Environmental DNA assays for Laricobius beetles (Coleoptera: Derodontidae), biocontrol agents of the hemlock woolly adelgid in North America.

The hemlock woolly adelgid, Adelges tsugae Annand (Hemiptera: Adelgidae), is an invasive pest causing significant ecological and economic damage to certain hemlock tree (Tsuga (Endlicher) Carrière, Pinales:Pinaceae) species. In response to this invasive threat, biological control strategies have been implemented, introducing natural predators such as Laricobius nigrinus Fender (Coleoptera: Derodontidae) and, more recently, Laricobius osakensis Montgomery and Shiyake (Coleoptera: Derodontidae), as specialist predators against A. tsugae. However, the genetic and morphological similarities between L. osakensis and both L. nigrinus and the native beetle, Laricobius rubidus LeConte (Coleoptera: Derodontidae), pose challenges in their identification. Effective monitoring of released predators is integral to evaluating the success of biological control measures. Environmental DNA (eDNA) holds potential for various detection applications, including species monitoring. In this study, we developed specific primers and probes targeting the mitochondrial cytochrome oxidase 1 gene sequences, achieving high specificity despite their 95% sequence similarity. With an optimal annealing temperature of 60 °C, our tools effectively differentiated L. osakensis from the other 2 beetles and demonstrated eDNA detection sensitivity down to 2 copies/µl. This research underscores the potential of precise molecular tools for advancing biological control and biodiversity assessment against invasive threats like A. tsugae.

Observation of key phenological stages of hemlock woolly adelgid (Hemiptera: Adelgidae): using citizen science as a tool to inform research and management.

Increasing efficiency of data gathering at the landscape scale on the growing number of pests and pathogens threatening forests worldwide has potential to improve management outcomes. Citizen science is expanding, with growing support and utility in environmental and conservation fields. We present a case study showing how citizen science observations can be used to inform research and management of a devastating forest pest. Hemlock woolly adelgid, Adelges tsugae Annand (Hemiptera: Adelgidae), was introduced to eastern North America, leading to decline and mortality of eastern [Tsuga canadensis (L.) Carrière] (Pinales: Pinaceae) and Carolina hemlock (Tsuga caroliniana Engelmann) trees. Management activities, most notably biological control, rely on observations of A. tsugae phenology to inform the timing of releases and monitoring surveys of their highly synchronized specialist predators. In this article, we outline a citizen science program and report phenological observations on A. tsugae. Additionally, we report data comparing A. tsugae estivation break in Virginia (VA) and New York (NY) State, revealing that estivation break is synchronized between NY and VA. This observation is supported by 6 years of citizen scientist observations, showing similar patterns throughout NY, with egg laying shown to be much more variable than estivation break.

Phenology of Leucotaraxis argenticollis, a specialist predator of the invasive hemlock woolly adelgid, in the eastern United States.

In hemlock stands within eastern US forests, classical biological control has been one of the main strategies used to manage the hemlock woolly adelgid, Adelges tsugae Annand. Specialist predator species may offer a management solution to help regulate A. tsugae populations. In the Pacific Northwest, a suite of specialist predators has been a focus of research and includes 2 species of silver fly, Leucotaraxis argenticollis (Zetterstedt) and Leucotaraxis piniperda (Malloch) (Diptera: Chamaemyiidae). Leucotaraxis spp. phenology has been documented in the Pacific Northwest, but the phenology of either western Leucotaraxis species is unknown in the eastern United States. This study sought to document the phenology of Le. argenticollis in NY in 2021 and in VA in 2021 and 2022. Nylon mesh cages were applied over eastern hemlock branches infested with A. tsugae to contain Le. argenticollis adults. Biweekly and monthly branch samples were taken in 2021 and 2022, documenting all life stages of A. tsugae and of Le. argenticollis that were observed. In 2021 and 2022, Le. argenticollis adults and eggs were present during the oviposition stage of the 2 generations of A. tsugae. In addition, Le. argenticollis larvae were present when A. tsugae ovisacs had eggs and while A. tsugae nymphs of both generations were present. These observations indicate that Le. argenticollis phenology is well synchronized with A. tsugae in the eastern United States.

Estivation and Postestivation Development of Hemlock Woolly Adelgid (Adelges tsugae) (Hemiptera: Adelgidae) at Different Temperatures.

Hemlock woolly adelgid (Adelges tsugae) is the most important pest of hemlocks in the eastern United States, where it completes three generations a year. We investigated the impact of temperature (8, 12, 16, and 20°C) on the estivation and postaestivation stages of the A. tsugae sistens generation. Temperature significantly impacted development and survival of this generation. The highest mortality occurred at the coolest temperature (8°C). Adelges tsugae developed rapidly as the temperature increased and optimum temperatures for development ranged between 17 and 22°C for the different instars. The estimated lower temperature threshold was 0°C for second instar nymphs and 3 -5°C for the other instars and the preoviposition period. Estivating first-instar sistentes resumed development (as evidenced by segments becoming visible) after 40-100 d at the constant temperatures (fastest at 16°C) then required only 105 degree-days (DD) for 50% of the individuals to molt. Subsequent instars developed rapidly (another 470 DD total to reach adult), and oviposition began at ~623 DD from the time the first instars resumed development. This study provides valuable data required to develop an annual phenology model for A. tsugae which will assist in timing monitoring and control treatments.

Hymenopteran Parasitoids of Leucotaraxis argenticollis (Diptera: Chamaemyiidae) and Leucotaraxis piniperda: Implications for Biological Control of Hemlock Woolly Adelgid (Hemiptera: Adelgidae).

The hemlock woolly adelgid (Adelges tsugae (Annand)) is a serious invasive pest of hemlock trees in eastern North America. Multiple biological control agents have been the focus of research aimed at pest management and conserving hemlock communities. Three promising A. tsugae specialist predators are the beetle Laricobius nigrinus (Fender) (Coleoptera: Derodontidae) and flies in the genus Leucotaraxis (Diptera: Chamaemyiidae), Leucotaraxis argenticollis (Zetterstedt), and Leucotaraxis piniperda (Malloch). However, these flies are vulnerable to parasitism by wasps in the genera Pachyneuron (Walker) (Hymenoptera: Pteromalidae) and Melanips (Walker) (Hymenoptera: Figitidae). This study explores parasitoid wasp interactions with these Leucotaraxis species in their native western North American range and potential impacts on the biological control program in the East. Leucotaraxis, La. nigrinus, and parasitoid emergences were observed from adelgid-infested foliage collected from Washington State and British Columbia in 2018, 2019, and 2020. Undescribed species of Pachyneuron and Melanips emerged from puparia as solitary parasitoids. Parasitoid emergence was positively correlated with Leucotaraxis emergence. Percent parasitism increased between February and July, with the months of June and July experiencing higher parasitoid emergence than Leucotaraxis. Differences in emergence patterns suggest that Pachyneuron may be more closely associated with Le. argenticollis as a host, and that Melanips may be associated with Le. piniperda. High parasitism in Leucotaraxis had no effect on La. nigrinus larval abundance, whereas the combined emergence of parasitoids and Leucotaraxis was positively correlated with La. nigrinus. This suggests that there is limited competition among these predators.

The Lari-Leuco Container: A Novel Collection Arena for Separating Insects Ascending or Descending From a Plant Foliage Sample.

Efficient separation of insects from plant material for quantification and collection is an important component of entomological research. This paper reports on a novel, easily replicable container designed to efficiently collect two different biological control agents dispersing from hemlock (Tsuga spp.) foliage infested with the invasive hemlock woolly adelgid (HWA), Adelges tsugae Annand (Hemiptera: Adelgidae). The container utilizes a simplified Berlese-style funnel design to collect Laricobius spp. (Coleoptera: Derodontidae) larvae dropping from the foliage into a removable bottom jar, a central jar to house the foliage sample, and a removable top jar to collect adult silver flies (Leucopis spp., Diptera: Chamaemyiidae) emerging from puparia on the twigs. The efficacy of two designs (with and without a funnel leading to the top collection jar) was evaluated using western hemlock [Tsuga heterophylla (Raf.) Sarg.] foliage naturally colonized with HWA and the two predator genera. All Laricobius larvae were effectively collected in the bottom jar, and the addition of an inverted funnel leading to the top collection jar increased the proportion of Leucopis flies reaching the target jar from 60% to 94%. This 'Lari-Leuco' container is presented as a research and motoring tool to benefit the integrated pest management program for HWA in eastern North America and for potential use in simultaneously separating ascending and descending life stages in other insect-plant or predator-prey systems.

Biological Control of Hemlock Woolly Adelgid: Implications of Adult Emergence Patterns of Two Leucopis spp. (Diptera: Chamaemyiidae) and Laricobius nigrinus (Coleoptera: Derodontidae) Larval Drop.

The hemlock woolly adelgid (Hemiptera: Adelgidae Adelges tsugae Annand) poses a serious threat to hemlocks in eastern North America, and ongoing research is focused on the identification and development of biological controls to protect and manage hemlock resources. Three predators native to the Pacific Northwest of North America that have been the focus of much research are Leucopis argenticollis (Zetterstedt), Leucopis piniperda (Malloch) (Diptera: Chamaemyiidae), and Laricobius nigrinus (Fender) (Coleoptera: Derodontidae). This study addresses the knowledge gap of adult Leucopis spp. emergence patterns, with comparisons to the timing of larval La. nigrinus drop for pupation. Adult Leucopis spp. emergence was observed in the lab from field-collected, adelgid-infested foliage from Washington state in 2019 and 2020. Adult Leucopis spp. were collected daily as they emerged from foliage collections and identified to species using morphological features; a subset was validated using DNA barcoding. Accumulated heating degree days were calculated to compare a standardized emergence timing across collections made at different locations and temperature regimes. The abundance of the two Leucopis spp. and of the combined Leucopis spp. and La. nigrinus varied among sites and years, and no species was consistently more abundant than the other. Evaluations of seasonal emergence trends of the three species determine the predator complex behaves in a temporally stratified and predictable way. Emergence of adult Le. argenticollis was observed first, followed by La. nigrinus larval drop, with Le. piniperda emerging at the end of larval drop, and finally a second emergence of Le. argenticollis.

Sticky traps as an early detection tool for crawlers of Adelges tsugae (Hemiptera: Adelgidae).

We developed an approach using sticky trap arrays as an early detection tool for populations of first-instar nymphs of the hemlock woolly adelgid (Adelges tsugae Annand), a pest of hemlocks (Tsuga spp. [Pinaceae]) in North America. We considered the detection rate of at least one nymph from trapping arrays consisting of one to six sticky panels, where we varied both the surface area of each trap that we assessed and the length of the trapping duration. We also estimated the time needed to set up, service, and assess groups of traps and attempted to relate capture of nymphs on traps to incidence and abundance of A. tsugae in the canopy above the traps. Arrays consisting of two traps provided a detection rate of 75% when 87.5% of the surface area of each trap was assessed, a process that required 38 min per array. The probability of detecting nymphs on traps left in the field for 5-6 d was similar to that for traps left for 12 d. The number of nymphs trapped in an array predicted the probability of finding A. tsugae in the canopy but only when all six traps were fully assessed. To reliably detect incipient A. tsugae infestations, we recommend placing arrays of traps at 1 km intervals along the perimeter of a stand during peak activity of first-instar sistentes nymphs and servicing these arrays every 5-7 d.