Structural differences in gut bacteria communities in developmental stages of natural populations of Lutzomyia evansi from Colombia's Caribbean coast.
Lutzomyia evansi, a phlebotomine insect endemic to Colombia's Caribbean coast, is considered to be the main vector of visceral and cutaneous leishmaniasis in the region. Although insects of this species can harbor pathogenic and non-pathogenic microorganisms in their intestinal microbiota, there is little information available about the diversity of gut bacteria present in Lutzomyia evansi. In this study, conventional microbiological methods and molecular tools were used to assess the composition of bacterial communities associated with Lutzomyia evansi guts in immature and adult stages of natural populations from the department of Sucre (Caribbean coast of Colombia).



Sand flies were collected from two locations (peri-urban and jungle biotype) in the Department of Sucre (Caribbean coast of Colombia). A total of 752 Lutzomyia evansi intestines were dissected. In this study, 125 bacterial strains were isolated from different culture media (LB Agar, MacConkey Agar). Different methods were used for bacterial identification, including ribosomal intergenic spacer analysis (RISA) and analysis of the 16S rRNA and gyrB gene sequences. The genetic profiles of the bacterial populations were generated and temporal temperature gradient gel electrophoresis (TTGE) was used to compare them with total gut DNA. We also used PCR and DNA sequence analysis to determine the presence of Wolbachia endosymbiont bacteria and Leishmania parasites.



The culture-dependent technique showed that the dominant intestinal bacteria isolated belong to Acinetobacter, Enterobacter, Pseudomonas, Ochrobactrum, Shinella and Paenibacillus in the larval stage; Lysobacter, Microbacterium, Streptomyces, Bacillus and Rummeliibacillus in the pupal stage; and Staphylococcus, Streptomyces, Brevibacterium, Acinetobacter, Enterobacter and Pantoea in the adult stage. Statistical analysis revealed significant differences between the fingerprint patterns of the PCR-TTGE bands in bacterial communities from immature and adult stages. Additionally, differences were found in bacterial community structure in fed females, unfed females, males and larvae. The intestinal bacteria detected by PCR-TTGE were Enterobacter cloacae and Bacillus thuringiensis, which were present in different life stages of Lu. evansi, and Burkholderia cenocepacia and Bacillus gibsonii, which were detected only in the larval stage. Wolbachia and Leishmania were not detected in gut samples of Lutzomyia evansi.



The analyses conducted using microbiological and molecular approaches indicated significant variations in the bacterial communities associated with the gut of Lu. evansi, depending on the developmental stage and food source. We propose that these elements affect microbial diversity in L. evansi guts and may in turn influence pathogen transmission to humans bitten by this insect.
Grupo de Investigación en Sistemática Molecular, Universidad Nacional de Colombia, Street 59 A # 63-20, Medellín, Postal Code 050003, Colombia. rajovigo2001@yahoo.com.|PECET (Programa de Estudio y Control de Enfermedades Tropicales), Universidad de Antioquia, Street 62 # 52-59, SIU-Sede de Investigación Universitaria, Laboratory 632, Medellín, Postal Code 050003, Colombia. rajovigo2001@yahoo.com.|Grupo de Microbiodiversidad y Bioprospección, Laboratorio de Biología Celular y Molecular, Universidad Nacional de Colombia sede Medellín, Street 59 A # 63-20, Medellín, Postal Code 050003, Colombia. rajovigo2001@yahoo.com.
Rafael José
RJ
Vivero
Grupo de Microbiodiversidad y Bioprospección, Laboratorio de Biología Celular y Molecular, Universidad Nacional de Colombia sede Medellín, Street 59 A # 63-20, Medellín, Postal Code 050003, Colombia.
Natalia Gil
NG
Jaramillo
Grupo de Microbiodiversidad y Bioprospección, Laboratorio de Biología Celular y Molecular, Universidad Nacional de Colombia sede Medellín, Street 59 A # 63-20, Medellín, Postal Code 050003, Colombia.
Gloria
G
Cadavid-Restrepo
Grupo de Investigación en Sistemática Molecular, Universidad Nacional de Colombia, Street 59 A # 63-20, Medellín, Postal Code 050003, Colombia.|PECET (Programa de Estudio y Control de Enfermedades Tropicales), Universidad de Antioquia, Street 62 # 52-59, SIU-Sede de Investigación Universitaria, Laboratory 632, Medellín, Postal Code 050003, Colombia.
Sandra I Uribe
SI
Soto
Grupo de Microbiodiversidad y Bioprospección, Laboratorio de Biología Celular y Molecular, Universidad Nacional de Colombia sede Medellín, Street 59 A # 63-20, Medellín, Postal Code 050003, Colombia.
Claudia Ximena Moreno
CX
Herrera
D063147:Animal Distribution; D000818:Animals; D001419:Bacteria / Q000145:classification* / Q000302:isolation & purification*; D003105:Colombia; D005260:Female; D041981:Gastrointestinal Tract / Q000382:microbiology*; D008018:Life Cycle Stages; D008297:Male; D011576:Psychodidae / Q000382:microbiology*
Adults; Gut; Immature; Microbiota; Sand flies; Vector