Measuring of free endotoxin in alum-precipitated vaccine of haemorrhagic septicaemia by limulus amebocyte lysate test

Summary Haemorrhagic septicaemia (HS) vaccine which is prepared in Razi Institute is used in endemic areas of Iran. Aluminum-hydroxide gel was used as adjuvant for preparing this vaccine. Post-vaccinal shock reactions were the main complaint after use of this vaccine. In a previous study, we could improve the vaccine by alum-precipitation Pasteurella multocida cells and removing the liquid phase. In this study, the amount of free endotoxin in aluminum-hydroxide and alum-HS vaccines was determined. It was found that endotoxin level was considerably decreased from 0.22 EU/ml to 0.03 EU/ml after alum-precipitation

A Serological and Molecular study on Toxoplasma gondii infection in sheep and goat in Tabriz

Toxoplasmosis is an important zoonotic diseases in human and animals. The disease caused by the protozoan Toxoplasma gondii. To determine the infection rate of toxoplasmosis in sheep and goats in east Azarbaijan province of Iran a total of 186 sera, 13 fetal brains, 13 cotyledons and 34 whole blood samples were collected from sheep and goats in Tabriz abattoir during the year 2010. Serum samples were tested for IgG antibodies against Toxoplasma gondii by Enzyme Linked Immunosorbent Assay (ELISA). This test is used in conjunction with direct and nested PCR techniques for detection of T. gondii DNA in blood,cotyledon and brain of fetuses, using 4 pairs of universal and specific primers, 18SrRNA, ITS-1, (Tg1,Tg2)and (Tg3,Tg4). Antibodies against T. gondii were found in 18.3% serum samples. DNA of T. gondii was detected in 69% of fetal brains, 23% of cotyledons and 14.7% of blood samples. The prevalence rate of T. gondii in sheep was 24.8% and in goats was 10.6% and the significant difference was observed between sheep and goats groups (p≤0.05). The prevalence of toxoplasmosis antibodies was significantly higher in adult sheep and goats than younger animals. There was significant difference between male (10.5%) andfemale (19.2%) sheep and goat was observed (p≤0.05). Detection of T. gondii DNA using PCR minimizes the problems which the researcher may face when using serological methods and facilitates diagnosis incomplex cases. The presence of T. gondii in blood, fetal tissues and neonatal specimens strongly suggests active and or congenital infection

Isolation of Anthrax Spores from Soil in Endemic Regions of Isfahan, Iran

Summary To isolate and detect anthrax spores from soil in different regions of Isfahan, Iran a total of 60 environmental specimens were collected during 2003. Bacterial endospores were extracted via flotation in distilled water and were cultured on blood agar and selective PLET media. Bacillus anthracis was identified using bacteriological and biological tests. Viable Bacillus anthracis spores were isolated from 9 (15%) soil samples of the 60 collected specimens in which 6 (66%) of isolates were encapsulated. The isolated bacteria and their virulence were confirmed with polymerase chain reaction (PCR) using specific primers. Its recommend that because of the existence of highly virulent strain of Bacillus anthracis in this region, a review on implementation of control programs such as regular vaccination of all susceptible livestock and surveillance of the disease in animals and human in such endemic areas is required

A Study on molecular characterization of Razi Bacillus anthracis Sterne 34F2 substrain in Iran

Anthrax, a zoonotic disease caused by Bacillus anthracis, has affected humans since ancient times. For genomic characterization of Razi B. anthracis Sterne 34F2 substrain, single nucleotide polymorphism (SNP) genotyping method developed by Van Erth, variable-number tandem-repeat (VNTR)-8 analysis proposed by Keim, and multiple-locus VNTR analysis (MLVA)-3 introduced by Levy were employed. In the SNPs typing system, where the nucleotide content of the genome at 13 evolutionary canonical loci was collectively analyzed, the originally South African 34F2 substrain was categorized in the A.Br.001/002 subgroup. In the VNTR-8 analysis, fragments with lengths of 314, 229, 162, 580, 532, 158, and 137 bp were identified at the following loci: vrrA, vrrB1, vrrB2, vrrC1, vrrC2, CG3, and pxO1, respectively. In addition, application of Levy's MLVA-3 genotyping method revealed that the genome of this strain carried 941, 451, and 864 bp fragments at AA03, AJ03, and AA07 loci, respectively. The present findings are undoubtedly helpful in meeting the requirements set by the World Organization for Animal Health (OIE) and World Health Organization (WHO) for anthrax vaccine manufacturers including Razi Institute. However, further similar studies are required to promote the current epidemiological knowledge of anthrax in Iran

A DNA vaccine candidate for B. anthracis immunization, pcDNA3.1+PA plasmid, induce Th1/Th2 mixed responses and protection in mice

The protective antigen (PA) of Bacillus anthracis (B. anthracis) is a potent immunogen and a candidate subunit vaccine. To address the question whether antibodies raised against PA following injection of pcDNA3.1+PA plasmid, encoding PA, can protect against virulent B. anthracis two different regimens of PA based vaccines (DNA and live spore) were used. The groups of BALB/c mice that received live spores of the Sterne strain, naked pcDNA3.1 and naked pcDNA3.1+PA were compared to control groups. All groups were injected three times with 30-day intervals. Two weeks after the last immunization, all mice were subjected to challenge with a pathogenic strain of B. anthracis (C2). Blood samples were taken before each injection and challenge. Evaluation of the sera by ELISA method showed that DNA immunization using pcDNA3.1+PA plasmid resulted in an antibody profile representative of a mixed Th1 and Th2 response, with a skewing to a Th1 response. The group which received the naked pcDNA3.1+PA had a survival rate of >80%. This challenge assay revealed that antibodies raised following DNA vaccination against PA can confer strong protection, and resistance against virulent species of B. anthracis

A DNA vaccine candidate for B. anthracis immunization, pcDNA3.1+PA plasmid, induce Th1/Th2 mixed responses and protection in mice

The protective antigen (PA) of Bacillus anthracis (B. anthracis) is a potent immunogen and a candidate subunit vaccine. To address the question whether antibodies raised against PA following injection of pcDNA3.1+PA plasmid, encoding PA, can protect against virulent B. anthracis two different regimens of PA based vaccines (DNA and live spore) were used. The groups of BALB/c mice that received live spores of the Sterne strain, naked pcDNA3.1 and naked pcDNA3.1+PA were compared to control groups. All groups were injected three times with 30-day intervals. Two weeks after the last immunization, all mice were subjected to challenge with a pathogenic strain of B. anthracis (C2). Blood samples were taken before each injection and challenge. Evaluation of the sera by ELISA method showed that DNA immunization using pcDNA3.1+PA plasmid resulted in an antibody profile representative of a mixed Th1 and Th2 response, with a skewing to a Th1 response. The group which received the naked pcDNA3.1+PA had a survival rate of >80%. This challenge assay revealed that antibodies raised following DNA vaccination against PA can confer strong protection, and resistance against virulent species of B. anthracis