Facultative diapause is a life history trait that allows insects to undergo continuous development when conditions are favorable or to enter diapause when they are not. Insect voltinism can have an impact on the success of a weed-biological control agent because additional generations can increase agent population growth and reduce late-season recovery in the target weed. The most common factors that cue diapause are photoperiod and temperature; however, the role of nutrition is increasingly being recognized. We conducted a laboratory experiment to examine the effects of photoperiod and foliage age on diapause induction, pupal weight, and pupal development time in Hypena opulenta (Christoph) (Lepidoptera: Erebidae), a biological control agent for invasive swallow-worts in North America. A factorial experimental design was employed whereby H. opulenta was reared at long (16:8 h light:dark) and short (12:12 h) photoperiods on young and old swallow-wort foliage (Vincetoxicum rossicum Kleopow) Barbar. (Apocynaceae). Photoperiod was the only factor that affected diapause induction in H. opulenta. While foliage age did not affect diapause induction, it did affect pupal weight and pupal development time, with older foliage resulting in lower pupal weight and extended pupal phase. In field conditions, these impacts could affect pupal mortality through reduced winter preparedness and increased exposure to predators. These results support H. opulenta voltinism models based on photoperiods and indicate that the tendency of captive-reared H. opulenta to enter diapause after 2 or 3 generations, even in the absence of short photoperiods, is not a result of changing foliage age.

Field releases of the stem-boring moths Archanara neurica and Lenisa geminipuncta (Lepidoptera: Noctuidae) as classical biological control agents for introduced Phragmites australis australis (Cav.) Trin. ex Steud. began in Canada during 2019. Since then, we have developed and tested release methods for eggs and larvae of both agents. Experiments here include comparisons of: (1) egg survival from exposed "egg card" and protected "egg cup" release methods to assess the risk of agent predation and test mitigation strategies; (2) performance of egg cups overwintered at the release site and those placed shortly before emergence in spring to test for any potential phenological asynchrony between biological control agents and the target weed; and (3) effectiveness of egg cup and larval-inoculated stem releases at a large geographic scale across 15 field sites in southern Ontario. Egg card releases experienced high egg loss likely due to predation (∼90% of eggs), but egg cups were successful at reducing these losses to near zero. Releasing eggs in cups during the spring made it easier to place releases near phenologically synchronized microsites, minimized the risk of damage to the egg cups, and avoided a 16% reduction in egg hatch likely associated with desiccation. Both larval-inoculated stems and egg cups produced highly encouraging feeding damage in the first season of release, with 7.7-fold higher damage from stem larvae compared to egg cups. Overall, we recommend a combination of larval-inoculated stems and spring egg cups for the primary release strategies of A. neurica and L. geminipuncta as biological control agents of introduced Phragmites. Both methods offer protection from predation and varying degrees of control over phenological mismatches. The release of larval-inoculated stems is a more labour-intensive but highly effective method in terms of initial feeding damage whereas egg cups are a more efficient mass-release method than stems with a lower amount of initial feeding damage per agent released.

Invasive plants can gain a foothold in new environments by manipulating soil conditions through allelopathy or through the disruption of associations between native plants and their mycorrhizal associates. The resulting changes in soil conditions can affect the recovery of habitats long after the invasive plant has been removed. We conducted a series of greenhouse experiments to examine the effects of soil conditioned by pale swallow-wort [Vincetoxicum rossicum (Kleopow) Barbarich; Apocynaceae], on the growth of native plants. Additionally, we tested the effects of aqueous extracts of common milkweed (Asclepias syriaca L.; Apocynaceae), a related plant with known allelopathic effects, on the regrowth of V. rossicum from transplanted root crowns. Soil from a 15-yr-old V. rossicum infestation reduced seedling emergence in A. syriaca as well as in V. rossicum itself. Conversely, the same soil had no effect on the growth of mature A. syriaca plants. Soil conditioned by V. rossicum growth in the greenhouse had no effect on the biomass and percentage cover generated by two restoration seed mixes. Soil conditioned by A. syriaca, however, yielded lower biomass and percentage cover from both seed mixes. In contrast to the allelopathic effects of A. syriaca on seedlings, aqueous extracts of A. syriaca increased aboveground plant growth in V. rossicum. Our results suggest that the effects of V. rossicum-conditioned soil on native plants are concentrated at the seedling establishment phase. Additionally, the use of diverse native seed mixes shows great potential for restoring productivity to ecosystems affected by V. rossicum.

Introduced Phragmites (Phragmites australis australis (Cav.) Trin. Ex Steud.) is one of the most invasive plants in North America. To supplement existing management tools, a classical biological control program began in Canada in 2019 using two host-specific stem-boring moths, Archanara neurica (Hübner) and Lenisa geminipuncta (Haworth) (Lepidoptera: Noctuidae). In this article, we summarize the first three years of monitoring data for L. geminipuncta and A. neurica as biological control agents for introduced Phragmites. First, we assess agent presence and activity in the initial years post-release based on feeding damage from long-term monitoring data across 30 release sites initiated between 2019 and 2023. Second, we investigate the within-site distribution of agent feeding damage to improve future monitoring and agent collection from nurse sites. Third, we report the results of an experiment to determine optimal release densities of A. neurica larvae. We found agent feeding damage at 92% of initial release sites in the first year and agent activity persisted at all of these sites into years two and three post-release. Patterns of agent feeding damage suggest that the agents disperse quickly through the patch following release, favouring the interior area over the edges of introduced Phragmites stands. Finally, releasing intermediate densities of 40 A. neurica larvae per release point was more efficient than releasing either units of 20 or 80 larvae. The results of the first three years of monitoring are highly encouraging for the introduced Phragmites biological control program. Insights from these early monitoring results will be used to refine optimal release strategies, improve our ability to locate egg-bearing stems at nurse sites to facilitate the collection and redistribution of agents to new release locations, and inform protocols for longer-term monitoring of impacts on the target weed once agents are established.

Although awareness of the influence of earthworms on soil seed banks in Canadian forests is growing, there have been few direct field measurements. We used a novel pairing of field-collected earthworms from a central Great Lakes forest in Ontario with a laboratory seed egestion assay to obtain a snapshot of the number of seeds passing through earthworms compared with seeds found in the surrounding soil. We identified a pool of seeds egested by earthworms that accounted for 2.4% of all seeds found in the earthworms and the top 0–10 cm of soil. Individual earthworms contained 0–5 seeds. The large-bodied adult anecic non-native Dew Worm or Common Nightcrawler (Lumbricus terrestris) egested a disproportionate number of seeds for its abundance (50% of egested seeds from 17% of earthworms), but smaller earthworms were also an important source of egested seeds (the other 50%). This small-scale proof-of-concept study demonstrates a method of directly measuring earthworm–seed interactions in the field. It can also detect seeds egested by earthworms below ground that would otherwise be missed by other seed accounting methods and it highlights the importance of granivory by non-surface casting earthworms.

Temperature plays an important role in winter diapause of temperate insects. Its effects can cause problems for biological control programs, both for the establishment of insects in novel climates and for the mass rearing of insects in the laboratory. Hypena opulenta (Christoph) (Lepidoptera: Erebidae), a biological control agent for invasive swallow-wort species in North America, has been observed to enter diapause in mid-summer in its introduced range. Additionally, H. opulenta has proved complicated to mass-rear, in part because individuals sometimes enter diapause unexpectedly, meaning that diapausing pupae may need to be stored over winter for varying lengths of time. We conducted pupal storage experiments to determine the effects of high pre-winter temperatures, like those experienced by early diapausing individuals in the field, on lipid store depletion. We conducted a second experiment to compare the effects of cold treatments of different lengths (1, 3, and 6 months), during diapause, on pupal mortality, development times, and the synchrony of adult emergence. Exposure to warm periods ranging from 25 to 75 days during early diapause did not affect pupal weight, adult emergence rates, or lipid store depletion in H. opulenta. Conversely, the length of cold exposure (between 1 and 6 months) during diapause clearly affected moth mortality, and the timing and consistency of adult emergence. Longer cold periods (3 and 6 months) resulted in earlier, more synchronous adult emergence, and lower diapause mortality, than shorter cold periods (1 month). Our results indicate that in terms of energy depletion, early diapause will not affect the fitness of emerging adults, which is important for the viability of univoltine populations in the southernmost part of the introduced range. Additionally, our results will assist with the H. opulenta mass rearing program, indicating the timing required to synchronize adult emergence.

The European root-boring moth, Chamaesphecia empiformis (Lepidoptera: Sesiidae), was released in Ontario, Canada, in 1971 and 1989 as a biological control agent for the perennial invasive weed cypress spurge, Euphorbia cyparissias Linnaeus (Euphorbiaceae). Until recently, the moth was believed to have failed to establish. Beginning in 2015, images posted on citizen science platforms, including BugGuide, iNaturalist, and Facebook groups, indicated that C. empiformis was in fact established. We have confirmed its presence in eastern Ontario by morphological and molecular identification of field-collected adults and eggs. This may be the longest-known lapse between the release of a weed biological control agent and confirmation of its establishment. Citizen science may provide valuable records documenting the establishment and dispersal of biological control agents.

Ingested soil grit is thought to enhance the grinding action of the earthworm gizzard but its role in earthworm -seed interactions is unknown. This study used feeding trials to investigate how different levels of supplemental soil grit (+0 %, +25 %, and +50 % sand additions by weight) influenced the impacts of the cosmopolitan anecic earthworm Lumbricus terrestris L. on seed ingestion, egestion, seed coat damage, and germination of garlic mustard (Alliaria petiolata). Added grit increased the amount of seed coat damage on garlic mustard seeds egested by L. terrestris. Earthworm egestion also increased the speed of garlic mustard germination (similar to 20 days control, similar to 14 days with L. terrestris and +0 % grit, similar to 11 days with L. terrestris and +25-50 % grit). The results demonstrate how earthworm ingestion and soil grit can modify the impacts of earthworms on seeds and highlight the importance of considering soil texture in field and laboratory earthworm experiments.

The psyllid Aphalara itadori Shinji (Hemiptera: Psyllidae) is a biocontrol agent against invasive knotweed species (Polygonaceae) in Europe and North America. Despite an extensive release program, successful establishment of the agent has never been confirmed. The slow establishment of A. itadori in its introduced range appears to be predominantly the result of nymphal mortality, either through desiccation, predation, or the adaptation of captive populations to low-stress environments. In 2016, CABI UK collected a new line of psyllids from Murakami, Niigata Prefecture, in Japan, where it was observed causing severe leaf-roll galling on one of the many varieties of native knotweeds. The induction of leaf-roll galls by Murakami psyllids on invasive Bohemian knotweed, Reynoutria x bohemica, has the potential to alleviate some or all of the observed nymphal mortality factors. We conducted a series of growth chamber experiments to explore the psyllid behaviors that initiate leaf-roll galls, and the ways in which the galls might benefit the psyllids and inhibit invasive knotweed. Reynoutria x bohemica exposed to A. itadori for 4 weeks suffered reduced stem elongation and leaf area compared to control plants. Artificial leaf-roll galls were consistently utilized by developing A. itadori nymphs, and contributed to their survival. Leaf-roll galling in R. x bohemica was predominantly initiated by early instar nymphs of A. itadori, and only occurred when leaves were attacked at a very early stage in their development. Our results can inform the knotweed biocontrol program in several key ways. Releases of A. itadori should be timed so that developing nymphs have access to newly emerging leaves. Monitoring efforts at release sites should focus on the growing tips of R. x bohemica, where the highly mobile nymphs appear to migrate. Finally, our results provide motivation to seek additional lines of A. itadori that may affect other invasive knotweed species in similar ways.

Effective management of the introduced invasive grass common reed [Phragmites australis (Cav.) Trin. ex Steud.] requires the ability to differentiate between the introduced and native subspecies found in North America. While genetic tools are useful for discriminating between the subspecies, morphological identification is a useful complementary approach that is low to zero cost and does not require specialized equipment or technical expertise. The objective of our study was to identify the best morphological traits for rapid and simple identification of native and introduced P. australis. A suite of 22 morphological traits were measured in 21 introduced and 27 native P. australis populations identified by genetic barcoding across southern Ontario, Canada. Traits were compared between the subspecies to identify measurements that offered reliable, diagnostic separation. Overall, 21 of the 22 traits differed between the subspecies, with four offering complete separation: the retention of leaf sheaths on dead stems; a categorical assessment of stem color; the base height of the ligule, excluding the hairy fringe; and a combined measurement of leaf length and lower glume length. Additionally, round fungal spots on the stem occurred only on the native subspecies and never on the sampled introduced populations. The high degree of variation observed in traits within and between the subspecies cautions against a "common wisdom" approach to identification or automatic interpretation of intermediate traits as indicative of aberrant populations or hybridization. As an alternative, we have compiled the five best traits into a checklist of simple and reliable measurements to identify native and introduced P. australis. This guide will be most applicable for samples collected in the late summer and fall in the Great Lakes region but can also inform best practices for morphological identification in other regions as well.

Common obstacles for establishment of newly‐introduced biological control agents include climate and the activity of native antagonists. In temperate climates, these obstacles can disproportionately affect overwintering life‐stages because they are exposed to low winter temperatures, and may rely on passive defence from predators. We conducted a series of field exposure experiments with predator‐exclusion treatments, in Ontario, Canada, to identify mortality factors for the pupae of Hypena opulenta , a biological control agent for invasive swallow‐worts in North America. During two winters, predation rates in containers with large holes, that enabled predation, were relatively low (mean: 23.75%) but non‐predation mortality in closed containers was high (mean: 66.25%), particularly during the colder of the two winters (87.5% vs. 52.5%). During the summer, non‐predation mortality in closed containers was low (mean: 7.5%) but predation rates in containers with large holes were higher than during the winter (mean: 53.33%), increasing as the summer progressed. Predation in containers with large holes was 70% during late summer, compared with 25% during the spring. Across all seasons, pupal predation was dominated by large non‐arthropod predators. Hypena opulenta can complete 2 generations per year. Photoperiods that induce diapause occur earlier in the introduced range than in the native range, however, and H. opulenta individuals in parts of the introduced range are likely to enter diapause early after a single generation. Our results highlight additional vulnerabilities encountered by such individuals, and can contribute to models predicting population dynamics of H. opulenta across its introduced range.

A wide variety of organisms use the regular seasonal changes in photoperiod as a cue to align their life cycles with favorable conditions. Yet the phenological consequences of photoperiodism for organisms exposed to new climates are often overlooked. We present a conceptual approach and phenology model that maps voltinism (generations per year) and the degree of phenological mismatch that can arise when organisms with a short-day diapause response are introduced to new regions or are otherwise exposed to new climates. Our degree-day-based model combines continent-wide spatialized daily climate data, calculated date-specific and latitude-specific day lengths, and experimentally determined developmental responses to both photoperiod and temperature. Using the case of the knotweed psyllid Aphalara itadori, a new biological control agent being introduced from Japan to North America and Europe to control an invasive weed, we show how incorporating a short-day diapause response will result in geographic patterns of attempted voltinism that are strikingly different from the potential number of generations based on degree-days alone. The difference between the attempted and potential generations represents a quantitative measure of phenological mismatch between diapause timing and the end of the growing season. We conclude that insects moved from lower to higher latitudes (or to cooler climates) will tend to diapause too late, potentially resulting in high mortality from inclement weather, and those moved from higher to lower latitude (to warmer climates) may be prone to diapausing too early, therefore not fully exploiting the growing season and/or suffering from insufficient reserves for the longer duration in diapause. Mapped output reveals a central region with good phenology match that shifts north or south depending on the geographic source of the insect and its corresponding critical photoperiod for diapause. These results have direct relevance for efforts to establish populations of classical biocontrol agents. More generally, our approach and model could be applied to a wide variety of photoperiod- and temperature-sensitive organisms that are exposed to changes in climate, including resident and invasive agricultural pests and species of conservation concern.

Pale swallow-wort, Vincetoxicum rossicum (Kleopow) Barbar., is an invasive weed in the lower Great Lakes Basin of North America. We investigated the relationship between V. rossicum productivity and several environmental variables across 54 established V. rossicum populations in southern Ontario. Variables included climate measurements, soil characteristics (pH, texture, and nutrient status), habitat type, plant community, and the diversity of root-associated fungi. Vincetoxicum rossicum roots were collected at all 54 sites, and associated fungi were compared using terminal restriction fragment length polymorphism. Aboveground biomass of V. rossicum was measured at 23 sites with similar light regimes, allowing comparison between populations. Results suggested that abiotic soil variables, and habitat type have little effect on V. rossicum productivity. Aboveground biomass production was significantly correlated with precipitation, and the presence of fungal pathogens and dark septate endophytes. Annual precipitation and soil clay content were also positively correlated with the abundance of soil pathogens. Accumulation of these microbes may negatively affect co-occurring native plants and associated fungal partners. The presence of these fungal species in soil could be used as an indicator of site susceptibility to invasion by V. rossicum and assist in the development of management plans for this exotic vine.