TB drug susceptibility testing in high fluoroquinolone resistance settings.

BACKGROUND: The insurgence of resistance to key drugs of the BPaLM (bedaquiline + pretomanid + moxifloxacin) regimen is a major concern. In settings with widespread resistance to fluoroquinolones (FQs), like Pakistan, new technologies, such as Xpert® MTB/XDR, may ensure drug resistance upfront screening. This study aims to assess MTB/XDR's performance in detecting FQs and isoniazid resistance, proposing a renewed diagnostic algorithm for drug-resistant TB (DR-TB). METHODS: This cross-sectional prospective study, approved by the local ethical committee, collected samples from people newly and previously diagnosed with TB over 6 months. Xpert® MTB/RIF Ultra, MTB/XDR, Genotype® MTBDRplus, Genotype® MTBDRsl, culture, and phenotypic drug susceptibility testing (pDST) for relevant drugs (including bedaquiline and levofloxacin) were performed. Next-generation sequencing (NGS) resolved discordances between MTB/XDR and pDST results. RESULTS: The analysis showed that MTB/XDR has 91.5% and 88.2% sensitivity and 99.5% and 97.7% specificity in detecting respectively isoniazid (INH) and resistance to FQs, demonstrating that MTB/XDR meets the WHO targets for INH resistance detection at the peripheral level. NGS effectively resolved discordances between MTB/XDR and pDST results. CONCLUSIONS: The obtained results allowed designing the proposed diagnostic algorithm for rapid identification of DR-TB, ensuring rapid and equitable access to drug susceptibility testing for TB, ultimately improving TB care and control.

CONTEXTE: La recrudescence de la résistance aux médicaments clés du régime BPaLM (bédaquiline + prétomanide + moxifloxacine) est une préoccupation majeure. Dans les contextes où la résistance aux fluoroquinolones (FQ) est répandue, comme le Pakistan, de nouvelles technologies, telles que Xpert® MTB/XDR, peuvent assurer un dépistage initial de la résistance aux médicaments. Cette étude vise à évaluer la performance de MTB/XDR dans la détection des FQ et de la résistance à l'isoniazide, en proposant un algorithme de diagnostic renouvelé pour la TB pharmacorésistante (DR-TB, pour l'anglais «drug-resistant TB ¼ ). MÉTHODES: Cette étude prospective transversale, approuvée par le comité d'éthique local, a recueilli des échantillons de personnes nouvellement diagnostiquées et précédemment diagnostiquées avec la TB pendant 6 mois. Xpert® MTB/RIF Ultra, MTB/XDR, le GenoType® MTBDRplus, le GenoType® MTBDRsl, la culture et des tests phénotypiques de sensibilité aux médicaments (pDST, pour l'anglais «phenotypic drug susceptibility testing ¼ ) pour les médicaments pertinents (y compris la bédaquiline et la lévofloxacine) ont été effectués. Le séquençage de nouvelle génération (NGS, pour l'anglais «next-generation sequencing ¼ ) a résolu les discordances entre les résultats MTB/XDR et pDST. RÉSULTATS: L'analyse a montré que le MTB/XDR a une sensibilité de 91,5% et 88,2% et une spécificité de 99,5% et 97,7% dans la détection respectivement de l'isoniazide et de la résistance aux FQ, démontrant que le MTB/XDR répond aux objectifs de l'OMS pour la détection de la résistance à l'isoniazide au niveau périphérique. NGS a efficacement résolu les discordances entre les résultats MTB/XDR et pDST. CONCLUSIONS: Les résultats obtenus ont permis de concevoir l'algorithme de diagnostic proposé pour l'identification rapide de la DR-TB, garantissant un accès rapide et équitable aux tests de sensibilité aux médicaments pour la TB, améliorant ainsi la prise en charge et le contrôle de la TB.

Preparation and evaluation of cytotoxic potential of paclitaxel containing poly-3-hydroxybutyrate-co-3-hydroxyvalarate (PTX/PHBV) nanoparticles.

Paclitaxel (PTX) is a potent anticancer drug. In the present study, PTX was loaded in poly-3-hydroxybutyrate-co-3-hydroxyvalarate (PHBV) to fabricate the PTX/PHBV (drug-loaded) nanoparticles via the nanoprecipitation method. Blank PHBV nanoparticles were also prepared. The drug-encapsulation efficiency of PTX/PHBV nanoparticles was 45±0.4%. The PTX/PHBV nanoparticles exhibited a pH-sensitive release profile and followed a quasi-Fickian diffusion mechanism. Cytotoxic properties of PHBV and PTX/PHBV nanoparticles were checked against the MCF-7 and Caco-2 cell lines. The PHBV nanoparticle did not inhibit the proliferation of MCF-7 and Caco-2 cell lines, thus depicting their non-toxic and biocompatible nature. On the other hand, the PTX/PHBV nanoparticles demonstrated 1.03-fold higher cytotoxicity and 1.61-fold enhanced apoptosis after treatment with the PTX/PHBV nanoparticles versus free PTX. In summary, the PHBV nanoparticles could be a potential candidate for the delivery of PTX for cancer treatment.

Primary drug resistance in extra-pulmonary tuberculosis: a hospital-based prospective study from Pakistan.

SETTING: Tertiary level specialised tuberculosis (TB) hospital.OBJECTIVE: To determine the prevalence of antimicrobial resistance in new extra-pulmonary tuberculosis (EPTB) cases.DESIGN: Prospective cross-sectional study. Presumptive EPTB patients with enlarged lymph nodes or pleural effusion having no history of TB treatment were enrolled. Specimens were tested for smear, Xpert® MTB/RIF and culture. Indirect drug susceptibility testing (DST) was performed using MGIT 960 and line-probe assays (LPA).RESULTS: Among 671 cases, 255 were bacteriologically confirmed and 185 DSTs were performed. Multidrug resistance (MDR-TB) was reported in 2.2% (95%CI 0.6-5.4), any resistance to rifampicin (RMP) in 2.7% (95%CI 0.9-6.2), isoniazid (INH) in 7.6% (95%CI 4.1-12.4), ethambutol in 1.1% (95%CI 0.1-3.9), pyrazinamide in 2.2% (95%CI 0.9-5.5) and fluoroquinolones (FQ) in 6.0% (95%CI 3.0-10.4). The sensitivity and specificity of LPA-DST was 100% and >98.8% respectively for RMP, INH and FQ. Among 82 cases with RMP of the results available for all three methods used, five were reported to be resistant on Xpert but all five were susceptible on MGIT 960 and four on MTBDRplus.CONCLUSION: Prevalence of RMP resistance in new EPTB cases is 2.7% (95%CI 0.9-6.2). Caution is warranted for RMP resistance detected using Xpert in EPTB samples with a very low bacterial load.

An immunohistochemical study of neurotransmitter profiles in developing human visual cortex.

The temporal pattern of development and distribution of gamma aminobutyric acid, serotonin, substance P and neuropeptide Y immunoreactive profiles was studied in the human visual cortex from 16 to 26 weeks of gestation, using an immunohistochemical technique. The immunoreactive profiles showed an increase in number and a change in their morphology and distribution pattern over the time period studied. A large number of neurons, fibers and terminals were stained with GABA antibody at 17-18 weeks and were distributed throughout the five zones of the developing visual cortex. GABA neurons were non-pyramidal and bipolar in form at 17-18 weeks while at 18-19 and 20-21 weeks the cells of subplate and intermediate zones were multipolar. Substance P and serotonin immunopositive fibers were present mainly in the intermediate zone at 16 and 17-18 weeks, where they were oriented in a horizontal manner. At subsequent ages they invaded the other zones also. Substance P positive neurons could be visualized only at 26 weeks of gestation in the intermediate, subventricular and ventricular zones; no cell bodies, however, stained with serotonin antibody. Neuropeptide Y immunoreactive cells and fibers were first seen in the intermediate zone but later were found to be distributed in other zones too. The observations indicate that the intermediate zone of the visual cortex in which the transmitters and peptides appear earlier assumes importance in the normal development as also noted in other mammals.

Early development of visual cortex in human fetuses.

Prenatal development of visual cortex (area 17) was studied in human fetuses of 8-9, 13-15 and 16-18 weeks of gestation, with a view to analyse the early critical events. Under light microscope, five zones of development were seen in all the age groups. The total thickness of cortex of area 17 as well as that of its cortical plate was measured with the help of camera lucida. It was observed that the total thickness of the cortex increased with increase in age. Diversity in the shape, size, staining intensity and arrangement of neurons was noted in the different zones. Most of the cells were found to have a thin rim of cytoplasm and a prominent nucleus with multiple nucleoli. The cells in subventricular zone and cortical plate were regularly arranged in vertical rows while in other zones, they were irregularly scattered. Several mitotic figures were seen in the ventricular zone at 8-9 weeks but later they were also noticed in subventricular and intermediate zones. In the later ages the mitotic figures were observed to be fewer in the ventricular zone. No mitosis was seen in cortical plate at any age period.

Xpert(®) MTB/RIF under routine conditions in diagnosing pulmonary tuberculosis: a study in two hospitals in Pakistan.

Xpert(®) MTB/RIF testing was offered to consecutive patients with presumptive tuberculosis (TB) attending two hospitals in Pakistan during April-May 2012, in addition to routine diagnostic protocol (smear microscopy, chest radiography and clinical judgement). We assessed the relative contribution of each tool in detecting pulmonary TB under routine conditions. Of 606 participants, 121 (20%) were detected as pulmonary TB: 46 (38%) by microscopy, 38 (31%) by Xpert alone and 37 (31%) on clinical and radiological grounds; 41 (65%) were detected by both Xpert and microscopy. One patient had rifampicin resistance. Although Xpert detected approximately twice as many TB cases as microscopy (n = 79, 65%), clinical judgement remained favoured by clinicians even when smear and Xpert were negative.

En plus d'un protocole de diagnostic de routine (examen microscopique des frottis, cliché thoracique et évaluation clinique), on a offert Xpert® MTB/RIF à des patients consécutifs suspects de tuberculose (TB) qui présentaient à deux hôpitaux du Pakistan au cours de la période avril­mai 2012. Nous avons évalué la contribution relative de chaque outil à la détection de la TB pulmonaire dans les conditions de routine. Sur 606 participants, 121 (20%) ont été diagnostiqués comme TB pulmonaire : 46 (38%) par l'examen microscopique, 38 (31%) par Xpert seul, et 37 (31%) sur une base clinique et radiologique ; 41 (65%) ont été détectés par l'examen microscopique et Xpert. Chez un patient, on a trouvé une résistance à la rifampicine. Quoique l'Xpert ait détecté approximativement deux fois le nombre de cas de TB détectés par l'examen microscopique (n = 79, 65%), le jugement clinique reste favorisé par les cliniciens, même lorsque le résultat du frottis et de l'Xpert est négatif.

En dos hospitales de Paquistán se propuso a un grupo de pacientes con presunción de tuberculosis (TB) que acudieron de manera consecutiva entre abril y mayo del 2012 la prueba Xpert® MTB/RIF, además del protocolo diagnóstico corriente (que comportaba la baciloscopia, la radiografía de tórax y la evaluación clínica). Se analizó la contribución relativa de cada instrumento en el diagnóstico de la TB pulmonar en las condiciones corrientes. De los 606 participantes, en 121 (20%) se estableció el diagnóstico de TB pulmonar de la siguiente manera: en 46 casos (38%) por microscopia, en 38 (31%) mediante la prueba Xpert sola y en 37 casos (31%) con base en las características clínicas y radiográficas; 41 (65%) fueron detectados por microscopia y Xpert. Un paciente exhibió resistencia a rifampicina. Si bien la prueba Xpert permitió el diagnóstico de cerca del doble de casos de tuberculosis que la baciloscopia (n = 79, 65%), el juicio clínico predomina aun en la decisión de los médicos, incluso frente a un resultado negativo de la baciloscopia y la prueba Xpert.

Comparison of two imaging modalities: F-speed film and digital images for detection of osseous defects in patients with interdental vertical bone defects.

OBJECTIVES: The aim of the present study was to compare the diagnostic accuracy of F-speed conventional film, unenhanced digital images and inversion-enhanced digital images for the detection of osseous defects in patients with vertical bone defects. METHODS: 23 vertical osseous defects in the mandible were evaluated. Intrasurgical measurements were made from the cementoenamel junction (CEJ) to the deepest extension of the osseous defects by one of the researchers. Radiographic measurements were obtained on conventional F-speed film, unenhanced digital images and inversion-enhanced digital images by six examiners. From each measure the corresponding probe measure was subtracted to form a difference score. RESULTS: Significant differences in means of difference scores were found among examiners within each imaging modality, and among the modalities within five of the six examiners. A significant (P<0.001) interaction term for the ANOVA indicated that differences among modality means were not the same across all examiners. The difference means were significantly different from zero for five of the six examiners with conventional F-speed film, four of six with inversion enhanced digital images, but for only one of six for unenhanced digital images. The reliability coefficient computed on a per examiner basis was 0.90 for conventional F-speed film, 0.94 for unenhanced digital image and 0.79 for inversion-enhanced digital image. CONCLUSIONS: In this study, unenhanced digital imaging was found to be superior to conventional F-speed film and inversion-enhanced digital images for accurately imaging periodontal osseous defects in patients.