Abstract INTRODUCTION: Insect cell cultures play an essential role in understanding arboviral replication. However, the replicative efficiency of some of these viruses such as dengue (DENV), yellow fever (YFV), and chikungunya (CHIKV) in a new cellular substrate (Lulo) and in the other two recognized cell lines has not been comparatively assessed. METHODS: Vero, C6/36, and Lulo cell lines were infected with DENV, YFV, and CHIKV. The viral progeny was quantified through plaque assays and quantitative reverse transcription-polymerase chain reaction, while for DENV2, the findings were confirmed by immunofluorescence antibody assay. RESULTS: The higher DENV2 titer (from multiplicity of infection 0.001) was obtained on day four post-infection in C6/36 and on day six in Vero cells, while the Lulo cell line was almost impossible to infect under the same conditions. However, C6/36 showed the highest values of viral RNA production compared to Vero cells, while the quantification of the viral RNA in Lulo cells showed high levels of viral genomes, which had no correlation to the infectious viral particles. CONCLUSIONS: C6/36 was the most efficient cell line in the alpha and flavivirus production, followed by Vero cells. Thus, Lulo cells may be a useful substrate to study the mechanisms by which cells evade viral replication.

Insect cell cultures are an important biotechnological tool for basic and applied studies. The objective of this work was to establish and characterise a new cell line from Culex quinquefasciatus embryonic tissues. Embryonated eggs were taken as a source of tissue to make explants that were seeded in L-15, Grace's, Grace's/L-15, MM/VP12, Schneider's and DMEM culture media with a pH range from 6.7-6.9 and incubated at 28ºC. The morphological, cytogenetic, biochemical and molecular characteristics of the cell cultures were examined by observing the cell shapes, obtaining the karyotypes, using a cellulose-acetate electrophoretic system and performing random amplified polymorphic DNA-polymerase chain reaction analysis, respectively. The Grace's/L-15 medium provided the optimal nutritional conditions for cell adhesion and proliferation. Approximately 40-60 days following the explant procedure, a confluent monolayer was formed. Cellular morphology in the primary cultures and the subcultures was heterogeneous, but in the monolayer the epithelioid morphology type predominated. A karyotype with a diploid number of six chromosomes (2n = 6) was observed. Isoenzymatic and molecular patterns of the mosquito cell cultures matched those obtained from the immature and adult forms of the same species. Eighteen subcultures were generated. These cell cultures potentially constitute a useful tool for use in biomedical applications.

Introducción: Lucilia sericata es una especie de importancia médica y forense, utilizada en terapia larval para curar heridas crónicas y en estudios médico-legales empleada en la estimación del intervalo post mórtem y el traslado de cadáveres. No existen registros de las características citogenéticas de esta mosca en el neotrópico. El objetivo principal de este trabajo fue identificar las características morfométricas cromosómicas y las estructuras primarias del cariotipo, a partir de especímenes de L. sericata de la cepa Bogotá, Colombia. Materiales y métodos: Se tomaron huevos embrionados, que fueron previamente esterilizados en su superficie, se maceraron y luego fueron sembrados en el medio de cultivo L-15, suplementado con 20% de SFB, e incubados a una temperatura de 28 °C, sin atmosfera de C0(2). La preparación de los cromosomas se obtuvo de monocapas celulares semiconfluentes, empleando diversas soluciones: antimitótica (Colchicina), hipotónica (KCl 0,075 M) y fijadora (Carnoy: metanol y ácido acético; 3:1). Se llevó a cabo la técnica de bandeo C para la identificación de regiones cromosómicas de heterocromatina constitutiva. Resultados: Se obtuvieron parámetros morfométricos de cada par cromosómico. El número diploide del cariotipo obtenido de los cultivos celulares fue 2n = 12; éstos se clasificaron morfológicamente, de acuerdo con patrones previamente establecidos, así: los pares I, II, IV y V fueron metacéntricos, y el par III fue submetacéntrico. A su vez, el par sexual fue heteromórfico, siendo el cromosoma X metacéntrico y el cromosoma Y submetacéntrico. El bandeo C fue positivo para todos los pares cromosómicos. Conclusiones: Se establecieron las características citogenéticas de L. sericata, cepa Bogotá, Colombia, relacionadas con número, forma, tamaño, posición del centrómero y regiones heterocromáticas de los cromosomas.

Objective: Lucilia sericata is an important species for medical and forensic purposes, it is used in maggot therapy in the treatment of chronic wounds and in medical-legal studies for establishing the post-mortem interval and the transfer of corpses. Currently there are no records of the cytogenetic characteristics of this fly in Neotropical region. The main objective of this study was to identify morphometric characteristics and primary structures from karyotype of L. sericata strain Bogota, Colombia. Methods and materials: Embryonated eggs were taken, which were previously surface sterilized, macerated and then seeded in L-15 medium culture, supplemented with 20% FBS and incubated at 28 °C, without C0(2) atmosphere. The preparation of chromosomes was obtained from semiconfluent monolayers, pretreated with various solutions: antimitotic (Colchicine), hypotonic (KCl 0.075 M) and fixative (Carnoy, methanol and acetic acid, 3:1). C-banding technique was carried out to identify chromosomal regions of constitutive heterochromatin. Results: Morphometric parameters were obtained from each pair of chromosomes. The diploid karyotype number obtained from cell cultures was 2n = 12; they were classified morphologically, according to patterns established previously, as follows: pairs I, II, IV and V were metacentric and pair III was submetacentric. On the other hand, the sexual pair was heteromorphic, being X chromosome metacentric and Y chromosome submetacentric. C banding was positive for all chromosome pairs. Conclusions: The cytogenetic characteristics of L. sericata, strain Bogotá, were established according to number, shape, centromer position and heterochromatic regions.

Introdução: Lucilia sericata é uma espécie de importência médica e forense, utilizada em terapia larval para curar feridas crônicas e em estudos médico-legais empregada na estimação do intervalo post morteme o traslado de cadáveres. Não existem registros das características citogéneticas desta mosca no neotrópico. O objetivo principal deste trabalho foi identificar as características morfométricas cromossômicas e as estruturas primárias do cariótipo, a partir de especímenes de L. sericata da cepa Bogotá, Colômbia. Materiais e métodos: Tomaram-se ovos embrionados, que foram previamente esterilizados em sua superfície, se maceraram e depois foram semeados no médio de cultivo L-15, suplementado com 20% de SFB, e incubados a uma temperatura de 28 °C, sem atmosfera de C02. A preparação dos cromossomas obteve-se de monocamadas celulares semiconfluentes, utilizando diversas soluções: antimitótica (Colchicina), hipotônica (KCl 0,075 M) e fixadora (Carnoy: metanol y ácido acético; 3:1). Levou-se a cabo a técnica de bandas C para a identificação de regiões cromossômicas de heterocromatina constitutiva. Resultados: Se obtiveram parâmetros morfométricos de cada par cromossômico. O número diplóide do cariótipo obtido dos cultivos celulares foi 2n = 12; estes se classificaram morfologicamente, de acordo com patrões previamente estabelecidos, assim: os pares I, II, IV e V foram metacêntricos, e o par III foi submetacéntrico. Por sua vez, o par sexual foi heteromórfico, sendo o cromossoma X me-tacêntrico e o cromossoma Y submetacêntrico. As bandas C foram positivas para todos os pares cromossômicos. Conclusões: Se estabeleceram as características citogenéticas de L. sericata, cepa Bogotá, Colômbia, relacionadas com número, forma, tamanho, posição do centrômero e regiões heterocromáticas dos cromossomas.

The objective of this work was to establish, under experimental laboratory conditions, a colony of Lucilia sericata, Bogotá-Colombia strain, to build life tables and evaluate two artifcial diets. This blowfy is frequently used in postmortem interval studies and in injury treatment. The parental adult insects collected in Bogotá were maintained in cages at 22°C±1 average temperature, 60%±5 relative humidity and 12 h photoperiodicity. The blowfies were fed on two artifcial diets that were evaluated over seven continuous generations. Reproductive and population parameters were assessed. The life cycle of the species was expressed in the number of days of the different stages: egg = 0.8±0.1, larvae I = 1.1±0.02, larvae II = 1.94±0.16, larvae III = 3.5±0.54, pupae = 6.55±0.47, male adult = 28.7±0.83 and female adult = 33.5±1.0. Total survival from egg stage to adult stage was 91.2% for diet 1, while for diet 2 this parameter was 40.5%. The lifetime reproductive output was 184.51±11.2 eggs per female. The population parameters, as well as the reproductive output of the blowfies that were assessed, showed relatively high values, giving evidence of the continuous increase of the strain over the different generations and making possible its maintenance as a stable colony that has lasted for more than two years.

La terapia larval es una técnica que permite aplicar las larvas de algunas especies de dípteros sobre heridas crónicas o sobre focos de infección localizados, con la finalidad de restaurar el tejido afectado. La velocidad y la eficacia del tratamiento, sumadas a la casi nula pérdida de tejido sano durante el proceso de reparación tisular, han hecho de la terapia larval o biocirugía una alternativa viable para la curación de heridas asociadas con entidades como pie diabético, úlceras venosas, úlceras crónicas de la piel y quemaduras, así como de ciertos tipos de tumores benignos, abscesos y osteomielitis. Aunque la medicina moderna ha sido, en muchos casos, reticente a la aplicación de terapias de esta índole, eventos como la resistencia a los antibióticos y las alteraciones en el proceso de cicatrización en las heridas crónicas han permitido modificar la posición inicial de muchos médicos al respecto.

Larval therapy is a technique that allows the use of larva of some diptera species on chronic wounds or localized infected focuses, in order to promote the affected tissue restoration. The treatment velocity and efficacy, with almost no healthy tissue loses during the process, have made larval therapy (sometimes called biosurgery) a viable alternative for healing wounds associated with entities as diabetic foot, venous ulcers, cutaneous chronic ulcers and burns, and some benign tumors, abscesses and osteomyelitis. Although modern medicine has been, in many cases, reticent on the application of this kind of therapies, events as antibiotic resistance and alterations in cicatrization process in chronic wounds, have modified the respective initial point of view of many doctors.