Proteome Analysis and Serological Characterization of Surface-Exposed Proteins of Rickettsia heilongjiangensis

Abstract

Background Rickettsia heilongjiangensis, the agent of Far-Eastern spotted fever (FESF), is an obligate intracellular bacterium. The surface-exposed proteins (SEPs) of rickettsiae are involved in rickettsial adherence to and invasion of host cells, intracellular bacterial growth, and/or interaction with immune cells. They are also potential molecular candidates for the development of diagnostic reagents and vaccines against rickettsiosis.Methods R. heilongjiangensis SEPs were identified by biotin-streptavidin affinity purification and 2D electrophoreses coupled with ESI-MS/MS. Recombinant SEPs were probed with various sera to analyze their serological characteristics using a protein microarray and an enzyme-linked immune sorbent assay (ELISA).ResultsTwenty-five SEPs were identified, most of which were predicted to reside on the surface of R. heilongjiangensis cells. Bioinformatics analysis suggests that these proteins could be involved in bacterial pathogenesis. Eleven of the 25 SEPs were recognized as major seroreactive antigens by sera from R. heilongjiangensis-infected mice and FESF patients. Among the major seroreactive SEPs, microarray assays and/or ELISAs revealed that GroEL, OmpA-2, OmpB-3, PrsA, RplY, RpsB, SurA and YbgF had modest sensitivity and specificity for recognizing R. heilongjiangensis infection and/or spotted fever.ConclusionsMany of the SEPs identified herein have potentially important roles in R. heilongjiangensis pathogenicity. Some of them have potential as serodiagnostic antigens or as subunit vaccine antigens against the disease.

Diversity of Rickettsiales in the Microbiome of the Lone Star Tick, Amblyomma americanum

“…The predicted haplotype of sca0 from our six ticks was identical to 15 previously reported sca0 sequences from Missouri (GenBank accession numbers EU544293 and EU544294), Tennessee (EU544295), and Maryland (EF450685 to EF450696 [36]) and had only rare differences with the rest. Thus, there is a surprising absence of diversity in this gene despite the fact that its product is highly immunogenic (36,37).…”

Detecção de proteínas imunorreativas de Rickettsia sp. cepa Mata Atlântica

“…Pornwiroon et al (2009) identificaram novas proteínas imunorreativas para R. parkeri, entre elas reconhecidas por soro de pacientes humanos, caracterizando-as como possíveis alvos potenciais para o diagnóstico e prevenção da doença. Qi et al (2013) identificaram proteínas de superfície expostas (SEPs) em Rickettsia heilongjiangensis, através de soro de camundongos infectados, onde OmpA-2 e GroEL obtiveram maior sororreatividade, demonstrando assim o potencial para o desenvolvimento de testes sorológicos eficazes.…”

Rickettsia rickettsiiouter membrane protein YbgF induces protective immunity in C3H/HeN mice

“…YbgF, a periplasmic protein, is an abundant surface protein and major sero-reactive antigen of rickettsiae. 16,17 It is thought to function in maintaining the cell envelope integrity of Gram-negative bacteria. 18 TolC is thought to be a bioactive transport molecule that couples with different inner membrane/ periplasmic proteins.…”

“…Each serum sample was serially diluted 2-fold for the determination of specific antibody titers by ELISA. 17 The cutoff value was set according to a previous study. 14 …”

“…17 YbgF and TolC on the surface of R. rickettsii were detected using IFA as previously described. 17 For TEM analysis, R. rickettsii cultured in Vero cells were fixed (0.5% glutaraldehyde, 4% paraformaldehyde, 0.1 M sodium cacodylate, 0.3% picric acid, pH 7.4) for 4 h on ice. After four washes in 0.1 M PBS, fixed cells were dehydrated with alcohol and permeabilized with a mix of LR White (Spi Supplies) and alcohol.…”