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Klebsiella pneumoniae capsular polysaccharide depolymerizing enzyme and application thereof

A technology of Klebsiella pneumoniae and capsular polysaccharide, applied in the field of medicine

Active Publication Date: 2021-12-07
SHANGHAI RUIZHOU BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, due to the host spectrum specificity of phages, the depolymerases discovered earlier could not produce satisfactory capsular cleavage on K19 capsular CRKP

Method used

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  • Klebsiella pneumoniae capsular polysaccharide depolymerizing enzyme and application thereof
  • Klebsiella pneumoniae capsular polysaccharide depolymerizing enzyme and application thereof
  • Klebsiella pneumoniae capsular polysaccharide depolymerizing enzyme and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0099] Example 1 Isolation and Identification of Bacterial Strains

[0100] Klebsiella pneumoniae 6570 isolated from the South Hospital of Renji Hospital Affiliated to Shanghai Jiaotong University School of Medicine was used as the host bacteria, and other Klebsiella pneumoniae clinical strains were from the South Hospital of Renji Hospital and Huashan Hospital Affiliated to Fudan University. According to Brisse (Wzi gene sequencing, a rapid method for determination of capsular type for Klebsiella strains. J. Clin. Microbiol. 51, 4073-4078.) method for the determination and typing of strain capsule types. These clinical isolates were cultured overnight in LB broth at 37°C and stored at -80°C with 50% glycerol.

[0101] Results: 24 clinical strains were isolated, all of which were resistant to multiple antibiotics, including cefuroxime (CXM), cefotaxime (CTX), amikacin (AMK), cefazolin (CZO), meropenem ( MEM), imipenem (IMP), gentamicin (GEN) and piperacillin (PIP). First,...

Embodiment 2

[0102] Example 2 Isolation of phage (SH-KP156570) infected with Klebsiella pneumoniae 6570 strain

[0103] Phages were isolated from biologically treated wastewater using Klebsiella pneumoniae clinical isolate 6570. The specific operation is as follows: the untreated water sample was centrifuged at 5000 rpm for 15 min, passed through a 0.22 μm filter membrane, and cultured overnight at 37°C with the bacterial solution infected with Klebsiella pneumoniae 6570 in LB liquid medium. After centrifugation the next day, filter the supernatant containing phages with a 0.22 μm filter, drop 5 μL of the filtrate on a double-layer agar plate covered with Klebsiella pneumoniae 6570, and culture overnight. Phage plaques with halos were found on the plate Piece. Plaques were picked with SM buffer (100 mM NaCl, 8 mM MgSO 4 ·7H 2 O, 50 mM Tris-HCl, pH = 7.5) after gradient dilution, incubated with the bacterial solution for five minutes, and then plated on 1.5% agar LB medium. After the upp...

Embodiment 3

[0105] Example 3 Determination of host spectrum

[0106] The host spectrum of SH-KP156570 was detected on 24 isolated clinical strains by spot test and double-layer agar plate test. Briefly, 400 μL of bacterial culture was mixed thoroughly with 4 mL of 0.5% agar LB, and plated on top of 1.5% agar LB medium. After the upper layer of agar solidified, 5 μL of the purified phage suspension was dropped on the bacterial lawn, and after overnight incubation at 37°C, the effect of the phage on the bacteria was observed.

[0107] Results: All bacteria produced plaques and halos after infection with phages, but phages could not lyse the Klebsiella pneumoniae K19 strain. The results showed that the SH-KP156570 phage was highly sensitive and specific to the Klebsiella pneumoniae K19 strain (Table 1).

[0108] Table 1. Cleavage profiles of phage SH-KP156570 and depolymerases Dpo7, Dpo41, Dpo42, Dpo43

[0109]

[0110] Note: ﹢ means there is an effect, - means no effect.

[0111] Dpo...

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Abstract

The invention provides a klebsiella pneumoniae capsular polysaccharide depolymerizing enzyme and application thereof. The klebsiella pneumoniae capsular polysaccharide depolymerizing enzyme provided by the invention can effectively decompose K19 type klebsiella pneumoniae exopolysaccharide and capsular, so that the contact between a drug or immune cell capable of killing klebsiella pneumoniae and the klebsiella pneumoniae is facilitated, and the drug effect is further exerted.

Description

technical field [0001] The invention relates to the field of medicine, in particular, the invention relates to Klebsiella pneumoniae capsular polysaccharide depolymerase and application thereof. Background technique [0002] Klebsiella pneumoniae is a common opportunistic pathogen and an important hospital-acquired infection pathogen. According to the epidemiological survey from January to June 2018, Klebsiella pneumoniae accounted for nearly 14% of the main clinical isolates, ranking second only to Escherichia coli. In addition, studies on the isolation rate of major Gram-negative bacilli over the years have found that the isolation rate of Klebsiella pneumoniae has shown an increasing trend. It can be colonized in the mouth, skin, intestinal tract and other parts, and can cause sepsis, pneumonia, urinary tract infection, and has important clinical research value. [0003] Due to the extensive use of antibiotics in clinical practice in recent years, a variety of bacteria ...

Claims

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Application Information

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IPC IPC(8): C12N9/18C12N15/56C12P19/14A61K38/46A61P31/04
CPCC12N9/18C12P19/14A61P31/04A61K38/00
Inventor 祝先潮何平马瑞景徐磊毛意芝方红燕
Owner SHANGHAI RUIZHOU BIOTECH CO LTD
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