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The process of differentiation of Trypanosoma cruzi in the triatomine’s intestine involves interactions that can be influenced by the morphology of the insect gut or by molecules such as proteins and glycoproteins; and it has been observed that virulence of isolates is related to vector species. Therefore the aim of this study was to compare the morphology and profile of proteins and glycoproteins middle and posterior intestine of male and female of Meccus pallidipennis and Triatoma barberi. Histological analysis of intestines stained with PAS was performed by light microscopy; and the electrophoretic pattern of SDS-PAGE and lectin affinity for recognition by Western blot residue N-acetyl-D-glucosamine, N-acetyl-D-galactosamine and a -2,3-sialic acid was observed. Morphological differences between middle and posterior intestine were observed in all experimental conditions, most important being the presence of intracellular PAS positive granules and a PAS positive extracellular secretion in the gut epithelium; components 61 and 64 kDa were identified in the electrophoretic pattern with residues of N-acetyl-D-glucosamine, which in previous studies have been associated with the digestion of the insect. Another hand, a component of 46kDa was identified with residues of N-acetyl-D-galactosamine, which had been previously shown to be involved in the process intestinal-parasite recognition by vector. The observed differences in morphology and the electrophoretic pattern of glycoproteins may influence the development of Trypanosoma cruzi in the intestinal tract of the insect.
ABSTRACT
OBJECTIVES
REFERENCES• Albuquerque, J. M., Gonzalez, M. S., Garcia, E. S., Mello, C. B., Azambuja, P., Almeida, J. C.,
de Souza, W., Nogueira, N. F. S. (2009). Cytochemical characterization of microvillar and perimicrovillar membranes in the posterior midgut epithelium of Rhodnius prolixus. Arthropod Structure & Development, 38, 31–44.
• Alves, C. R., Albuquerque-Cunha, J. M., Mello, C. B., Garcia, E. S., Nogueira, N. F., Bourguingnon, S. C., de Souza, W., Azambuja, P., Gonzalez, M. S. (2007). Trypanosoma cruzi: attachment to perimicrovillar membrane glycoproteins of Rhodnius prolixus. Experimental Parasitology, 116, 44–52.
• Rivas, E. (2014). Comparación del perfil de glicoproteínas presentes en el intestino medio y posterior de Meccus pallidipennis y Triatoma barberi. Tesis, Facultad de Química, UNAM.
• To identify macroscopic and microscopic differences in the intestine by light microscopy.
• To identify proteins of the triatominae gut by SDS-PAGE.
• To identify glycoproteins using WGA, PNA and MAA by Western blot.
(1) Microbiología y Parasitología, Facultad de Medicina, UNAM, Distrito Federal, México. (2) Laboratorio de Inmunología Comparada de Piel y Mucosas, Facultad de Medicina, UNAM, Distrito Federal, México. (3) Centro de Investigación, UNAM-UABJO, Facultad de
Odontologia, UABJO, Oaxaca, México.
Profile comparison of glycoprotein's present in the middle and posterior intestine of Meccus pallidipennis and Triatoma barberi
Elizabeth Jaqueline Rivas Medina (1), Margarita Cabrera Bravo (1), Gloria Rojas Wastavino (1), Mauro Vences Blanco (1) , Pérez Torres Armando (2), Carlos Josué Solorzano Mata (3), Paz-
María Salazar-Schettino (1).
MATERIALS AND METHODS
Experiments were divided into two sections: histological characterization of middle and posterior intestine in different feeding condition’s, and protein and glycoprotein analysis.
For the fist section, intestines of adults of Meccus pallidipennis and Triatoma barberi were dissected and processed to obtain histological sections which were stained with hematoxilin-eosin and Periodic Acid- Schiff.
For the second section, intestines of adults of Meccus pallidipennis and Triatoma barberi were dissected in order to made electrophoresis and Western blot, to identify proteins and glycoproteins.
Histological characterization.
Based in morphology of its epithelium, the triatominae’s intestine was divided into three main sections: anterior middle intestine, posterior middle intestine and rectum. Anterior and posterior middle intestine had pseudostratified columnar epithelium while rectum had simple cuboidal epithelium.
Main differences observed were epithelium thickness, PAS positive cytoplasmic granules, PAS positive extracellular secretion and membrane folding with different deepness (Figures 1 and 2).
Protein profile.
Protein profile of middle and posterior intestine was similar between each specie, there were three components of 64, 61 and 14 kDa, which were relevant in both species and showed differences in his expression on middle and posterior intestine.
Glycoprotein’s identification.
Three glycoproteins with residues of N-acetylglucosamine and N-acetylgalactosamine were recognised by WGA and PNA lectins on the intestine of males and females of Meccus pallidipennis and Triatoma barberi (Figure 3), nevertheless there wasn’t recognition of any residue of a-2,3-sialic acid by MAA lectin.
• Morphological differences were observed in the intestine of male and female of Meccus pallidipennis and Triatoma barberi in different meal conditions.
• Cytoplasmatic PAS positive granules and PAS positive extracelular secretion were more remarkable on the rectum of Triatoma barberi female.
• Two components of 61 and 64 kDa were observed in the electrophoretic pattern, which also presented N-acetylglucosamine residues recognised by WGA. Those components had been showed importance on digestion process in other triatominae species.
• A component of 46 kDa was identified in the protein profile and showed N-acetylgalactosamine residues recognised by PNA. That component had been related to the linkeage between Trypanosoma cruzi and its vectors.
• MAA didn’t recognise any component with a-2,3-sialic acid residues.
ACKNOWLEDGMENTS AND CONTACTThis project was made with the financial support of DGAPA (PAPIIT) UNAM.MX IN211613.
Acknowledgements to Biol. Irma Elena López Martínez and Biol. Ivonne Sánchez Cervantes, of Laboratorio de Inmunología Comparada de Piel y Mucosas from Facultad de Medicina, UNAM; for their support with the histological techniques.
Contact.
[email protected] and [email protected]
CONCLUSIONS
RESULTS
Figure 1. Microphohography of intestine of triatominae, stained with hematoxylin-
eosine.
Figure 2. Microphohography of intestine of triatominae, stained with PASS.
64 kDa 61 kDa 46 kDaIM: Middle intestineIP: posterior intestine
Figure 3. Western-blot with WGA and PNA lectins.
WGA PNA
