Gomez-Polo P., Alomar O., Castane C., Aznar-Fernandez T., Lundgren J.G., Pinol J., Agusti N. (2015) Understanding trophic interactions of Orius spp. (Hemiptera: Anthocoridae) in lettuce crops by molecular methods. Pest Management Science. : 0-0.LinkDoi: 10.1002/ps.3989
BACKGROUND: The aphid Nasonovia ribisnigri (Mosley) (Hemiptera: Aphididae) and the thrips Frankliniella occidentalis (Pergande) (Thysanoptera: Thripidae) are common pests in Mediterranean lettuce crops, where Orius spp. are common generalist predators. Predation by Orius spp. was studied in a lettuce plot by conventional polymerase chain reaction (PCR) and real-time PCR analyses using specific primers of both main pests. Also, high-throughput sequencing was used to have a wider approach of the diet of these predators in natural field conditions. RESULTS: Molecular analyses indicated a higher predation on N. ribisnigri in spring and on F. occidentalis in summer. Predation on alternative prey, like Collembola, was also found in both seasons. Real-time PCR was more sensitive than conventional PCR in showing the target trophic links, whereas high-throughput sequencing revealed predation on other natural enemies - intraguild predation (IGP), showing other trophic interactions of Orius majusculus within the studied ecosystem. CONCLUSIONS: This study gives important information about the trophic relationships present in Mediterranean lettuce crops in different periods of the year. The detected predation by Orius spp. on alternative prey, as well as on other natural enemies, should be further investigated to clarify whether it adds or detracts to the biological control of N. ribisnigri and F. occidentalis. © 2015 Society of Chemical Industry.
Gomez-Polo P., Alomar O., Castane C., Lundgren J.G., Pinol J., Agusti N. (2015) Molecular assessment of predation by hoverflies (Diptera: Syrphidae) in Mediterranean lettuce crops. Pest Management Science. 71: 1219-1227.LinkDoi: 10.1002/ps.3910
BACKGROUND: Hoverflies (Diptera: Syrphidae) are generalist predators of a great variety of pests. Nasonovia ribisnigri (Hemiptera: Aphididae) and Frankliniella occidentalis (Thysanoptera: Thripidae) are two common pests in Mediterranean lettuce crops, where they occur alongside alternative prey (e.g. Collembola). A semi-field experiment was conducted in an experimental lettuce plot where hoverfly predation on N. ribisnigri, F. occidentalis and Collembola was studied by conventional PCR and qPCR using specific primers, as well as by next-generation sequencing (NGS) in order to reveal other potential trophic interactions. RESULTS: Trophic linkages between hoverflies and N. ribisnigri were the strongest both in spring and summer. F. occidentalis and Collembolans were also detected in both seasons, but with less frequency. qPCR detected a higher frequency of consumption than conventional PCR when both tests were run at optimal conditions. NGS analyses showed intraguild predation on other hoverfly species, as well as on anthocorids, spiders and even aphid parasitoids. CONCLUSIONS: Conventional PCR and qPCR provided important insights into Mediterranean hoverfly species predation on target pest and non-pest prey. NGS gave a complementary approach revealing a broader diet of these predators within the studied ecosystem. © 2014 Society of Chemical Industry.
Pinol J., Mir G., Gomez-Polo P., Agusti N. (2015) Universal and blocking primer mismatches limit the use of high-throughput DNA sequencing for the quantitative metabarcoding of arthropods. Molecular Ecology Resources. : 0-0.LinkDoi: 10.1111/1755-0998.12355
The quantification of the biological diversity in environmental samples using high-throughput DNA sequencing is hindered by the PCR bias caused by variable primer-template mismatches of the individual species. In some dietary studies, there is the added problem that samples are enriched with predator DNA, so often a predator-specific blocking oligonucleotide is used to alleviate the problem. However, specific blocking oligonucleotides could coblock nontarget species to some degree. Here, we accurately estimate the extent of the PCR biases induced by universal and blocking primers on a mock community prepared with DNA of twelve species of terrestrial arthropods. We also compare universal and blocking primer biases with those induced by variable annealing temperature and number of PCR cycles. The results show that reads of all species were recovered after PCR enrichment at our control conditions (no blocking oligonucleotide, 45 °C annealing temperature and 40 cycles) and high-throughput sequencing. They also show that the four factors considered biased the final proportions of the species to some degree. Among these factors, the number of primer-template mismatches of each species had a disproportionate effect (up to five orders of magnitude) on the amplification efficiency. In particular, the number of primer-template mismatches explained most of the variation (~3/4) in the amplification efficiency of the species. The effect of blocking oligonucleotide concentration on nontarget species relative abundance was also significant, but less important (below one order of magnitude). Considering the results reported here, the quantitative potential of the technique is limited, and only qualitative results (the species list) are reliable, at least when targeting the barcoding COI region. © 2014 John Wiley & Sons Ltd.
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