Salvaging the supernatant: next generation cytopathology for solid tumor mutation profiling.

With the expanding role of targeted therapy in patients with solid tumors, pathologists face the daunting task of having to maximize limited volume tissue obtained by fine needle aspiration for a variety of molecular tests. While most molecular studies on fine needle aspiration samples have been reported using cellular material, recent studies have shown that a substantial amount of DNA can be retrieved from the supernatant fluid of aspirate needle rinses after cell pelleting for cytospin or cell block preparations. In routine clinical workflow, the supernatant is discarded; however this fluid may provide a complementary source of DNA for tumor mutational profiling. In this study, we evaluated the post-centrifuged supernatant from 25 malignant and 10 benign fine needle aspiration needle rinses. The mean and median DNA yields from the supernatants were 445 ng and 176.4 ng (range, 15.1-2958 ng), respectively. Next generation sequencing using the Ion AmpliSeq Cancer Hotspot Panel v2 detected somatic mutations in all 25 malignant samples. No mutations were detected in any of the benign samples tested. When available, mutations detected in the supernatant fluid were compared to the next generation sequencing analysis performed on a prior or concurrent surgical specimen from the same patient and showed 100% concordance. In a subset of cases (n = 19) mutations in EGFR, KRAS, BRAF, PIK3CA, and NRAS were successfully confirmed by droplet digital PCR, providing an orthogonal platform for mutation analysis. In summary, in this study we show that post centrifuged supernatants from fine needle aspiration needle rinses can provide a robust substrate for expanded mutation profiling by next generation sequencing, as well as hotspot mutation testing by droplet digital PCR. The ability to detect somatic mutations from otherwise discarded supernatant fluids offers the ability to triage and effectively utilize limited volume fine needle aspiration samples when multiple molecular tests are requested, without the need to re-biopsy for additional tissue samples.

[Meckel's diverticulitis, an uncommon cause of acute abdomen]. / Diverticulitis de Meckel, una causa poco frecuente de abdomen agudo.

Meckel's diverticulum corresponds to the aberrant involution of the omphalo-mesenteric canal or vitelline duct, which is located at the level of the antimesenteric border of the terminal ileum. It is the most common structural anomaly of the gastrointestinal tract, it is almost always asymptomatic and its diagnosis is usually incidental, however the complication with diverticulitis is an unusual condition. We describe the case of a 65-year-oldman, who was admitted from another institution with a diagnosis of acute abdomen. On physical examination, he presented signs of peritoneal irritation with evidence of leukocytosis and neutrophilia in the admission blood count. Computerized tomography of the abdomen with intra venous contrast was performed, which was interpreted as complicated Meckel's diverticulitis, being corroborated during the surgical act and confirmed by pathological anatomy. Meckel's diverticulitis is a rare entity, however it is important to recognize it within the differential diagnoses of acute abdomen, which will allow prompt intervention and a favorable outcome.

El divertículo de Meckel (DM) corresponde a la involución aberrante del canal onfalo-mesentérico o conducto vitelino, el cual se ubica a nivel del borde antimesentérico del íleon terminal. Es la anomalía estructural más común del tracto gastrointestinal, casi siempre es asintomático y su diagnóstico por lo general es incidental, sin embargo, la complicación con diverticulitis es una condición poco usual. Describimos el caso de un hombre de 65 años, que ingresó referido de otra institución con diagnóstico de abdomen agudo, al examen físico presentó signos de irritación peritoneal con evidencia de leucocitosis y neutrofilia en hemograma de ingreso. Se realizó tomografía computarizada de abdomen con contraste endovenoso, la cual se interpretó como diverticulitis de Meckel complicada, siendo corroborado durante el acto quirúrgico y confirmado mediante anatomía patológica. La diverticulitis de Meckel es una entidad rara, sin embargo, es importante reconocerla dentro de los diagnósticos diferenciales de abdomen agudo, lo cual permitirá una pronta intervención y un favorable desenlace.

Anisakid Nematodes and Potential Risk of Human Anisakiasis through the Consumption of Hake, Merluccius spp., Sold Fresh in Spanish Supermarkets.

Nematode parasite species belonging to the Anisakis simplex complex are the most important cause of human anisakiasis through the consumption of (mainly) undercooked, previously not frozen, or conveniently treated fish. In Spain, the consumption of hake has been recognized as an important source of this parasitosis. With the aim of shedding light on the risk factors that can influence the potential risk of human anisakiasis in Spain through the consumption of fresh hake sold by nationwide supermarket chains, a total of 536 small hake specimens belonging to the species Merluccius bilinearis caught off the Northeast American coasts and Merluccius merluccius caught in the Northeast Atlantic and Mediterranean waters was analysed. Anisakids morphologically identified as Anisakis type I were found as the most prevalent and the most abundant species and were considered the main potential cause of human anisakiasis. Intrinsic and extrinsic factors concerning the hake, such as its origin and season of capture, its size, as well as the days passed between its capture and consumption, should be taken into account to avoid this human parasitosis. It is essential that consumers have access to fish label information which should include, as regulated by the European Commission, traceability data.

Diverticulitis de Meckel, una causa poco frecuente de abdomen agudo / Meckel's diverticulitis, an uncommon cause of acute abdomen

Resumen El divertículo de Meckel (DM) corresponde a la involución aberrante del canal onfalo-mesentérico o conducto vitelino, el cual se ubica a nivel del borde antimesentérico del íleon terminal. Es la anomalía estructural más común del tracto gastrointestinal, casi siempre es asintomático y su diagnóstico por lo general es incidental, sin embargo, la complica ción con diverticulitis es una condición poco usual. Describimos el caso de un hombre de 65 años, que ingresó referido de otra institución con diagnóstico de abdomen agudo, al examen físico presentó signos de irritación peritoneal con evidencia de leucocitosis y neutrofilia en hemograma de ingreso. Se realizó tomografía computarizada de abdomen con contraste endovenoso, la cual se interpretó como diverticulitis de Meckel complicada, siendo corroborado durante el acto quirúrgico y confirmado mediante anatomía patológica. La diverticulitis de Meckel es una entidad rara, sin embargo, es importante reconocerla dentro de los diagnósticos diferenciales de abdomen agudo, lo cual permitirá una pronta intervención y un favorable desenlace.

Abstract Meckel's diverticulum corresponds to the aberrant invo lution of the omphalo-mesenteric canal or vitelline duct, which is located at the level of the antimesenteric border of the terminal ileum. It is the most common structural anomaly of the gastrointestinal tract, it is almost always asymptomatic and its diagnosis is usually incidental, how ever the complication with diverticulitis is an unusual con dition. We describe the case of a 65-year-oldman, who was admitted from another institution with a diagnosis of acute abdomen. On physical examination, he presented signs of peritoneal irritation with evidence of leukocytosis and neutrophilia in the admission blood count. Computerized tomography of the abdomen with intra venous contrast was performed, which was interpreted as complicated Meckel's diverticulitis, being corroborated during the surgical act and confirmed by pathological anatomy. Meckel's diverticulitis is a rare entity, however it is important to recognize it within the differential diagnoses of acute abdomen, which will allow prompt intervention and a favorable outcome.

Trypanosoma cruzi en donantes que acuden al banco de sangre "Dr. Julio García Álvarez" del hospital Dr. Luis Razetti, estado Barinas, Venezuela / Trypanosoma cruzi in donors who go to the "Dr. Julio Garcia Alvarez" Blood Bank of the Dr. Luis Razetti Hospital, in the Barinas city, Venezuela

Trypanosoma cruzi is the causative agent of Chagas disease; this is considered one of the main parasitosis in Latin America. Blood transfusions occupy the second place of transmission of this parasite, after vector transmission and usually occur in urban areas through infected donors from endemic rural areas, generally asymptomatic and ignorant of their condition. In Venezuela, Barinas state is recognized as an important endemic area of T. cruzi transmission. In this sense, was to evaluate parasitological and serological techniques in donors who go to the "Dr. Julio Garcia Alvarez" Blood Bank in the Barinas city. To this end, 240 blood samples were collected from an equal number of donors and were assessed with fresh examination techniques, capillary microcentrifugation, colored smears, and ELISA anti-T. cruzi IgG. In none of the samples circulating parasites were observed, while 3 donors resulted positive with ELISA anti-T. cruzi IgG (1.25% seroprevalence). Direct parasitological tests were not effective for the determination of T. cruzi because there was no evidence of blood trypomastigotes in any of the cases studied

Infecciones helmínticas intestinales diagnosticadas en el ambulatorio urbano Los Curos, Mérida / Intestinal helminth infections diagnosed in the ambulatory urban Los Curos, Merida

Las infecciones helmínticas intestinales que afectan al hombre están íntimamente ligadas a condiciones ecológicas, humanas y/o sociales que pueden favorecer o no su persistencia. El objetivo del presente trabajo fue presentar la casuística de las infecciones helmínticas intestinales que se diagnosticaron en el Laboratorio Clínico Asistencial del Ambulatorio Urbano Tipo III Los Curos entre los años 2012 a 2015. Para tal fin se realizó una investigación retrospectiva donde se estudiaron para el período evaluado los registros del laboratorio. Los resultados evidenciaron 61,7% de individuos parasitadas, 1,9% con algún helminto (Ascaris lumbricoides, Trichuris trichiura, Strongyloides stercoralis, Enterobius vermicularis, Hymenolepis nana o Ancylostomideos). No se evidenció diferencia entre la presencia de helmintos y los parámetros demográficos edad o género. Se concluye que las condiciones ecoepidemiológicas de Los Curos no son lo suficientemente aptas para el mantenimiento de infecciones helmínticas intestinales.

The intestinal helminth infections that affect man are intimately linked to ecological, human and / or social conditions that may or not allow their persistence. The objective of the present study was to present the casuistry of the intestinal helminth infections that were diagnosed in the Clinical Laboratory of the Ambulatory Urban III Los Curos between the years 2012 to 2015. For that purpose, a retrospective investigation was carried out where they were studied for the period laboratory records were evaluated. The results evidenced 61.7% of parasitized individuals, 1.9% with some helminth (Ascaris lumbricoides, Trichuris trichiura, Strongyloides stercoralis, Enterobius vermicularis, Hymenolepis nana or Ancylostomideos). There was no difference between the presence of helminths and the demographic parameters age or sex. It is concluded that the ecoepidemiological conditions of Los Curos are not sufficiently adequate for the maintenance of intestinal helminth infections.

Detección de transmisión transplacentaria de Anaplasma Marginale en bovinos asintomáticos / Detection of Anaplasma marginale Transplacental Transmission in Asymptomatic Cattle / The presence of Anaplasma marginale in calves descendants from cows with asymptomatic infections, was detected. Polymerase chain reaction (PCR) assays carried out in blood samples from 31 healthy cows and their respective calves, revealed the presence of A. marginale-specific DNA in 70 and 40% of them, respectively. The detection of a portion of A. marginale genome in newborn calves, suggests a transplacental mother-offspring transmission. The epizootiological implications of A. marginale-infection in asymptomatic animals and the potential of transplacental transmission in the maintenance of A. marginale were considered. Results lead to conclude that A. marginale transplacental transmission might be of frequent occurrence in the study area, so more specific techniques, such as PCR, are suggested for a more thorough diagnosis.

Se detecta la presencia de Anaplasma marginale en becerros hijos de vacas con infección asintomática. Ensayos de reacción en cadena de la polimerasa (PCR) llevados a cabo en muestras de sangre de 31 vacas lactantes, en aparente buen estado físico y sus respectivas crías, revelaron la presencia de ADN específico de A. marginale en el 70 y 40%, respectivamente. La detección de parte del genoma de A. marginale en becerros recién nacidos, hijos de vacas asintomáticas PCR positivas, sugiere la transmisión de la infección madre-hijo por vía transplacentaria. Se discute las implicaciones epizootiológicas de la infección por A. marginale en animales asintomáticos y se advierte sobre la potencialidad de esta forma de transmisión en el mantenimiento del ciclo de este organismo. Se concluye que la transmisión por vía transplacentaria podría ser un evento de frecuente ocurrencia en el área de estudio y se sugiere la utilización de técnicas producto de la nueva biotecnología como la PCR para incrementar la especificidad en el diagnóstico.

Uso de proteínas como alternativa diagnóstica para discriminar infecciones entre Trypanosoma vivax y Trypanosoma evansi / Use of proteins as diagnostic tool to discriminate between Trypanosoma vivax and Trypanosoma evansi infections

Tripomastigotes sanguícolas de Trypanosoma vivax y Trypanosoma evansi obtenidos de infecciones experimentales en ovejas y ratones respectivamente, fueron utilizados para purificar y caracterizar proteínas citosólicas mediante el método de partición con Tritón X-114. Los resultados revelan diferencias en los patrones proteicos entre las dos especies. Asimismo, las reacciones antigénicas mediante Western blot utilizando suero de animales naturalmente infectados, permitió discriminar las infecciones entre ambos parásitos. Se sugiere la utilización de esta metodología como una prueba diagnóstica confiable.

Detección de infecciones subclínicas por Trypanosoma vivax en bovinos de fincas ganaderas de Mérida Venezuela / Detection of Trypanosoma vivax subclinic infections in cattle from Mérida Venezuela

Se registra la detección de infecciones subclínicas por Trypanosoma vivax en bovinos asintomáticos de fincas ganaderas de Mérida, Venezuela. El diagnóstico fue realizado utilizando métodos parasitológicos (examen microscópico de muestra fresca, micro-capilar y láminas coloreadas); bioquímicos (Western blot) y moleculares (PCR) en muestras sanguíneas. Se advierte sobre el riesgo potencial que infecciones, ocultas o inaparentes pudieran representar para los rebaños bovinos. Se discute su significado y su importancia epizootológica para regiones ganaderas del país.