Preventing the introduction and establishment of forest invasive alien species (FIAS) such as the Asian gypsy moth (AGM) is a high-priority goal for countries with extensive forest resources such as Canada. The name AGM designates a group of closely related Lymantria species (Lepidoptera: Erebidae: Lymantriinae) comprising two L. dispar subspecies (L. dispar asiatica, L. dispar japonica) and three closely related Lymantria species (L. umbrosa, L. albescens, L. postalba), all considered potential FIAS in North America. Ships entering Canadian ports are inspected for the presence of suspicious gypsy moth eggs, but those of AGM are impossible to distinguish from eggs of innocuous Lymantria species. To assist regulatory agencies in their identification of these insects, we designed a suite of TaqMan® assays that provide significant improvements over existing molecular assays targeting AGM. The assays presented here can identify all three L. dispar subspecies (including the European gypsy moth, L. dispar dispar), the three other Lymantria species comprising the AGM complex, plus five additional Lymantria species that pose a threat to forests in North America. The suite of assays is built as a “molecular key” (analogous to a taxonomic key) and involves several parallel singleplex and multiplex qPCR reactions. Each reaction uses a combination of primers and probes designed to separate taxa through discriminatory annealing. The success of these assays is based on the presence of single nucleotide polymorphisms (SNPs) in the 5’ region of mitochondrial cytochrome c oxidase I (COI) or in its longer, 3’ region, as well as on the presence of an indel in the “FS1” nuclear marker, generating North American and Asian alleles, used here to assess Asian introgression into L. dispar dispar. These assays have the advantage of providing rapid and accurate identification of ten Lymantria species and subspecies considered potential FIAS.

Anthropogenic activities have a major impact on the global environment. Canada's natural resources are threatened by the spread of fungal pathogens, which is facilitated by agricultural practices and international trade. Fungi are introduced to new environments and sometimes become established, in which case they can cause disease outbreaks resulting in extensive forest decline. Here, we describe how a nationwide sample collection strategy coupled to next-generation sequencing (NGS) (i.e., metagenomics) can achieve fast and comprehensive screening for exotic invasive species. This methodology can help provide guidance to phytopathology stakeholders such as regulatory agencies. Several regulated invasive species were monitored by processing field samples collected over 3 years (2013 to 2015) near high-risk areas across Canada. Fifteen sequencing runs were required on the Ion Torrent platform to process 398 samples that yielded 45 million reads. High-throughput screening of fungal and oomycete operational taxonomic units using customized fungi-specific ribosomal internal transcribed spacer 1 barcoded primers was performed. Likewise, Phytophthora-specific barcoded primers were used to amplify the adenosine triphosphate synthase subunit 9-nicotinamide adenine dinucleotide dehydrogenase subunit 9 spacer. Several Phytophthora spp. were detected by NGS and confirmed by species-specific quantitative polymerase chain reaction (qPCR) assays. The target species Heterobasidion annosum sensu stricto could be detected only through metagenomics. We demonstrated that screening target species using a variety of sampling techniques and NGS-the results of which were validated by qPCR-has the potential to increase survey capacity and detection sensitivity, reduce hands-on time and costs, and assist regulatory agencies to identify ports of entry. Considering that early detection and prevention are the keys in mitigating invasive species damage, our method represents a substantial asset in plant pathology management.

Monochamol (2-undecyloxy-1-ethanol) is a male-produced aggregation pheromone for several Monochamus Dejean (Coleoptera: Cerambycidae) species. We conducted trapping experiments in Canada, Poland, and China to test whether monochamol was attractive to additional Monochamus species and if attraction was synergised by plant volatiles and bark beetle (Coleoptera: Curculionidae) pheromones. We provide the first evidence of attraction for M. urussovii (Fischer) and M. saltuarius (Gebler) to monochamol or monochamol+kairomones. The highest numbers of M. urussovii were captured in traps baited with monochamol+plant volatiles (Manuka oil, ethanol and (95/5±) α−pinene). Captures of M. saltuarius were highest in traps baited with monochamol, with the addition of cubeb oil tending to reduce captures. The highest numbers of M. scutellatus (Say) were captured in traps baited with monochamol+kairomones. A similar pattern in trap captures was found for M. notatus (Drury), M. marmorator Kirby, M. carolinensis (Olivier), and M. mutator LeConte. Detection rates, that is, proportion of traps capturing at least one specimen, was highest for traps baited with monochamol plus kairomones, particularly for less-common species. These results support the emerging hypothesis that pheromone compounds can attract related cerambycid species with cumulative evidence for attraction to monochamol for 12 species of Monochamus worldwide.

Introductions of some forest invasive alien species result in important economic, environmental, and ecological impacts. One approach used by the Canadian Food Inspection Agency to improve the detection of these species is to collect logs from trees in declining health at high risk sites of introduction and to incubate them to obtain insects, if present. Trichoferus campestris (Faldermann) (Coleoptera: Cerambycidae) adults emerged, and live larvae were extracted, from one of two logs taken from a dying Norway maple, Acer platanoides Linnaeus (Sapindaceae), in Mississauga, Ontario, Canada. In its native range, eastern Asia, this beetle is polyphagous, however, in North America there is no host record despite numerous interceptions of larvae in wood packaging material and captures of adults at various post-entry sites. An examination of the feeding damage caused by T. campestris in that maple suggests this insect is not a primary pest of trees in Ontario.

The attractiveness of ultra high release ethanol lures to ambrosia beetles in Slovakian oak forests was tested from 2010 to 2012. A total of 24,705 specimens were captured during this three year period with Xyleborinus saxesenii (Ratzeburg, 1837) representing 49.28% (12,174 specimens) of the total. Other dominant species captured in the traps were Anisandrus dispar (F., 1792) (27.84%), Xyleborus monographus (F., 1792) (9.72%) and Trypodendron signatum (F., 1792) (6.04%). During this experiment, Xylosandrus germanus (Blandford, 1894) was detected for the first time in Slovakia with an increase in capture each year (19, 40 and 77 specimens, respectively). Flight period for ambrosia beetles in Slovakia occurs from the beginning of April through the end of September. This is the first time that ethanol baited traps were deployed in Slovakian oak forests and the lures were an effective tool for monitoring native and non-native ambrosia beetles.