Molecular diversity of hpd gene in clinical isolates of Haemophilus influenzae

Infections due to Haemophilus influenzae result in tremendous global morbidity. The conjugated vaccines against H. influenzae type b (Hib) have dramatically reduced the incidence of invasive Hib disease in the routine immunization of infants. The several proteins used as vaccine candidates for this pathogen, but they don't produce efficient immune in animal models against all strains of H. influenzae. This study aimed to determine the diversity of hpd gene nucleotide sequences of Iranian native clinical isolates of H. influenzae as a native vaccine candidate compared to standard strains. Twenty isolates of H. influenzae recovered from different clinical specimens of patients admitted to Milad and Imam Khomeini hospitals, Tehran, Iran. Then, isolates detected and identified as H. influenzae using biochemical tests, and further confirmation through omp6 gene PCR. The hpd gene was amplified by PCR using gene-specific primers, and the amplicons digested with EcoR1. For four isolates, the Amplicon of hpd gene sequenced, and the sequences aligned with sequences harbored in GenBank. Subsequently, sequences were submitted to the EMBL site (http://www.ebi.ac.uk/embl/). EcoR1 restriction enzyme pattern was the same among the 19 clinical isolates, and only one isolate was different. That different one with 3 out of 19 isolates were sequenced. The results showed that the nucleotide sequences and the deduced amino acid sequences for protein D in clinical isolates were highly conserved with similarities >95. In conclusion, regarding high similarity up to 99 in clinical isolates, protein D can be a novel vaccine candidate against all types of H. influenza from Iran. This finding should be proved with more isolates, and also, evaluate the immunological features of protein D in animal models. © 201

Biological and immunological characteristics of Brucella abortus S99 major outer membrane proteins

Introduction and objective: Outer membrane proteins (OMPs) of Brucella are considered as immunogenic structures which can be used to design and develop a subunit vaccine for human brucellosis. Brucella abortus S99 OMPs promote the synthesis of high levels of specific anti-Brucella IgG molecules in rabbits when administrated with lipopolysaccharide (LPS). The objective of this study is evaluation of the efficacy of B. abortus major OMPs with LPS in the induction of immune response against brucellosis. Materials and methods: OMPs were derived from B. abortus by sequential extraction of sonicated cells with ultracentrifugation and predigestion with lysozyme. Proteins could be separated by anion-exchange chromatography and gel-filtration. Based on SDS-PAGE profiles, porins have been dominantly purified among three different classes of B. abortus OMPs. Sera of immunized rabbits against B. abortus porins were analyzed by enzymelinked immunosorbent assay (ELISA). LPS of B. abortus and complete Freud's adjuvant (CFA) were also applied to elicit higher levels of anti-Brucella antibodies. Results: ELISA confirmed the potency of porins and porins combination with CFA and LPS to promote humoral specific response. Among the above-mentioned compounds, a combination of porins + LPS or porins + CFA has been the most potent immunogenic compound to induce higher titer of antibody against B. abortus S99 in the animal model. Conclusion: The application of a complex of Brucella LPS and porins as an effective method to elicit protective and long-lasting immunity against Brucella infection and would be studied to design and develop a subunit vaccine for human brucellosis

Association of microbiota-derived propionic acid and Alzheimer�s disease; bioinformatics analysis

Purpose: Microbiota-derived metabolites could alter the brain tissue toward the neurodegeneration disease. This study aims to select the genes associated with Propionic acid (PPA) and compromise Alzheimer�s disease (AD) to find the possible roles of PPA in AD pathogenesis. Methods: Microbiota-derived metabolites could alter the brain tissue toward the neurodegeneration disease. This study aims to select the genes associated with Propionic acid (PPA) and compromise Alzheimer�s disease (AD) to find the possible roles of PPA in AD pathogenesis. Results: Amongst all genes associated with PPA and AD, 284 genes to be shared by searching databases and were subjected to further analysis. AD-PPA genes mainly involved in cancer, bacterial and virus infection, and neurological and non-neurological diseases. Gene Ontology and pathway analysis covered the most AD hallmark, such as amyloid formation, apoptosis, proliferation, inflammation, and immune system. Network analysis revealed hub and bottleneck genes. MCODE analysis also indicated the seed genes represented in the significant subnetworks. ICAM1 and CCND1 were the hub, bottleneck, and seed genes. Conclusions: PPA interacted genes implicated in AD act through pathways initiate neuronal cell death. In sum up, AD-PPA shared genes exhibited evidence that supports the idea PPA secreted from bacteria could alter brain physiology toward the emerging AD signs. This idea needs to confirm by more future investigation in animal models. © 2020, Springer Nature Switzerland AG

Preparation and Evaluation of a New Lipopolysaccharide-based Conjugate as a Vaccine Candidate for Brucellosis

Objectives: Development of an efficacious vaccine against brucellosis has been a challenge for scientists for many years. At present, there is no licensed vaccine against human brucellosis. To overcome this problem, currently, antigenic determinants of Brucella cell wall such as Lipopolysaccharide (LPS) are considered as potential candidates to develop subunit vaccines. Methods: In this study, Brucella abortus LPS was used for conjugation to Neisseria meningitidis serogroup B outer membrane vesicle (OMV) as carrier protein using carbodiimide and adipic acid-mediated coupling and linking, respectively. Groups of eight BALB/c mice were injected subcutaneously with 10μg LPS alone, combined LPS+OMV and conjugated LPS-OMV on 0 days, 14 days, 28 days and 42 days. Anti-LPS IgG was measured in serum. Results: The yield of LPS to OMV in LPS-OMV conjugate was 46.55, on the basis of carbohydrate content. The ratio for LPS to OMV was 4.07. The LPS-OMV conjugate was the most immunogenic compound that stimulated following the first injection with increased IgG titer of ~5-fold and ~1.3-fold higher than that produced against LPS and LPS in noncovalent complex to OMV (LPS+OMV), respectively. The highest anti-LPS IgG titer was detected 2 weeks after the third injection (Day 42) of LPS-OMV conjugate. The conjugated compound elicited higher titers of IgG than LPS+OMV, that showed a 100-120-fold rise of anti-LPS IgG in mice. Conclusion: These results indicate that our conjugated LPS-OMV can be used as a brucellosis vaccine, but further investigation is required. © 2014

Variability in gene cassette patterns of class 1 and 2 integrons associated with multi drug resistance patterns in Staphylococcus aureus clinical isolates in Tehran-Iran

Background: To investigate antibiotic resistance, the occurrence and distribution of class 1 and 2 integrons in multidrug- resistant Staphylococcus aureus isolates from hospitals in Tehran, Iran. The isolates were examined for susceptibility to antimicrobial agents. The mecA gene, class 1 and 2 integrons were detected by PCR. Integrase positive strains were further analysed for the presence of resistance gene cassettes using specific primers and were sequenced. Results: Among 139S.aureus isolates, 109 (78.4 ) and 112 (80.5 ) strains were considered as multidrug resistant and mecA positive, respectively. Class 1 integrons and internal variable regions were found in 72.6 (101/139) and 97 (98/101) and class 2 integrons and variable regions also in 35.2 (49/139) and 65.3 (32/49) of S.aureus clinical isolates, respectively. Twelve distinct cassette arrays were found, containing genes encoding resistance to β-lactams, aminoglycosides, streptothricin, trimethoprim, chloramphenicol,a putative glucose dehydrogenase precursor and a protein with unknown function. Gene cassette arrays aadB, aadA2 and dhfrA1-sat2-aadA1 were common in S.aureus isolates. We detected a completely new gene cassettes which contained aadB, oxa2, aacA4, orfD-aacA4-catB8, aadB-catB3, orfD-aacA4 and aadB-aadA1-cmlA6 of class 1 and dhfrA1-sat2-aadA1, dhfrA11, dhfrA1-sat2 of class 2 integrons. Conclusions: This is the first study to report carriage of class 1 and 2 integrons and associated gene cassettes among in S.aureus isolates from Iran. © 2015 Mostafa et al

Gut Microbiota and Serum Biomarker Analyses in Obese Patients Diagnosed with Diabetes and Hypothyroid Disorder

Background: Variations of serum biomarkers and bacterial diversity of the gastrointestinal tract in obese patients with diabetes or hypothyroid are poorly understood. The aim of this study was to provide recent findings in this regard. Methods: A total of 119 obese patients 17 with diabetes, 23 with hypothyroid, and 79 patients without either diabetes or hypothyroid (control) were recruited in this study. Serum biomarkers such as biochemical, hormonal (insulin and glucagon), and cytokine levels interleukin (IL)-6, IL-1β, tumor necrosis factor-alpha, IL-10, and transforming growth factor beta-1 (TGF-β1) were measured under fasting conditions. Bacterial abundance of gut microbiota was also quantitated by real-time polymerase chain reaction using 16S rRNA gene-based specific primers. Results: Average value of blood sugar (P: 0.0184), hemoglobin A1c, insulin, homeostasis model assessment insulin resistance, TGF-β 1, IL-6, IL-1β, interferon gamma (Pfor each < 0.001), and phylum Actinobacteria odds ratio (OR): 1.5, P: 0.032 was significantly higher in diabetic versus control group. In contrast, the levels of IL-10 (P < 0.001), Firmicutes (OR: 0.6, P: 0.058), and Akkermansia muciniphila (OR: 0.4, P: 0.053) were significantly lower in diabetic versus control group. However, there was no statistically significant difference between the values in hypothyroid versus control group either in crude or adjusted models. Conclusion: While there are some relationships between serum biomarkers or bacterial abundance with diabetes prediction in obese patients, this prognostication is less likely in obese patients with hypothyroid. Further investigation is warranted in the application of identified preclinical biomarkers in the diagnosis of diabetes or hypothyroid in obese patients. © Copyright 2021, Mary Ann Liebert, Inc., publishers

Genetic diversity of Mycobacterium tuberculosis isolates causing pulmonary and extrapulmonary tuberculosis in the capital of Iran

Objectives: Evaluation of the genetic diversity of Mycobacterium tuberculosis (M.tb) and determining if the association between a specific genotype and the site of infection is crucial. Accordingly, the current study aimed at comparing predominant M.tb genotypes in pulmonary (PTB) and extrapulmonary tuberculosis (EPTB) isolates circulating in the capital of Iran. Methods: The genetic diversity of culture-confirmed PTB and EPTB isolates were evaluated by Spoligotyping and MIRU-VNTR (mycobacterial interspersed repetitive-unit�variable-number tandem-repeat) typing methods. Genotyping data were analyzed with SITVIT, MIRU-VNTRplus, and TBminer databases. To assess adjusted associations, chi-square/the Fisher exact test and multiple logistic regression model were applied. Results: URAL2 (NEW-1) (28/88; 31.8) and CAS1-DELHI (25/84; 29.8) genotypes were predominant in EPTB and PTB strains, respectively. Based on MIRU-VNTR typing, 158 different MIRU-VNTR patterns were identified. Clustering rate and minimum estimate of the proportion of TB caused by recent transmission was 4.1 and 8.1, respectively. Conclusions: The current study provided new insight into circulating genotypes of M.tb in PTB and EPTB patients in Tehran, Iran. This low percentage of TB transmission rate, demonstrated that mode of TB transmission was mainly associated with reactivation of latent TB rather than recently transmitted infection in this region. There was no significant difference in the association between the genotypes of M.tb strains and the site of the disease. © 2018 Elsevier Inc

Extraction and molecular determination of major outer membrane proteins of Brucella abortus S99

Background: Brucellosis is one of the five common bacterial zoonoses caused by a gram negative, non-spore forming, and facultative intracellular bacterial organism belonging to the genus Brucella. Although brucellosis is considered as a health problem for both men and domestic animals in many countries, any licensed human vaccine has not been designed and produced for it yet. To overcome the problem, currently, antigenic determinants of Brucella cell wall e.g. outer membrane proteins OMPs and lipopolysaccharide LPS are considered as potential candidates to develop subunit vaccines. Materials and Methods: Brucella abortus S99 used in the present study is obtained from the standard bacterial collection of Institute Pasteur of Iran. OMPs were extracted by deoxycholate extraction technique and further purification performed by sequential centrifugation and ultracentrifugation. Protein concentration was determined using the Nanodrop NDâ��10000 spectrophotometery. SDS-polyacrylamied gel electrophoresis ((SDS–PAGE) was performed to determine the electerophoretic pattern and the molecular weight of the extracted OMP samples. Results: OMPs concentration of B.abortus S99 has been measured and reported as 6.27 mg/ml. SDS-PAGE analysis indicated one protein band in the range of 36-38 kDa which would be classified as the porins of B.abortus S99. Conclusion: Extraction of B.abortus S99 OMPs with the applied method in the present study produced a satisfactory yield of OMPs. These proteins belonging to the second group of OMPs, called porins

Comparison of Three Different Methods for Detection of IL28 rs12979860 Polymorphisms as a Predictor of Treatment Outcome in Patients with Hepatitis C Virus

Objectives: This study aimed to evaluate the specificity, sensitivity, cost, and turn-around time of three methods of gene polymorphism analysis and to study the relationship between IL28B rs12979860 and SVR rate to pegIFN-α/RVB therapy among patients with chronic hepatitis C. Methods: A total of 100 samples from chronic hepatitis C patients were analyzed in parallel using the three methods: direct sequencing, real-time polymerase chain reaction (PCR), amplification refractory mutation system (ARMS)-PCR. Results: The different profiles for IL28B rs12979860 alleles (CC, CT, and TT) obtained with PCR-RFLP, ARMS-PCR, and direct sequencing were consistent among the three methods. Prevalence of rs12979860 genotypes CC, CT and TT in HCV genotype 1a was 10(19.6), 35(68.6), and six (11.8), respectively, and in HCV genotype 31, it was 13(26.5), 31(63.3), and five (10.2), respectively. No significant difference was seen between rs12979860 genotype and HCV genotype (p = 0.710). Conclusion: Screening by ARMS - PCR SNOP detection represents the most efficient and reliable method to determine HCV polymorphisms in routine clinical practice. © 2016