Characterization of Three Cases of Primary Hypogalactia in Jalisco, Mexico.

INTRODUCTION: Human lactation should be taken into account as an important issue for the international agenda. Despite advances in lactation information and knowledge, insufficient milk production is still a concern for mothers and health practitioners, including International Board Certified Lactation Consultants and others. Primary hypogalactia, or insufficient milk production is uncommon, but should be considered when there is poor weight gain and decreased urine output in infants despite good latch-on and suckling, or anatomic differences in the physical exam of the lactating breast. MAIN ISSUE: This case series presents three cases illustrating insufficient milk production resulting in infants who experienced significant dehydration and poor weight gain. MANAGEMENT: Primary hypoplasia was diagnosed by means of a thorough interview and physical examination that entailed a consultation with a physician who was also an International Board Certified Lactation Consultant. CONCLUSION: Awareness of an infant's feeding needs and proper evaluation of a child's health status is paramount if health care providers are to identify the important factors contributing to breastfeeding problems. In some instances, breastfeeding goals cannot be achieved, and then the provider's role becomes support in coming to terms with persistent insufficient milk production, and coordinating appropriate supplementation to meet each baby's nutritional needs.

Frequency of urinary pesticides in children: a scoping review.

Pesticides are any mix of ingredients and substances used to eliminate or control unwanted vegetable or animal species recognized as plagues. Its use has been discussed in research due to the scarcity of strong scientific evidence about its health effects. International literature is still insufficient to establish a global recommendation through public policy. This study aims to explore international evidence of the presence of pesticides in urine samples from children and their effects on health through a scoping review based on the methodology described by Arksey and O'Malley. The number of articles resulting from the keyword combination was 454, and a total of 93 manuscripts were included in the results and 22 were complementary. Keywords included in the search were: urinary, pesticide, children, and childhood. Children are exposed to pesticide residues through a fruit and vegetable intake environment and household insecticide use. Behavioral effects of neural damage, diabetes, obesity, and pulmonary function are health outcomes for children that are commonly studied. Gas and liquid chromatography-tandem mass spectrometry methods are used predominantly for metabolite-pesticide detection in urine samples. Dialkylphosphates (DAP) are common in organophosphate (OP) metabolite studies. First-morning spot samples are recommended to most accurately characterize OP dose in children. International evidence in PubMed supports that organic diets in children are successful interventions that decrease the urinary levels of pesticides. Several urinary pesticide studies were found throughout the world's population. However, there is a knowledge gap that is important to address (public policy), due to farming activities that are predominant in these territories.

Non-traditional risk factors of progression of chronic kidney disease in adult population: a scoping review.

Chronic kidney disease (CKD) has become a public health concern over the last several years. Nowadays developed countries spend around 3% of their annual health-care budget on patients with CKD. According to the scientific community the most remarkable risk factors for CKD are diabetes and hypertension. Unknown CKD etiology has been reported as a global phenomenon including uncommon risk factors such as: dehydration, leptospirosis, heat stress, water quality, and others. This study aims to report non-traditional risk factors for ESRD based on a scoping review methodology. The scoping review methodology described by Arksey and O'Malley was used by performing an extensive review of the information. A total of 46 manuscripts were reviewed. The non-traditional ESRD risk factors are depicted based on six categories. Gender and ethnicity have been considered as risk factors for ESRD. Erythematous systemic lupus (ESL) is reported as an important risk factor for ESRD. Pesticide use has been an significant risk factor due to its effects on human and environmental health. Some compounds commonly used in homes against insects and plants are related to ESRD. Congenital and hereditary diseases in the urinary tract have been studied as a cause of ESRD in children and young adults. End-stage renal disease is a major concern for public health on a global level. As it can be seen, non-traditional risk factors are several and have different etiologies. It is necessary to put the issue on the table and add it to the public agenda in order to find multidisciplinary solutions.

Seasonal Urinary Levels of Glyphosate in Children From Agricultural Communities.

BACKGROUND: Glyphosate is one of the most commonly used pesticides in agricultural activities worldwide. For the last 20 years, its use has increased rapidly becoming a public health concern. The IARC classified glyphosate as probably carcinogenic; however, the reported evidence is not enough to establish a statement. OBJECTIVE: This work aimed to measure glyphosate levels in the urine of children from a single rural community in an endemic region of chronic kidney disease and malnutrition. METHODS: A cross-sectional study was carried out in a rural community in western Mexico. The study included 95 children between the ages of 6 and 16. A urine sample (first-morning spot) was obtained from children and processed to measure glyphosate levels using high-performance liquid chromatography coupled with tandem mass spectrometry. RESULTS: All samples tested positive for glyphosate levels. Urine glyphosate levels were related to the season and the age of the children. CONCLUSION: Glyphosate is present in children of all ages in the community even if they were not in direct contact with it. No toxicity cases were reported, nor were there other health problems related to glyphosate. However, more long-term studies should be done that provide a statement regarding the harmful effects that glyphosate has on public health.

Microbiological profile of urinary tract infections in a tertiary medical facility in Western Mexico: An update.

Urinary tract infections (UTIs) are one of the most important issues in modern medicine. In developing countries, the use of antibiotics is a common practice, and due to this, antibiotic resistance has increased. The objective of this research was to update and report on the microbiological profile of urinary tract infections based on the number of positive urine cultures (UCs), resistance, sensitivity, and the prevalence of bacterial strains. The results were obtained from the database of a tertiary medical facility in Western Mexico. The number of positive UCs was 1769 from inpatients and outpatients who were users of medical services in the hospital from January to December of 2017. The most commonly isolated microorganism was E. coli, with 1225 cases, of which 603 (49.2%) were ESBL (Extended-Spectrum Beta-Lactamase-producing bacteria) strains. The resistance rate of nitrofurantoin was 36.6%, and meropenem showed the most promising results with a resistance rate of only 7.1%. Resistances to quinolones and cephalosporins among the isolates investigated were 51%-67%. Based on our results, it is necessary to increase controls and to improve management protocols in order to achieve better medical practices by reducing antibiotic resistance.

PTPN22 gene functional polymorphism (rs2476601) in older adults with frailty syndrome.

The frailty syndrome is a common clinical marker of vulnerability in older adults conducive to an overall decline in inflammatory stress responsiveness; yet little is known about the genetic risk factors for frailty in elderly. Our aim was to investigate the association between the rs2476601 polymorphism in PTPN22 gene and susceptibility to frailty in Mexican older adults. Data included 630 subjects 70 and older from The Coyoacán cohort, classified as frail, pre-frail, and non-frail following Fried's criteria. Sociodemographic and clinical characteristics were compared between groups at baseline and after a multivariate analysis. The rs2476601 polymorphism was genotyped by TaqMan genotyping assay using real-time PCR and genotype frequencies were determined for each frailty phenotype in all participants and subsets by age range. Genetic association was examined using stratified and interaction analyses adjusting for age, sex and variables selected in the multivariate analysis. Disability for day-life activities, depression and cognitive impairment were associated with the risk of pre-frailty and frailty at baseline and after adjustment. Carrying the T allele increased significantly the risk of frailty in patients 76 and older (OR 5.64, 95% CI 4.112-7.165) and decreased the risk of pre-frailty under no clinical signs of depression (OR 0.53; 95% CI 0.17-1.71). The PTPN22 polymorphism, rs2476601, could be a genetic risk factor for frailty as subject to quality of life. This is the first study analyzing such relationship in Mexican older adults. Confirming these findings requires additional association studies on wider age ranges in populations of older adults with frailty syndrome.

Microbiome alterations are related to an imbalance of immune response and bacterial translocation in BDL-rats.

OBJECTIVES: Bacterial translocation in patients with cirrhosis is an important triggering factor for infections and mortality. In the bile duct ligation (BDL) model, crucial players of bacterial translocation are still unknown. This study aims to determine the interrelation between microbiome composition in the colon, mesenteric lymph nodes, and liver, as well as the local inflammatory microenvironment in the BDL model. MATERIALS AND METHODS: Liver damage was assayed by Masson trichrome staining, and hepatic enzymes. The diversity of microbiota in colon stools, mesenteric lymph nodes, and liver was determined by 16S rRNA pyrosequencing. Cytokine expression in mesenteric lymph nodes was analyzed by qRT-PCR. RESULTS: Our results show that Proteobacteria was the predominant phylum found to translocate to mesenteric lymph nodes and liver in cirrhotic rats. Bile duct ligation induces a drastic intestinal dysbiosis, revealed by an increased relative abundance of Sarcina, Clostridium, Helicobacter, Turicibacter, and Streptococcus genera. However, beneficial bacteria, such as Lactobacillus, Prevotella and Ruminococcus were found to be notably decreased in BDL groups. Mesenteric pro-inflammatory (TNF-α, IL-1ß, IL-6, TLR-4) and regulatory (TGF-ß, Foxp3, and IL-10) molecules at 30 days post-BDL were significantly increased. Conversely, TGF-ß and Foxp3 were significantly augmented at 8 days post-BDL. CONCLUSION: Dysbiosis in the colon and mesenteric lymph nodes is linked to an imbalance in the immune response; therefore, this may be an important trigger for bacterial translocation in the BDL model.

[Characterization of gene expression associated with both the AcrAB/TolC system and the membrane permeability, in Salmonella spp isolates with and without gyrA mutation]. / Caracterización de la expresión de genes asociados con el sistema AcrAB/TolC y la permeabilidad de membrana en aislados de Salmonella spp. con y sin mutación en gyrA.

Introduction. The marA, soxS, ramA, acrB and ompF genes have been studied in order to characterize mechanisms of AcrAB-TolC active efflux pumps and membrane permeability alterations that reduce fluoroquinolones susceptibility in Salmonella spp. Methods. Mutations in marA, soxS, ramA, acrB and ompF genes were detected, as well as their expression levels in presence and absence of ciprofloxacin, calculating the level of change between them by qPCR. Data were analysed by using SPSS 19.0. Results. No mutations in these genes were found, but both AcrAB-TolC regulatory genes and structural acrB gene expression were affected by ciprofloxacin in both mutant strains and wild type bacterial strains (WT). The activation of the marA gene in presence of drug was higher in WT strains (level of change 0.823) than in mutants strains (level of change 0.158; p=0.049). In gyrA mutants, a reduction in ompF gene expression in presence of ciprofloxacin was found (level of change -0.949 p=0.017). Conclusion. The reduction of fluoroquinolones susceptibility in Salmonella spp is a complex process, in which several different bacterial mechanisms are involved. This study has found a high difference in the degree of participation among studied mechanisms, between bacterial strains with and without gyrA mutation. Whereas WT strains activated efflux pumps especially through marA gene, mutants supressed ompF gene expression related to porins.

Caracterización de la expresión de genes asociados con el sistema AcrAB/TolC y la permeabilidad de membrana en aislados de Salmonella spp. con y sin mutación en gyrA / Characterization of gene expression associated with both the AcrAB/TolC system and the membrane permeability, in Salmonella spp isolates with and without gyrA mutation

Introducción. Se han estudiado los genes marA, soxS, ramA, acrB y ompF para caracterizar los mecanismos del sistema de expulsión activa AcrAB/TolC y las alteraciones en la permeabilidad de membrana que reducen la sensibilidad a fluoroquinolonas en Salmonella spp. Métodos. Se detectaron las mutaciones en los genes marA, soxS, ramA, acrB y ompF y se cuantificó su nivel de expresión en presencia y ausencia de ciprofloxacino calculando su valor de nivel de cambio por qPCR. Los datos se analizaron estadísticamente mediante el programa SPSS 19.0. Resultados. No se encontraron mutaciones en ninguno de los genes, pero la expresión de los genes reguladores de AcrAB/TolC y del gen estructural acrB se vieron afectados por la presencia de ciprofloxacino tanto en las cepas con mutación en gyrA como en las cepas silvestres. La activación del gen marA en presencia del fármaco fue mayor en las cepas silvestres (nivel de cambio de 0,823) que en las mutantes (nivel de cambio de 0,158; p=0,049). Se vio una disminución de la expresión del gen ompF en presencia de ciprofloxacino en cepas con mutación (nivel de cambio de -0,949 p=0,017). Conclusión. La disminución de la sensibilidad a fluoroquinolonas en Salmonella spp es un proceso complejo, donde intervienen diferentes mecanismos bacterianos. Este estudio encuentra gran diferencia en el grado de participación de los mecanismos estudiados entre las cepas con y sin mutación en gyrA. Mientras que las cepas silvestres activan el sistema de expulsión activa, especialmente a través del gen marA, las cepas con mutación reprimen la expresión del gen ompF, relacionado con las porinas (AU)

Introduction. The marA, soxS, ramA, acrB and ompF genes have been studied in order to characterize mechanisms of AcrAB-TolC active efflux pumps and membrane permeability alterations that reduce fluoroquinolones susceptibility in Salmonella spp. Methods. Mutations in marA, soxS, ramA, acrB and ompF genes were detected, as well as their expression levels in presence and absence of ciprofloxacin, calculating the level of change between them by qPCR. Data were analysed by using SPSS 19.0. Results. No mutations in these genes were found, but both AcrAB-TolC regulatory genes and structural acrB gene expression were affected by ciprofloxacin in both mutant strains and wild type bacterial strains (WT). The activation of the marA gene in presence of drug was higher in WT strains (level of change 0.823) than in mutants strains (level of change 0.158; p=0.049). In gyrA mutants, a reduction in ompF gene expression in presence of ciprofloxacin was found (level of change -0.949 p=0.017). Conclusion. The reduction of fluoroquinolones susceptibility in Salmonella spp is a complex process, in which several different bacterial mechanisms are involved. This study has found a high difference in the degree of participation among studied mechanisms, between bacterial strains with and without gyrA mutation. Whereas WT strains activated efflux pumps especially through marA gene, mutants supressed ompF gene expression related to porins (AU)

Frecuencia de la resistencia a fluoroquinolonas en Mycobacterium tuberculosisen Elche (Alicante) / Frequency of fluoroquinolone resistance in Mycobacterium tuberculosis in Southeast Spain

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Plasmid-mediated quinolone resistance (PMQR) and mutations in the topoisomerase genes of Salmonella enterica strains from Brazil.

The objective of this study was to identify mutations in the Quinolone Resistance Determining sources Regions (QRDR) of the gyrA, gyrB, parC, and parE genes and to determine if any of the qnr variants or the aac(6')-Ib-cr variant were present in strains of Salmonella spp. isolated in Brazil. A total of 126 Salmonella spp. strains from epidemic (n = 114) and poultry (n = 12) origin were evaluated. One hundred and twelve strains (88.8%) were resistant to nalidixic acid (NAL) and 29 (23.01%) showed a reduced susceptibility to ciprofloxacin (Cip). The mutations identified were substitutions limited to the QRDR of the gyrA gene in the codons for Serine 83, Aspartate 87 and Alanine 131. The sensitivity to NAL seems to be a good phenotypic indication of distinguishing mutated and non-mutated strains in the QRDR, however the double mutation in gyrA did not cause resistance to ciprofloxacin. The qnrA1 and qnrB19 genes were detected, respectively, in one epidemic strain of S. Enteritidis and one strain of S. Corvallis of poultry origin. Despite previous detection of qnr genes in Brazil, this is the first report of qnr gene detection in Salmonella, and also the first detection of qnrB19 gene in this country. The results alert for the continuous monitoring of quinolone resistance determinants in order to minimize the emergence and selection of Salmonella spp. strains showing reduced susceptibility or resistance to quinolones.

Expression of the marA, soxS, acrB and ramA genes related to the AcrAB/TolC efflux pump in Salmonella entérica strains with and without quinolone resistance-determining regions gyrA gene mutations

Several studies have been conducted in recent years to elucidate the structure, function and significance of AcrB, MarA, SoxS and RamA in Salmonella enterica. In this study, the relative quantification of acrB, soxS, marA and ramA genes expression was evaluated in 14 strains of S. enterica, with or without accompanying mutations in the quinolone resistance-determining regions of the gyrA gene, that were exposed to ciprofloxacin during the exponential growth phase. The presence of ciprofloxacin during the log phase of bacterial growth activated the genes marA, soxS, ramA and acrB in all S. enterica strains analyzed in this study. The highest expression levels for acrB were observed in strains with gyrA mutation, and marA showed the highest expression in the strains without mutation. Considering only the strains with ciprofloxacin minimum inhibitory concentration values < 0.125 [1]g/mL (sensitive to ciprofloxacin), the most expressed gene in the strains both with and without mutations was acrB. In the strains with ciprofloxacin minimum inhibitory concentration values > 0.125 [1]g/mL (low susceptibility), with and without mutations in gyrA, the most expressed gene was marA. In this study, we observed that strains resistant to nalidixic acid may express genes associated with the efflux pump and the expression of the AcrAB-TolC pump genes seems to occur independently of mutations in gyrA.

Expression of the marA, soxS, acrB and ramA genes related to the AcrAB/TolC efflux pump in Salmonella enterica strains with and without quinolone resistance-determining regions gyrA gene mutations.

Several studies have been conducted in recent years to elucidate the structure, function and significance of AcrB, MarA, SoxS and RamA in Salmonella enterica. In this study, the relative quantification of acrB, soxS, marA and ramA genes expression was evaluated in 14 strains of S. enterica, with or without accompanying mutations in the quinolone resistance-determining regions of the gyrA gene, that were exposed to ciprofloxacin during the exponential growth phase. The presence of ciprofloxacin during the log phase of bacterial growth activated the genes marA, soxS, ramA and acrB in all S. enterica strains analyzed in this study. The highest expression levels for acrB were observed in strains with gyrA mutation, and marA showed the highest expression in the strains without mutation. Considering only the strains with ciprofloxacin minimum inhibitory concentration values<0.125 µg/mL (sensitive to ciprofloxacin), the most expressed gene in the strains both with and without mutations was acrB. In the strains with ciprofloxacin minimum inhibitory concentration values ≥ 0.125 µg/mL (low susceptibility), with and without mutations in gyrA, the most expressed gene was marA. In this study, we observed that strains resistant to nalidixic acid may express genes associated with the efflux pump and the expression of the AcrAB-TolC pump genes seems to occur independently of mutations in gyrA.

Plasmid-mediated quinolone resistance (PMQR) and mutations in the topoisomerase genes of Salmonella enterica strains from Brazil

The objective of this study was to identify mutations in the Quinolone Resistance Determining sources Regions (QRDR) of the gyrA, gyrB, parC, and parE genes and to determine if any of the qnr variants or the aac(6')-Ib-cr variant were present in strains of Salmonella spp. isolated in Brazil. A total of 126 Salmonella spp. strains from epidemic (n = 114) and poultry (n = 12) origin were evaluated. One hundred and twelve strains (88.8%) were resistant to nalidixic acid (NAL) and 29 (23.01%) showed a reduced susceptibility to ciprofloxacin (Cip). The mutations identified were substitutions limited to the QRDR of the gyrA gene in the codons for Serine 83, Aspartate 87 and Alanine 131. The sensitivity to NAL seems to be a good phenotypic indication of distinguishing mutated and nonmutated strains in the QRDR, however the double mutation in gyrA did not cause resistance to ciprofloxacin. The qnrA1 and qnrB19 genes were detected, respectively, in one epidemic strain of S. Enteritidis and one strain of S. Corvallis of poultry origin. Despite previous detection of qnr genes in Brazil, this is the first report of qnr gene detection in Salmonella, and also the first detection of qnrB19 gene in this country. The results alert for the continuous monitoring of quinolone resistance determinants in order to minimize the emergence and selection of Salmonella spp. strains showing reduced susceptibility or resistance to quinolones.

Comparison of the bactericidal activity of various fluoroquinolones against Mycobacterium tuberculosis in an in vitro experimental model.

OBJECTIVES: To compare the bactericidal activity of various fluoroquinolones against Mycobacterium tuberculosis in the latent and exponential growth phases. METHODS: Ciprofloxacin, levofloxacin and moxifloxacin were tested against 16 M. tuberculosis clinical isolates (4 resistant and 12 susceptible to fluoroquinolones) from Elche, Spain, isolated between 1992 and 2009. To study bactericidal activity, an inoculum of approximately 10(5) cfu of each isolate was cultured in Middlebrook 7H9 broth. The broth was previously acidified to pH 4.6 to obtain the microorganism in the stationary phase. Cultures with different concentrations (0.1 to 50 mg/L) of antibiotic and antibiotic-free controls were incubated for 48 h then plated onto Middlebrook 7H11 to detect bacterial killing. In all stages of the process the M. tuberculosis strain ATCC 41323 was included as a quality control to ensure reproducible results. RESULTS: Moxifloxacin and levofloxacin were found to exhibit bactericidal activity at lower concentrations and against more strains in both the latent and the exponential growth phases compared with ciprofloxacin. The bactericidal activity of moxifloxacin was greater than that of levofloxacin against microorganisms in the exponential growth phase, but the opposite was true in the latent phase. CONCLUSIONS: Our data confirm the usefulness of moxifloxacin in the treatment of tuberculosis and suggest that levofloxacin may be used as an alternative drug in the treatment of latent tuberculosis when it is not possible to use isoniazid. Based on the results presented, ciprofloxacin appears to be a poor choice.

Plasmid-mediated quinolone resistance by genes qnrA1 and qnrB19 in Salmonella strains isolated in Brazil.

Considering the importance of the mechanisms involved in quinolone resistance, this study evaluate the presence of PMQR in 126 epidemic and not epidemic strains of Salmonella spp. It was noted that presence of PMQR, by itself, did not generate resistance to ciprofloxacin; but detection of qnr genes in Salmonella spp. and the identification of the qnrB19 variant in a strain of poultry origin alert for the indiscriminate use of quinolones in poultry production, that can result in a pressure for mutant selection of resistant strains with a clinical limitation use of FQs in the near future. And last but not least, is the need to continued study of resistance mechanisms and to monitor the microbial resistance profile of epidemiological strains.

Interaction between linezolid and Mycobacterium tuberculosis in an experimental in vitro model.

As linezolid is emerging as a useful alternative in the treatment of multi-resistant tuberculosis, we developed an in vitro model to elucidate certain aspects of the interaction that takes place between the microorganism and the antibiotic. We found that the drug does not generate resistant mutants following repeated exposure at subinhibitory concentrations and that it exhibits bactericidal activity against most of the susceptible strains in vitro at high concentrations (83.3% of the strains at concentrations of 50 µg/mL) in the exponential growth phase. This bactericidal activity is much less if the microorganism is in nonreplicating phase, even in the case of susceptible strains in vitro. Therefore, linezolid appears to be an alternative as far as generation of resistance is concerned in the treatment of multi-resistant tuberculosis. However, it has limited bactericidal capacity against strains in nonreplicant phase, which makes it necessary to use linezolid in combination with other drugs that act on these microorganisms.

Identification of bacterial DNA in noninfectious pleural fluid with a highly sensitive PCR method.

BACKGROUND: Bacterial DNA due to bacterial translocation has been identified in noninfectious ascitic fluid samples. OBJECTIVE: This study investigated the possible presence of bacterial DNA in the pleural fluid of patients with pleural effusions of noninfectious origin, using a highly sensitive PCR-based method. METHODS: Pleural fluid samples from 175 patients (average age ± SD: 69 ± 14 years) with noninfectious pleural effusion (62 transudates, 113 exudates) were analyzed. Bacterial DNA was detected using nested PCR with amplification of a fragment of the gene r16S, with 2 amplification protocols, i.e. low sensitivity (10 and 40 cycles) and high sensitivity (40 and 40 cycles). RESULTS: With the less sensitive amplification process, only 1 sample was positive (Haemophilus parainfluenzae in a patient with hepatic hydrothorax). With the highly sensitive nested PCR method, bacterial DNA was identified in the pleural fluid, of both transudative and exudative origin, of 75 of the 175 patients (43%). In cases of isolation of a single bacterium, the more frequent were Escherichia coli, Salmonella enterica and Streptococcus pneumoniae. CONCLUSIONS: Regardless of its origin, bacterial DNA can be identified in almost half of noninfectious pleural effusions by using a highly sensitive PCR-based method. The possible clinical significance or prognostic value of these findings deserves to be evaluated.

Comparison of methods of DNA extraction for real-time PCR in a model of pleural tuberculosis.

Molecular methods have been reported to have different sensitivities in the diagnosis of pleural tuberculosis and this may in part be caused by the use of different methods of DNA extraction. Our study compares nine DNA extraction systems in an experimental model of pleural tuberculosis. An inoculum of Mycobacterium tuberculosis was added to 23 pleural liquid samples with different characteristics. DNA was subsequently extracted using nine different methods (seven manual and two automatic) for analysis with real-time PCR. Only two methods were able to detect the presence of M. tuberculosis DNA in all the samples: extraction using columns (Qiagen) and automated extraction with the TNAI system (Roche). The automatic method is more expensive, but requires less time. Almost all the false negatives were because of the difficulty involved in extracting M. tuberculosis DNA, as in general, all the methods studied are capable of eliminating inhibitory substances that block the amplification reaction. The method of M. tuberculosis DNA extraction used affects the results of the diagnosis of pleural tuberculosis by molecular methods. DNA extraction systems that have been shown to be effective in pleural liquid should be used.