Melting curve analysis for detection and identification of ghost parasitoids in host carcasses a month after host death-fluorescence data

1. Incidence of parasitism is often underestimated because ‘ghost’
parasitoids (deadnunemerged parasitoids or those that have emerged leaving
the host carcasses) are difficult to detect and identify. This study
demonstrates that the use of melting curve analysis (MCA) of host
carcasses can detect and identify DNA of ghost parasitoids even a month
after host death. 2. The coccinellid hosts Cycloneda sanguinea, Eriopis
connexa, Harmonia axyridis and Hippodamia convergens were sampled
from cole crops in 2017 and 2018 in the Midwest of Brazil, and reared and
observed daily for parasitoid emergence. Dead coccinellids were held for
30 days after death before storage and only host carcasses with parasitoid
emergence observed were analysed. Species-specific primers were designed
for the identification of the parasitoid species that emerged during host
rearing: Dinocampus coccinellae, Homalotylus mirabilis, Ho.
terminalis, Strongygaster triangulifera and Phalacrotophora sp.
The melting temperatures (Tm) of their amplicons were used as positive
controls in MCA post-amplification in qPCR. 3. Detection of parasitoid DNA
in host carcasses was possible for D. coccinellae, Ho. mirabilis and
Phalacrotophora sp. with a limit of detection (LOD) for all the
parasitoids <1 pg of DNA, except for Phalacroptopthora sp. (LOD =
1.9 ng). Parasitoids were detected in 31 out of 70 host carcasses (44.3%)
with detection ranging from 0% to 71%, depending on the parasitoid
species. 4. Synthesis and applications. This work demonstrates that MCA
could be used to detect and rapidly identify the DNA of some parasitoid
species (e.g. Ho. mirabilis, D. coccinellae and Phalacrotophora
sp.) in host carcasses up to a month after parasitoid emergence with high
sensitivity and specificity. MCA can be used on any field-collected host
and, for some parasitoid species, it should lead to more accurate
estimates of the incidence of parasitism. Our approach can be easily
adapted and applied to study other parasitoid–host interactions.