A new test offers an improved PCR-based method for the identification of human-, nonhuman mammal-, and bird-derived mosquito blood meals.
West Nile virus (WNV), malaria, yellow fever, and Zika are some mosquito-borne diseases that affect hundreds of millions of people each year. The identification of host feeding sources and preferences are essential for studies into the dynamics of transmission of mosquito-borne diseases.
An underlying issue is that a mosquito can feed upon multiple hosts, making the identification of host feeding source problematic. PCR-based methods, such as multiplex PCR, provides the most robust host identification allowing a clear distinction between humans and other nonhuman mammalian blood meals. Yet for viruses such as WNV that use birds as their natural host, a multiplex PCR method becomes a two-step process. A second step includes the use of a single primer assay to examine bird blood, creating a costly and time-consuming sequence identification process.
In a paper published in the Journal of Medical Entomology, scientists at Iowa State University have developed a multiplex PCR assay able to distinguish human-, nonhuman mammal-, or bird-derived blood meals from a single PCR reaction. The new assay enables the accurate distinction between potential blood meal hosts, more specifically the addition of bird species, without cross-contamination and eliminates the need for costly sequence identification.
In brief, the authors designed a multiplex PCR assay to target mitochondrial cytochrome b sequences for the identification of human, bird, and nonhuman mammalian host blood meals. The primer set consisted of a universal reverse primer that could amplify across all species and a forward primer that was specific for each species group. Each primer set was explicitly designed to get a PCR amplicon that varied in size but correlated to its respective group; nonhuman mammals (386 bp), human (216 bp), and bird (518 bp), thus eliminating the need for group sequence identification.
To show the effectiveness of this assay, the authors tested their primers on whole blood samples taken from a human, bird (tree sparrow), and nonhuman mammal (sheep). Each sample produced distinguishable bands of 216, 518, or 386 bp, respectively. Positive bands were also seen when the assay was applied across multiple nonhuman mammalian species (386 bp), including cat, cow, dog, mouse pig, and sheep, and different bird species (518 bp). Furthermore, the assay produced positive results for field-caught mosquitoes that had previously been examined for known blood meal host sources.
According to the authors, this new multiplex PCR assay offers an improved method for the identification of mosquito blood meals, enabling researchers to determine the blood host quickly using different sized amplicons for human, bird, and nonhuman mammals from a single PCR reaction.
Human whole blood samples used in this study were supplied by StemExpress.
Field EN et al. (2019) An Improved Multiplex Polymerase Chain Reaction (PCR) Assay for the Identification of Mosquito (Diptera: Culicidae) Blood Meals. J Med Entomol. Doi: 10.1093/jme/tjz182.