Construction of Enterococcus faecium -based recombinant Ef-PA0057 vaccine of Pseudomonas aeruginosa and its protection in mice
10.19405/j.cnki.issn1000–1492.2026.03.018
- VernacularTitle:屎肠球菌介导的铜绿假单胞菌重组Ef-PA0057疫苗的构建及其保护力观察
- Author:
Wengui LI
1
;
Xingkun OU
1
;
Ailin HE
1
Author Information
1. Chongqing Key Laboratory of Infectious and Parasitic Diseases， Institute of Infections and Parasitic Diseases， The First Affiliated Hospital， Chongqing Medical University， Chongqing 400016
- Publication Type:Journal Article
- Keywords:
Enterococcus faecium;
Pseudomonas aeruginosa;
PA0057;
vaccine;
humoral immunity
- From:
Acta Universitatis Medicinalis Anhui
2026;61(3):518-523
- CountryChina
- Language:Chinese
-
Abstract:
ObjectiveTo construct an Enterococcus faecium （Ef）-based recombinant Ef-PA0057 vaccine of Pseudomonas aeruginosa（Pa）and to study its protective immune mechanism in mice. MethodsThe PA0057 gene was cloned from the genomic DNA of PA01 strain ATCC9027 by PCR and inserted into pGEX-1λT to construct pGEX-PA0057. The recombinant plasmid was electroporated into TX0016 strain to construct rEf-PA0057 vaccine. The plasmid was extracted from rEf for PCR. The rEf vaccine was expressed through IPTG induction， and the expression of protein was analyzed by SDS-PAGE and Western blot. BALB/c mice were immunized intragastrically with 5×108 CFU rEf-PA0057 vaccine 3 times per week for 3 weeks. 4 weeks after the first immunization， mice were challenged intranasally with 5×107 CFU of PA01 strain. 2 weeks after challenge， mice were sacrificed， and their lungs were separated. Bacteria in lungs were incubated and colonies were counted. Sera were collected at 0， 4， and 6 weeks after the first immunization. The IgG and its subclasses， and IgE were detected by ELISA. ResultsThe 900 bp PA0057 gene was successfully cloned by PCR. PCR showed that PA0057 gene was amplified when the extracted plasmid from rEf as template； the relative molecular mass （Mr） of the expressed PA0057-GST fusion protein was approximately 58 ku， detected by SDS-PAGE. The amount of the expressed protein was 18% of the total bacterial proteins. Western blot showed that the target protein could be recognized by Pa sera. The colony numbers of lung tissue in rEf-PA0057 vaccine group， blank vector group and Ef control group were （0.297±0.011）×108 CFU， （7.576±0.206）×108 CFU and （7.551±0.185）×108 CFU， respectively， the difference was statistically significant （P0.01）. The levels of IgG， IgG1， IgG2b， IgG3 and IgE increased. At the same time point， there was a significant difference compared with the two control groups （P0.01）. ConclusionThe rEf-PA0057 vaccine is successfully constructed. It may induce mice to produce humoral response against challenge with PA01.
