to promote the immune system to kill acinetobacter baumannii of KL3 type. Disclosure of Invention The invention provides a Baoman acinetobacter baumannii bacteriophage and a depolymerizing enzyme as well as a preparation method and application thereof, and aims to develop a product for specifically degrading capsules to promote an immune system to kill KL3 type Baoman acinetobacter baumannii. The Acinetobacter baumannii bacteriophage P919 provided by the invention has strong specific cracking capacity on the Acinetobacter baumannii of the KL3 type, and the encoded depolymerizing enzyme has excellent temperature stability below 70 ℃, so that the capsular polysaccharide of the Acinetobacter baumannii of the KL3 type can be effectively degraded. The invention provides an Acinetobacter baumannii phage (Acinetobacter baumanniiphage) P919, wherein the Acinetobacter baumannii phage P919 is preserved in China center for type culture collection (China center for type culture collection) on 10-month 11 of 2025, and the preservation number is CCTCC NO: M20252192. The Acinetobacter baumannii bacteriophage P919 provided by the invention has strong specific cracking capacity on the Acinetobacter baumannii of the KL3 type, and the coded depolymerizing enzyme has excellent temperature stability below 70 ℃, can effectively degrade capsular polysaccharide of the Acinetobacter baumannii of the KL3 type, and promotes phagocytosis of macrophages on the Acinetobacter baumannii of the KL3 type. The invention also provides a phage depolymerizing enzyme gene which is separated from the acinetobacter baumannii phage P919, and the sequence of the phage depolymerizing enzyme gene is shown as SEQ ID NO. 1. The invention also provides a phage depolymerizing enzyme P919DPO, which is encoded by the phage depolymerizing enzyme gene, and the amino acid sequence of the phage depolymerizing enzyme is shown as SEQ ID NO. 2. The invention also provides a preparation method of phage depolymerizing enzyme P919DPO, which comprises the following steps: Cloning a phage depolymerizing enzyme P919DPO gene fragment shown in SEQ ID No.1, connecting the P919DPO gene fragment to a pET28a plasmid vector to obtain a recombinant plasmid, transferring the recombinant plasmid into a host cell, screening to obtain a recombinant host cell containing the recombinant plasmid, culturing the recombinant host cell to obtain a culture solution, and collecting liquid after solid-liquid separation of the culture solution to obtain the phage depolymerizing enzyme P919DPO. Further, the host cell is E.coli DE3. Further, the culture conditions are 37-38 ℃, 180-200 rpm. Further, the phage depolymerizing enzyme P919DPO gene fragment shown in SEQ ID NO.1 is obtained by amplifying phage P919 genome DNA serving as a template and SEQ ID NO. 3-SEQ ID NO.4 serving as primers. The invention also provides application of