Role of the first WHO mutation catalogue in the diagnosis of antibiotic resistance in Mycobacterium tuberculosis in the Valencia Region, Spain: a retrospective genomic analysis.

BACKGROUND: In June, 2021, WHO published the most complete catalogue to date of resistance-conferring mutations in Mycobacterium tuberculosis. Here, we aimed to assess the performance of genome-based antimicrobial resistance prediction using the catalogue and its potential for improving diagnostics in a real low-burden setting. METHODS: In this retrospective population-based genomic study M tuberculosis isolates were collected from 25 clinical laboratories in the low-burden setting of the Valencia Region, Spain. Culture-positive tuberculosis cases reported by regional public health authorities between Jan 1, 2014, and Dec 31, 2016, were included. The drug resistance profiles of these isolates were predicted by the genomic identification, via whole-genome sequencing (WGS), of the high-confidence resistance-causing variants included in the catalogue and compared with the phenotype. We determined the minimum inhibitory concentration (MIC) of the isolates with discordant resistance profiles using the resazurin microtitre assay. FINDINGS: WGS was performed on 785 M tuberculosis complex culture-positive isolates, and the WGS resistance prediction sensitivities were: 85·4% (95% CI 70·8-94·4) for isoniazid, 73·3% (44·9-92·2) for rifampicin, 50·0% (21·1-78·9) for ethambutol, and 57·1% (34·0-78·2) for pyrazinamide; all specificities were more than 99·6%. Sensitivity values were lower than previously reported, but the overall pan-susceptibility accuracy was 96·4%. Genotypic analysis revealed that four phenotypically susceptible isolates carried mutations (rpoB Leu430Pro and rpoB Ile491Phe for rifampicin and fabG1 Leu203Leu for isoniazid) known to give borderline resistance in standard phenotypic tests. Additionally, we identified three putative resistance-associated mutations (inhA Ser94Ala, katG Leu48Pro, and katG Gly273Arg for isoniazid) in samples with substantially higher MICs than those of susceptible isolates. Combining both genomic and phenotypic data, in accordance with the WHO diagnostic guidelines, we could detect two new multidrug-resistant cases. Additionally, we detected 11 (1·6%) of 706 isolates to be monoresistant to fluoroquinolone, which had been previously undetected. INTERPRETATION: We showed that the WHO catalogue enables the detection of resistant cases missed in phenotypic testing in a low-burden region, thus allowing for better patient-tailored treatment. We also identified mutations not included in the catalogue, relevant at the local level. Evidence from this study, together with future updates of the catalogue, will probably lead in the future to the partial replacement of culture testing with WGS-based drug susceptibility testing in our setting. FUNDING: European Research Council and the Spanish Ministerio de Ciencia.

Population-based sequencing of Mycobacterium tuberculosis reveals how current population dynamics are shaped by past epidemics.

Transmission is a driver of tuberculosis (TB) epidemics in high-burden regions, with assumed negligible impact in low-burden areas. However, we still lack a full characterization of transmission dynamics in settings with similar and different burdens. Genomic epidemiology can greatly help to quantify transmission, but the lack of whole genome sequencing population-based studies has hampered its application. Here, we generate a population-based dataset from Valencia region and compare it with available datasets from different TB-burden settings to reveal transmission dynamics heterogeneity and its public health implications. We sequenced the whole genome of 785 Mycobacterium tuberculosis strains and linked genomes to patient epidemiological data. We use a pairwise distance clustering approach and phylodynamic methods to characterize transmission events over the last 150 years, in different TB-burden regions. Our results underscore significant differences in transmission between low-burden TB settings, i.e., clustering in Valencia region is higher (47.4%) than in Oxfordshire (27%), and similar to a high-burden area as Malawi (49.8%). By modeling times of the transmission links, we observed that settings with high transmission rate are associated with decades of uninterrupted transmission, irrespective of burden. Together, our results reveal that burden and transmission are not necessarily linked due to the role of past epidemics in the ongoing TB incidence, and highlight the need for in-depth characterization of transmission dynamics and specifically tailored TB control strategies.

High-resolution mapping of tuberculosis transmission: Whole genome sequencing and phylogenetic modelling of a cohort from Valencia Region, Spain.

BACKGROUND: Whole genome sequencing provides better delineation of transmission clusters in Mycobacterium tuberculosis than traditional methods. However, its ability to reveal individual transmission links within clusters is limited. Here, we used a 2-step approach based on Bayesian transmission reconstruction to (1) identify likely index and missing cases, (2) determine risk factors associated with transmitters, and (3) estimate when transmission happened. METHODS AND FINDINGS: We developed our transmission reconstruction method using genomic and epidemiological data from a population-based study from Valencia Region, Spain. Tuberculosis (TB) incidence during the study period was 8.4 cases per 100,000 people. While the study is ongoing, the sampling frame for this work includes notified TB cases between 1 January 2014 and 31 December 2016. We identified a total of 21 transmission clusters that fulfilled the criteria for analysis. These contained a total of 117 individuals diagnosed with active TB (109 with epidemiological data). Demographic characteristics of the study population were as follows: 80/109 (73%) individuals were Spanish-born, 76/109 (70%) individuals were men, and the mean age was 42.51 years (SD 18.46). We found that 66/109 (61%) TB patients were sputum positive at diagnosis, and 10/109 (9%) were HIV positive. We used the data to reveal individual transmission links, and to identify index cases, missing cases, likely transmitters, and associated transmission risk factors. Our Bayesian inference approach suggests that at least 60% of index cases are likely misidentified by local public health. Our data also suggest that factors associated with likely transmitters are different to those of simply being in a transmission cluster, highlighting the importance of differentiating between these 2 phenomena. Our data suggest that type 2 diabetes mellitus is a risk factor associated with being a transmitter (odds ratio 0.19 [95% CI 0.02-1.10], p < 0.003). Finally, we used the most likely timing for transmission events to study when TB transmission occurred; we identified that 5/14 (35.7%) cases likely transmitted TB well before symptom onset, and these were largely sputum negative at diagnosis. Limited within-cluster diversity does not allow us to extrapolate our findings to the whole TB population in Valencia Region. CONCLUSIONS: In this study, we found that index cases are often misidentified, with downstream consequences for epidemiological investigations because likely transmitters can be missed. Our findings regarding inferred transmission timing suggest that TB transmission can occur before patient symptom onset, suggesting also that TB transmits during sub-clinical disease. This result has direct implications for diagnosing TB and reducing transmission. Overall, we show that a transition to individual-based genomic epidemiology will likely close some of the knowledge gaps in TB transmission and may redirect efforts towards cost-effective contact investigations for improved TB control.

Black tongue associated with linezolid.

Darkening of the tongue and oral mucosa is a reaction pattern that can be related to a number of physiologic, metabolic, and toxic disorders, and medications and exogenous substances. Black discoloration of the tongue should be distinguished from black "hairy" tongue, which is characterized by hypertrophy of the filiform papillae. We report a case of a 42-year-old man presented with a black discoloration of his tongue during treatment with linezolid for spondylodiscitis. So in conclusion, tongue discoloration is a benign and reversible condition and a probable adverse event associated with linezolid. We present this case to increase clinicians' awareness of a new potential adverse effect of linezolid.

[Multiple inflammatory liver pseudotumors and Enterococcus durans bacteremia]. / Seudotumor inflamatorio hepático múltiple y bacteriemia por Enterococcus durans.

Inflammatory pseudotumors are rare benign lesions that can occur throughout the body. These masses are usually associated with fever, pain, and mass effect, and are frequently mistaken for malignant neoplasms. Liver pseudotumors are especially rare, with 150 cases reported up to 2007, since the original report of Pack and Baker in 1953. We describe the case of a patient with suspected multiple hepatic abscesses, who was finally diagnosed with an inflammatory pseudotumor by percutaneous biopsy. The patient received prolonged antibiotic therapy, with complete resolution of the pseudotumor. The differential diagnosis and management of this entity is discussed.

Seudotumor inflamatorio hepático múltiple y bacteriemia por Enterococcus durans / Multiple inflammatory liver pseudotumors and Enterococcus durans bacteremia

Los seudotumores inflamatorios son lesiones raras, benignas y con diferentes localizaciones. Habitualmente asocian fiebre, dolor y efecto masa, y con frecuencia pueden diagnosticarse erróneamente como neoplasias. Los seudotumores hepáticos son especialmente infrecuentes, con menos de 150 casos descritos hasta 2007 tras la primera descripción de Pack y Baker en 1953. Describimos el caso de un paciente con sospecha de abscesos hepáticos múltiples, diagnosticado finalmente de seudotumor inflamatorio hepático mediante biopsia de la lesión. Recibió tratamiento antibiótico prolongado hasta la resolución del cuadro. Se discutirá el manejo clínico y el diagnóstico diferencial(AU)

Inflammatory pseudotumors are rare benign lesions that can occur throughout the body. These masses are usually associated with fever, pain, and mass effect, and are frequently mistaken for malignant neoplasms. Liver pseudotumors are especially rare, with 150 cases reported up to 2007, since the original report of Pack and Baker in 1953. We describe the case of a patient with suspected multiple hepatic abscesses, who was finally diagnosed with an inflammatory pseudotumor by percutaneous biopsy. The patient received prolonged antibiotic therapy, with complete resolution of the pseudotumor. The differential diagnosis and management of this entity is discussed(AU)

[In vitro activity of caspofungin against fluconazole-resistant Candida isolates from patients with HIV infection]. / Actividad in vitro de caspofungina frente a aislados clínicos de Candida resistentes a fluconazol en pacientes con infección por el VIH.

INTRODUCTION: Antifungal therapy for mucosal candidiasis caused by fluconazole-resistant Candida species is problematic. The aim of this study was to investigate the in vitro activity of caspofungin against Candida strains with reduced susceptibility to fluconazole isolated from HIV-infected patients. METHODS: The in vitro activity of caspofungin was assessed in 28 fluconazole-resistant Candida isolates obtained from the oral cavity of a cohort of 174 consecutive HIV-infected patients. Minimum inhibitory concentrations (MICs) were determined by a standardized broth microdilution method, as recommended by the NCCLS. RESULTS: Overall, caspofungin MICs ranged from < or = 0.06 microg/ml to 1 microg/ml. MICs at which 50% (MIC50) and 90% (MIC90) of isolates were inhibited were 0.25 microg/ml and 0.5 microg/ml, respectively. MICs ranged from < or = 0.06 microg/ml to 0.5 microg/ml for Candida albicans (n = 11), and < 0.06 microg/ml to 1 microg/ml or Candida glabrata (n = 11). MICs for the two strains of Candida krusei were 0.125 microg/ml and 1 microg/ml. The range of MICs for Candida tropicalis and Candida inconspicua strains was 0.25 microg/ml to 0.5 microg/ml. CONCLUSION: Caspofungin was very active in vitro against a variety of fluconazole-resistant Candida strains recovered from a clinical cohort of HIV-infected patients. The MIC50 values and MIC ranges were slightly higher for Candida glabrata than for Candida albicans.