Rapid tests for the detection of the Mycobacterium abscessus subsp. bolletii strain responsible for an epidemic of surgical-site infections in Brazil

A single strain of Mycobacterium abscessus subsp. bolletii, characterised by a particular rpoB sequevar and two highly related pulsed field gel electrophoresis patterns has been responsible for a nationwide outbreak of surgical infections in Brazil since 2004. In this study, we developed molecular tests based on polymerase chain reaction restriction-enzyme analysis (PRA) and sequencing for the rapid identification of this strain. Sequences of 15 DNA regions conserved in mycobacteria were retrieved from GenBank or sequenced and analysed in silico. Single nucleotide polymorphisms specific to the epidemic strain and located in enzyme recognition sites were detected in rpoB, the 3' region of the 16S rDNA and gyrB. The three tests that were developed, i.e., PRA-rpoB, PRA-16S and gyrB sequence analysis, showed 100%, 100% and 92.31% sensitivity and 93.06%, 90.28% and 100% specificity, respectively, for the discrimination of the surgical strain from other M. abscessus subsp. bolletii isolates, including 116 isolates from 95 patients, one environmental isolate and two type strains. The results of the three tests were stable, as shown by results obtained for different isolates from the same patient. In conclusion, due to the clinical and epidemiological importance of this strain, these tests could be implemented in reference laboratories for the rapid preliminary diagnosis and epidemiological surveillance of this epidemic strain.

Comparison of two rapid tests for anti-phenolic glycolipid-I serology in Brazil and Nepal

The diagnosis of leprosy continues to be based on clinical symptoms and early diagnosis and treatment are critical to preventing disability and transmission. Sensitive and specific laboratory tests are not available for diagnosing leprosy. Despite the limited applicability of anti-phenolic glycolipid-I (PGL-I) serology for diagnosis, it has been suggested as an additional tool to classify leprosy patients (LPs) for treatment purposes. Two formats of rapid tests to detect anti-PGL-I antibodies [ML immunochromatography assay (ICA) and ML Flow] were compared in different groups, multibacillary patients, paucibacillary patients, household contacts and healthy controls in Brazil and Nepal. High ML Flow intra-test concordance was observed and low to moderate agreement between the results of ML ICA and ML Flow tests on the serum of LPs was observed. LPs were "seroclassified" according to the results of these tests and the seroclassification was compared to other currently used classification systems: the World Health Organization operational classification, the bacilloscopic index and the Ridley-Jopling classification. When analysing the usefulness of these tests in the operational classification of PB and MB leprosy for treatment and follow-up purposes, the ML Flow test was the best point-of-care test for subjects in Nepal and despite the need for sample dilution, the ML ICA test yielded better performance among Brazilian subjects. Our results identified possible ways to improve the performance of both tests.

Risk factors for positive tuberculin skin tests among migrant and resident children in Lausanne, Switzerland

Résumé en français Cadre : Policlinique pédiatrique à Lausanne en Suisse, pays rencontrant une proportion importante de tuberculose au sein de la population de migrants. But : Déterminer les facteurs de risque associés à un test tuberculinique positif (ou test de Mantoux), notamment l'influence du BCG (Bacille Calmette Guérin) et d'un contact avec un personne ayant une tuberculose active. Les patients concernés étaient des enfants examinés dans le cadre d'un contrôle de santé ou dans le cadre d'une étude d'entourage d'un cas déclaré de tuberculose. Méthode : Etude descriptive comprenant des enfants ayant eu un test tuberculinique (2 unités RT23) entre novembre 2002 et avril 2004. L'âge, le sexe, l'anamnèse de contact avec une personne ayant une tuberculose active, la vaccination par le BCG, le pays d'origine et le lieu de naissance (en Suisse ou hors de la Suisse) étaient répertoriés. Résultats : Parmi les 234 enfants de l'étude, 176 (75%) avaient une réaction tuberculinique égal à zéro et 31 (13%) avaient une réaction positive (> 10mm). Dans le modèle de régression linéaire, la taille de la réaction tuberculinique variait significativement selon l'anamnèse de contact avec une personne ayant une tuberculose active, l'âge, l'incidence de la tuberculose dans le pays d'origine et la vaccination par le BCG. Le sexe ou le lieu de naissance n'influençait pas la taille de la réaction. Dans le modèle de régression logistique incluant toutes les valeurs répertoriées, les paramètres significativement associés avec un Mantoux positif étaient l'âge (Odds Ratio = 1.21, 95% CI 1.08 ; 1.35), l'anamnèse de contact avec une personne ayant une tuberculose active (OR = 7.31, 95% CI 2.23 ; 24) et l'incidence de la tuberculose dans le pays d'origine (OR = 1.01, 95% CI 1.00 ; 1.02). Le sexe (OR = 1.18, 95% CI 0.50 ; 2.78) et la vaçcination par le BCG (OR = 2.97, 95% CI 0.91 ; 9.72) n'étaient pas associés avec une réaction tuberculinique positive. Conclusions : L'incidence de la tuberculose dans le pays d'origine, la vaccination par le BCG et l'âge influencent le test de Mantoux (taille ou proportion de réaction > 10mm). Toutefois, le facteur de risque le plus important d'avoir une réaction tuberculinique positive est l'anamnèse de contact avec. une personne ayant une tuberculose active.

Prozone effects in microscopic agglutination tests for leptospirosis in the sera of mice infected with the pathogenic Leptospira interrogans serovar Canicola

Mice experimentally infected with a pathogenic strain of Leptospira interrogans serovar Canicola produced false negative results (prozone effect) in a microscopic agglutination test (MAT). This prozone effect occurred in several serum samples collected at different post-infection times, but it was more prominent in samples collected from seven-42 days post-infection and for 1:50 and 1:100 sample dilutions. This phenomenon was correlated with increased antibody titres in the early post-infection phase. While prozone effects are often observed in serological agglutination assays for the diagnosis of animal brucellosis and human syphilis, they are not widely reported in leptospirosis MATs.

Bilateral sequential nonarteritic anterior ischemic optic neuropathy: a comparison of visual outcomes in fellow eyes using quantitative analysis of goldmann visual fields.

OBJECTIVE To better define the concordance of visual loss in patients with nonarteritic anterior ischemic optic neuropathy (NAION). METHODS The medical records of 86 patients with bilateral sequential NAION were reviewed retrospectively, and visual function was assessed using visual acuity, Goldmann visual fields, color vision, and relative afferent papillary defect. A quantitative total visual field score and score per quadrant were analyzed for each eye using the numerical Goldmann visual field scoring method. RESULTS Outcome measures were visual acuity, visual field, color vision, and relative afferent papillary defect. A statistically significant correlation was found between fellow eyes for multiple parameters, including logMAR visual acuity (P = .01), global visual field (P < .001), superior visual field (P < .001), and inferior visual field (P < .001). The mean deviation of total (P < .001) and pattern (P < .001) deviation analyses was significantly less between fellow eyes than between first and second eyes of different patients. CONCLUSIONS Visual function between fellow eyes showed a fair to moderate correlation that was statistically significant. The pattern of vision loss was also more similar in fellow eyes than between eyes of different patients. These results may help allow better prediction of visual outcome for the second eye in patients with NAION.

Rivaroxaban: Quantification by anti-FXa assay and influence on coagulation tests: a study in 9 Swiss laboratories.

INTRODUCTION: Rivaroxaban (RXA) is licensed for prophylaxis of venous thromboembolism after major orthopaedic surgery of the lower limbs. Currently, no test to quantify RXA in plasma has been validated in an inter-laboratory setting. Our study had three aims: to assess i) the feasibility of RXA quantification with a commercial anti-FXa assay, ii) its accuracy and precision in an inter-laboratory setting, and iii) the influence of 10mg of RXA on routine coagulation tests. METHODS: The same chromogenic anti-FXa assay (Hyphen BioMed) was used in all participating laboratories. RXA calibrators and sets of blinded probes (aim ii.) were prepared in vitro by spiking normal plasma. The precise RXA content was assessed by high-pressure liquid chromatography-tandem mass spectrometry. For ex-vivo studies (aim iii), plasma samples from 20 healthy volunteers taken before and 2 - 3hours after ingestion of 10mg of RXA were analyzed by participating laboratories. RESULTS: RXA can be assayed chromogenically. Among the participating laboratories, the mean accuracy and the mean coefficient of variation for precision of RXA quantification were 7.0% and 8.8%, respectively. Mean RXA concentration was 114±43μg/L .RXA significantly altered prothrombin time, activated partial thromboplastin time, factor analysis for intrinsic and extrinsic factors. Determinations of thrombin time, fibrinogen, FXIII and D-Dimer levels were not affected. CONCLUSIONS: RXA plasma levels can be quantified accurately and precisely by a chromogenic anti-FXa assay on different coagulometers in different laboratories. Ingestion of 10mg RXA results in significant alterations of both PT- and aPTT-based coagulation assays.

Combination of trastuzumab and letrozole after resistance to sequential trastuzumab and aromatase inhibitor monotherapies in patients with estrogen receptor-positive, HER-2-positive advanced breast cancer: a proof-of-concept trial (SAKK 23/03).

A sequential treatment design was chosen in this trial to ensure complete resistance to single-agent non-steroidal aromatase inhibitor (AI) and trastuzumab both given as monotherapy before receiving the combination of a non-steroidal AI and trastuzumab. Key eligibility criteria included postmenopausal patients with advanced, measurable, human epidermal growth factor receptor-2 (HER-2)-positive disease (assessed by FISH, ratio (≥2)), hormone receptor (HR)-positive disease, and progression on prior treatment with a non-steroidal AI, e.g. letrozole or anastrozole, either in the adjuvant or in the advanced setting. Patients received standard dose trastuzumab monotherapy in step 1 and upon disease progression continued trastuzumab in combination with letrozole in step 2. The primary endpoint was clinical benefit rate (CBR) in step 2. Totally, 13 patients were enrolled. In step 1, six patients (46%) achieved CBR. Median time to progression (TTP) was 161 days (95% confidence interval (CI): 82-281). In step 2, CBR was observed in eight out of the 11 evaluable patients (73%), including one patient with partial response. Median TTP for all the 11 patients was 188 days (95% CI: 77-not reached). Results of this proof-of-concept trial suggest that complete resistance to both AI and trastuzumab can be overcome in a proportion of patients by combined treatment of AI and trastuzumab, as all patients served as their own control. Our results appear promising for a new treatment strategy that offers a chemotherapy-free option for at least a subset of patients with HR-positive, HER-2-positive breast cancer over a clinically relevant time period.

The use of rapid dengue diagnostic tests in a routine clinical setting in a dengue-endemic area of Colombia

There is insufficient evidence of the usefulness of dengue diagnostic tests under routine conditions. We sought to analyse how physicians are using dengue diagnostics to inform research and development. Subjects attending 14 health institutions in an endemic area of Colombia with either a clinical diagnosis of dengue or for whom a dengue test was ordered were included in the study. Patterns of test-use are described herein. Factors associated with the ordering of dengue diagnostic tests were identified using contingency tables, nonparametric tests and logistic regression. A total of 778 subjects were diagnosed with dengue by the treating physician, of whom 386 (49.5%) were tested for dengue. Another 491 dengue tests were ordered in subjects whose primary diagnosis was not dengue. Severe dengue classification [odds ratio (OR) 2.2; 95% confidence interval (CI) 1.1-4.5], emergency consultation (OR 1.9; 95% CI 1.4-2.5) and month of the year (OR 3.1; 95% CI 1.7-5.5) were independently associated with ordering of dengue tests. Dengue tests were used both to rule in and rule out diagnosis. The latter use is not justified by the sensitivity of current rapid dengue diagnostic tests. Ordering of dengue tests appear to depend on a combination of factors, including physician and institutional preferences, as well as other patient and epidemiological factors.

Molecular testing for fragile X: analysis of 5062 tests from 1105 fragile X families--performed in 12 clinical laboratories in Spain.

Fragile X syndrome is the most common inherited form of intellectual disability. Here we report on a study based on a collaborative registry, involving 12 Spanish centres, of molecular diagnostic tests in 1105 fragile X families comprising 5062 individuals, of whom, 1655 carried a full mutation or were mosaic, three cases had deletions, 1840 had a premutation, and 102 had intermediate alleles. Two patients with the full mutation also had Klinefelter syndrome. We have used this registry to assess the risk of expansion from parents to children. From mothers with premutation, the overall rate of allele expansion to full mutation is 52.5%, and we found that this rate is higher for male than female offspring (63.6% versus 45.6%; P < 0.001). Furthermore, in mothers with intermediate alleles (45-54 repeats), there were 10 cases of expansion to a premutation allele, and for the smallest premutation alleles (55-59 repeats), there was a 6.4% risk of expansion to a full mutation, with 56 repeats being the smallest allele that expanded to a full mutation allele in a single meiosis. Hence, in our series the risk for alleles of <59 repeats is somewhat higher than in other published series. These findings are important for genetic counselling.

An in-house real-time polymerase chain reaction: standardisation and comparison with the Cobas Amplicor HBV monitor and Cobas AmpliPrep/Cobas TaqMan HBV tests for the quantification of hepatitis B virus DNA

This study aimed to standardise an in-house real-time polymerase chain reaction (rtPCR) to allow quantification of hepatitis B virus (HBV) DNA in serum or plasma samples, and to compare this method with two commercial assays, the Cobas Amplicor HBV monitor and the Cobas AmpliPrep/Cobas TaqMan HBV test. Samples from 397 patients from the state of São Paulo were analysed by all three methods. Fifty-two samples were from patients who were human immunodeficiency virus and hepatitis C virus positive, but HBV negative. Genotypes were characterised, and the viral load was measure in each sample. The in-house rtPCR showed an excellent success rate compared with commercial tests; inter-assay and intra-assay coefficients correlated with commercial tests (r = 0.96 and r = 0.913, p < 0.001) and the in-house test showed no genotype-dependent differences in detection and quantification rates. The in-house assay tested in this study could be used for screening and quantifying HBV DNA in order to monitor patients during therapy.

Prediction and discrimination of osteoporotic hip fracture in postmenopausal women.

Osteoporotic hip fractures increase dramatically with age and are responsible for considerable morbidity and mortality. Several treatments to prevent the occurrence of hip fracture have been validated in large randomized trials and the current challenge is to improve the identification of individuals at high risk of fracture who would benefit from therapeutic or preventive intervention. We have performed an exhaustive literature review on hip fracture predictors, focusing primarily on clinical risk factors, dual X-ray absorptiometry (DXA), quantitative ultrasound, and bone markers. This review is based on original articles and meta-analyses. We have selected studies that aim both to predict the risk of hip fracture and to discriminate individuals with or without fracture. We have included only postmenopausal women in our review. For studies involving both men and women, only results concerning women have been considered. Regarding clinical factors, only prospective studies have been taken into account. Predictive factors have been used as stand-alone tools to predict hip fracture or sequentially through successive selection processes or by combination into risk scores. There is still much debate as to whether or not the combination of these various parameters, as risk scores or as sequential or concurrent combinations, could help to better predict hip fracture. There are conflicting results on whether or not such combinations provide improvement over each method alone. Sequential combination of bone mineral density and ultrasound parameters might be cost-effective compared with DXA alone, because of fewer bone mineral density measurements. However, use of multiple techniques may increase costs. One problem that precludes comparison of most published studies is that they use either relative risk, or absolute risk, or sensitivity and specificity. The absolute risk of individuals given their risk factors and bone assessment results would be a more appropriate model for decision-making than relative risk. Currently, a group appointed by the World Health Organization and lead by Professor John Kanis is working on such a model. It will therefore be possible to further assess the best choice of threshold to optimize the number of women needed to screen for each country and each treatment.

Safety of falciparum malaria diagnostic strategy based on rapid diagnostic tests in returning travellers and migrants: a retrospective study.

BACKGROUND: Rapid diagnostic tests for malaria (RDTs) allow accurate diagnosis and prompt treatment. Validation of their usefulness in travellers with fever was needed. The safety of a strategy to diagnose falciparum malaria based on RDT followed by immediate or delayed microscopy reading at first attendance was evaluated in one referral hospital in Switzerland. METHODS: A retrospective study was conducted in the outpatient clinic and emergency ward of University Hospital, covering a period of eight years (1999-2007). The study was conducted in the outpatient clinic and emergency ward of University Hospital. All adults suspected of malaria with a diagnostic test performed were included. RDT and microscopy as immediate tests were performed during working hours, and RDT as immediate test and delayed microscopy reading out of laboratory working hours. The main outcome measure was occurrence of specific complications in RDT negative and RDT positive adults. RESULTS: 2,139 patients were recruited. 1987 had both initial RDT and blood smear (BS) result negative. Among those, 2/1987 (0.1%) developed uncomplicated malaria with both RDT and BS positive on day 1 and day 6 respectively. Among the 152 patients initially malaria positive, 137 had both RDT and BS positive, four only BS positive and five only RDT positive (PCR confirmed) (six had only one test performed). None of the four initially RDT negative/BS positive and none of the five initially BS negative/RDT positive developed severe malaria while 6/137 of both RDT and BS positive did so. The use of RDT allowed a reduction of a median of 2.1 hours to get a first malaria test result. CONCLUSIONS: A malaria diagnostic strategy based on RDTs and a delayed BS is safe in non-immune populations, and shortens the time to first malaria test result.