Biology Reference
In-Depth Information
likely caused by contamination by minute amounts of
Wolbachia
DNA on
M. occidentalis
surfaces or by
undigested, prey-derived
Wolbachia
present in the guts due to insufficient starvation periods used in prior
studies (Wu and Hoy 2012a). Furthermore, when a strain of
M. occidentalis
was discovered to lack
Cardinium
naturally and the appropriate crosses were conducted, reproductive incompatibility was
observed, indicating that
Cardinium
was the likely agent of the incompatibility rather than
Wolbachia
(Wu and Hoy 2012b).
The data illustrate several points: predators can retain prey (and the prey symbiont DNA) in their guts for a
long time, resulting in false positives in their predator. Thus, when studying symbionts that appear to occur
both in predator and prey (and parasitoid and host?), a long series of starvation times should be tested
using sensitive PCR protocols. Because WGA and high-fidelity PCR are so sensitive (able to detect as few
as one or two DNA molecules), PCR can pick up very small amounts of contamination. The false-positive
egg data probably are due to external contamination of the eggs by
T. urticae Wolbachia
DNA. Resolving the
sensitivity of your PCRs with sensitivity analyses (PCR amplification of cloned gene fragments mixed with
arthropod DNA that has been serially diluted) will clarify just how sensitive your PCR is (Jeyaprakash and
Hoy 2000). Failure to know about the presence of other putative symbionts, however, also can result in
erroneous conclusions. Surveys for symbionts in both predator and prey (or host and parasitoids) should
be conducted so that function(s) of the symbionts can be attributed correctly. For example, Plantard et al.
(2012) recently found that
Wolbachia
thought to occur in a tick was actually in a parasitoid of the tick.
Finally, external decontamination of predators should be conducted to reduce false positives.
References Cited
Agusti, N., de Vicente, M.C., and Gabarra, R., (2000). Developing SCAR markers to study predation
on
Trialeurodes vaporariorum
.
Insect Mol. Biol.
9
: 263-268.
Aljanabi, S.M., and Martinez, I., (1997). Universal and rapid salt-extraction of high quality genomic
DNA for PCR-based techniques.
Nucleic Acids Res.
25
: 4692-4693.
Al-Soud, W.A., and Radstrom, P., (1998). Capacity of nine thermostable DNA polymerases to
mediate DNA amplification in the presence of PCR-inhibiting samples.
Appl. Environ.
Microbiol.
64
: 3748-3753.
Anderson, J.F., Andreadis, T.G., Vossbrinck, C.R., Tirrell, S., Wakem, E.M., and French, R.A.,
et al. (1999). Isolation of West Nile virus from mosquitoes, crows and a Cooper's hawk in
Connecticut.
Science
286
: 2331-2333.
Aransay, A.M., Scoulica, E., and Tselentis, Y., (2000). Detection and identification of
Leishmania
DNA within naturally infected sand flies by seminested PCR on minicircle kinetoplastic DNA.
Appl. Environ. Microbiol.
66
: 1933-1938.
Arnheim, N., and Erlich, H., (1992). Polymerase chain reaction strategy.
Annu. Rev. Biochem.
61
:
131-156.
Arnheim, N., White, T., and Rainey, W.E., (1990). Application of PCR: organismal and population
biology.
BioScience
40
: 174-182.
Asgari, S., Hardy, J.R.E., Sinclair, R.G., and Cooke, B.D., (1998). Field evidence for mechanical
transmission of rabbit haemorrhagic disease virus (RHDV) by flies (Diptera: Calliphoridae)
among wild rabbits in Australia.
Virus Res.
54
: 123-132.
Austin, A.D., and Dillon, N., (1997). Extraction and PCR of DNA from parasitoid wasps that have
been chemically dried.
Aust. J. Entomol.
36
: 241-244.
Austin, J.J., Smith, A.B., and Thomas, R.H., (1997a). Palaeontology in a molecular world: the search
for authentic ancient DNA.
Trends Evol. Ecol.
12
: 303-306.
Austin, J.J., Ross, A.J., Smith, A.B., Fortey, R.A., and Thomas, R.H., (1997b). Problems of
reproducibility-does geologically ancient DNA survive in amber-preserved insects?
Proc. R. Soc.
Lond. B
264
: 467-474.
