Manual de laboratorio para el diagnóstico por PCR en tiempo real en laboratorios descentralizados y vigilancia genómica de SARS-CoV-2 / Laboratory manual for real-time PCR diagnosis in decentralized laboratories and genomic surveillance of SARS-CoV-2

En los antecedentes, brinda detalles técnicos de enfermedades previas al COVID-19 y sigue esta consigna internacional: "Sin embargo, a la luz de la posible introducción de un caso sospechoso relacionado con el 2019-nCoV en la Región de las Américas, la Organización Panamericana de la Salud / Organización Mundial de la Salud (OPS / OMS) recomienda a los Estados Miembros garantizar su identificación oportuna, el envío de las muestras a laboratorios Nacionales o de referencia y la implementación del protocolo de detección molecular para 2019-nCoV, según la capacidad del laboratorio." Aborda las tres mutaciones del virus conocidas hasta la fecha del documento y las tres variantes conocidas: Reino Unido e Irlanda del Norte, República de Sudáfrica y Brasil, siendo la de mayor transmisibilidad, según el documento, la del Reino Unido. El primero de los objetivos del documento es: "Brindar los lineamientos generales para la toma de muestra, conservación, transporte y diagnóstico del SARS-Cov-2 en los laboratorios descentralizados bajo la técnica de Reacción en Cadena de la Polimerasa (PCR) en tiempo Real, en atención a pacientes que cumplen con la definición de caso y se tipifica como "sospechoso inusitado"." Incluye como anexo 2 el documento: "Lineamientos para el abordaje de casos positivos para SARS-COV-2 de viajeros que ingresan al país por el aeropuerto internacional "La Aurora" (AILA)"

E. coli nitroreductase NfsA is a reporter gene for non-invasive PET imaging in cancer gene therapy applications.

The use of reporter genes to non-invasively image molecular processes inside cells has significant translational potential, particularly in the context of systemically administered gene therapy vectors and adoptively administered cells such as immune or stem cell based therapies. Bacterial nitroreductase enzymes possess ideal properties for reporter gene imaging applications, being of non-human origin and possessing the ability to metabolize a range of clinically relevant nitro(hetero)cyclic substrates. Methods: A library of eleven Escherichia coli nitroreductase candidates were screened for the ability to efficiently metabolize 2-nitroimidazole based positron emission tomography (PET) probes originally developed as radiotracers for hypoxic cell imaging. Several complementary methods were utilized to detect formation of cell-entrapped metabolites, including various in vitro and in vivo models to establish the capacity of the 2-nitroimidazole PET agent EF5 to quantify expression of a nitroreductase candidate. Proof-of-principle PET imaging studies were successfully conducted using 18F-HX4. Results: Recombinant enzyme kinetics, bacterial SOS reporter assays, anti-proliferative assays and flow cytometry approaches collectively identified the major oxygen-insensitive nitroreductase NfsA from E. coli (NfsA_Ec) as the most promising nitroreductase reporter gene. Cells expressing NfsA_Ec were demonstrably labelled with the imaging agent EF5 in a manner that was quantitatively superior to hypoxia, in monolayers (2D), multicellular layers (3D), and in human tumor xenograft models. EF5 retention correlated with NfsA_Ec positive cell density over a range of EF5 concentrations in 3D in vitro models and in xenografts in vivo and was predictive of in vivo anti-tumor activity of the cytotoxic prodrug PR-104. Following PET imaging with 18F-HX4, a significantly higher tumor-to-blood ratio was observed in two xenograft models for NfsA_Ec expressing tumors compared to the parental tumors thereof, providing verification of this reporter gene imaging approach. Conclusion: This study establishes that the bacterial nitroreductase NfsA_Ec can be utilized as an imaging capable reporter gene, with the ability to metabolize and trap 2-nitroimidazole PET imaging agents for non-invasive imaging of gene expression.

Qualitative and quantitative analysis of formaldehyde in samples of hair straighteners and suitability of the labels

Formaldehyde is an active compound, irregularly used in hair products, that has the property of straighten and waterproofing the wires. However, it is highly toxic and can stimulate dermatological hypersensitivity and cancer. In this context it is of fundamental importance the inspection of these products that can be used in safe conditions for the consumer, without formaldehyde in concentrations higher than the allowed. Thus, the aim of this research was the qualitative and quantitative identification formaldehyde in samples of hair straighteners that was obtained by donation in the beauty salons of Araraquara-SP. In addition, the analysis of the packaging labels of the products tested were conducted, following the requirements of the national legislation - RDC 07/2015 which defines the mandatory labeling standards for cosmetic products. A qualitative analysis for formaldehyde identification is based on the formation of a purple colored complex. The quantitative analysis was performed by spectrophotometry. The qualitative and quantitative formaldehyde analysis methods were applied to 13 bottles of hair straighteners. When submitted to qualitative analysis, all samples showed formaldehyde presence. The quantitative analysis demonstrated that the samples identified as B, C, D, E, G, H, I, J and M presented formaldehyde concentration of 3.5 to 14.5%, which is above of the limit recommended by the National Health Surveillance Agency (ANVISA), of 0.2%. In the label analysis, in all samples were found irregularities.(AU)

Methylene blue dye for identification of processus vaginalis during hydrocele repair: experience in a teaching hospital.

BACKGROUND: The aim of this study is to investigate the use of methylene blue in perioperative identification of the patent processus vaginalis in a group of boys presenting with congenital or recurrent hydrocele where surgery was performed by junior surgeons in training. METHODS: We retrospectively reviewed the notes of 22 boys with hydrocele, of which two recurrences, who were operated via a standard inguinal approach, by trainees. Methylene blue 0.3-0.5 mL was injected into the hydrocele fluid through the scrotal wall. A processus vaginalis was identified as a blue line. RESULTS: Methylene blue injection clearly identified a patent processus vaginalis in 91% of patients. In 9% (N.=2), of which one recurrence, methylene blue injection demonstrated a hydrocele with an obliterated processus vaginalis. There were no intraoperative complications. No testicular atrophy was recorded. CONCLUSIONS: Injection of methylene blue into the hydrocele sac may be considered a useful aid for a clearer identification of a difficult patent processus vaginalis. In the present series, there were no complications, and thus we believe that this technique might be suitable and especially helpful, in cases of recurrent hydrocele, and for junior surgeons in training.

[Cervical cancer: Current situation and management in Morocco]. / Le cancer du col de l'utérus : état des lieux et prévention au Maroc.

With more than 3300 new cases and almost 2500 deaths each year, cervical cancer (CC) ranks second among female cancers in Moroccan women. The majority of cases occurs in women aged 50 and over. In absence of a national cancer registry, data published in Morocco are limited to the number of cases recorded in some oncology centers, so the incidence of this cancer is likely much higher than estimated. A Moroccan national program against CC based on the practice of visual inspection after application of acetic acid was set up in 2010, allowing both screening and possibly immediate treatment of (pre)cancerous lesions. However, this program has not been implemented in all regions of the country. The CC develops slowly and most often without any symptoms, and so it is diagnosed at an advanced stage of the disease. Virtually, all CC are associated with persistent infection of high risk human papillomavirus (HPV), particularly HPV16 and 18. For more than ten years, two prophylactic vaccines targeting these two HPV genotypes have been marketed. They have proved their excellent immunogenicity and efficacy and they are well tolerated. However, HPV vaccine is not yet recommended by health authorities in Morocco. In this literature review, we focused on the current situation of CC, the prevalence of HPV infection and the prevention strategies against CC in Morocco.

The use of high-frequency short bipolar pulses in cisplatin electrochemotherapy in vitro.

Background In electrochemotherapy (ECT), chemotherapeutics are first administered, followed by short 100 µs monopolar pulses. However, these pulses cause pain and muscle contractions. It is thus necessary to administer muscle relaxants, general anesthesia and synchronize pulses with the heart rhythm of the patient, which makes the treatment more complex. It was suggested in ablation with irreversible electroporation, that bursts of short high-frequency bipolar pulses could alleviate these problems. Therefore, we designed our study to verify if it is possible to use high-frequency bipolar pulses (HF-EP pulses) in electrochemotherapy. Materials and methods We performed in vitro experiments on mouse skin melanoma (B16-F1) cells by adding 1-330 µM cisplatin and delivering either (a) eight 100 µs long monopolar pulses, 0.4-1.2 kV/cm, 1 Hz (ECT pulses) or (b) eight bursts at 1 Hz, consisting of 50 bipolar pulses. One bipolar pulse consisted of a series of 1 µs long positive and 1 µs long negative pulse (0.5-5 kV/cm) with a 1 µs delay in-between. Results With both types of pulses, the combination of electric pulses and cisplatin was more efficient in killing cells than cisplatin or electric pulses only. However, we needed to apply a higher electric field in HF-EP (3 kV/cm) than in ECT (1.2 kV/cm) to obtain comparable cytotoxicity. Conclusions It is possible to use HF-EP in electrochemotherapy; however, at the expense of applying higher electric fields than in classical ECT. The results obtained, nevertheless, offer an evidence that HF-EP could be used in electrochemotherapy with potentially alleviated muscle contractions and pain.

INFLUENCE OF INVERTED INTERNAL LIMITING MEMBRANE FLAP TECHNIQUE ON THE OUTER RETINAL LAYER STRUCTURES AFTER A LARGE MACULAR HOLE SURGERY.

PURPOSE: To examine the influence of inverted internal limiting membrane (ILM) flap technique on the outer retinal layer structures after macular hole (MH) surgery. METHODS: We included 24 eyes of 24 patients who underwent vitrectomy for large MHs (≥400 µm) with successful MH closure and observed for at least 6 months. Fourteen eyes were treated with inverted ILM flap technique (inverted group) and 10 with conventional ILM peeling (ILMP group). We evaluated the postoperative recovery rate of the external limiting membrane and ellipsoid zone and the best-corrected visual acuity. RESULTS: The postoperative recovery rates of the external limiting membrane and ellipsoid zone in the inverted group were lower than those in the ILMP group (21.4 vs. 70.0%, P = 0.024; 0 vs. 30.0, P = 0.059, respectively). The external limiting membrane recovery period in the inverted group was significantly longer than that in the ILMP group (11.0 ± 1.7 vs. 3.4 ± 2.8 months, P = 0.015). The best-corrected visual acuity change (letters) in the inverted group was significantly smaller than that in the ILMP group (9.0 vs. 22.5, P = 0.040). CONCLUSION: The poorer anatomical and visual results associated with inverted ILM flap compared with ILM peeling suggest the limitation of the ILM flap technique to repair refractory MHs.

New Vital Dye Injection Technique With Vitrectomy Probe.

BACKGROUND AND OBJECTIVE: To evaluate a new, safer way to inject vital dye during chromovitrectomy for dye-assisted macular peeling. PATIENTS AND METHODS: Prospective consecutive case series. Enrolled patients underwent macular surgery with a new technique of staining the epiretinal membrane and the internal limiting membrane with vital dyes. Twenty eyes of 20 patients (eight men, 12 women) were affected by idiopathic epiretinal membrane and underwent 25-gauge via pars plana vitrectomy for epiretinal membrane removal. Ten eyes were stained with Membrane Blue-Dual (Brilliant Blue G + trypan blue; DORC, Zuidland, The Netherlands) and 10 eyes were stained with Brilliant Peel (Brillant Blue G; Fluoron GmbH, Ulm, Germany). Preoperatively and postoperatively (1, 3, and 6 months) all patients received a complete ophthalmologic examination with best-corrected visual acuity (BCVA) assessment, swept-source optical coherence tomography, and multifocal electroretinography (mfERG). RESULTS: The mean age of the patients was 67.0 years ± 8.6 years (range: 55 years to 78 years). No statistically significant difference in BCVA improvement or central foveal thickness decrease was observed in either group (P > .05). The mfERG showed an increase in electrical response densities 3 months after surgery. CONCLUSION: The authors describe a new technique to inject vital dye during chromovitrectomy for dye-assisted macular peeling that may help to improve the overall safety of macular surgery. [Ophthalmic Surg Lasers Imaging Retina. 2018;49:528-533.].

Circular stripes were more common in Barrett's esophagus after acetic acid staining.

BACKGROUND: The diagnosis of Barrett's esophagus (BE) is disturbed by numerous factors, including correct gastroesophageal junction judgment, the initial location of the Z-line and the biopsy result above it. The acetic acid (AA) could help to diagnose BE better than high resolution imaging technology or magnifying endoscopy, by providing enhanced contrast of different epithelium. We have noticed AA could produce multiple white circular lines, forming circular stripes (CS), at lower esophagus, which hasn't been reported by others. This study aimed to investigate whether the CS is a special marker in BE patients. METHODS: A total of 47 BE patients and 63 healthy people were enrolled from March 2016 to October 2016, and 2% AA staining had been operated routinely at lower esophagus under high resolution gastroscopy. We observed whether there were CS after AA staining and the images were compared between the two groups. RESULTS: CS were confirmed in 42 patients (89.36%) in the BE group and 5 (7.94) in the control group ((χ2 = 72.931, P < 0.001)). The average width of CS was 0.76 ± 0.25 cm in BE group, which was similar to that in the control group (0.88 ± 0.11 cm). Villous or punctate or reticular pattern usually existed above or below the CS. CONCLUSIONS: CS could be found at lower esophagus in most BE patients with AA staining, and this special feature might be valuable in diagnosing, evaluating and following up of BE patients.

Accuracy of combinations of visual inspection using acetic acid or lugol iodine to detect cervical precancer: a meta-analysis.

BACKGROUND: Visual inspection of the cervix with acetic acid (VIA) or with Lugol's iodine (VILI) have been evaluated for cervical cancer screening in developing countries. OBJECTIVES: To assess the diagnostic accuracy and clinical utility of visual methods to detect cervical intraepithelial neoplasia grade 2+ (CIN2+) using: (1) VIA alone; (2) VILI alone; (3) co-testing; and (4) VILI as a triage test of a positive VIA result. SEARCH STRATEGY: PubMed, EMBASE, and the Cochrane Library were searched up to May 2016. SELECTION CRITERIA: All reports on the accuracy of VIA and VILI, or combinations of VIA/VILI, to detect CIN2+ were identified. Histology and colposcopy when no biopsy was taken were used as the reference standard. DATA COLLECTION AND ANALYSIS: Selected studies were scored on methodological quality, and sensitivity and specificity were computed. Clinical utility was assessed from the positive predictive value (PPV) and the complement of the negative predictive value (cNPV). MAIN RESULTS: We included 23 studies comprising 101 273 women. The pooled sensitivity and specificity of VILI was 88 and 86%, respectively. VILI was more sensitive, but not less specific, compared with VIA (relative sensitivity = 1.11; 95% confidence interval, 95% CI, 1.06-1.16; relative specificity = 0.98; 95% CI 0.95-1.01). Co-testing was hardly more sensitive, but significantly less specific, than VILI alone. VILI to triage VIA-POSITIVE women was not less sensitive, but more specific, compared with VIA alone (relative sensitivity = 0.98, 95% CI 0.96-1.01; relative specificity = 1.04, 95% CI 1.02-1.05). The average PPVs were low (range 11-16%), whereas the cNPV varied between 0.3% (VILI, co-testing) and 0.6% (triage). CONCLUSIONS: Although imperfect, VILI alone appeared to be the most useful visual screening strategy. TWEETABLE ABSTRACT: VILI alone seems to be the most useful visual screening test for cervical cancer screening.

Development and validation of a training module on the use of acetic acid for the detection of Barrett's neoplasia.

Background and study aims Acetic acid chromoendoscopy (AAC) enhances the ability to correctly identify Barrett's neoplasia, and is increasingly used by both expert and nonexpert endoscopists. Despite its increasing use, there is no validated training strategy to achieve competence. The aims of our study were to develop a validated training tool in AAC-assisted lesion recognition, to assess endoscopists' baseline knowledge of AAC-assisted lesion recognition, and to evaluate the efficacy and impact of this training tool. Methods A validated assessment of 40 images and 20 videos was developed. A total of 13 endoscopists with experience of Barrett's endoscopy but no formal training in AAC were recruited to the study. Participants underwent: baseline assessment 1, online training, assessment 2, interactive seminar, assessment 3. Results Baseline assessment demonstrated a sensitivity of 83 % and a negative predictive value (NPV) of 83 %. The online training intervention significantly improved sensitivity to 95 % and NPV to 94 % (P < 0.01). Further improvement was seen after a 1-day interactive seminar including live cases, with sensitivity increasing to 98 % and NPV to 97 %. Conclusions The data demonstrate the need for training in AAC-assisted lesion recognition as baseline performance, even by Barrett's experts, was poor. The online training and testing tool for AAC for Barrett's neoplasia was successfully developed and validated. The training intervention improved performance of endoscopists to meet ASGE PIVI standards. The training tool increases the endoscopist's degree of confidence in the use of AAC. The training tool also leads to shift in attitudes of endoscopists from Seattle protocol towards AAC-guided biopsy protocol for Barrett's surveillance.

Evaluation of Provider Skills in Performing Visual Inspection with Acetic Acid in the Cervical Cancer Screening Program in the Meknes-Tafilalet Region of Morocco.

BACKGROUND: This study documented the performance of providers of visual inspection with acetic acid (VIA) at primary health centers, assessing their compliance with the VIA skills checklist and determinants of non-compliance, and exploring their perceptions of VIA training sessions. MATERIALS AND METHODS: A cross- sectional study was conducted among VIA providers in the Meknes-Tafilalet region of Morocco. Structured observation of their performance was conducted through supervisory visits and multiple focus group discussions (FGDs). RESULTS: Performance of all the recommended steps for effective communication was observed in a low proportion of procedures (36.4%). Midwives/nurses had higher compliance than general practitioners (GPs) (p<0.001). All recommended steps for VIA examination were performed for a high proportion of procedures (82.5%). Compliance was higher among midwives/nurses than among GPs (p<0.001) and among providers in rural areas than those in urban areas (p<0.001). For pre-VIA counselling, all recommended steps were performed for only 36.8% of procedures. For post-VIA counseling, all recommended steps were performed in a high proportion (85.5% for VIA-negative and 85.1% for VIA-positive women). Midwives/nurses had higher compliance than GPs when advising VIA-positive women (p=0.009). All infection prevention practices were followed for only 14.2% of procedures, and compliance was higher among providers in rural areas than those in urban areas (p<0.001). Most FGD participants were satisfied with the content of VIA training sessions. However, they suggested periodic refresher training and supportive supervision. CONCLUSIONS: Quality assurance of a cervical cancer screening program is a key element to ensure that the providers perform VIA correctly and confidently.

Contrast-Induced Acute Kidney Injury: Comparison of Preventative Therapies.

Contrast medium is used daily for diagnostic and interventional procdures as a means to visualize blood vessels. The administration of contrast dye, however, can lead to an acute reduction in kidney function. This complication can impact length of hospital stay, risk of dialysis, and increased hospital mortality. Common preventative measures include N-acetylcysteine and intravenous hydration. The evidence reviewed revealed hydration to be the more effective treatment to reduce the risk of acute kidney injury.

mHealth to Train Community Health Nurses in Visual Inspection With Acetic Acid for Cervical Cancer Screening in Ghana.

OBJECTIVE: There is a shortage of trained health care personnel for cervical cancer screening in low-/middle-income countries. We evaluated the feasibility and limited efficacy of a smartphone-based training of community health nurses in visual inspection of the cervix under acetic acid (VIA). MATERIALS AND METHODS: During April to July 2015 in urban Ghana, we designed and developed a study to determine the feasibility and efficacy of an mHealth-supported training of community health nurses (CHNs, n = 15) to perform VIA and to use smartphone images to obtain expert feedback on their diagnoses within 24 hours and to improve VIA skills retention. The CHNs completed a 2-week on-site introductory training in VIA performance and interpretation, followed by an ongoing 3-month text messaging-supported VIA training by an expert VIA reviewer. RESULTS: Community health nurses screened 169 women at their respective community health centers while receiving real-time feedback from the reviewer. The total agreement rate between all VIA diagnoses made by all CHNs and the expert reviewer was 95%. The mean (SD) rate of agreement between each CHN and the expert reviewer was 89.6% (12.8%). The agreement rates for positive and negative cases were 61.5% and 98.0%, respectively. Cohen κ statistic was 0.67 (95% CI = 0.45-0.88). Around 7.7% of women tested VIA positive and received cryotherapy or further services. CONCLUSIONS: Our findings demonstrate the feasibility and efficacy of mHealth-supported VIA training of CHNs and have the potential to improve cervical cancer screening coverage in Ghana.

Clinical sensitivity of HPV assays for the detection of high grade cervical disease in cervical samples treated with glacial acetic acid.

BACKGROUND: Lysis of bloody liquid based cytology (LBC) specimens with glacial acetic acid (GAA) is performed to aid cytological interpretation. However, the influence of GAA treatment on HPV detection is not fully understood and in studies designed to assess this, few cases of high-grade disease have been included. OBJECTIVES: To assess the sensitivity of HPV molecular tests for the detection of high grade cervical disease in GAA treated samples STUDY DESIGN: A total of 207 specimens associated with high grade dyskaryosis and treated with GAA were collated prospectively. Overall 140 specimens had underlying CIN2+, including 88 CIN3. All specimens were tested with the Abbott RealTime High Risk HPV test (rtHPV) and the Qiagen Hybrid Capture 2High Risk HPV DNA test (HC2). Specimens associated with a CIN2+ that were negative by either assay were genotyped. RESULTS: The sensitivity of rtHPV for CIN2+ and CIN3+ was 92.8% (87.2, 96.5) and 94.3% (87.2, 98.1) respectively. Sensitivity of the HC2 for CIN2+ and CIN3+ was 97.2% (92.8, 99.2) and 96.6% (90.3, 99.2) respectively. The sensitivity of both assays in GAA treated specimens was thus consistent with the level required for clinical application. HPV negative, CIN2+ specimens were generally attributable to HPV types outside the explicit analytical range of the assays. CONCLUSIONS: The data indicate that GAA treatment has little impact on the detection of CIN2+ by HPV testing in LBC specimens.

HIGH-RESOLUTION MULTIMODAL IMAGING AFTER IDIOPATHIC EPIRETINAL MEMBRANE SURGERY.

PURPOSE: To investigate the changes of the vitreomacular interface during a 1-year follow-up after idiopathic epiretinal membrane (iERM) surgery. METHODS: Six patients affected by fovea-attached iERM were recruited in this pilot study. Pars plana vitrectomy associated with epiretinal membrane peeling was performed uneventfully in all cases. In four cases, the inner limiting membrane was removed using Brilliant blue G. En face high-resolution adaptive optics and cross-sectional spectral domain optical coherence tomography retinal imaging were performed before and at 1, 3, 6, and 12 months after surgery. The microstructures of vitreomacular interface in high-resolution adaptive optics images were correlated to the cross-sectional spectral domain optical coherence tomography data. RESULTS: Preoperatively, adaptive optics images showed multiple abnormalities of the vitreomacular interface, such as macrofolds, microfolds, and hyperreflective microstructures. We identified two subtypes of iERM according to the distribution of microfolds over the foveal area, which included the radial-type and the grid-type iERM. After surgery, the morphology of the vitreomacular interface changed compared with the preoperative state. The number of both macrofolds and microfolds was reduced in all cases. The hyperreflective structures were still resolvable in all cases, however presenting different shape and morphology than preoperatively. In addition, they showed marked differences between eyes that had internal limiting membrane removal and eyes that did not. CONCLUSION: Adaptive optics imaging gives new insight into the changes of vitreomacular interface after iERM surgery. Enhanced multimodal imaging of the vitreomacular interface and retinal structures can be valuable to monitor treatment outcome of iERM.

Analysis of the intra- and intertumoral heterogeneity of hypoxia in pancreatic cancer patients receiving the nitroimidazole tracer pimonidazole.

BACKGROUND: Hypoxia is thought to be an adverse feature of pancreatic cancer, but direct measurement in patients is technically challenging. To address this, we characterised the intra/interpatient heterogeneity of hypoxia in surgical specimens from patients who received the 2-nitroimidazole tracer pimonidazole pre-operatively. METHODS: Pimondazole was given intravenously 16-20 h before pancreatectomy, and the extent and intratumoral heterogeneity of hypoxia determined by image analysis applied to multiple tissue blocks stained by immunohistochemistry. Intra/interpatient heterogeneity was estimated by variance component analysis. RESULTS: Pimonidazole staining was analysed in 10 tumours. The extent of labelling varied amongst patients (0-26%), with a broader range of hypoxia in the epithelial (1-39%) compared with the stromal (1-13%) compartments. Variance component analysis demonstrated greater inter- than intrapatient variability of hypoxia, and that multiple (4-5) tumour sections are required to provide a consistent evaluation of its extent in individual tumours. CONCLUSIONS: There is significant intra- and intertumoral heterogeneity of hypoxia in pancreatic cancers, and these do not appear to be generally more hypoxic than other cancer types. This study establishes the feasibility to assess hypoxia in pancreatic cancer patients using pimonidazole, but questions the reliability of measurements made using a single tissue section.

Efficacy of calcein and Coomassie Blue dyeing of shell growing-edges and micro growth-bands: Ageing juvenile of Pinctada mazatlanica (Pterioida: Pteriidae) / Eficiencia de la tinción de bandas de crecimiento con calceina y azul de coomassie, para estimar la edad de jóvenes de Pinctada mazatlanica (Pterioida: Pteriidae)

Age validation is the first step to determine shellfish species age determination. This information is vital for different inferential models used in marine ecosystem management activities. In spite that various validation techniques are used for marking carbon calcium structures, the calcein marking technique for oysters had never been used for age validation in Pinctada mazatlanica. Thus the objectives of this study included: the evaluation of calcein to mark a shell growing-edge, and the efficacy of Coomassie Blue staining on posterior shell growth, to produce visible micro growth-bands that would enable age validation of juvenile mother-of- pearl oysters. Oysters were collected and cultivated at The Perlas del Cortez S. de R. L. MI. pearl-farming opera tion, in Pichilingue, La Paz Bay, Baja California Sur, Mexico; a total of 36 oysters (shell height 11.5-36.4mm) were injected with calcein (0.125g/L), and another 50 oysters (shell height 14.8-42.7mm) were submersed in calcein (0.4 and 0.7g/L). Shell slices of calcein-marked oysters were posteriourly stained with Coomassie Blue R-25 for micro growth-band recognition. Our results showed that Calcein marking only worked by submersion and produced a concise bright lime-green florescent band along the growing-edge with clear boundaries for both concentrations. However, marks resulted better at the lower calcein concentration (0.4g/L) with more “perfect” and “good” marks on the growing-edge (p=0.0012). Commassie Blue staining technique was successful, and allowed to conclude that one micro growth-band was laid down per day, similar to other oyster species. Mean 15-d increment of shell growth height was slightly greater at the lower calcein concentration ( =0.735mm) than at the higher one ( =0.577mm) (not significant difference, p=0.198). Calcein marking of shell growing- edges and Commassie Blue staining of posterior shell growth, as a method for age validation is recommended for shellfish shell growth-band counts. This will allow back-dating for estimation of very precise colonization dates, both spatially and temporally in future work.

La validación de la edad es el primer paso para determinar las edades de las especies de moluscos, esta información es de vital importancia para los diferentes modelos de inferencia utilizados en actividades de gestión de los ecosistemas marinos. Diversas técnicas de valida- ción se utilizan para marcar estructuras de carbonato de calcio, aunque la técnica de marcado de calceína en ostras nunca se había utilizado para la validación de la edad de P. mazatlanica. Los objetivos de este estudio fueron: evaluar la calceína como marcador interno de la concha y la eficiencia del azul de Coomassie en la tinción de la matriz proteica de la concha, para facilitar la observación y conteo de micro bandas de crecimiento que permiten validar la edad de las ostras juveniles de madre perla. Las ostras fue- ron recolectadas en la costa de la empresa Perlas del Cortez S. de RL MI., en Pichilingue en Bahía de La Paz, Baja California Sur, México. Se inyectaron 36 ostras (altura de concha 11.5-36.4mm) (0.125g/L de calceína) y otras 50 ostras (altura de la concha 14.8-42.7mm) se sumergieron (0.4 y 0.7g/L de calceína). Secciones de la concha marcadas con calceína fueron teñidos con azul de Coomassie R-25 para el reconocimiento de las micro bandas de crecimiento. El marcado con calceína fue exitoso por inmersión y produjo una banda fluorescente de color verde lima brillante con- cisa a lo largo del crecimiento interno de la concha. Sin embargo, las marcas fueron mejores a una concentración de calceína inferior (0.4g/L), con mayor cantidad de marcas “buenas” y “perfectas” (p=0.0012). La técnica de tinción con azul de Commassie también fue exitosa. Se detectó un crecimiento diario por micro banda, similar a lo encontrado en otras especies de ostras. La diferencia del crecimiento medio en relación a la altura de la concha en un lapso de 15 días, fue ligeramente mayor con una concentración de calceína inferior ( =0.735mm) que con la de mayor concentración ( =0.577mm), pero no significativamente (p=0.198). El marcado de conchas con calceína y tinción de matrices proteicas con azul de Coomassie posterior a su crecimiento, es recomendando como un método para la validación de la edad facilitando el conteo de micro bandas de crecimiento internas de la concha. Además, permitirá estimar edades con el fin de predecir fechas de colonización y ubicación de bancos naturales.

A comparison of two visual inspection methods for cervical cancer screening among HIV-infected women in Kenya.

OBJECTIVE: To determine the optimal strategy for cervical cancer screening in women with human immunodeficiency virus (HIV) infection by comparing two strategies: visual inspection of the cervix with acetic acid (VIA) and VIA followed immediately by visual inspection with Lugol's iodine (VIA/VILI) in women with a positive VIA result. METHODS: Data from a cervical cancer screening programme embedded in two HIV clinic sites in western Kenya were evaluated. Women at a central site underwent VIA, while women at a peripheral site underwent VIA/VILI. All women positive for cervical intraepithelial neoplasia grade 2 or worse (CIN 2+) on VIA and/or VILI had a confirmatory colposcopy, with a biopsy if necessary. Overall test positivity, positive predictive value (PPV) and the CIN 2+ detection rate were calculated for the two screening methods, with biopsy being the gold standard. FINDINGS: Between October 2007 and October 2010, 2338 women were screened with VIA and 1124 with VIA/VILI. In the VIA group, 26.4% of the women tested positive for CIN 2+; in the VIA/VILI group, 21.7% tested positive (P < 0.01). Histologically confirmed CIN 2+ was detected in 8.9% and 7.8% (P = 0.27) of women in the VIA and VIA/VILI groups, respectively. The PPV of VIA for biopsy-confirmed CIN 2+ in a single round of screening was 35.2%, compared with 38.2% for VIA/VILI (P = 0.41). CONCLUSION: The absence of any differences between VIA and VIA/VILI in detection rates or PPV for CIN 2+ suggests that VIA, an easy testing procedure, can be used alone as a cervical cancer screening strategy in low-income settings.