USO DE MICROORGANISMOS NUTRICIONALES EN CONTROL BIOLOGICO DE MOSQUITOS DE IMPORTANCIA SANITARIA

DIAZ NIETO LM; PEROTTI MA; BERÓN CM

Mar del Plata

Congreso; X Congreso Argentino de Microbiología General SAMIGE 2014; 2014

Sociedad Argentina de Microbiología General

During the last decades, there has been a growing interest in symbionts and nutritional microorganisms of mosquitoes and other disease vectors, with the aim of searching for novel methods of control. Some pathogenic bacteria are able to produce proteins toxic to mosquitoes, and therefore these are being used for the control of this insectcs. By transforming free-living organisms adapted to mosquito`s environmental conditions, it is possible to express bacterial mosquitocidal toxins, and obtain more stability of these toxins in the wild. In a previous work, we evaluated the use of different microorganisms (cyanobacteria, algae, yeasts and intestinal bacteria of mosquito) as natural food of Culex pipiens larvae. Of the four microorganisms used, only yeasts allowed mosquitoes to complete development until adult stage. We followed the occurrence of fluorescent marked yeast by binding Saccharomyces cerevisiae with green fluorescent protein (GFP). S. cerevisiae GFP+ was screened from different stages of mosquito development. Mosquito`s four larval instars, pupae, adult as well as offspring eggs (F1) were screened for yeasts by its growth in appropriate culture media, by experiments of fluorescence microscopy detecting GFP+ and molecular methodologies such as PCR amplification of the GFP gene and of the yeast 18S rRNA with generic primers, and we only detected fluorescent yeast until fourth instar larvae and GFP gene until adult females. F1 eggs did not indicate the presence of yeasts (not detected by PCR). However, we still obtained some microorganisms that were grown in a suitable culture medium. The aim of this work was to identify those microorganisms and analyse if they are inoculated by the mosquito female in the eggs in order to provide the first nutrients for the neonate larvae. The sequences obtained by the amplification of the DNA fragments with specific and degenerate 18S rDNA primers for yeast did not allow the identification of any yeasts, but by the amplification of the 16S rDNA gene of these microorganisms let us recognize bacteria belonging to the genus Acinetobacter and Klebsiella. In addition, we detected both bacteria genera in mosquito guts, by the amplification of 16S rDNA fragments. In order to detect if these bacteria were inoculated in the eggs by the mosquito female, we reared mosquitoes in sterile and controlled conditions, to eliminate the possibility of the contamination of the eggs by environmental bacteria. However, we could only confirm the presence of both bacteria genera in laid eggs. Further experiments using other rearing techniques should be performed to confirm if these bacteria are inoculated by females, and thus determine whether these organisms could be used in the biological control programs of mosquito populations. This can be carried out by elimination of fundamental bacteria in mosquito nutrition present in the guts, or by expression of mosquitocidal toxins in those microorganisms.