Table 1.
Primer sequences used in this study.
Table 2.
Primer combinations used in this study.
Fig 1.
Blood meal scoring scheme, modified from Detinova [34], to estimate the extent of digestion in blood fed mosquitoes.
Fresh blood meals were categorized as BF1, and were characterized by a large size, bright red color, and the absence of developing ovaries. Blood meals categorized as BF2 were intermediate in digestion extent, and had darker red blood meals with developing ovaries not taking up more than 50% of the abdomen. Blood meals categorized as BF3 were in later stages of digestion, brown in color, and had ovaries that took up more than 50% of the abdomen. This is illustrated with blood fed Aedes aegypti killed between 0 and 72 h post-feeding.
Table 3.
Amplification success of vertebrate host mosquito only DNA templates and no DNA controls.
Fig 2.
Mean (±SEM) DNA concentration (nM) of PCR products, measured by Qubit fluorometer, of the six template categories (Mammalia, Aves, Reptilia, Amphibia, Mosquito, negative control [No DNA]) by three primer combinations (Mod_RepCOI_F + Mod_RepCOI_R, VertCOI_7194_F + Mod_RepCOI_R, Mod_RepCOI_F + VertCOI_7216_R).
Tukey’s HSD test detected pairwise differences between mean DNA concentration for each primer combination and host class. Significant differences between groups are indicated by letters. Mean DNA concentration of groups that have the same letter are not significantly different from each other, while mean DNA concentration of groups that do not share a common letter are significantly different. Comparisons were considered significant if P < 0.05. Error bars represent standard error of the mean.
Table 4.
Results of fully crossed analysis of variance (ANOVA).
Table 5.
Results of blood meal analysis performed using newly designed cytochrome c subunit I primers.
Table 6.
Summary of results and factors for consideration in designing a hierarchical approach to PCR using three combinations of new vertebrate-specific cytochrome c oxidase subunit I primers.