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Resumen Se han encontrado asociaciones entre polimorfismos genéticos y parámetros reproductivos, este abordaje adicionalmente permitiría proponer explicaciones a diferentes fenómenos estudiados. FSHr, InhA, AMH y AMHr son genes asociados al desarrollo folicular, que tienen una alta homología y la misma función en bovinos. El objetivo del presente trabajo fue evaluar en bovinos muy relacionados si se encuentran algunos polimorfismos asociados a parámetros reproductivos reportados en otras especies. Durante el 2018, en el Magdalena Medio Colombiano fueron tomadas muestras de sangre anticoagulada de 50 búfalas mestizas (Bubalus bubalis) y 50 vacas cebú comercial (Bos indicus). Se registró la paridad, días abiertos, intervalo parto primer servicio y edad al primer parto de cada animal, sin alteraciones anatómicas, con edad y peso similar, mantenidos en las mismas condiciones de alimentación y manejo. Se extrajo el ADN por el método de Salting out y se evaluaron los polimorfismos de acuerdo con lo reportado en la literatura. Se compararon los datos usando la prueba Mann Whitney, se consideró significativo un valor de p<0,05. Se encontró que los búfalos tienen menores niveles de AMH, edad al primer parto, intervalo ente partos que las vacas (p<0,001), mayor paridad (p=0,005). No se confirmaron los po- limorfismos reportados en Holstein y en humanos en ninguna de las muestras y especies analizadas. No confirmar los hallazgos en bovinos de otras razas y espe- cies, aunque las especies tengan diferentes parámetros reproductivos, muestra la necesidad de replantear el abordaje del estudio de la asociación de los fenómenos reproductivos y genéticos.
Abstract Associations have been found between genetic polymorphisms and reproductive parameters, this approach additionally would allow us to propose explanations to the phenomena studied. FSHr, InhA, AMH and AMHr are genes associated with follicular development, which have a high homology and the same function in bovines. The objective of this work was to evaluate if some polymorphisms described in other species associated with reproductive parameters are found in two closely related bovines. During 2018, in the Colombian Magdalena Medio anticoagulated blood samples from 50 buffaloes (Bubalus bubalis) and 50 commercial zebu cows (Bos indicus) were taken. All animals without anatomical abnormalities and similar weight and age were taken and the. parity, intercalving period and age at first calving were also recorded. DNA were extracted using Salting out method and the reported polymorphisms of the above mentioned genes were evaluated. Data were compared using Mann Whitney test and p<0.05 value were considered significant. Buffaloes have lower AMH levels, age at first calving, calving interval than Bos indicus cows (p<0.001) and higher parity (p=0.005). The polymorphisms reported in Holstein and humans were not confirmed in any of the samples and species analyzed. The results, are in some way paradoxical because there are differences in reproductive parameters but nothing different in the studies genes. It shows the need to rethink the approach of the study of the association of reproductive and genetic phenomena
Resumo Foram encontradas associações entre polimorfismos genéticos e parâmetros repro- dutivos; essa abordagem também permitiria propor explicações para os diferentes fenômenos estudados. FSHr, InhA, AMH e AMHr são genes associados ao desenvol- vimento folicular, que apresentam alta homologia e a mesma função em bovinos. O objetivo do presente trabalho foi avaliar em bovinos intimamente relacionados se alguns polimorfismos associados a parâmetros reprodutivos relatados em outras espécies. Em 2018, 50 vacas de búfalo (Bubalus bubalis) e 50 de zebu comerciais (Bos indicus) foram amostradas na Magdalena Medio colombiana. Foram registra- das paridade, dias abertos, intervalo de nascimento do primeiro serviço e idade no primeiro nascimento de cada animal, sem alterações anatômicas, com idade e peso semelhantes, mantidas nas mesmas condições de alimentação e manuseio. O DNA foi extraído pelo método Salting out e os polimorfismos foram avaliados de acordo com o relatado na literatura. Os dados foram comparados pelo teste de Mann Whitney, sendo considerado significativo um valor de p <0,05. Verificou-se que os búfalos apresentam níveis mais baixos de AMH, idade do primeiro nascimento, intervalo entre os nascimentos que as vacas (p <0,001), maior paridade (p = 0,005). Os polimorfis- mos relatados em Holstein e em humanos não foram confirmados em nenhuma das amostras e espécies analisadas. A não confirmação dos achados em bovinos de outras raças e espécies, embora as espécies possuam diferentes parâmetros repro- dutivos, mostra a necessidade de repensar a abordagem do estudo da associação dos fenômenos reprodutivos e genéticos.
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Objective: To investigate the aerodynamic characteristics of the nebulized recombinant human inter- feron α1b(rhIFN α1b)injection and its delivery in different respiratory modes both in vitro. Methods: The particle size distribution and aerodynamic properties of the nebulized rhIFN α1b injection for inhalation were evaluated with Spraytec STP5313 and the next generation pharmaceutical impactor(NGI). The total delivered dose and delivery rate were deter- mined using a breathing simulator. Results:After atomization, the D50 of rhIFN α1b droplets was 2.74 μm, the fine par- ticle fraction(FPF)was 77.49%, the mass median aerodynamic diameter(MMAD)was 3.26 μm, and the geometric standard deviation(GSD)was 1.93. In neonatal, infant, and child breathing modes, the delivered total amount of rhIFN α1b by spraying for 220 seconds was 2.10, 2.44, and 3.51 μg, respectively. Conclusion: After atomization, the particle size of rhIFN α1b injection was small enough to be transmitted to the lung, and the total delivered dose and delivery rate showed a tendency of increase in turn in the neonatal, infant, and child breathing modes, indicating that the effective dose of the drug and the age of patients should be considered when formulating the clinical treatment plan.
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Background@#According to the First National Tuberculosis (TB) Prevalence Survey in Mongolia the prevalence of bacteriologically-confirmed pulmonary TB among adults was 559.6 (95% CI: 454.5–664.7) per 100000 population in 2014–2015. This was three times as high as previously estimated. Nationwide anti-tuberculosis (TB) drug resistance survey was conducted in 1999 and 2007 in Mongolia. Share of multidrug resistant TB (MDR-TB) cases among newly notified TB cases increased from 1.0% in 1999 to 1.4% in 2007. Accordingly, we aimed to perform drug susceptibility test on strains isolated from TB Prevalence Survey and to determine the prevalence of drug resistant TB.@*Material and Methods@#All 242 MTB strains isolated from the survey TB cases were tested GenoTypeMTBDRplus test and conventional 1st line DST on solid medium. @*Result@#Conventional DST and GenoTypeMTBDRplus tests done for 93.8% (227/242) of them and 6.2% (15/242) were tested by GenoTypeMTBDRplus only. A 61.6% (95%CI 55.3-67.4) of all cases were susceptible to first line anti-TB drugs, any drug resistance and MDR-TBdetected as 38.4% (95% CI 32.5-44.7)and 9.5% (95% CI 6.4-13.9), respectively. Prevalence of MDR-TB was7.8% (95% CI 4.9-12.4) among new and 17.9% (95% CI 9.0-32.7) among previously treated cases. The 64 strains were identified as a resistant to isoniazid, 32.8% (42/64) and 65.6% (21/64) were katG, and inhAmutation, respectively. One isolate (1.6%) was mutations in both the inhAand katGgenes.The predominant mutations detected in therpoB were S531L (91.3%) among rifampicin resistant isolates and the mutation in inhAwas C–15T (100%) and katG mutation was S315T1 (100%) among isoniazid-resistant isolates. @*Conclusion@#Prevalence of cases with DR-TB is high among prevalent TB cases, especially prevalence of MDR-TB among new cases. In comparison to previous studies, DR-TB cases seem to be increased. Rifampicin resistant strains have a mutation of the rpoBand resistance to isoniazid is predominantly associated with the inhA mutation.
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BACKGROUND: The purpose of this study was to analyze the relationship between the gene mutation patterns by the GenoType MTBDRplus (MTBDRplus) assay and the phenotypic drug susceptibility test (pDST) results of isoniazid (INH) and prothionamide (Pto). METHODS: A total of 206 patients whose MTBDRplus assay results revealed katG or inhA mutations were enrolled in the study. The pDST results were compared to mutation patterns on the MTBDRplus assay. RESULTS: The katG and inhA mutations were identified in 68.0% and 35.0% of patients, respectively. Among the 134 isolated katG mutations, three (2.2%), 127 (94.8%) and 11 (8.2%) were phenotypically resistant to low-level INH, high-level INH, and Pto, respectively. Among the 66 isolated inhA mutations, 34 (51.5%), 18 (27.3%) and 21 (31.8%) were phenotypically resistant to low-level INH, high-level INH, and Pto, respectively. Of the 34 phenotypic Pto resistant isolates, 21 (61.8%), 11 (32.4%), and two (5.9%) had inhA, katG, and both gene mutations. CONCLUSION: It is noted that Pto may still be selected as one of the appropriate multidrug-resistant tuberculosis regimen, although inhA mutation is detected by the MTBDRplus assay until pDST confirms a Pto resistance. The reporting of detailed mutation patterns of the MTBDRplus assay may be important for clinical practice, rather than simply presenting resistance or susceptibility test results.
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Humanos , Bioensayo , Susceptibilidad a Enfermedades , Genotipo , Isoniazida , Mycobacterium , Mycobacterium tuberculosis , Protionamida , Proyectos de Investigación , Tuberculosis Resistente a Múltiples MedicamentosRESUMEN
ABSTRACT Tuberculosis is leading cause of death among the global bacterial infections. The main causative for tuberculosis is Mycobacterium tuberculosis, which will survive in its host human being for decades in latent or chronic levels. In addition, the late multidrug resistance at a disturbing rate accompanies the appearance of tuberculosis. The quick spread of resistance to initial stage treatment medications has redirected the focus of the medical community in the creation of an array of new drug against Mycobacterium tuberculosis. The InhA protein is a component of Fatty acid synthetase (FAS) II and exhibits an NADH reliant enoyl-ACP reductase activity. InhA is a vital enzyme of M.tuberculosis in control of cell wall synthesis, which can turn out to be a great focus for the synthesis of anti-tubercular treatment. Inspired from the offering biological actions of phytoconstituents from Allium sativum, the current research concentrates on looking at novel lead compounds from the plant. Molecular docking studies were carried out employing specific phytoconstituents from A.sativum with the protein InhA target. Ajoene shows much more encouragingresults with a Mol Dock rating of 80.6047Kcal/mol, as opposed to the typical initial line drug isoniazid (Moldock score: -58.7028 Kcal/mol). Molecular docking prediction indicate that Ajoene could be formulated into a possible treatment drug for Mycobacterium tuberculosis.
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Objective To analyze the characteristics of the rpoB, KatG and inhA genes mutations in rifampin and isoniazid resistant Mycobacterium tuberculosis (MTB) clinical isolates in Baise district, Guangxi autonomous region. Methods 128 MTB clinical strains were collected and isolated for drug susceptibility testing, and drug resistant strain DNA was subtracted for rpoB, KatG and inhA genes mutation analysis. Results 75%(27/36)isolates carried mutations in the rpoB gene,and 59.3%(16/27)isolates carried mutations in 531 sites. 44.1%(15/34) isolates carried mutations in KatG or inhA, and 66.7%(10/15) isolates appeared in KatG 315 site, with two new mutations found in KatG 279 and 427 site. In these mutation isolates, 13.3%(2/15) mutations appeared in inhA 5, 6.7%(1/15) in inhA 16, and 20%(3/15) in both katG and inhA. Conclusions The mutation of rpoB, katG and inhA genes in TB is highly correlated with its resistance to rifampin and isoniazid in Baise district, Guangxi autonomous region. The study will provide a basis for further understanding the anti-bacterium mechanism and quick diagnostic methods for drug-resistant tuberculosis.
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Objective:To observe the efficacy of Chuankezhi injection combined with budesonide and formoterol fumarate powder for inhalation in the treatment of chronic obstructive pulmonary disease ( COPD) . Methods:Totally 120 patients with COPD were ran-domly divided into two groups:the observation group inhaled budesonide and formoterol fumarate powder for inhalation combined with intramuscular injection of Chuankezhi injection, and the control group inhaled budesonide and formoterol fumarate powder for inhala-tion. The treatment course was 28 days. The pulmonary function indices [ FEV1% expected value, FEV1 /FVC, 6 min walking dis-tance (6MWD)], the times of acute exacerbation and the adverse drug reactions during the treatment were observed and compared be-tween the two groups. Results:After the treatment, the lung function and 6MWD were improved in the two groups, and the improve-ment of FEV1% expected value in the treatment group was more obvious than that in the control group [(59. 7+12. 1)% vs (49. 8+11. 3)%, P<0. 05]. The number of patients with at least once of acute exacerbation was significantly reduced in the treatment group when compared with that in the control group (35% vs 62%, P<0. 01). The incidence of adverse drug reactions in the two groups showed no significant difference. Conclusion:Chuankezhi injection combined with budesonide and formoterol fumarate powder for in-halation can notably improve the lung function in the patients with COPD and reduce the times of acute exacerbation without effect on the incidence of adverse drug reactions.
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Background and Objectives: Multidrug-resistant (MDR) Mycobacterium tuberculosis strains are serious threats to the control of tuberculosis and comprise an increasing public health problem. Rapid detection of such strains is quite critical in timely management of such issues. The study was performed with an objective to compare Genotype MTBDRplus reverse hybridization probe assay (Hain Lifescince, GmBH, Nehern, Germany) with culture based proportion method for rapidly identifying MDR-TB strains from suspected multi drug resistant cases, referred to GENETUP Kathmandu, Nepal. Methodology: A commercially available new Genotype MTBDRplus assay was evaluated for its ability to detect mutations in Mycobacterial isolates conferring resistance to rifampicin (RMP) and isoniazid (INH). A total of 64 MDR isolates (i.e., at least resistant to RMP and INH), 5 fully susceptible strains and 1 RMP sensitive strains by conventional proportion method were analyzed using Genotype MTBDRplus assay. MTBDRplus assay is designed to detect the mutations in the hot spot region of rpoB gene, katG and regulatory region of inhA gene. Results: The MTBDRplus assay detected 59 of 61 RMP resistant strains (96.72%) with mutations on 81-bp hot spot region of rpoB gene and 60 of 63 INH resistant strains (95.23%) with mutation in codon 315 katG and regulatory region of inhA. The sensitivity and specificity for the detection of RMP resistance were 96.72% and 100% respectively. While, value of the same two variables for INH resistance were 95.23% and 100%, respectively. Conclusions: The new Genotype MTBDRplus assay represents a rapid, reliable, upgraded tool with high sensitivity and specificity for the detection of INH and RMP resistance strains that can readily be included in a routine laboratory work for the early diagnosis and control of MDR-TB.
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BACKGROUND: In Korea, tuberculosis is resistant to isoniazid (INH) and/or rifampicin (RIF) in more than 10% of cases. To prevent the spread of resistant Mycobacterium tuberculosis strains, it is crucial to develop more rapid resistance detection methods. METHODS: To determine the feasibility of using direct sequencing for detecting INH- and RIF-resistant strains, the katG gene, the regulatory region of the inhA gene, and the 81-bp hot-spot region of the rpoB gene from 95 culture isolates and 46 respiratory specimens were sequenced. Total 141 culture isolates were classified by conventional drug susceptibility testing (DST) as INH(R)/RIF(R) (N=30), INH(R)/RIF(S) (N=23), INH(S)/RIF(R) (N=15), and INH(S)/RIF(S) (N=73). RESULTS: Compared with phenotypic DST, the overall sensitivity and specificity of sequencing were 83.0% (44/53) and 96.6% (85/88), respectively, for INH resistance, and 93.3% (42/45) and 100% (96/96), respectively, for RIF resistance. The rates were similar between culture isolates and respiratory specimens. Interestingly, three specimens with inhA -15C>T mutation were susceptible to INH by conventional DST. CONCLUSIONS: Detection of mutations in the katG codon 315, the inhA regulatory region, and the hot-spot region of rpoB would be useful for rapid detection of INH and RIF resistance in Korea.
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Humanos , Antituberculosos/farmacología , Proteínas Bacterianas/genética , Catalasa/genética , Farmacorresistencia Bacteriana Múltiple , Genotipo , Isoniazida/farmacología , Mutación , Mycobacterium tuberculosis/genética , Oxidorreductasas/genética , República de Corea , Rifampin/farmacología , Sensibilidad y Especificidad , Análisis de Secuencia de ADN/métodosRESUMEN
BACKGROUNDS: Mutations of katG and inhA (ORF and promoter) are known to be related to isoniazid (INH) resistance of Mycobacterium tuberculosis. Because reports on these mutations in Korean isolates are limited (i.e. only the frequency of katG codon 463 was evaluated.), we tried to know the kinds of mutations of two genes and their frequencies in INH resistant Korean M. tuberculosis strains. METHODS: PCR was performed to amplify katG (2,223 bp), inhA ORF (-77~897, 975 bp), and inhA promoter (-168~80, 248 bp) from 29 multidrug resistant M. tuberculosis (MDR-TB) DNAs prepared by bead beater-phenol method. Their sequences were determined and analyzed by ABI PRISM 3730 XL Analyzer and MegAlign package program, respectively. RESULTS: All of the isolates had more than one mutation in katG or inhA gene. Twenty seven (93%) of 29 tested strains had katG mutations, which suggests that katG is a critical gene determining INH resistance of M. tuberculosis. Amino acid substitutions, such as Arg463Leu and Ser315Thr, due to point mutations of the katG were the most frequent (62.1% and 55.2%) mutations. In addition, deletion of the katG gene was frequently observed (17.2%). Analyzed Korean MDR-TB isolates also had variable inhA mutations. Point mutation of inhA promoter region, such as -15 (C-->T) was frequently found. Substitution of amino acid (Lsy8Asn) due to point mutation (AAA-->AAC) of inhA ORF was found in 1 isolate. Interestingly, 14 point mutated types that were not previously reported were newly found. While four types resulted in amino acid change, the others were silent mutations. CONCLUSIONS: Although it is not clear that the relationship of these newly found mutations with INH resistance, they show marked diversity in Korean MDR-TB strains. It also suggests their feasibility as a molecular target to supplement determining the INH resistance of clinical isolates because of the possible existence of low-level INH resistant strains.
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Animales , Sustitución de Aminoácidos , Codón , ADN , Ectima Contagioso , Isoniazida , Mycobacterium tuberculosis , Mutación Puntual , Reacción en Cadena de la Polimerasa , Regiones Promotoras Genéticas , TuberculosisRESUMEN
BACKGROUNDS: Mutations of katG and inhA (ORF and promoter) are known to be related to isoniazid (INH) resistance of Mycobacterium tuberculosis. Because reports on these mutations in Korean isolates are limited (i.e. only the frequency of katG codon 463 was evaluated.), we tried to know the kinds of mutations of two genes and their frequencies in INH resistant Korean M. tuberculosis strains. METHODS: PCR was performed to amplify katG (2,223 bp), inhA ORF (-77~897, 975 bp), and inhA promoter (-168~80, 248 bp) from 29 multidrug resistant M. tuberculosis (MDR-TB) DNAs prepared by bead beater-phenol method. Their sequences were determined and analyzed by ABI PRISM 3730 XL Analyzer and MegAlign package program, respectively. RESULTS: All of the isolates had more than one mutation in katG or inhA gene. Twenty seven (93%) of 29 tested strains had katG mutations, which suggests that katG is a critical gene determining INH resistance of M. tuberculosis. Amino acid substitutions, such as Arg463Leu and Ser315Thr, due to point mutations of the katG were the most frequent (62.1% and 55.2%) mutations. In addition, deletion of the katG gene was frequently observed (17.2%). Analyzed Korean MDR-TB isolates also had variable inhA mutations. Point mutation of inhA promoter region, such as -15 (C-->T) was frequently found. Substitution of amino acid (Lsy8Asn) due to point mutation (AAA-->AAC) of inhA ORF was found in 1 isolate. Interestingly, 14 point mutated types that were not previously reported were newly found. While four types resulted in amino acid change, the others were silent mutations. CONCLUSIONS: Although it is not clear that the relationship of these newly found mutations with INH resistance, they show marked diversity in Korean MDR-TB strains. It also suggests their feasibility as a molecular target to supplement determining the INH resistance of clinical isolates because of the possible existence of low-level INH resistant strains.
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Animales , Sustitución de Aminoácidos , Codón , ADN , Ectima Contagioso , Isoniazida , Mycobacterium tuberculosis , Mutación Puntual , Reacción en Cadena de la Polimerasa , Regiones Promotoras Genéticas , TuberculosisRESUMEN
OBJECTIVE To develop a new DNA chip for rapid detection of rpoB,katG and inhA gene mutation in rifampin and/or isoniazid-resistant Mycobacterium tuberculosis isolated from clinical laboratorial specimen.METHODS We designed 16 oligonucleotide probes specific for detection of the mutant sequences in genes rpoB,katG and inhA.RESULTS Mutations were found in 26 strains(100%) of 26 randomly selected rifampin-resistant M.tuberculosis and 24 strains(80%) of 30 randomly selected isoniazid-resistant M.tuberculosis by DNA chip.CONCLUSIONS DNA chip technology has high sensitivity and specificity in detection of rifampin-resistant M.tuberculosis and may be applied in clinical diagnosis.
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BACKGROUND: Development of rapid drug susceptibility testing provides the opportunity for rapid identification of individuals with drug resistant tubercle bacilli, allowing selection of appropriate therapeutic regimens. METHODS: A total of 502 drug resistant isolates were subjected to reverse blot hybridization assay to detect mutations within genes (rpoB, katG, inhA, and ahpC) associated with rifampicin (RMP) and isoniazid (INH) resistance. RESULTS: Among the 264 RMP resistant strains (RMPR) tested, the most prevalent mutation was the Ser531Leu seen in 121 strains (46%). The second common mutation occurred in 84 strains (32%) at codon 526. And 27 strains (10%) showed the mutation at codon 516. Among all 469 INH resistant strains (INHR), the katG mutation was responsible for INH. The inhA mutation was present in 88 strains (19%). In 11 isolates (2%), coexisting of the katG and inhA mutations were identified. Reverse hybridization assay successfully detected over 80% of INHR and over 92% of RMPR among Korean isolates. CONCLUSION: Reverse hybridization was useful for rapid detection of INHR and RMPR.