Untargeted profiling of urinary steroid metabolites after testosterone ingestion: opening new perspectives for antidoping testing.

AIM: Antidoping procedures are expected to greatly benefit from untargeted metabolomic approaches through the discovery of new biomarkers of prohibited substances abuse. RESULTS: Endogenous steroid metabolites were monitored in urine samples from a controlled elimination study of testosterone undecanoate after ingestion. A platform coupling ultra-high pressure LC with high-resolution quadrupole TOF MS was used and high between-subject metabolic variability was successfully handled using a multiblock data analysis strategy. Links between specific subsets of metabolites and influential genetic polymorphisms of the UGT2B17 enzyme were highlighted. CONCLUSION: This exploratory metabolomic strategy constitutes a first step toward a better understanding of the underlying patterns driving the high interindividual variability of steroid metabolism. Promising biomarkers were selected for further targeted study.

Sequence-Based Hepatitis B Virus Antiviral Resistance Testing in Switzerland

BACKGROUND: A growing number of patients with chronic hepatitis B is being treated for extended periods with nucleoside and/or nucleotide analogs. In this context, antiviral resistance represents an increasingly common and complex issue. METHODS: Mutations in the hepatitis B virus (HBV) reverse transcriptase (rt) gene and viral genotypes were determined by direct sequencing of PCR products and alignment with reference sequences deposited in GenBank. RESULTS: Plasma samples from 60 patients with chronic hepatitis B were analyzed since March 2009. The predominant mutation pattern identified in patients with virological breakthrough was rtM204V/I ± different compensatory mutations, conferring resistance to L-nucleosides (lamivudine, telbivudine, emtricitabine) and predisposing to entecavir resistance (n = 18). Complex mutation patterns with a potential for multidrug resistance were identified in 2 patients. Selection of a fully entecavir resistant strain was observed in a patient exposed to lamivudine alone. Novel mutations were identified in 1 patient. Wild-type HBV was identified in 9 patients with suspected virological breakthrough, raising concerns about treatment adherence. No preexisting resistance mutations were identified in treatment-naïve patients (n = 13). Viral genome amplification and sequencing failed in 16 patients, of which only 2 had a documented HBV DNA > 1000 IU/ml. HBV genotypes were D in 28, A in 6, B in 4, C in 3 and E in 3 patients. Results will be updated in August 2010 and therapeutic implications discussed. CONCLUSIONS: With expanding treatment options and a growing number of patients exposed to nucleoside and/or nucleotide analogs, sequence-based HBV antiviral resistance testing is expected to become a cornerstone in the management of chronic hepatitis B.

Evaluation of indirect susceptibility testing of Mycobacterium tuberculosis to the first- and second-line, and alternative drugs by the newer MB/BacT system

In order to evaluate the Organon Teknika MB/BacT system used for testing indirect susceptibility to the alternative drugs ofloxacin (OFLO), amikacin (AMI), and rifabutin (RIF), and to the usual drugs of standard treatment regimes such as rifampin (RMP), isoniazid (INH), pyrazinamide (PZA), streptomycin (SM), ethambutol (EMB), and ethionamide (ETH), cultures of clinical specimens from 117 patients with pulmonary tuberculosis under multidrug-resistant investigation, admitted sequentially for examination from 2001 to 2002, were studied. Fifty of the Mycobacterium tuberculosis cultures were inoculated into the gold-standard BACTEC 460 TB (Becton Dickinson) for studying resistance to AMI, RIF, and OFLO, and the remaining 67 were inoculated into Lowenstein Jensen (LJ) medium (the gold standard currently used in Brazil) for studying resistance to RMP, INH, PZA, SM, EMB, and ETH. We observed 100% sensitivity for AMI (80.8-100), RIF (80.8-100), and OFLO (78.1-100); and 100% specificity for AMI (85.4-100), RIF (85.4-100), and OFLO (86.7-100) compared to the BACTEC system. Comparing the results obtained in LJ we observed 100% sensitivity for RMP (80-100), followed by INH - 95% (81.8-99.1), EMB - 94.7% (71.9-99.7), and 100% specificity for all drugs tested except for PZA - 98.3 (89.5-99.9) at 95% confidence interval. The results showed a high level of accuracy and demonstrated that the fully automated, non-radiometric MB/BacT system is indicated for routine use in susceptibility testing in public health laboratories.