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Immuno-fluorescence Assay of Leptospiral Surface-exposed Proteins
Published on: July 1, 2011
Cloning and sequencing of the ompL37 gene present in Leptospira interrogans, a surface protein in pathogenic
Elaheh Rezaei1, Pejvak Khaki1, Soheila Moradi Bidhendi1
1Department of Microbiology, Razi Vaccine & Serum Research Institute, Agricultural Research, Education and Extension Organization (AREEO), Karaj, Iran.
Background And Objectives:
Leptospirosis, an infection caused by pathogenic leptospires, is associated with insufficient sanitation and poverty. Leptospira is transmitted through contact with contaminated urine of reservoir animals. The primary objective of this study was to clone and sequence the ompL37 gene present in local and vaccine serovars.
Materials And Methods:
A total of 16 Leptospira interrogans serovars were cultured in EMJH liquid medium. After growing, genomic DNA was extracted using phenol-chloroform method. Primer pair was synthesized to amplify the 996 bp ompL37 sequence. The amplified ompL37 gene was cloned into pTZ57R/T vector. The sequences obtained from this study were compared with an only recorded sequence in the Genbank by the Meg Align software.
Results:
PCR products showed an amplified 996bp ompL37 gene product belonging to pathogenic serovars, while no ompL37 products were amplified in non-pathogenic serovars. Sequences comparison tests from 16 native serotypes examined in this study displayed a similarity range of 84% to 99.5% among serovars used. The results showed that two serotypes of L. interrogans including Serjoehardjo (RTCC2810 and RTCC2821) had the highest identity up to 95.5%. Two serovars of L. interrogans including Pomona (RTCC2822) and Icterohaemorrhagiae (RTCC2823) had the lowest identity about 84%.
Conclusion:
As the results showed, ompL37, present on the surface of such bacteria, showed a conserved sequence. ompL37, as a key role in cell adhesion and pathogenicity, can be used for designing diagnostic tests and vaccines. Furthermore, sequencing of various sites in ompL37 gene, including binding sites and immunogenic epitopes, can be valuable alternatives for future studies.
Insights
This study cloned and sequenced the Leptospira interrogans ompL37 gene, finding it conserved in pathogenic serovars. This ompL37 gene is crucial for developing new diagnostic tests and vaccines for leptospirosis.
Area of Science:
- Microbiology
- Genetics
- Molecular Biology
Background:
- Leptospirosis is a bacterial infection caused by pathogenic Leptospira.
- Transmission occurs via contact with contaminated animal urine, often linked to poor sanitation.
- The outer membrane protein L37 (OmpL37) is a potential virulence factor.
Purpose of the Study:
- To clone and sequence the ompL37 gene from local and vaccine Leptospira serovars.
- To analyze the genetic variability of ompL37 among different serovars.
Main Methods:
- Genomic DNA extraction from 16 Leptospira interrogans serovars.
- Polymerase Chain Reaction (PCR) amplification of the 996 bp ompL37 gene.
- Cloning of the amplified gene into a pTZ57R/T vector and sequence comparison using MegAlign software.
Main Results:
- The ompL37 gene was successfully amplified in pathogenic serovars but not in non-pathogenic ones.
- Sequence analysis revealed high similarity (84%–99.5%) among the ompL37 genes of the examined serovars.
- Specific serovars like Serjoehardjo showed higher identity (95.5%), while Pomona and Icterohaemorrhagiae showed lower identity (84%).
Conclusions:
- The ompL37 gene exhibits a conserved sequence across pathogenic Leptospira serovars.
- OmpL37 plays a key role in bacterial cell adhesion and pathogenicity.
- The conserved nature of ompL37 makes it a promising target for developing novel diagnostic tools and vaccines against leptospirosis.
