Manoochehr Rasouli; Ahmad Zavaran Hoseini; Bahram Karimi; Abdolvahab Alborzi; Simin Kiany
Volume 6, Issue 2 , June 2009, , Pages 75-86
Abstract
Background: Heat shock protein 70 (HSP70) is present in all organisms studied so far, and is a major immunogen in infections caused by pathogens including Leishmania spp. Objective: The aim of this study was to clone and express HSP70 from L. infantum strain MCAN/IR/96/LON-49 and evaluate antibody response ...
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Background: Heat shock protein 70 (HSP70) is present in all organisms studied so far, and is a major immunogen in infections caused by pathogens including Leishmania spp. Objective: The aim of this study was to clone and express HSP70 from L. infantum strain MCAN/IR/96/LON-49 and evaluate antibody response against HSP70 in visceral leishmaniasis (VL). Methods: The L. infantum HSP70 gene segment was amplified by specific primers. It was cloned into pTZ57R vector and subcloned into pET32a (+) expression vector. The new construct was transformed in the E.coli Rosetta strain, and HSP70 protein was expressed in the presence of 1 mM IPTG and purified using a HiTrap chelating column. Antibody responses against HSP70 were determined by ELISA in 37 patients with visceral leishmaniasis and 63 healthy controls. Results: Expression of HSP70 protein was confirmed using SDS-PAGE electrophoresis and dot blot with an anti-His tag antibody. There was no difference between the sequence of nucleotides of the HSP70 gene in the present study and other reported sequences. The ELISA results indicated that the sera of 81.1% (30/37) of the patients and 6.3% (5/63) of controls reacted to L. infantum HSP70. Conclusion: The conservative nature of the HSP70 molecule is an advantage in vaccine studies, because of minor differences (6%) between the nucleotide sequences and consequently the similarity in amino acid sequences in various strains of L. infantum. It could therefore be used in vaccine research against leishmaniasis and also as a tool for serodiagnosis.
Manoochehr Rasouli; Simin Kiany; Abdolvahhab Alborzi
Volume 2, Issue 4 , December 2005, , Pages 226-231
Abstract
Background: Brucella is a gram-negative bacterium, causing acute and chronic infection in humans and animals. Cell-mediated immunity is the main protective immune response against Brucella spp. Activation of macrophages by IFN-γ and generation of reactive oxygen intermediates and nitric oxide are ...
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Background: Brucella is a gram-negative bacterium, causing acute and chronic infection in humans and animals. Cell-mediated immunity is the main protective immune response against Brucella spp. Activation of macrophages by IFN-γ and generation of reactive oxygen intermediates and nitric oxide are the main immunologic mechanisms responsible for control of Brucella infection. Objective: To investigate the correlation between IFN-γ gene polymorphism and brucellosis. Methods: 195 patients with brucellosis, 186 healthy patients' family members and 82 healthy farmers who kept infected animals and consumed their contaminated dairy products were selected to take part in the study. IFN-γ genotyping at position +874 (T→A) was carried out by allele specific polymerase chain reaction (AS-PCR) method. Results: The frequency of AT and TT genotypes significantly increased in farmers compared to patients with brucellosis (P=0.03) while there was no significant difference in genotype distribution between patients and their healthy family members. Conclusion: IFN-γ (+874) AA genotype is probably a genetic factor that contributes to the susceptibility of the individuals to brucellosis.