Evaluation of immune responses induced by polymeric OMP25-BLS Brucella antigen

Yousefi, Soheil; Abbassi-Daloii, Tooba; Sekhavati, Mohammad Hadi; Tahmoorespur, Mojtaba

doi:10.1016/j.micpath.2017.12.045

Cited by 16 publications

(13 citation statements)

References 61 publications

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“…The molecular genotyping techniques used for the differentiation of Brucella species showed a significant DNA polymorphism. One of these loci is the omp2 gene, which encodes the 36 kDa outer membrane protein 2 of Brucella species (Yousefi et al, 2016;Yousefi et al, 2018). This gene reveals diversities among different isolates and special primers have been designed for the identification of Brucella species, biovars, and strains (Cloeckaert et al, 2002;He, 2012;Kaden et al, 2017).…”

Section: Discussionmentioning

confidence: 99%

Molecular Detection of Outer Membrane Protein 2 Gene in Isolated Brucella from Sheep and Goats in Sharazor/Kurdistan Region of Iraq

Saeed¹

2019

PVJ

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Brucellosis is an economically important zoonotic disease that is endemic in different parts of Iraq. An outbreak of abortion occurred in sheep and goats in Sharazor subdistrict, Sulaymaniyah, Iraq and Brucella infection was suspected to be the cause. Between November 1, 2014, and January 31, 2015, 47 tissue and vaginal swab samples were collected from 17 herds. Samples from each herd were mixed together to make 17 pooled samples. The Brucella genus-specific primer Bruc was used to amplify the 223 bp amplicon of bcsp31 gene. Fifteen of the 17 samples (88%) were positive for Brucella genus. Thereafter, five samples were randomly selected to identify the species by designing four sets of Brucella-species-specific primers. Sequencing of the Outer Membrane Protein 2 (omp2) gene was performed and the sequences were aligned with others in the GenBank databases. The results showed that the omp2 gene of the five isolates from Sharazor had 99-100% similarities with Brucella melitensis bv. 2 that was isolated from United States (accession number CP007789) and B. melitensis strain 20236 (accession # CP008750) isolated from human in China. The amino acid sequences of the omp2a gene in the five isolates were similar with other isolates in GenBank with the exception of B. suis bv. 2 (accession # CP007720). This study reported the occurrence of B. melitensis for the first time in Sharazor subdistrict, Sulaymaniyah, northern Iraq, using molecular techniques. ©2019 PVJ. All rights reserved Key words: Brucella melitensis Brucellosis Outer membrane proteins Phylogenetic analysis To Cite This Article: Saeed NM, Talib SM and Dyary HO, 2019. Molecular Detection of outer membrane protein 2 gene in isolated Brucella from sheep and goats in Sharazor/Kurdistan Region of Iraq. Pak Vet J, 39(3): 383-388. http://dx.

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Section: Discussionmentioning

confidence: 99%

Molecular Detection of Outer Membrane Protein 2 Gene in Isolated Brucella from Sheep and Goats in Sharazor/Kurdistan Region of Iraq

Saeed¹

2019

PVJ

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“…The development of recombinant DNA techniques along with protein engineering science could develop fourth-generation vaccines, which only use immunogenic component of the pathogens. These recombinant vaccines, through their ability to stimulate appropriate responses and immunization, can be considered as good alternatives compared to traditional vaccines (15). Brucella is able to circumvent the host's innate and adaptive immunity and as an intercellular pathogen, it can disrupt T cell energy and TCL through different pathways.…”

Section: Discussionmentioning

confidence: 99%

Engineering, Cloning and Expression of DNA Sequence Coding of OMP31 Epitope of Brucella melitensis linked to IL-2 in Escherichia coli

et al. 2018

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Brucellosis is a zoonetic disease that is caused by Brucella melitensis, which affects human health and animal husbandry industry. Omp31 is the most exposed outer membrane proteins (OMPs) and is a dominant antigen in smooth strains of B. melitensis, thus it is considered a desirable protein in subunit vaccine against brucellosis disease. Besides, IL-2 acts as a growth and differentiation stimulator of T-cells in cellular immune responses, therefore, it can serve as an adjuvant in subunit vaccines against intracellular pathogens. It seems that chimera scaffold, consisting of some copies of OMP31 experimental epitope region and IL-2, can provide a good immunization. In this regard, these tow fragments are linked together through using rigid linker and splicing overlap extension (SOE) PCR technique. Successful PCR products were TA cloned and then sub-cloned into pET-22b (+) vector as an expression vector. Recombinant protein was expressed in Escherichia coli BL21 (DE3) as an expressing lineage bacterium and then confirmed by SDS-PAGE and Western-blotting analysis. Secondary and tertiary structure prediction of recombinant protein and its immunity characterization were evaluated using in silico process. Further studies on the immunogenicity properties of this protein are expected to be carried out in the future to confirm the results in vivo.

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“…Antibody responses, cytokines determination, and lymphocyte proliferation assay were performed as described previously [16]. Briefly, the mice were slaughtered 2 weeks after lasted injection for serum collection by centrifugation at 3000 g for 20 min and the supernatant was stored at −80°C.…”

Section: Immunogenicity Assessmentmentioning

confidence: 99%

“…Cell culture supernatants were collected at the end of the incubation and centrifuged at 300 g for 10 min. Interferon gamma (INF-γ), tumor necrosis factor alpha (TNFα), and interleukin-4 (IL-4) levels were measured by a sandwich ELISA according to the manufacturer's instructions (Mabtech, Nacka, Sweden) [16]. To lymphocyte proliferation assay, spleens were dissected from the mice and suspended in sterile cold PBS containing 2% FBS.…”

Section: Immunogenicity Assessmentmentioning

confidence: 99%

“…Red blood cells were lysed with lysis buffer and the single-cell suspension was adjusted to 3 × 10 6 cells/ml and dispensed into a 96-well plate in triplicate and incubated with 10 µg/ ml of the recombinant proteins for 48 h. Incubation was continued by 20 µl 3-4,5-dimethylthiazol-2-yl-2,5-diphenyltetrazolium bromide (MTT) (Sigma), followed by 100 µl of dimethyl sulfoxide (Sigma). Absorbance was measured using a spectrophotometric plate reader at 590 nm [16].…”

Section: Immunogenicity Assessmentmentioning

confidence: 99%

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The antigenicity performance of divalent recombinant B. melitensis vaccines versus univalent ones

Abbassi-Daloii

Yousefi

Tahmoorespur

et al. 2019

Alexandria Journal of Medicine

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Brucellosis is an infectious disease caused by different Brucella species. The outer membrane proteins 25 and 31 play a significant role in stimulation of immunity against Brucella. Herein, the humoral and cellular immune responses of selected recombinant proteins emulsified in chitosan nanoparticles as individual (univalent) and simultaneous (divalent) injections were assessed. The humoral and cellular immune responses were measured by enzyme-linked immunosorbent assay in 12 groups (individual and simultaneous injections of rOMP25 and rOMP31 in different protein concentrations) and lymphocyte proliferation was measured using 3-4,5-dimethylthiazol-2-yl2,5-diphenyltetrazolium bromide (MTT) assay based on average Optical density (OD) of stimulated cells/average OD of unstimulated cells. Finally, data were analyzed using one-way analysis of variance. rOMP25 + rOMP31 group stimulated higher titer of INF-γ than other groups, whilst there were no statistically significant differences between all uni/divalent immunized groups. Tumor necrosis factor alpha titer showed no significant difference between divalent immunized groups (except rOMP31 + rOMP25(1) and rOMP31 + rOMP25(3)) and positive control group. Interleukin-4 analysis results demonstrated that there were no significant differences between positive control and uni/divalent vaccinated groups. In addition, analysis of antibody responses revealed rOMP25 + rOMP31, rOMP25 + rOMP31(2), and rOMP25 + rOMP31(3) groups induced higher level of total antibody compared to other immunized groups, although both univalent and divalent immunized mice induced higher IgG2a titer than IgG1 with the mean of IgG2a/IgG1 ratio~1.01 indicating strong bent of Th1 immune response. The cell proliferation assay demonstrated the vaccination with rOMP25 + rOMP3, rOMP25 + rOMP31(2), and rOMP25 + rOMP31(3) elicited vigorous antigen-specific cell proliferative. rOMP25 + rOMP3, rOMP25 + rOMP31(2), and rOMP25 + rOMP31(3) treatments could be used as potential candidates for developing new subunit vaccines.

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Evaluation of immune responses induced by polymeric OMP25-BLS Brucella antigen

Cited by 16 publications

References 61 publications

Molecular Detection of Outer Membrane Protein 2 Gene in Isolated Brucella from Sheep and Goats in Sharazor/Kurdistan Region of Iraq

Molecular Detection of Outer Membrane Protein 2 Gene in Isolated Brucella from Sheep and Goats in Sharazor/Kurdistan Region of Iraq

Engineering, Cloning and Expression of DNA Sequence Coding of OMP31 Epitope of Brucella melitensis linked to IL-2 in Escherichia coli

The antigenicity performance of divalent recombinant B. melitensis vaccines versus univalent ones

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