Differential identification of Ascogregarina species (Apicomplexa: Lecudinidae) in Aedes aegypti and Aedes albopictus (Diptera: Culicidae) by polymerase chain reaction.
We report 2 polymerase chain reaction (PCR)-based methods for distinguishing morphologically similar gregarine species based on amplification of variable regions of the internal transcribed spacer region of ribosomal DNA. The gregarines we investigated were Ascogregarina barretti (Vavra), A. culicis (Ross), and A. taiwanensis (Lien and Levine), parasites of the mosquitoes Ochlerotatus triseriatus (Say), Aedes aegypti (Linnaeus), and Ae. albopictus (Skuse), respectively. These 3 important vector mosquitoes often utilize the same container habitats, where larval development and infection by the parasite occurs, leaving ample opportunity for cross-species gregarine infection. Because previous studies have shown that the parasites A. culicis and A. taiwanensis variably affect fitness in both normal and abnormal mosquito hosts, distinguishing parasite infection and species is important. The task is complicated by the fact that these 2 parasite species are virtually identical in morphology, whereas A. barretti is morphologically distinct. Of the 2 PCR-based assays reported here, the first provides a rapid, sensitive, and straight-forward means of general ascogregarine detection based on a single PCR amplification. The second method provides a means of differentiation between A. culicis and A. taiwanensis based on a species-specific PCR assay. Together, these assays allow whole mosquitoes to be tested for the presence of Ascogregarina species as well as identification of both A. culicis and A. taiwanensis singly or in dual infections.
Morales
Maria E
ME
Interdisciplinary Program in Molecular and Cellular Biology, School of Public Health and Tropical Medicine, Tulane University Health Sciences Center, New Orleans, Louisiana, 70112, USA.
Ocampo
Clara B
CB
Cadena
Horacio
H
Copeland
Claudia S
CS
Termini
Michael
M
Wesson
Dawn M
DM
D000330:Aedes / Q000469:parasitology*; D000818:Animals; D016782:Apicomplexa / Q000235:genetics / Q000302:isolation & purification*; D001483:Base Sequence; D016384:Consensus Sequence; D017931:DNA Primers / Q000737:chemistry; D018509:DNA, Helminth / Q000737:chemistry*; D004275:DNA, Ribosomal / Q000737:chemistry*; D021903:DNA, Ribosomal Spacer / Q000737:chemistry; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods*; D012337:RNA, Ribosomal, 18S / Q000235:genetics; D012339:RNA, Ribosomal, 28S / Q000235:genetics; D012340:RNA, Ribosomal, 5.8S / Q000235:genetics; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D016415:Sequence Alignment; D013045:Species Specificity
D017931:DNA Primers; D018509:DNA, Helminth; D004275:DNA, Ribosomal; D021903:DNA, Ribosomal Spacer; D012337:RNA, Ribosomal, 18S; D012339:RNA, Ribosomal, 28S; D012340:RNA, Ribosomal, 5.8S