Pushing the limits of whole genome amplification: successful sequencing of RADseq libraries from single micro-hymenoptera (Chalcidoidea, Trichogramma)

A major obstacle to high-throughput genotyping of micro-hymenoptera is their small size. As species are difficult to discriminate and because complexes may exist, the sequencing of a pool of specimens is hazardous. Thus, one should be able to sequence pangenomic markers (e.g. RADtags) from a single specimen. To date, whole genome amplification (WGA) prior to library construction is still a necessity as only ca 10ng of DNA can be obtained from single specimens. However this amount of DNA is not compatible with manufacturer’s requirements for commercialised kits. Here we tested the accuracy of the GenomiPhi kit V2 on Trichogramma wasps by comparing RAD libraries obtained from the WGA of single specimens (generation F0 and F1, ca 1 ng input DNA for the WGA) and a biological amplification of genomic material (the pool of the progeny of the F1 generation). Globally, we found that ca 99% of the examined loci (up to 48,189; 109 bp each) were compatible with the mode of reproduction of the studied model (haplodiploidy) or a Mendelian inheritance of alleles. The remaining 1% (ca 0.01% of the analysed nucleotides) could represent WGA bias or other experimental / analytical bias. This study shows that the multiple displacement amplification method on which the GenomiPhi kit relies, could also be of great help for the high-throughput genotyping of micro-hymenoptera used for biological control or other organisms from which only a very low amount of DNA can be extracted such as human disease vectors (e.g. sand flies, fleas, ticks etc.)

Investigating Biological Control Agents for Controlling Invasive Populations of the Mealybug Pseudococcus comstocki in France

WOS: 000378865200032PubMed ID: 27362639Pseudococcus comstocki (Hemiptera: Pseudococcidae) is a mealybug species native to Eastern Asia and present as an invasive pest in northern Italy and southern France since the start of the century. It infests apple and pear trees, grapevines and some ornamental trees. Biocontrol programmes against this pest proved successful in central Asia and North America in the second half of the 20th century. In this study, we investigated possible bio-control agents against P. comstocki, with the aim of developing a biocontrol programme in France. We carried out systematic DNA-barcoding at each step in the search for a specialist parasitoid. First we characterised the French target populations of P. comstocki. We then identified the parasitoids attacking P. comstocki in France. Finally, we searched for foreign mealybug populations identified a priori as P. comstocki and surveyed their hymenopteran parasitoids. Three mealybug species (P. comstocki, P. viburni and P. cryptus) were identified during the survey, together with at least 16 different parasitoid taxa. We selected candidate biological control agent populations for use against P. comstocki in France, from the species Allotropa burrelli (Hymenoptera: Platygastridae) and Acerophagus malinus (Hymenoptera: Encyrtidae). The coupling of molecular and morphological characterisation for both pests and natural enemies facilitated the programme development and the rejection of unsuitable or generalist parasitoids.French "Agence Nationale de la Recherche"French National Research Agency (ANR) [ANR-10-JCJC-1708 BICORAMICS]; EUEuropean Union (EU) [324475 COLBICS, 265865 PURE]; Turkey-France Cooperation grant CNRS-TUBITAK; INRA Plant Health and Environment DivisionInstitut National de la Recherche Agronomique (INRA)This work was funded by the French "Agence Nationale de la Recherche" (grant ANR-10-JCJC-1708 BICORAMICS), by the EU Seventh Framework Programme (grants Marie-Curie IAPP #324475 COLBICS and KBBE #265865 PURE), by a Turkey-France Cooperation grant CNRS-TUBITAK, and by the INRA Plant Health and Environment Division. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript

Genetic diversity of armored scales (Hemiptera: Diaspididae) and soft scales (Hemiptera: Coccidae) in Chile

Scale insects (Sternorrhyncha: Coccoidea) are one of the most invasive and agriculturally damaging insect groups. Their management and the development of new control methods are currently jeopardized by the scarcity of identification data, in particular in regions where no large survey coupling morphological and DNA analyses have been performed. In this study, we sampled 116 populations of armored scales (Hemiptera: Diaspididae) and 112 populations of soft scales (Hemiptera: Coccidae) in Chile, over a latitudinal gradient ranging from 18 degrees S to 41 degrees S, on fruit crops, ornamental plants and trees. We sequenced the COI and 28S genes in each population. In total, 19 Diaspididae species and 11 Coccidae species were identified morphologically. From the 63 COI haplotypes and the 54 28S haplotypes uncovered, and using several DNA data analysis methods (Automatic Barcode Gap Discovery, K2P distance, NJ trees), up to 36 genetic clusters were detected. Morphological and DNA data were congruent, except for three species (Aspidiotus nerii, Hemiberlesia rapax and Coccus hesperidum) in which DNA data revealed highly differentiated lineages. More than 50% of the haplotypes obtained had no high-scoring matches with any of the sequences in the GenBank database. This study provides 63 COI and 54 28S barcode sequences for the identification of Coccoidea from Chile

No inbreeding depression in laboratory-reared individuals of the parasitoid wasp Allotropa burrelli

Inbreeding depression is a major concern in almost all human activities relating to plant and animal breeding. The biological control of pests with natural enemies is no exception, because populations of biocontrol agents experience a series of bottlenecks during importation, rearing, and introduction. A classical biological control program for the Comstock mealybug Pseudococcus comstocki (Hemiptera: Pseudococcidae) was initiated in France in 2008, based on the introduction of an exotic parasitoid, Allotropa burrelli Mues. (Hymenoptera: Platygastridae), a haplodiploid parasitoid imported from Japan. We evaluated the sensitivity of A. burrelli to inbreeding, to optimize rearing and release strategies. We compared several morphological and life-history traits between the offspring of siblings and the offspring of unrelated parents. We took into account the low level of genetic variability due to the relatively small size of laboratory-reared populations by contrasting two types of pedigree: one for individuals from a strain founded from a single field population, and the other generated by hybridizing individuals from two strains founded from two highly differentiated populations. Despite this careful design, we obtained no evidence for a negative impact of inbreeding on laboratory-reared A. burrelli. We discussed the results in light of haplodiploid sex determination and parasitoid mating systems, and classical biological control practices

Diversity and geographic distribution of the indigenous and exotic parasitoids of the olive fruit fly, Bactrocera oleae (Diptera: Tephritidae), in Southern France

International audienceThe olive fruit fly, Bactrocera oleae (Dipt., Tephritidae), is the most important pest of olive crops in the world. Economic losses associated to the limited efficiency of pesticides and natural regulation require the development of new alternatives. A classical biological control program was thus implemented in 2007 in France with two main objectives : (1) test the efficiency of a new exotic parasitoid, Psyttalia lounsburyi (Hym., Braconidae) on the olive fruit fly populations and (2) understand how intraspecific hybridization could affect the demographic success of exotic biocontrol agents and, more generally, invasive species. In 2008, more than 43,000 P. lounsburyi were consequently introduced in 60 sites located in Southern France, covering the whole geographic distribution of olive crops in this country. The pluri-annual surveys realised between 2007 and 2010 gave the opportunity to better document the dynamics of olive fruit fly populations as well as the associated communities of parasitoids. Main results on these two topics are outlined here in a view to stimulate collaborative research and more precisely document the community ecology of B. oleae and its natural enemies in the Mediterranean area and elsewhere