Canadian multicenter laboratory study for standardized second-line antimicrobial susceptibility testing of Mycobacterium tuberculosis

The purpose of this study was to establish a standardized protocol for second-line antimicrobial susceptibility testing of Mycobacterium tuberculosis using the Bactec MGIT 960 system in Canadian laboratories. Four Canadian public health laboratories compared the susceptibility testing results of 9 second-line antimicrobials between the Bactec 460 and Bactec MGIT 960 systems. Based on the data generated, we have established that the Bactec MGIT 960 system provides results comparable to those obtained with the previous Bactec 460 method. The critical concentrations established for the testing of the antimicrobials used are as follows: amikacin, 1 μg/ml; capreomycin, 2.5 μg/ml; ethionamide, 5 μg/ml; kanamycin, 2.5 μg/ml; linezolid, 1 μg/ml; moxifloxacin, 0.25 μg/ml; ofloxacin, 2 μg/ml; p-aminosalicylic acid, 4 μg/ml; rifabutin, 0.5 μg/ml.     

An improved PCR method for detection of HIV-1 proviral DNA of a wide range of subtypes and recombinant forms circulating globally

Proviral DNAs are being measured increasingly as a marker of the efficacy of highly active anti-retroviral therapy (HAART) and is accepted for the early diagnosis of perinatal HIV-1 infections. This requires a standardized test which enables the detection of a wide range of subtypes worldwide including O, N and circulating recombinant forms (CRFs). Based on a previous publication, a PCR - Test for HIV-1 provirus detection in peripheral blood mononuclear cells (PBMCs) was developed. Blood samples from 80 individuals infected with HIV-1 and 20 persons negative for HIV-1&2 from Africa and Germany were tested for the presence of HIV-1 provirus DNA. The primer system used enables the detection of proviral DNA despite the high concentrations of human DNA. The limit of detection was determined to be 5 copies per 10(5) cells. All 20 samples from persons negative for HIV were negative for HIV-1 proviral DNA while provirus DNA was amplified from 76 of the 80 (95%) samples from persons infected with HIV. The amplified products were detected by gel-electrophoresis, flow cytometry and real-time PCR. All three detection systems provided the same results.     

The L76V mutation in HIV-1 protease is potentially associated with hypersusceptibility to protease inhibitors Atazanavir and Saquinavir: is there a clinical advantage?

Background

We aimed to determine the reliability and validity of a hepatitis symptom inventory and to identify predictors of hepatitis C (HCV) treatment initiation in a cohort of HIV-infected patients.

Methods

Prospective clinic based study that enrolled patients referred for HCV therapy consideration. A hepatitis symptom inventory and the Center for Epidemiologic Studies Depression Scale (CES-D) were administered to HIV/HCV individuals. The symptom inventory was factor analyzed and subscale reliability estimated with Cronbach's alpha. Predictive validity was evaluated using generalized estimating equations (GEE). Predictors of HCV treatment were identified using logistic regression.

Results

Between April 2008 to July 2010, 126 HIV/HCV co-infected patients were enrolled in the study. Factor analysis using data from 126 patients yielded a three-factor structure explaining 60% of the variance for the inventory. Factor 1 (neuropsychiatric symptoms) had 14 items, factor 2 (somatic symptoms) had eleven items, and factor 3 (sleep symptoms) had two items, explaining 28%, 22% and 11% of the variance, respectively. The three factor subscales demonstrated high intrinsic consistency reliability. GEE modeling of the 32 patients who initiated HCV therapy showed that patients developed worsening neuropsychiatric and somatic symptoms following HCV therapy with stable sleep symptoms. Bivariate analyses identified the following as predictors of HCV therapy initiation: lower HIV log₁₀ RNA, lower scores for neuropsychiatric, somatic and sleep symptoms, lower CES-D scores and white ethnicity. In stepwise multiple logistic regression analysis, low neuropsychiatric symptom score was the strongest independent predictor of HCV therapy initiation and HIV log₁₀ RNA was inversely associated with a decision to initiate HCV treatment.

Conclusions

A 41-item hepatitis-related symptom inventory was found to have a clinically meaningful 3-factor structure with excellent internal consistency reliability and predictive validity. In adjusted analysis, low neuropsychiatric symptom scores and controlled HIV infection were independent predictors of HCV treatment initiation. The usefulness of the HCV symptom inventory in monitoring HCV treatment should be evaluated prospectively.     

DNA methylation studies on imprinted loci in a male monozygotic twin pair discordant for Beckwith–Wiedemann syndrome

Tierling S, Souren NY, Reither S, Zang KD, Meng‐Hentschel J, Leitner D, Oehl‐Jaschkowitz B, Walter J. DNA methylation studies on imprinted loci in a male monozygotic twin pair discordant for Beckwith–Wiedemann syndrome. Beckwith–Wiedemann syndrome (BWS) is one of the most prevalent congenital disorders predominantly caused by epigenetic alterations. Here we present an extensive case study of a monozygotic monochorionic male twin pair discordant for BWS. Our analysis allows to correlate BWS symptoms, like a protruding tongue, indented ears and transient neonatal hypoglycaemia, to an abnormal methylation at the KvDMR1. DNAs extracted from peripheral blood, skin fibroblasts, saliva and buccal swab of both twins, their sister and parents were analysed at 11 differentially methylated regions (DMRs) including all four relevant DMRs of the BWS region. The KvDMR1 was exclusively found to be hypomethylated in all cell types of the affected BWS twin, while the unaffected twin and the relatives showed normal methylation in fibroblasts, buccal swab and saliva DNA. Interestingly, the twins share a common blood‐specific hypomethylation phenotype most probably caused by a feto‐fetal transfusion between both twins. Because microsatellite analysis furthermore revealed a normal biparental karyotype for chromosome 11, our results point to an exclusive correlation of the observed BWS symptoms to locally restricted epimutations at the KvDMR1 of the maternal chromosome.     

West Nile virus infection rates in Culex nigripalpus (Diptera: Culicidae) do not reflect transmission rates in Florida

We describe the first documented field transmission of West Nile (WN) virus by a North American mosquito. WN was first detected in northern Florida in 2001. An intensive mosquito trapping and surveillance program was conducted in this region for four nights to assess mosquito transmission of WN. Four mosquito traps, each with a single sentinel chicken, were placed at five different locations on each of four nights. A total of 11,948 mosquitoes was collected, and 14 mosquito pools were found to contain WN, giving a minimum infection rate between 1.08 and 7.54 per 1,000. Only one of the 80 sentinel chickens seroconverted to WN, demonstrating a single mosquito transmission event during the study and a mosquito transmission rate of between 0.8 and 1 per 1,000. Culex nigripalpus Theobald was responsible for WN transmission to the sentinel chicken, although both Cx. nigripalpus and Culex quinquefasciatus Say were found infected with WN. Mosquito transmission rates are reported in this study for the first time for a WN outbreak. This information is essential to determine risk of human and animal infection.     

Impact of robotics on the outcome of elderly patients with endometrial cancer

Objective

To evaluate the impact of introducing a robotics program on clinical outcome of elderly patients with endometrial cancer.

Methods

Evaluation and comparison of peri-operative morbidity and disease-free interval in 163 consecutive elderly patients (≥ 70 years) with endometrial cancer undergoing staging procedure with traditional open surgery compared to robotic surgery.

Results

All consecutive patients ≥ 70 years of age with endometrial cancer who underwent robotic surgery (n = 113) were compared with all consecutive patients ≥ 70 years of age (n = 50) before the introduction of a robotic program in December 2007. Baseline patient characteristics were similar in both eras. Patients undergoing robotic surgery had longer mean operating times (244 compared with 217 minutes, p = 0.009) but fewer minor adverse events (17% compared with 60%, p < 0.001). The robotics cohort had less estimated mean blood loss (75 vs 334 mL, p < 0.0001) and shorter mean hospital stay (3 vs 6 days, p < 0.0001). There was no difference in disease-free survival (p = 0.61) during the mean follow-up time of 2 years.

Conclusion

Transitioning from open surgery to a robotics program for the treatment of endometrial cancer in the elderly has significant benefits, including lower minor complication rate, less operative blood loss and shorter hospitalization without compromising 2-year disease-free survival.     

Oocyte morphology on day 0 correlates with aneuploidy as detected by polar body biopsy and FISH

Purpose

For better selection of oocytes and embryos, preimplantation genetic screening (PGS) was introduced. As from the beginning of IVF, morphology was used as selection criteria; we investigated the combination of both. If there was a correlation between phenotype and genotype, invasive PGS might be replaced.

Method

Therefore, 104 cycles with PGS were done by biopsy of the first polar body and FISH with five chromosomes. Morphology of the oocyte was recorded digitally and noted for 12 categories in 4–13 values; evaluation of the chromosomes was noted for five chromosomes in five values. Morphology and genetics were correlated to each other.

Result

Correlations between morphology and genetics for day 0 were found: oocytes with an irregular or dark zona are less probable to have a normal chromosome 13 (80 vs. 53 %, p = 0.001). A medium amount of detritus in the perivitelline space makes it more probable to have a normal chromosome 18 (94 vs. 78 %, p = 0.001). A halo in the cytoplasm makes it less probable to be euploid for chromosome 22 (56 vs. 75 %, p = 0.018). For day 1, pattern “1, 2, 3 and fine” in the pronuclei makes it more probable to be euploid for chromosome 22 (78 vs. 63 %, p = 0.002).

Conclusion

There are correlations between the oocyte genome and its morphology also on day 0. These correlations are not sufficient to replace PGS.     

Impact on postmenopausal symptoms of adding continuous C-21 versus C-19 progestin to estrogen

OBJECTIVE: To compare the effects of (i) continuous low dosage C-19 progestin (dl-norgestrel, NG) plus cyclical conjugated estrogen (CEE) versus (ii) continuous low dosage C-21 progestin [medroxyprogesterone acetate (MPA)] plus CEE on postmenopausal vaginal bleeding, mood and somatic, psychosomatic and psychological symptoms. METHODS: Nine hypercholesterolemic postmenopausal women with intact uteri were randomly assigned in a prospective, double-blind, two-period cross-over study of CEE (25/28 days) plus either (i) NG, (0.05 mg/day) or (ii) MPA (2.5 mg/day) for 1 year and after an appropriate wash-out period were switched to the alternative regimen for another year. Four hysterectomized control subjects received the CEE only. RESULTS: Administration of CEE + MPA versus CEE + NG resulted in a significantly higher percent of cycles which were free of vaginal bleeding (97 vs 85%), spotting (92 vs 79%) and either spotting or bleeding (92 vs 76%, P < 0.01). All three regimens significantly reduced the overall combined scores for postmenopausal somatic, psychosomatic and psychological symptoms (P < 0.05). CONCLUSIONS: Vaginal bleeding and/or spotting were significantly less frequent with CEE + MPA versus CEE + NG. However, each of the three hormonal regimens improved mood and significantly reduced postmenopausal symptoms in comparison to untreated control values.     

Head-to-Head Evaluation of Five Chlamydia Tests Relative to a Quality-Assured Culture Standard

Nucleic acid amplification tests offer superior sensitivity for the detection of Chlamydia trachomatis infection, but many laboratories still use nonamplification methods because of the lower cost and ease of use. In spite of their availability for more than a decade, few studies have directly compared the nonamplification tests. Such comparisons are still needed in addition to studies that directly compare individual nonamplification and amplification tests. The purpose of this study was to evaluate and compare the performance characteristics relative to culture of five different tests for the detection of C. trachomatis with and without confirmation of positive results. The tests were applied to endocervical specimens from 4,980 women attending family planning clinics in the northwestern United States. The five nonculture tests included Chlamydiazyme (Abbott), MicroTrak direct fluorescent antibody (DFA) (Syva), MicroTrak enzyme immunoassay (EIA) (Syva), Pace 2 (Gen-Probe), and Pathfinder EIA (Sanofi/Kallestad). All positive results obtained with a nonculture test (except MicroTrak DFA) were confirmed by testing the original specimens with a blocking antibody test (Chlamydiazyme), a cytospin DFA (MicroTrak EIA and Pathfinder EIA), and a probe competition assay (Pace 2). The prevalence of culture-proven chlamydia was 3.9%. The sensitivities of the nonculture tests were in a range from 62 to 75%, and significant differences between tests in terms of sensitivity were observed. The positive predictive value for each test was 0.85 or higher. The specificities of the nonculture tests without performance of confirmations were greater than 99%. Performing confirmatory tests eliminated nearly all of the false positives.