Rezaei Elaheh, Khaki Pejvak, Moradi Bidhendi Soheila, Noofeli Mojtaba, Soltani Maryam Sadat
Department of Microbiology, Razi Vaccine & Serum Research Institute, Agricultural Research, Education and Extension Organization (AREEO), Karaj, Iran.
Department of Human Bacterial Vaccines, Razi Vaccine & Serum Research Institute, Agricultural Research, Education and Extension Organization (AREEO), Karaj, Iran.
Iran J Microbiol. 2019 Oct;11(5):373-378.
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 gene present in local and vaccine serovars.
A total of 16 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 sequence. The amplified 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.
PCR products showed an amplified 996bp gene product belonging to pathogenic serovars, while no 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 including Serjoehardjo (RTCC2810 and RTCC2821) had the highest identity up to 95.5%. Two serovars of including Pomona (RTCC2822) and Icterohaemorrhagiae (RTCC2823) had the lowest identity about 84%.
As the results showed, , present on the surface of such bacteria, showed a conserved sequence. , as a key role in cell adhesion and pathogenicity, can be used for designing diagnostic tests and vaccines. Furthermore, sequencing of various sites in gene, including binding sites and immunogenic epitopes, can be valuable alternatives for future studies.
钩端螺旋体病是由致病性钩端螺旋体引起的一种感染性疾病，与卫生条件差和贫困有关。钩端螺旋体通过接触储存宿主动物的污染尿液传播。本研究的主要目的是克隆和测序本地血清型和疫苗血清型中存在的基因。
共16个血清型在EMJH液体培养基中培养。培养后，采用酚氯仿法提取基因组DNA。合成引物对以扩增996 bp的序列。扩增的基因克隆到pTZ57R/T载体中。本研究获得的序列通过Meg Align软件与Genbank中唯一记录的序列进行比较。
PCR产物显示扩增出996 bp的属于致病性血清型的基因产物，而非致病性血清型未扩增出产物。本研究检测的16种本地血清型的序列比较试验显示，所用血清型之间的相似性范围为84%至99.5%。结果表明，两种血清型的，包括Serjoehardjo（RTCC2810和RTCC2821）具有高达95.5%的最高同一性。两种血清型的，包括波摩那型（RTCC2822）和出血性黄疸型（RTCC2823）具有约84%的最低同一性。
结果显示，存在于此类细菌表面的，显示出保守序列。作为细胞黏附和致病性的关键作用因子，可用于设计诊断试验和疫苗。此外，对基因中各个位点进行测序，包括结合位点和免疫原性表位，可能是未来研究中有价值的选择。
