Real-time PCR optimization to identify Mycobacterium tuberculosis complex strains in clinical samples.

Resumen del artículo publicado en FEBS Journal,http://dx.doi.org/10.1111/febs.12919During recent years several molecular techniques have become available for Mycobacterium tuberculosis complex (MTC) detection,both for clinical samples and for isolates. One of the techniques more widely used is real time PCR in combination with nucleic acid amplification protocols. There are numerous studies based on PCR for the diagnosis of tuberculosis although the different protocols and primers used in the laboratory, together with the variability in the diagnostic performance of the methods tested, require that a comparative study be performed. Furthermore,the fact that the detection from clinical samples requires using highly sensitive targets suggests that this type of study should include multicopy targets to compare their efficiency with respect to the single copy. Our aim was to identify the members of the MTC using real-time PCR assays based on SYBR Green,among a large panel of isolated bacterial strains and clinical samples.We chose three targets (IS6110, senx3-regx3 and cfp32) and the optimal values for each PCR assay were empirically defined by testing in triplicate different concentrations of MgCl2 and primer sets and different annealing temperatures. These conditions were determined based on the specific amplification reactions that showed a lower Ct value, higher fluorescence and absence of non-specific PCR products. The analytical sensitivity was evaluated by ten-fold serial dilutions of DNA from MTC and the specificity was tested by 62 different microorganisms, including bacteria related with the MTC. The diagnostic yield was evaluated in 66 specimens from patients with suspected tuberculosis;30 had tuberculosis and 36 (control group) had different diseases.Under the conditions that resulted in optimization, standard curves showed that senx3-regx3 assay was the most efficient, followed by IS6110 and cfp32. However, the detection of bacterial DNA was faster with the repetitive element IS6110, with Ct values of up to 3 and 9 cycles of difference with respect to senx3-regx3 and cfp32. The analytical specificity, done only with the senx3-regx3 and IS6110 targets, was in the order of 100 and 93.5%, since IS6110 amplified various non-tuberculous micobacteria.For all the clinical samples studied, the sensitivity of both assays was identical (93.3%) but the specificity of senx3-regx3(100%) was higher than that of IS6110 (94.7%). In conclusion,real time PCR assay-SYBR Green based on the targets senx3-regx3 is highly reproducible and more sensitive and specific than the assays based on IS6110 or cfp32. The protocol developed in this study provides an appropriate and rapid tool to identify the strains of MTC in different clinical isolates and specimens.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Spinal infection: state of the art and management algorithm

Spinal infection is a rare pathology although a concerning rising incidence has been observed in recent years. This increase might reflect a progressively more susceptible population but also the availability of increased diagnostic accuracy. Yet, even with improved diagnosis tools and procedures, the delay in diagnosis remains an important issue. This review aims to highlight the importance of a methodological attitude towards accurate and prompt diagnosis using an algorithm to aid on spinal infection management. METHODS: Appropriate literature on spinal infection was selected using databases from the US National Library of Medicine and the National Institutes of Health. RESULTS: Literature reveals that histopathological analysis of infected tissues is a paramount for diagnosis and must be performed routinely. Antibiotic therapy is transversal to both conservative and surgical approaches and must be initiated after etiological diagnosis. Indications for surgical treatment include neurological deficits or sepsis, spine instability and/or deformity, presence of epidural abscess and upon failure of conservative treatment. CONCLUSIONS: A methodological assessment could lead to diagnosis effectiveness of spinal infection. Towards this, we present a management algorithm based on literature findings

Multi-messenger observations of a binary neutron star merger

On 2017 August 17 a binary neutron star coalescence candidate (later designated GW170817) with merger time 12:41:04 UTC was observed through gravitational waves by the Advanced LIGO and Advanced Virgo detectors. The Fermi Gamma-ray Burst Monitor independently detected a gamma-ray burst (GRB 170817A) with a time delay of ~1.7 s with respect to the merger time. From the gravitational-wave signal, the source was initially localized to a sky region of 31 deg2 at a luminosity distance of 40+8-8 Mpc and with component masses consistent with neutron stars. The component masses were later measured to be in the range 0.86 to 2.26 Mo. An extensive observing campaign was launched across the electromagnetic spectrum leading to the discovery of a bright optical transient (SSS17a, now with the IAU identification of AT 2017gfo) in NGC 4993 (at ~40 Mpc) less than 11 hours after the merger by the One- Meter, Two Hemisphere (1M2H) team using the 1 m Swope Telescope. The optical transient was independently detected by multiple teams within an hour. Subsequent observations targeted the object and its environment. Early ultraviolet observations revealed a blue transient that faded within 48 hours. Optical and infrared observations showed a redward evolution over ~10 days. Following early non-detections, X-ray and radio emission were discovered at the transient’s position ~9 and ~16 days, respectively, after the merger. Both the X-ray and radio emission likely arise from a physical process that is distinct from the one that generates the UV/optical/near-infrared emission. No ultra-high-energy gamma-rays and no neutrino candidates consistent with the source were found in follow-up searches. These observations support the hypothesis that GW170817 was produced by the merger of two neutron stars in NGC4993 followed by a short gamma-ray burst (GRB 170817A) and a kilonova/macronova powered by the radioactive decay of r-process nuclei synthesized in the ejecta

Pharmacokinetic drug interactions of antimicrobial drugs:a systematic review on oxazolidinones, rifamycines, macrolides, fluoroquinolones, and Beta-lactams

Like any other drug, antimicrobial drugs are prone to pharmacokinetic drug interactions. These drug interactions are a major concern in clinical practice as they may have an effect on efficacy and toxicity. This article provides an overview of all published pharmacokinetic studies on drug interactions of the commonly prescribed antimicrobial drugs oxazolidinones, rifamycines, macrolides, fluoroquinolones, and beta-lactams, focusing on systematic research. We describe drug-food and drug-drug interaction studies in humans, affecting antimicrobial drugs as well as concomitantly administered drugs. Since knowledge about mechanisms is of paramount importance for adequate management of drug interactions, the most plausible underlying mechanism of the drug interaction is provided when available. This overview can be used in daily practice to support the management of pharmacokinetic drug interactions of antimicrobial drugs

Nurses' perceptions of aids and obstacles to the provision of optimal end of life care in ICU

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