Epidemiology of Non-Tuberculous Mycobacteria isolated from clinical specimens in Madrid, Spain, from 2013 to 2017.

The epidemiology of non-tuberculous mycobacteria (NTM) in Spain is largely unknown because systematic reporting is not compulsory. The aim of our study was to describe the frequency and diversity of NTM species in our region and their distribution according to the source sample, gender, and age of the patients. We performed a multicenter study of all NTM isolated in 24 public hospitals in Madrid from 2013 to 2017. A total of 6.923 mycobacteria were isolated: 4535 (65.5%) NTM, and 2.388 (34.5%) Mycobacterium tuberculosis complex (MTB). Overall, 61 different NTM species were identified. The most frequently isolated species were Mycobacterium avium complex (47.7%), M. lentiflavum (12.2%), M. gordonae (9.2%), M. fortuitum (8.9%), and M. abscessus (3.9%). Whereas MTB cases were stable during the study period, the number of NTM isolates increased considerably from 930 isolates in 2013 to 1012 in 2017; a sharp increase occurred in the last year. The rise in NTM isolates was mostly due to M. lentiflavum, M. kansasii, and M. abscessus mainly isolated from respiratory specimens in patients older than 60. The increase in isolation rate of NTM in our region is consistent with the increasing rates reported worldwide in the last decades. The rise in NTM isolates was mainly attributed to M. lentiflavum but it also should be noted the increasing of species with high pathogenic potential such as M. kansasii and M. abscessus.

Combined analysis of copy number alterations by single-nucleotide polymorphism array and MYC status in non-metastatic breast cancer patients: comparison according to the circulating tumor cell status.

Recent technological advances have made it possible to detect circulating tumor cells (CTCs) as a prognostic marker in operable breast cancer patients. Whether the presence of CTCs in cancer patients correlates with molecular alterations in the primary tumor has not been widely explored. We identified 14 primary breast cancer specimens with known CTC status, in order to evaluate the presence of differential genetic aberrations by using SNP array assay. There was a global increase of altered genome, CNA, and copy-neutral loss of heterozygosity (cn-LOH) observed in the CTC-positive (CTC(+)) versus CTC-negative (CTC(-)) cases. As the preliminary results showed a higher proportion of copy number alteration (CNA) at 8q24 (MYC loci) and the available evidence supporting the role of MYC in the processes cancer metastases is conflicting, MYC status was determined in tissue microarray sections in a larger series of patients (n = 49) with known CTC status using FISH. MYC was altered in 62% (16/26) CTC(+) patients and in 43% (6/14) CTC(-) patients (p = 0.25). Based on the observation in our study, future studies involving a larger number of patients should be performed in order to definitively define if this correlation exists.

Environmental and biological factors influencing the UV-C resistance of Listeria monocytogenes.

In this investigation, the effect of microbiological factors (strain, growth phase, exposition to sublethal stresses, and photorepair ability), treatment medium characteristics (pH, water activity, and absorption coefficient), and processing parameters (dose and temperature) on the UV resistance of Listeria monocytogenes was studied. The dose to inactivate 99.99% of the initial population of the five strains tested ranged from 21.84 J/mL (STCC 5672) to 14.66 J/mL (STCC 4031). The UV inactivation of the most resistant strain did not change in different growth phases and after exposure to sublethal heat, acid, basic, and oxidative shocks. The pH and water activity of the treatment medium did not affect the UV resistance of L. monocytogenes, whereas the inactivation rate decreased exponentially with the absorption coefficient. The lethal effect of UV radiation increased synergistically with temperature between 50 and 60 °C (UV-H treatment). A UV-H treatment of 27.10 J/mL at 55 °C reached 2.99 and 3.69 Log10 inactivation cycles of L. monocytogenes in orange juice and vegetable broth, and more than 5 Log10 cycles in apple juice and chicken broth. This synergistic effect opens the possibility to design UV combined processes for the pasteurization of liquid foods with high absorptivity.

Long-term effect of hormone therapy on bone in early menopause: vertebral fractures after 20 years.

OBJECTIVE: The role of menopausal hormone therapy (HT) on vertebral fracture prevention after treatment discontinuation is controversial. The aim of this study was to assess the incidence of vertebral fracture in a group of women who received HT in early menopause compared with another group who did not receive such treatment after 20 years of follow-up. SUBJECTS AND METHODS: In 1990, we included 177 patients aged 43-57 years old (mean 49.1 ± 3.9 years) in a prospective study to evaluate the effect of different HT regimens on bone metabolism and mineral density. After 20-21 years, a total of 49 patients from the initial study were retrieved. These patients were divided into two groups: the first group included women who had taken HT, and those who constituted the control groups and had not taken HT formed the second group. Clinical and demographic data were analyzed and vertebral fracture was assessed by radiology using the Genant semiquantitative scale. RESULTS: Of the 49 patients enrolled, 32 (65.3%) received HT for an average of 5.5 (± 2.96) years while the 17 (34.7%) remaining belonged to the control group without treatment. A higher rate of vertebral fracture was observed in the group receiving HT (p = 0.03). Depending on the degree of fracture (Genant semiquantitative method), subsequent analysis by subgroups corroborated the higher rate in the group receiving HT in all cases (p < 0.05). Multivariate analysis ruled out the effect of the clinical and demographic variables (current age, age at menopause, body mass index, type of menopause and drugs for the treatment of osteoporosis) in the final result. CONCLUSION: In spite of the fact that this study does not have a large enough sample, our data suggest that HT used in the early years of menopause does not present a long-term protective effect on vertebral fracture after discontinuing treatment.

Impact of WALANT Hand Surgery in a Secondary Care Hospital in Spain. Benefits to the Patient and the Health System.

Purpose: The aim of this study is to compare patient benefits and economic costs of hand surgeries using the wide-awake local anesthesia no tourniquet (WALANT) technique versus a conventional major outpatient suite and review outcomes and complications in a series of cases of patients operated on using the WALANT technique. Methods: A prospective cohort study was first conducted comparing 150 cases of ambulatory hand surgery (carpal tunnel syndrome and trigger finger) using the WALANT technique and not requiring an operating room setting with 150 cases of outpatient surgery performed in an operating room involving a preoperative evaluation and the use of sedation and tourniquet. Preoperative, intraoperative, and postoperative pain was monitored, and days requiring postoperative analgesia were recorded. The resources and costs were evaluated. and patient satisfaction was assessed using a specific survey.Subsequently, 580 patient medical records were retrospectively reviewed, including 419 carpal tunnel syndrome and 197 trigger finger interventions (616 WALANT surgeries). Results: Intraoperative pain was equivalent for both groups, and postoperative pain was significantly lower in the WALANT group, with a reduced need for analgesics. Satisfaction was greater for the local anesthesia group. The use of personnel resources and hospital materials was reduced in the WALANT group, with a total estimated cost savings of 1.019 USD per patient.There were no complications related to the WALANT technique and the lidocaine and adrenaline combination. We found a complication rate of 5.58%, and, in line with the literature, most complications were minor, managed conservatively, and not related to the anesthetic technique. Conclusions: Procedures such as carpal tunnel and trigger finger surgeries can be safely performed using wide-awake surgery. Patient satisfaction is higher than with the conventional procedure performed in the operating room. Pain control is excellent, especially during the postoperative period. Clinical relevance: Hand surgery patients benefit from the WALANT technique in terms of comfort and timeliness because there is no need for preoperative tests or evaluations. In addition, it represents significant savings in hospital resources. In our case series, complications were in line with those previously reported with other anesthetic techniques.

Inactivation of Salmonella enterica by UV-C light alone and in combination with mild temperatures.

The aim of this investigation was to study the efficacy of the combined processes of UV light and mild temperatures for the inactivation of Salmonella enterica subsp. enterica and to explore the mechanism of inactivation. The doses to inactivate the 99.99% (4D) of the initial population ranged from 18.03 (Salmonella enterica serovar Typhimurium STCC 878) to 12.75 J ml(-1) (Salmonella enterica serovar Enteritidis ATCC 13076). The pH and water activity of the treatment medium did not change the UV tolerance, but it decreased exponentially by increasing the absorption coefficient. An inactivating synergistic effect was observed by applying simultaneous UV light and heat treatment (UV-H). A less synergistic effect was observed by applying UV light first and heat subsequently. UV did not damage cell envelopes, but the number of injured cells was higher after a UV-H treatment than after heating. The synergistic effect observed by combining simultaneous UV and heat treatment opens the possibility to design combined treatments for pasteurization of liquid food with high UV absorptivity, such as fruit juices.

Interstitial disease associated with connective tissue disease and vasculitis.

Systemic autoimmune diseases comprise a complex, heterogeneous group of entities. Noteworthy among the pulmonary complications of these entities is interstitial involvement, which manifests with the same radiopathologic patterns as in idiopathic interstitial pneumonia. High-resolution computed tomography is the gold-standard imaging technique; it enables us to identify and classify the disease and to determine its extent, providing useful information about the prognosis. In this group of processes, the most common pattern of presentation is nonspecific interstitial pneumonia. It is essential for radiologists to work together in collaboration with other specialists to reach the correct diagnosis and enable appropriate, integrated treatment.

Cord occlusion followed by laser cord transection in monochorionic monoamniotic discordant twins.

OBJECTIVE: To compare the outcomes of a consecutive case series of monochorionic (MC) monoamniotic (MA) discordant twins treated with umbilical cord occlusion and transection, with those of a cohort of MC diamniotic (DA) twins treated with conventional cord occlusion. METHODS: This study included 17 MCMA twins (12 true MA and five iatrogenic) treated with cord occlusion and transection and a control group of 72 MCDA discordant twins treated during the same period with cord occlusion in a single center. Duration of surgery, rates of preterm delivery (PTD) or preterm premature rupture of membranes (PPROM) < 32 weeks and intrauterine fetal demise (IUFD), perinatal outcome and neonatal survival were prospectively recorded in both groups. RESULTS: Median durations of surgery were 28.5 (range, 14.0-74.0) min and 24.0 (3.0-60.0) min in the cord transection and control groups, respectively (P = 0.24). There were no significant differences between cord transection and control groups in the rates of PPROM (35.3% vs. 20.8%, P = 0.22), PTD (41.2% vs. 28.2%, P = 0.29), IUFD (0% vs. 2.8%, P = 1.0) and neonatal survival (76.5% vs. 80.6%, P = 1.0). Gestational age at delivery (median 35.0 (24.5-39.0) vs. 37.1 (26.2-41.0) weeks, P = 0.21) and fetal birth weight (2215 (800-3200) g vs. 2605 (588-3830) g, P = 0.51) were similar between study groups. CONCLUSION: Cord occlusion and transection in MCMA discordant twins resulted in similar perinatal outcomes to those of MCDA discordant twins treated with cord occlusion.

Characterization of Mycobacterium tuberculosis Beijing isolates from the Mediterranean area.

BACKGROUND: The Beijing lineage of Mycobacterium tuberculosis is causing concern due to its global distribution and its involvement in severe outbreaks. Studies focused on this lineage are mainly restricted to geographical settings where its prevalence is high, whereas those in other areas are scarce. In this study, we analyze Beijing isolates in the Mediterranean area, where this lineage is not prevalent and is mainly associated with immigrant cases. RESULTS: Only 1% (N = 26) of the isolates from two population-based studies in Spain corresponded to Beijing strains, most of which were pan-susceptible and from Peruvian and Ecuadorian patients. Restriction fragment length polymorphism typing with the insertion sequence IS6110 identified three small clusters (2-3 cases). Mycobacterial interspersed repetitive unit-variable number tandem repeat typing (MIRU-15) offered low discriminatory power, requiring the introduction of five additional loci. A selection of the Beijing isolates identified in the Spanish sample, together with a sample of Beijing strains from Italy, to broaden the analysis context in the Mediterranean area, were assayed in an infection model with THP-1 cells. A wide range of intracellular growth rates was observed with only two isolates showing an increased intracellular replication, in both cases associated with contained production of TNF-alpha. No correlation was observed between virulence and the Beijing phylogenetic group, clustered/orphan status, or resistance. The Beijing strain responsible for extensive spread on Gran Canaria Island was also identified in Madrid, but did not lead to secondary cases and did not show high infectivity in the infection model. CONCLUSIONS: The Beijing lineage in our area is a non-homogeneous family, with only certain highly virulent representatives. The specific characterization of Beijing isolates in different settings could help us to accurately identify the virulent representatives before making general assumptions about this lineage.

Everolimus plus exemestane in hormone-receptor-positive, HER2-negative locally advanced or metastatic breast cancer: incidence and time course of adverse events in the phase IIIb BALLET population.

BACKGROUND: The addition of everolimus to exemestane therapy significantly improves progression-free survival in postmenopausal patients with hormone-receptor (HR)-positive HER2-negative endocrine-resistant breast cancer. However, the safety profile of this schedule still might be optimized. METHODS: Patients included in the BALLET trial were assessed. The objectives of this analysis were to provide additional information on the safety profile of this schedule depending on prior anticancer therapies and to characterize the time course of adverse events (AEs) and serious AEs (SAEs) of clinical interest throughout the study period. Non-infectious pneumonitis (NIP), stomatitis, asthenia and weight loss were selected as AEs of clinical interest. RESULTS: The safety population of this analysis comprised 2131 patients. There were similar incidences of AEs and SAEs of clinical interest regardless of previous anticancer therapies. Most stomatitis and asthenia events occurred within the first three months. Incidence of weight loss appeared to plateau except in the case of grade 3-4 events, which occurred rarely. The incidence of any grade NIP (between 2 to 6%) and grade 3-4 NIP (between 0 to 1%) was low across the study, but steady. CONCLUSIONS: Everolimus plus exemestane is a well-known therapeutic option for aromatase inhibitor pretreated advanced breast cancer patients, and its toxicity profile is similar to that described in previous studies. Close monitoring, especially within the first three months, early intervention with preventive measures and patient education to help recognize the first signs and symptoms of AEs, will help to reduce their incidence and severity.

Clinical Consensus Recommendations Regarding Non-Invasive Respiratory Support in the Adult Patient with Acute Respiratory Failure Secondary to SARS-CoV-2 infection. / Recomendaciones de consenso respecto al soporte respiratorio no invasivo en el paciente adulto con insuficiencia respiratoria aguda secundaria a infección por SARS-CoV-2.

Coronavirus disease 2019 (COVID-19) is a respiratory tract infection caused by a newly emergent coronavirus, that was first recognized in Wuhan, China, in December 2019. Currently, the World Health Organization (WHO) has defined the infection as a global pandemic and there is a health and social emergency for the management of this new infection. While most people with COVID-19 develop only mild or uncomplicated illness, approximately 14% develop severe disease that requires hospitalization and oxygen support, and 5% require admission to an intensive care unit. In severe cases, COVID-19 can be complicated by the acute respiratory distress syndrome (ARDS), sepsis and septic shock, and multiorgan failure. This consensus document has been prepared on evidence-informed guidelines developed by a multidisciplinary panel of health care providers from four Spanish scientific societies (Spanish Society of Intensive Care Medicine [SEMICYUC], Spanish Society of Pulmonologists [SEPAR], Spanish Society of Emergency [SEMES], Spanish Society of Anesthesiology, Reanimation, and Pain [SEDAR]) with experience in the clinical management of patients with COVID-19 and other viral infections, including SARS, as well as sepsis and ARDS. The document provides clinical recommendations for the noninvasive respiratory support (noninvasive ventilation, high flow oxygen therapy with nasal cannula) in any patient with suspected or confirmed presentation of COVID-19 with acute respiratory failure. This consensus guidance should serve as a foundation for optimized supportive care to ensure the best possible chance for survival and to allow for reliable comparison of investigational therapeutic interventions as part of randomized controlled trials.

Prospective study on the application of a WALANT circuit for surgery of tunnel carpal syndrome and trigger finger. / Estudio prospectivo sobre la aplicación de un circuito WALANT para la cirugía del síndrome del túnel carpiano y dedo en resorte.

OBJECTIVES: To evaluate benefits for the patient and the economic impact for the implementation of a wide awake local anesthesia no tourniquet (WALANT) hand surgery compared to traditional major outpatient circuit. METHODS: A prospective cohort study was planned comparing 150 cases of ambulatory hand surgery (carpal tunnel and trigger finger) using WALANT technique intervention out from the operating room; with another 150 which underwent intervention, outpatient setting, with preoperative evaluation, sedation and tourniquet, in the operation room. Preoperative, intraoperative and postoperative pain was monitored, as well as the days that required postoperative analgesia.The resources used and costs were evaluated. Satisfaction was evaluated using a specific survey. RESULTS: The pain during the surgery was equivalent for both groups and was significantly lower postoperatively for the WALANT group, with less need for the use of analgesics. Satisfaction was greater for the local anesthesia group. The use of personnel resources and hospital material was less for the WALANT group, with total saving calculated by 1,019€ per patient. CONCLUSIONS: Procedures such as carpal tunnel surgery and trigger finger surgery can be safely performed using wide awake surgery. Patient satisfaction is higher to conventional procedure in the operation room. Pain control is excellent, especially during the postoperative period. WALANT technique for hand surgery represents a benefit for the patient in comfort, timeliness and no need for preoperative tests or evaluation. In addition, it represents a significant savings in hospital resources.

Genetic markers a landscape in prostate cancer.

Prostate cancer (PC) is one of the most common cancers worldwide. The observed variability in progression and responses to the same treatment between patients underlie the genetic heterogeneity of the disease. Nowadays, screening and follow-up biomarkers in PC are still having a deep lack of information, which makes difficult the cancer diagnosis, prognosis and the selection of the most suitable therapies. This is making that currently unnecessary biopsies, over-treatments and hormonoresistances have high rates of prevalence among patients. New biomarkers are urgently needed and in this sense genomic biomarkers could be the most suitable tools. These genetic markers will be helpful for improving the precision of prognostic and the predictive current tools which are employed in the clinical practice. A recent literature search up was conducted, including clinical trials and pre-clinical basic research studies. Keywords included germline variants, prostate cancer, biomarkers, androgen deprivation therapy, screening and liquid biopsy; among others. We have reviewed how germline variants, CNVs and repetitive regions are relevant to prostate carcinogenesis, treatment and progression. Moreover, we have also considered novel biomarkers for PC prognosis based on differentially expressed genes. Finally, we have included new strategies in recent markers of liquid biopsy or updated technologies for minimal samples analysis. The improvement of genetic markers use and their application to the clinical practice, will enhance the variability of simple, non-invasive, tools such as liquid biopsy and germline variants, these will reduce the number of PC needle biopsies and current over-treatments that are usual in the management of this cancer.

Fast update of undetected Mycobacterium chimaera infections to reveal unsuspected cases.

Mycobacterium chimaera is involved in a worldwide alert due to contaminated heater-cooler units. A real-time polymerase chain reaction (RT-PCR)-based procedure was implemented to survey undetected cases of M. chimaera infection. PCR was negative in the 59 prosthetic heart valves from patients with PCR-16SrRNA-negative infective endocarditis. PCR identified M. chimaera in one of 15 clinically significant retrospective Mycobacterium avium-Mycobacterium intracellulare complex isolates, which corresponded to a patient who had undergone heart valve replacement in a different institution. Whole-genome sequencing demonstrated that he was the first case in Spain with involvement of the strain responsible for the global outbreak. These results highlight the relevance of retrospective tracking for undetected M. chimaera infections.

How to: identify non-tuberculous Mycobacterium species using MALDI-TOF mass spectrometry.

BACKGROUND: The implementation of MALDI-TOF MS for microorganism identification has changed the routine of the microbiology laboratories as we knew it. Most microorganisms can now be reliably identified within minutes using this inexpensive, user-friendly methodology. However, its application in the identification of mycobacteria isolates has been hampered by the structure of their cell wall. Improvements in the sample processing method and in the available database have proved key factors for the rapid and reliable identification of non-tuberculous mycobacteria isolates using MALDI-TOF MS. AIMS: The main objective is to provide information about the proceedings for the identification of non-tuberculous isolates using MALDI-TOF MS and to review different sample processing methods, available databases, and the interpretation of the results. SOURCES: Results from relevant studies on the use of the available MALDI-TOF MS instruments, the implementation of innovative sample processing methods, or the implementation of improved databases are discussed. CONTENT: Insight about the methodology required for reliable identification of non-tuberculous mycobacteria and its implementation in the microbiology laboratory routine is provided. IMPLICATIONS: Microbiology laboratories where MALDI-TOF MS is available can benefit from its capacity to identify most clinically interesting non-tuberculous mycobacteria in a rapid, reliable, and inexpensive manner.

Molecular, epidemiological and infectivity characterisation of a Mycobacterium tuberculosis strain prevalent in Madrid.

The most prevalent strain of Mycobacterium tuberculosis in Madrid, Spain (strain 5) was recovered from 45 cases between 1997 and 2004 and showed a highly homogeneous genetic composition. This strain was not exclusive to Spain, and its spoligotyping signature (ST20) was found in entries from different countries in the SITVIT1 database. Patients infected with strain 5 were more frequently positive for human immunodeficiency virus and autochthonous, and had been in prison more frequently, but strain 5 did not show increased infectivity in an in-vitro model of infection.

Biglycan and Decorin Expression and Distribution in Palatal Adhesion.

Previous studies demonstrated that chondroitin sulfate proteoglycans (CSPGs) on apical surfaces of palatal medial edge epithelial (MEE) cells were necessary for palatal adhesion. In this study, we identified 2 proteoglycans, biglycan and decorin, that were expressed in the palatal shelves prior to adhesion. In addition, we established that these proteoglycans were dependent on transforming growth factor ß (TGFß) signaling. Laser capture microdissection was used to collect selected palatal epithelial cells from embryonic mouse embryos at various palate development stages. The expression of specific messenger RNA (mRNA) for biglycan and decorin was determined with quantitative real-time polymerase chain reaction. The TGFßrI kinase inhibitor (SB431542) was used in palatal organ cultures to determine if blocking TFGß signaling changed biglycan and decorin distribution. Immunohistochemistry of both biglycan and decorin revealed expression on the apical and lateral surfaces of MEE cells. Biglycan protein and mRNA levels peaked as the palatal shelves adhered. Decorin was less abundant on the apical epithelial surface and also had reduced mRNA levels compared to biglycan. Their proteins were not expressed on MEE cells of palates treated with SB431542, an inhibitor of TGFß signaling. The temporal expression of biglycan and decorin on the apical surface of MEE, combined with the evidence that these proteins were regulated through the TGFß pathway, indicated that they may be important for adhesion.

Circulating Tumor Cells: Markers and Methodologies for Enrichment and Detection.

Cancer is a leading cause of disease worldwide; however, nowadays many points of its initiation processes are unknown. In this chapter, we are focusing on the role of liquid biopsies in cancer detection and progression. CTCs are one of the main components of liquid biopsies, they represent a subset of tumor cells that have acquired the ability to disseminate from the primary tumor and intravasate to the circulatory system. The greatest challenge in the detection of CTCs is their rarity in the blood. Human blood consists of white blood cells (5-10 × 106/mL), red blood cells (5-9 × 109/mL), and platelets (2.5-4 × 108/mL); very few CTCs will be present even in patients with known metastatic disease, with often less than one CTC per mL of blood. CTCs are found in frequencies on the order of 1-10 CTCs per mL of whole blood in patients with metastatic disease, and it is reduced in half for non-metastatic stages. Therefore, accurate methodologies for their capture and analysis are really important. The main aim of the present chapter is to describe different methodologies for CTCs capturing and analysis.

Diagnostic performance of Anyplex II MTB/MDR/XDR for detection of resistance to first and second line drugs in Mycobacterium tuberculosis.

PURPOSE: Genotypic methods have considerably improved the diagnosis of multidrug-resistant (MDR) tuberculosis. One of these tests is Anyplex II MTB/MDR/XDR (Anyplex). Our aim was to evaluate the diagnostic performance of this multiplex PCR. METHODS: We conducted our study on 47 MDR tuberculosis and 14 pan-susceptible strains. We evaluated the ability of Anyplex to detect resistance mutations in rpoB (rifampin [RIF]), katG and inhA (isoniazid [INH]), gyrA (fluoroquinolones [FLQ]), and rrs and eis (aminoglycosides [AMG]). We used the agar proportion method as gold standard. We also studied concordance with GenoType MTBDRplus (first line drugs) and MTBDRsl (second line drugs). DNA sequencing was applied to clarify discrepancies. RESULTS: All pan-susceptible strains were susceptible by Anyplex. Sensitivity and specificity of Anyplex for detection of resistance mutations were 97.9% and 100%, respectively, for RIF, 91.5% and 100% for INH, 80% and 100% for FLQ, and 50% and 99.7% for AMG. Concordance with GenoType was perfect for RIF, INH, and FLQ (kappa score, k=1.0) and moderate for AMG (k=0.48). Sensitivity and specificity for detection of MDR tuberculosis were 89.4% and 100%, respectively. DNA sequencing of the phenotypically resistant strains considered as susceptible by Anyplex, confirmed no mutations in the corresponding genes. CONCLUSIONS: Anyplex is a reliable assay for the detection of MDR tuberculosis and shows excellent concordance with GenoType. Anyplex reduces the time to diagnosis of MDR tuberculosis strains, as it is recommended by current guidelines on control of tuberculosis.

A novel strategy based on genomics and specific PCR reveals how a multidrug-resistant Mycobacterium tuberculosis strain became prevalent in Equatorial Guinea 15 years after its emergence.

OBJECTIVE: Molecular epidemiology techniques in tuberculosis (TB) can identify high-risk strains that are actively transmitted. We aimed to implement a novel strategy to optimize the identification and control of multidrug-resistant (MDR) TB in a specific population. METHODS: We developed a strain-specific PCR tailored from whole genome sequencing (WGS) data to track a specific MDR prevalent strain in Equatorial Guinea (EG-MDR). RESULTS: The PCR was applied prospectively on remnants of GeneXpert reaction mixtures owing to the lack of culture facilities in Equatorial Guinea. In 147 (93%) of 158 cases, we were able to differentiate between infection by the EG-MDR strain or by any other strain and found that 44% of all rifampicin-resistant TB cases were infected by EG-MDR. We also analysed 93 isolates obtained from Equatorial Guinea 15 years ago, before MDR-TB had become the problem it is today. We found that two of the scarce historical MDR cases were infected by EG-MDR. WGS revealed low variability-six single nucleotide polymorphisms acquired by this strain over 15 years-likely because of the lack in the country of a specific program to treat MDR-TB. CONCLUSIONS: Our novel strategy, which integrated WGS analysis and strain-specific PCRs, represents a low-cost, rapid and transferable strategy that allowed a prospective efficient survey and fast historical analysis of MDR-TB in a population.