DNA metabarcoding dietary analyses of the wood mouse Apodemus speciosus on Innoshima Island, Japan, and implications for primer choice

We used DNA metabarcoding to assess the seasonal diets of the large Japanese wood mouse, Apodemus speciosus (Muridae, Rodentia), in forest edges adjacent to citrus orchards on Innoshima Island, Japan. We used one chloroplast and three mitochondrial DNA barcoding markers to determine mouse diets. Among the various plant and invertebrate diets, A. speciosus typically consumed Chinese cork oak (Quercus variabilis) in early spring (likely acorns preserved during winter) and gypsy moths (Lymantria dispar, a forest pest) in late spring and summer. In addition, we found that A. speciosus also preyed on orchard pests, including the gutta stink bug and other potentially harmful invertebrates. The season during which A. speciosus preyed on stink bugs corresponded with the harvest of orchard products. This study revealed several of the ecological roles of A. speciosus within the boundary zone between forest and human ecosystems. Furthermore, based on the performance of various mitochondrial markers in dietary profiling of invertebrate food items, we recommend the multi-locus DNA metabarcoding method to comprehensively assess the diet of A. speciosus.

Histologic lesions of experimental infection with Lymantria dispar multicapsid nucleopolyhedrovirus and Lymantria dispar cytoplasmic polyhedrosis virus in European gypsy moth caterpillars (Lymantria dispar dispar)

European gypsy moths ( Lymantria dispar dispar) are an invasive species in North America, and are listed by the International Union for the Conservation of Nature as one of the 100 most destructive invasive species worldwide. They have several known viruses, some of which are used as biological control agents. However, there are no detailed descriptions of many entomopathogenic viral infections, including in European gypsy moths, using bright-field microscopy. In this study, 11 European gypsy moth caterpillars were evaluated histologically: 4 were experimentally infected with Lymantria dispar multicapsid nucleopolyhedrovirus (LdMNPV; Baculoviridae); 4 were experimentally infected with Lymantria dispar cytoplasmic polyhedrosis virus (LdCPV; Reoviridae); 3 control animals were uninfected. A complete tissue set was evaluated in all animals from all groups using bright-field microscopy, including epidermis, cuticle, striated muscle, tracheae, foregut, midgut, hindgut, Malpighian tubules, hemocytes, fat body, and nervous system. LdMNPV-infected caterpillars had marked karyomegaly and intranuclear viral inclusions in cells of the epidermis, tracheae, fat body, and hemocytes. LdMNPV-infected caterpillars also had hyperplasia and hypertrophy of epidermal and tracheal epithelial cells. LdCPV-infected caterpillars had numerous granular eosinophilic intracytoplasmic viral inclusions in midgut epithelial cells. Both LdMNPV-infected and LdCPV-infected caterpillars had atrophy of fat body adipocytes; this change was more pronounced in LdCPV-infected caterpillars. This work provides the first detailed descriptions of these viral infections in European gypsy moth caterpillars using bright-field light microscopy and provides images of normal histology from control caterpillars.

Potential Differences and Methods of Determining Gypsy Moth Female Flight Capabilities: Implications for the Establishment and Spread in Novel Habitats

The introduction of the Asian gypsy moth into novel environments continues with frequent interceptions in North America. There is a concern that these subspecies will pose a greater threat to the forests and urban environments of North America than the established gypsy moths (Lymantria dispardispar L.), due to their greater capacity for female flight. Asian gypsy moth populations vary in many key traits, including female flight capabilities. The potential impacts of female flight, in combination with the other key traits, on the ecology and spread of this insect are first discussed in this communication. This also provides the first review of most of the current literature on the variations in flight capability and flight distance of gypsy moth populations, as well as variation in other traits of concern and the potential methods of identification, with special attention paid to the Asian subspecies Lymantria dispar japonica Motschulsky and Lymantria dispar asiatica Vinkovskij. There are currently good tools for identifying the general origin of introduced gypsy moth populations, but these do not provide enough information to effectively manage introductions. Gypsy moth key traits differ among populations, even within each subspecies of the gypsy moth, so introduction of gypsy moths from other world areas into locations where the gypsy moth is already present could result in unwanted changes in gypsy moth biology. It also appears that the introduction of flight-capable females could enhance a population’s dispersal capability and require modifications to management protocols used for flightless females. Therefore, rapid tools to assess key traits in introduced populations are needed to adequately plan for, or deal with, new introductions into novel habitats.

Defoliation by Gypsy Moth (Lepidoptera, Erebidae) Induces Differential Delayed Induction of Trichomes in Two Birch Species

Outbreaks of the gypsy moth (Lymantria dispar japonica Motschulsky) cause serious defoliation in birch. A single year of defoliation has no significant impact on the trees, whereas continuous defoliation events could be fatal. How birch species avoid serious damage caused by gypsy moth outbreak is yet to be revealed. Trichomes on leaf surfaces of birch trees are an effective antiherbivore defense strategy. We examined a 1-yr delayed induction of glandular (GT) and nonglandular trichomes (NGT) on leaf surfaces caused by stress in white birch (Betula platyphylla Sukaczev var. japonica [Miq.] Hara) and monarch birch (Betula maximowicziana Regel). Saplings were subjected to four treatments in June 2009: herbivory (50% of the leaf area was grazed by gypsy moths), mechanical cutting (50% of the leaf area was cut using scissors), shading (50% light shading with a black sheet), and control (covered with a net to prevent herbivory). Then, the density of GT and NGT on early leaves was determined in April 2010. In both birch species, the density of GT was higher in herbivory than in other treatments. The density of GT due to mechanical cutting was higher than that in the control, but only for white birch. However, the density of NGT was lower after mechanical cutting than in other treatments for white birch. There were no differences in the density of NGT among treatments for monarch birch. These results show that 1-yr delayed induction of GT by herbivory was stronger in white birch than in monarch birch.

Management of flying insects on expressways through an academic-industrial collaboration: evaluation of the effect of light wavelengths and meteorological factors on insect attraction

Insect outbreaks often occur in the absence of natural enemies and in the presence of excess suitable host materials. Outbreaks of gypsy moths are especially problematic in remote areas located in high-latitude regions in Japan because the majority of adults emerge during the short summer season and initiate synchronous mass flight toward artificial lights. The aggregation of moths in public facilities not only is an annoyance to visitors but also permits the establishment of new populations the following year. The aim of this study was to establish a method to reduce the numbers of large moths that are attracted to lights in the rest areas of expressways in Hokkaido based on the results of research on their behavioral ecology and physiology. First, we conducted extensive insect surveys using light traps that emit light at different wavelengths; the traps were set along the expressways in the summers of 2014–2018. The insects attracted to the light were roughly classified into those showing a preference for broadband light wavelengths (from UV-A to green) and short light wavelengths (from UV-A to blue). The former included aquatic insects and winged ants, and the latter included moths and beetles. Next, we analyzed correlations between moth emergence and daily meteorological data. When gypsy moths were abundant during an outbreak, the daily catch of gypsy moths was positively correlated with the highest ambient temperature on the catch day but not with the visibility range, wind speed, or moon phase. In contrast, the daily catch of oak silkmoths did not correlate with any of these parameters. Our results provide guidance for the management of forest insects inhabiting cool-temperate to subarctic regions based on light wavelengths with reference to weather variables.

Comparison of Survival and Development of Gypsy Moth Lymantria dispar L. (Lepidoptera: Erebidae) Populations from Different Geographic Areas on North American Conifers

Host utilization information is critical to managers for estimating the hosts at risk and potential geographic range for gypsy moths from different geographic origins. In this study, the development and survival of gypsy moths from all three subspecies on 13 North American conifers and three broadleaf hosts were compared. There was variation in the ability of gypsy moth larvae from different geographic origins to utilize (survive and develop on) key North American conifers. However, that variation was not consistent within gypsy moth subspecies, but instead was more consistent with populations from different origins being preadapted to utilize different hosts and having different biologic traits. Some Asian populations developed and survived well on some conifers while populations from Europe and North America gained weight faster and/or survived better than some Asian populations. Although development was slower and survival poorer on several of the conifers, first instar larvae were able to utilize conifers unless the needles were tough or feeding deterrents were present. Host phenology was also critical since the early instars fed preferentially on new foliage or buds. Gypsy moth larvae can utilize many hosts, so this makes it a very adaptable invasive species that warrants taking measures to prevent its spread.

Microsatellite and Morphological Analyses Reveal Unexpected Diversity in Lymantria dispar in China

The gyspy moth Lymantria dispar Linnaeus, a widely distributed leaf-eating pest, is considered geographically isolated in the world, with two Asian gypsy moth subspecies, Lymantria dispar asiatica and Lymantria dispar japonica. In China, only one subspecies, L. d. asiatica, has been observed. In this study, we characterized gypsy moth diversity and divergence using 427 samples covering a wide range of the species distribution, with a focus on sampling along a latitudinal gradient in China. We combine the quantitative analysis of male genitalia and the genetic diversity analysis of nine microsatellite loci of nuclear genes nuclear genes to study the structure of gypsy moth individuals in 23 locations in the world and the male genitalia of gypsy moths in some areas. In mixed ancestry model-based clustering analyses based on nuclear simple sequence repeats, gypsy moths were divided into three well-known subspecies, a unique North American cluster, and a southern Chinese cluster with differentiation between the Asian gypsy moth and European gypsy moth. We also found individuals identified as European gypsy moths in two distant regions in China. The results of a quantitative analysis of male genitalia characteristics were consistent with an analysis of genetic structure and revealed the differentiation of gypsy moths in southern China and of hybrids suspected to be associated with L. d. japonica in the Russian Far East. Admixture in gypsy moths can be explained by many factors such as human transport. In China, we detected European gypsy moths, and found unexpectedly high genetic diversity within populations across a wide range of latitudes.

Expansion of LINEs and species-specific DNA repeats drives genome expansion in Asian Gypsy Moths

Two subspecies of Asian gypsy moth (AGM), Lymantria dispar asiatica and L. dispar japonica, pose a serious alien invasive threat to North American forests. Despite decades of research on the ecology and biology of this pest, limited AGM-specific genomic resources are currently available. Here, we report on the genome sequences and functional content of these AGM subspecies. The genomes of L.d. asiatica and L.d. japonica are the largest lepidopteran genomes sequenced to date, totaling 921 and 999 megabases, respectively. Large genome size in these subspecies is driven by the accumulation of specific classes of repeats. Genome-wide metabolic pathway reconstructions suggest strong genomic signatures of energy-related pathways in both subspecies, dominated by metabolic functions related to thermogenesis. The genome sequences reported here will provide tools for probing the molecular mechanisms underlying phenotypic traits that are thought to enhance AGM invasiveness.

Relating Aerial Deposition of Entomophaga maimaiga Conidia (Zoopagomycota: Entomophthorales) to Mortality of Gypsy Moth (Lepidoptera: Erebidae) Larvae and Nearby Defoliation

We collected data on mortality of late-instar gypsy moth, Lymantria dispar (L.), from outbreak populations over 4 wk in June 2017 at 10 sites in the New England region of the United States, along with estimated rainfall at these sites. Deposition of airborne conidia of the fungal pathogen, Entomophaga maimaiga Humber, Shimazu & R.S. Soper, was measured at these same sites as well as at seven other locations in New England. We also quantified the geographical distribution of gypsy moth-caused defoliation in New England in 2017 and 2018 from Landsat imagery. Weekly mortality of gypsy moth larvae caused by E. maimaiga correlated with local deposition of conidia from the previous week, but not with rainfall. Mortality from this pathogen reached a peak during the last 2 wk of gypsy moth larval development and always exceeded that caused by LdNPV, the viral pathogen of gypsy moth that has long been associated with gypsy moth outbreaks, especially prior to 1989. Cotesia melanoscela (Ratzeburg) was by far the most abundant parasitoid recovered and caused an average of 12.6% cumulative parasitism, but varied widely among sites. Deposition of E. maimaiga conidia was highly correlated with percent land area defoliated by gypsy moths within distances of 1 and 2 km but was not significantly correlated with defoliation at distances greater than 2 km. This is the first study to relate deposition of airborne conidia of E. maimaiga to mortality of gypsy moths from that agent.