Genotyping of Mycobacterium Tuberculosis Isolated from Suspected Patients in Tehran in 2015-2017

2019 ◽  
Author(s):  
Saeed Zaker ◽  
Hanie Bagherifard
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Nucleotide Polymorphisms ◽  
Single Nucleotide ◽  
Lack Of Information ◽  
Iranian Studies ◽  
Pcr Products ◽  
Life Threatening ◽  
Epidemiological Pattern ◽  
Group 3 ◽  
Group 2

Background and Aims: Unlike many global efforts to eradicate tuberculosis caused by Mycobacterium, it remains as a life-threatening infection with a worldwide incidence of 1.5 million cases each year. However, due to the lack of information about Mycobacterium tuberculosis characterization, more studies are required to evaluate strain diversity and epidemiology of tuberculosis to improve the therapeutic approaches. This study aimed to genotype the Mycobacterium tuberculosis isolated from suspected patients in Tehran, Iran through 2015-2017.Materials and Methods: In the current study, 30 isolates (sputum, broncho-alveolar lavage and biopsy) were collected from different tuberculosis patients at Massoud Clinical Lab of Tehran from 2015 to 2017. To find the single nucleotide polymorphisms and mutated regions, polymerase chain reaction (PCR) was performed on all the isolates to amplify the katG and gyrA genes. Then, PCR products were sequenced and analyzed.Results: The majority of isolates were assigned to PGG2 (90%), followed by PGG3 (10%) but no isolate belonging to PGG1 was found.Conclusions: Our findings demonstrate a remarkable epidemiological pattern of tuberculosis in Tehran. In group 2, isolates showed a considerably higher frequency compared to isolates in group 3, which is consistent with other findings reported in Iran. However, in contrast to other Iranian studies, no isolated strains were categorized in principal PGG1.

Mycobacterium tuberculosis isolates belonging to katG gyrA group 2 are associated with clustered cases of tuberculosis in Italian patients

2004 ◽  
Vol 53 (2) ◽  
pp. 155-159 ◽  
Author(s):  
L. Dolzani ◽  
M. Rosato ◽  
B. Sartori ◽  
E. Banfi ◽  
C. Lagatolla ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Local Population ◽  
Rflp Analysis ◽  
Nucleotide Polymorphisms ◽  
Transmitted Infection ◽  
Single Nucleotide ◽  
North East ◽  
The North ◽  
Group 2 ◽  
Group 1

Fifty-one consecutive isolates of Mycobacterium tuberculosis, collected during a 2-year period in the north-east of Italy, were subjected to IS6110-RFLP analysis to detect the presence of clusters and assigned to one of the three genotypic groups delineated by single nucleotide polymorphisms in the genes katG and gyrA. All the isolates collected from the local population belonged to group 2 or 3, while group 1 isolates were found only in specimens collected from African immigrants. Clustered cases of tuberculosis, which are likely to be related to recently transmitted infection, were found to be significantly associated with katG gyrA group 2. In the local situation, strains belonging to this group may therefore present a higher risk of transmission.

scholarly journals 173 The role of an intronic single nucleotide polymorphism within the dopamine receptor type-2 gene on pituitary prolactin protein expression in bulls

2020 ◽  
Vol 98 (Supplement_2) ◽  
pp. 37-37
Author(s):  
Andrea N DeCarlo ◽  
Keelee J McCarty ◽  
Sarah K Richey ◽  
Nathan Long ◽  
Scott Pratt
Keyword(s):  
Protein Expression ◽  
Genotype Distribution ◽  
Nucleotide Polymorphisms ◽  
Genotypic Effect ◽  
Single Nucleotide ◽  
Detection Range ◽  
Cull Cows ◽  
Densitometry Analysis ◽  
Pcr Products

Abstract Detrimental effects to male reproductive physiology have been observed due to changes in prolactin (PRL) serum concentration. Regulation of PRL by dopamine binding to the dopamine type-2 receptor (DRD2) is well defined and associations between male physiology and single nucleotide polymorphisms (SNPs) within the DRD2 gene have been observed. The objective of the study was to evaluate association of a DRD2 SNP to PRL protein expression in bulls. Testis and epididymis were collected from bulls grazing a forage containing or lacking a dopamine agonist at the end of a 126 d study (n = 14). Bovine pituitaries (n = 587) were collected randomly over 3 mo from a local abattoir which processes cull cows and bulls. Sex of pituitaries was verified (n = 259 males) by duplex PCR for amplification of SRY and b-actin followed by Southern blotting of PCR products for selection of male. Prolactin protein expression was assessed in testis, epididymis, and pituitary by western blotting. Expression of PRL protein was below detection range in reproductive tissues but was present in pituitary, therefore experiments continued in pituitary. Restriction fragment length polymorphism genotyping was performed on pituitaries by amplification of the DRD2 SNP region followed by digestion with a Tfil enzyme. Digested of products produced 3,2, or 1 band (AG, AA, GG, respectively). A subset of male pituitaries was blotted by slot blot manifold and PRL protein expression assessed by immunodetection and densitometry analysis normalized to GAPDH expression. Pituitary genotype distribution was 17.4% AA (n = 16), 63% AG (n = 58), and 19.6% GG (n = 18). Prolactin protein expression in the pituitary was similar across genotype (P = 0.23). These findings indicate that the DRD2 SNP has no genotypic effect on PRL protein expression in bovine pituitary.

scholarly journals Screening of Three Different Alleles of mtDNA (G709A, G3496T, A3537G) in Subpopulation of UKM Students

2018 ◽  
Vol 5 (1) ◽  
pp. 37-40
Author(s):  
Seri Mirianti Ishar ◽  
Jeyaganesan Pillay a/l Balaraman ◽  
Muhammad Jefri Mohd Yusof ◽  
Khairul Osman ◽  
Lee Loong Chuen
Keyword(s):  
Uv Light ◽  
Forensic Genetics ◽  
Nucleotide Polymorphisms ◽  
Coding Region ◽  
Documentation System ◽  
Single Nucleotide ◽  
National University ◽  
Pcr Products ◽  
Malay Population ◽  
Mutation Allele

Human DNA consists of nucleus DNA (nDNA) and mitochondrial DNA (mtDNA). Both are valuable in medicine and forensic genetics but in this project, single nucleotide polymorphisms (SNPs) in mtDNA are used to trace the mutation occurred. Mutations in the sequence of alleles can lead to haplogroup variation and also certain diseases. The purpose of this study is to screen of mutations on alleles G709A, G3496T, and A3537G in Malay population of The National University of Malaysia (UKM) students. These SNPs lie in the ND1 (nitrogen dehydrogenase subunit 1) coding region, and the reports state that these three alleles are prone to mutate. From MitoMap Web site, the mutations of these alleles are reported to have potential in causing several diseases with the collaboration of other SNPs mutation. Allele G709A is reported to have an association with hearing loss and Leber Hereditary Optic Neuropathy (LHON) while allele G3496T is associated to LHON only. Allele A3537G is related to diabetes. A total of 100 DNA samples were collected from Malay students of UKM and preserved on FTA card to be purified later. The concentration of the DNA on the purified FTA card was between 10μM to 20μM. An attempt was made by amplifying those three loci from the genomic DNA. The amplified product was detected and separated using 1% gel electrophoresis. Before sequencing, the PCR products were visualized under UV light using gel documentation system. All PCR products were sequenced to detect the mutation on every single position chosen. From the alignment of sequencing results, allele G709A and allele G3496T showed no mutation. Meanwhile four samples from alleles A3537G has the mutation. From the results obtained, it seems that mutations are rare in all selected alleles. It is recommended to increase the sample size and alleles selected in the future to increase the strength of the study. This study also should be applied to other populations in Malaysia such as Chinese and Indian.  

scholarly journals Decreased Serum Cholinesterase Activity- A Reliable Diagnostic Aid for Tuberculosis

2021 ◽  
Author(s):  
Dipak Kumar Chattopadhyay
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Drug Therapy ◽  
Lung Disease ◽  
Disease Control ◽  
Normal Control ◽  
Serum Cholinesterase ◽  
Diagnostic Potential ◽  
Group 3 ◽  
Ferric Hydroxamate ◽  
Group 2

Introduction: Siderophore, the low molecular weight ferric iron chelator, is secreted extracellularly by Mycobacterium tuberculosis, an obligate aerobe. The pathogenic mycobacteria need iron as co-factor for the extracellularly secreted enzyme Superoxide Dismutase (SOD) for its pathogenicity as well as it requires iron for its metabolic functions like reduction of oxygen for synthesis of Adenosine Triphosphate (ATP), etc. The termination of impulse conduction is endorsed by the rapid hydrolysis of Acetylcholine (ACh) by Acetylcholinesterase (AChE) in the central as well as peripheral nervous system (cholinergic pathway). The inhibitors to Cholinesterase (ChE) might lead to accumulation of ACh, hyper stimulation of nicotinic and muscarinic receptors and also disruption of neurotransmission. Possibility of an inhibitor substance for AChE in Tuberculosis (TB) patients interestingly correlates with the symptom of night sweating in those subjects. Aim: To assay the level of serum ChE in normal control, lung disease control and tubercular subjects;circumvent the serum ChE level as a diagnostic potential in TB at an early stage. Materials and Methods: The study was conducted on total 124 subjects, and were divided into three groups: Group 1: normal control (n=31), Group 2: lung disease control (n=31) and Group 3: patients suffering from TB {3A: pulmonary TB (n=31) and 3B: extrapulmonary TB (n=31)}. Serum ChE activity for all the subjects were measured according to the method of Hestrin S. Serum ChE level was assayed for group 3 subjects after additional one month’s anti-TB drug treatment and also for group 2 subjects after one month with usual treatment. The level of significance was assessed using Student’s t-test. Results: There was a significant inhibition of serum ChE activity in both pulmonary and extrapulmonary TB patients in comparison to that of in normal control as well as lung disease control subjects (p<0.01). With the anti-tubercular drug therapy for one month, there was significant recovery in the serum ChE activity in pulmonary as well as extrapulmonary tubercular subjects (p<0.01). Conclusion: It appears that the high level of hydroxamate type of siderophores (secreted by Mycobacterium tuberculosis for acquiring iron) might form ACh ferric hydroxamate complex binding more strongly with serum ChE resulting in inhibition of serum ChE activity in tubercular subjects. With anti-tubercular drug therapy, there was decrease in serum ACh ferric hydroxamate complex level resulting in recovery of serum ChE activity.

scholarly journals Genetic Variants in JAK1 Gene and Susceptibility to Hepatitis C Viral Infection in Iraq

2016 ◽  
Vol 10 (1) ◽  
pp. 37-41
Author(s):  
Fatima Abood Chaloob
Keyword(s):  
Hepatitis C ◽  
Hcv Infection ◽  
Nucleotide Polymorphisms ◽  
Single Nucleotide ◽  
Global Challenge ◽  
Pcr Products ◽  
Chronic Hcv Infection ◽  
Chronic Hcv ◽  
Apparently Healthy

Infection with hepatitis C virus (HCV) imposes a global challenge with over 180 million cases worldwide. Only few patients spontaneously had their virus neutralized, while most patients develop chronic HCV infection. This implies a key role of genetic factors in viral clearance or persistence. The current study aimed at clarifying the effect of certain single nucleotide polymorphisms (SNPs) on individual's susceptibility to HCV infection.  A total of 60 patients with confirmed HCV infection and 35 apparently healthy individuals were enrolled in this study. Blood sample was obtained from each participant, from which DNA was extracted. The JAK1gene was amplified with conventional PCR technique using three sets of primers targeting three SNPs in this gene: rs2780895, rs4244165 and rs17127024. Restriction fragment length polymorphism (RFLP) was used for genotyping of PCR products. Each of rs2780895 and rs17127024 had two genotypes in both patients and controls, however, only the heterozygous genotype of the SNP rs2780895 (CT) significantly associated with the susceptibility to HCV. The SNP rs4244165 appeared in only with homozygous wild genotype (GG) in both patients and controls. It can be concluded that allele T of the SNP rs2780895 could be considered as a risk factor for infection with HCV

scholarly journals Characterization of Mycobacterium tuberculosis Isolates from Patients in Houston, Texas, by Spoligotyping

2000 ◽  
Vol 38 (2) ◽  
pp. 669-676 ◽  
Author(s):  
Hanna Soini ◽  
Xi Pan ◽  
Amol Amin ◽  
Edward A. Graviss ◽  
Anees Siddiqui ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Population Based ◽  
Nucleotide Polymorphisms ◽  
Genetic Groups ◽  
Tuberculosis Epidemiology ◽  
New Information ◽  
Catalase Peroxidase ◽  
A Subunit ◽  
Group 2 ◽  
Group 1

Mycobacterium tuberculosis isolates (n= 1,429) from 1,283 patients collected as part of an ongoing population-based tuberculosis epidemiology study in Houston, Texas, were analyzed by spoligotyping and IS6110 profiling. The isolates were also assigned to one of three major genetic groups on the basis of nucleotide polymorphisms located at codons 463 and 95 in the genes (katG and gyrA) encoding catalase-peroxidase and the A subunit of DNA gyrase, respectively. A total of 225 spoligotypes were identified in the 1,429 isolates. There were 54 spoligotypes identified among 713 isolates (n= 623 patients) assigned to 73 IS6110 clusters. In addition, among 716 isolates (n = 660 patients) with unique IS6110 profiles, 200 spoligotypes were identified. No changes were observed either in the IS6110 profile or in the spoligotype for the 281 isolates collected sequentially from 133 patients. Five instances in which isolates with slightly different spoligotypes had the same IS6110 profile were identified, suggesting that in rare cases isolates with different spoligotypes can be clonally related. Spoligotypes correlated extremely well with major genetic group designations. Only three very similar spoligotypes were shared by isolates from genetic groups 2 and 3, and none was shared by group 1 and group 2 organisms or by group 1 and group 3 organisms. All organisms belonging to genetic groups 2 and 3 failed to hybridize with spacer probes 33 to 36. Taken together, the results support the existence of three distinct genetic groups of M. tuberculosis organisms and provide new information about the relationship between IS6110 profiles, spoligotypes, and major genetic groups of M. tuberculosis.

Characterisation of the melanocortin-1 receptor gene in alpaca and identification of possible markers associated with phenotypic variations in colour

2009 ◽  
Vol 49 (8) ◽  
pp. 675 ◽  
Author(s):  
N. L. Feeley ◽  
K. A. Munyard
Keyword(s):  
Mus Musculus ◽  
Bos Taurus ◽  
Receptor Gene ◽  
Nucleotide Polymorphisms ◽  
Wild Type ◽  
Single Nucleotide ◽  
Melanocortin 1 Receptor ◽  
Mc1r Gene ◽  
Pcr Products ◽  
Phenotypic Variations

The aim of this study was to determine if any correlation exists between melanocortin-1 receptor (MC1R) polymorphisms and skin and fibre colour in alpacas. Primers capable of amplifying the entire alpaca MC1R gene were designed from a comparative alignment of Bos taurus and Mus musculus MC1R gene sequences. The complete MC1R gene of 41 alpacas exhibiting a range of fibre colours, and which were sourced from farms across Australia, was sequenced from PCR products. Twenty-one single nucleotide polymorphisms were identified within MC1R. Two of these polymorphisms (A82G and C901T) have the potential to reduce eumelanin production by disrupting the activity of MC1R. No agreement was observed between fibre colour alone and MC1R genotype in the 41 animals in this study. However, when the animals were assigned to groups based on the presence or absence of eumelanin in their fibre and skin, only animals that had at least one allele with the A82/C901 combination expressed eumelanin. We propose that A82/C901 is the wild-type dominant ‘E’ MC1R allele, while alpacas with either G82/T901 or G82/Y901 are homozygous for the recessive ‘e’ MC1R allele and are therefore unable to produce eumelanin.

scholarly journals Using Mycobacterium tuberculosis Single-Nucleotide Polymorphisms To Predict Fluoroquinolone Treatment Response

2019 ◽  
Vol 63 (7) ◽  
Author(s):  
Marva Seifert ◽  
Edmund Capparelli ◽  
Donald G. Catanzaro ◽  
Timothy C. Rodwell
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Single Nucleotide Polymorphisms ◽  
Specific Resistance ◽  
Therapeutic Targets ◽  
World Health ◽  
Population Pharmacokinetic ◽  
Nucleotide Polymorphisms ◽  
Single Nucleotide ◽  
Content Type ◽  
Level Increase

ABSTRACT Clinical phenotypic fluoroquinolone susceptibility testing of Mycobacterium tuberculosis is currently based on M. tuberculosis growth at a single critical concentration, which provides limited information for a nuanced clinical response. We propose using specific resistance-conferring M. tuberculosis mutations in gyrA together with population pharmacokinetic and pharmacodynamic modeling as a novel tool to better inform fluoroquinolone treatment decisions. We sequenced the gyrA resistance-determining region of 138 clinical M. tuberculosis isolates collected from India, Moldova, Philippines, and South Africa and then determined each strain’s MIC against ofloxacin, moxifloxacin, levofloxacin, and gatifloxacin. Strains with specific gyrA single-nucleotide polymorphisms (SNPs) were grouped into high or low drug-specific resistance categories based on their empirically measured MICs. Published population pharmacokinetic models were then used to explore the pharmacokinetics and pharmacodynamics of each fluoroquinolone relative to the empirical MIC distribution for each resistance category to make predictions about the likelihood of patients achieving defined therapeutic targets. In patients infected with M. tuberculosis isolates containing SNPs associated with a fluoroquinolone-specific low-level increase in MIC, models suggest increased fluoroquinolone dosing improved the probability of achieving therapeutic targets for gatifloxacin and moxifloxacin but not for levofloxacin and ofloxacin. In contrast, among patients with isolates harboring SNPs associated with a high-level increase in MIC, increased dosing of levofloxacin, moxifloxacin, gatifloxacin, or ofloxacin did not meaningfully improve the probability of therapeutic target attainment. We demonstrated that quantifiable fluoroquinolone drug resistance phenotypes could be predicted from rapidly detectable gyrA SNPs and used to support dosing decisions based on the likelihood of patients reaching therapeutic targets. Our findings provide further supporting evidence for the moxifloxacin clinical breakpoint recently established by the World Health Organization.

scholarly journals Genotyping Single-Nucleotide Polymorphisms by the Invader Assay with Dual-Color Fluorescence Polarization Detection

2001 ◽  
Vol 47 (8) ◽  
pp. 1373-1377 ◽  
Author(s):  
Tony M Hsu ◽  
Scott M Law ◽  
Shenghui Duan ◽  
Bruce P Neri ◽  
Pui-Yan Kwok
Keyword(s):  
Fluorescence Polarization ◽  
Cost Effective ◽  
Snp Markers ◽  
Nucleotide Polymorphisms ◽  
Single Nucleotide ◽  
Clear Cut ◽  
Pcr Product ◽  
Invader Assay ◽  
Pcr Products ◽  
Incorporation Assay

Abstract Background: The PCR-Invader® assay is a robust, homogeneous assay that has been shown to be highly sensitive and specific in genotyping single-nucleotide polymorphism (SNP) markers. In this study, we introduce two changes to improve the assay: (a) we streamline the PCR-Invader method by assaying both alleles for each SNP in one reaction; and (b) we reduce the cost of the method by adopting fluorescence polarization (FP) as the detection method. Methods: PCR product was incubated with Invader oligonucleotide and two primary probes at 93 °C for 5 min. Signal probes corresponding to the cleaved flaps of the primary probes [labeled with fluorescein and 6-carboxytetramethylrhodamine (TAMRA) dye] and Cleavase® VIII enzyme (a flap endonuclease) were then added to the mixture. This reaction mixture was incubated at 63 °C for 5 min. FP measurements were made with a fluorescence plate reader. Results: Eighty-eight individuals were genotyped across a panel of 10 SNPs, using PCR product as template, for a total of 880 genotypes. An average “no call” rate of 3.2% was observed after first round of experiments. PCR products were remade in those samples that failed to produce any genotype in the first round, and all gave clear-cut genotypes. When the genotypes determined by the PCR-Invader assay and template-directed dye-terminator incorporation assay with FP were compared, they were in 100% concordance for all SNP markers and experiments. Conclusions: The improvements introduced in this study make PCR-Invader assay simpler and more cost-effective, and therefore more suitable for high-throughput genotyping.

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