Nonlethal sampling of DNA from critically endangered diving beetles (Coleoptera: Dytiscidae) using a single antenna

Nonlethal DNA sampling is highly desirable in molecular genetic studies of protected and endangered species. To develop a demonstrably nonlethal method of obtaining DNA from endangered diving beetles (Dytiscus sharpi sharpi Wehncke, Cybister lewisianus Sharp and Cybister brevis Aubé), we amputated the antennae of these endangered diving beetles and investigated the impact of the amputation on reproductive behaviors, egg‐laying and lifespan. Diving beetles with either one or no antennae copulated without delay and laid eggs, comparable to the pairs of intact beetles under breeding conditions. The lifespan of antennae‐amputated D. sharpi sharpi was the same as that of the intact beetles. A single antenna was sufficient to allow polymerase chain reaction (PCR) detection of a mitochondrial DNA gene, cytochrome‐c oxidase subunit I (COI), and the sequence of the COI gene could be determined directly. The PCR‐ready genomic DNA was available both in fresh antennae isolated from living beetles and in old antennae from whole beetles preserved for at least 5–6 years in pure ethanol. These results suggest that an antenna is a good sampling site for isolating genomic DNA from endangered diving beetles without sacrificing and disturbing reproductive behaviors such as mating and egg‐laying, or lifespan.     

Evaluation of traps of six different designs to capture pine sawyer beetles (Coleoptera: Cerambycidae)

1 Sawyer beetles (Coleoptera: Cerambycidae) and metallic wood‐boring beetles (Coleoptera: Buprestidae) attack recently killed or felled trees. They are attracted to potential hosts by volatiles and possibly by visual cues. We examined traps of six different designs to determine the role of silhouette and shape in the capture of pine sawyer beetles, Monochamus L. 2 Traps with a round black cylinder, a single vane (clear or black), or cross vanes (clear or black) all mounted above a pan were compared to an open pan. Each pan contained soapy water to retain insects. 3 Traps with a black silhouette were significantly more effective in capturing male and female Monochamus scutellatus (Say) than traps with or without clear vanes. For both sexes of M. mutator LeConte, pan traps were as effective as traps with a black silhouette. Silhouette was not important for the capture of male and female M. notatus (Drury). Trap catches for the six trap types were not significantly different for male Buprestis maculativentris Say. 4 A pan trap with a black silhouette is recommended for general use in capturing wood‐boring cerambycids and buprestids.     

Beyond origami: using behavioural observations as a strategy to improve trap design

In contrast to ad hoc methods of developing traps for pest monitoring systems, a systematic approach using direct observation of animals allowed a greater understanding of the reasons why trap catch was significantly different in traps of basically similar design. The information gained using this approach could then be used to guide the further development of the trap. The same feature on two related designs of cockroach trap was varied: the slope of the ramp leading into the trap was either 60°, 30°, or 0°. The 30° ramp version of both traps caught significantly more Blattella germanica (L.) (Dictyoptera: Blattellidae). The 60° and 0° ramp versions both caught equal, lower, numbers but observation showed that these net catches were achieved by quite different means; few insects entered over the 60° ramps but none escaped, whereas all entered over the 0° ramps but half escaped. Similar approaches could be applied to other insect-trap systems.     

Disrupting spring colonization of Colorado potato beetle to nonrotated potato fields

Overwintering Colorado potato beetles (Leptinotarsa decemlineata (Say)) were concentrated primarily within woody borders, and mortality was lower in borders than in potato fields. After overwintering, only 15–44% of live beetles were in the potato fields. In experiments with small plots, colonization of fields from woody borders was reduced ∼60% by a trap crop, either treated with adulticide or with beetles collected daily. Such trap crops, or simply pitfall traps to prevent colonization from woody borders, could significantly reduce early-season adult numbers and subsequent larval populations. However, success is dependent on the local densities of overwintered beetles, and the prevalence of arrestment behavior in the case of trap crops.     

Migration to and dispersal from oilseed rape by the pollen beetle,Meligethes aeneus,in relation to wind direction

1 Host‐plant‐odour‐induced upwind anemotaxis is accepted as the main mechanism by which herbivorous insects find their host plant within an heterogenous environment, but field data supporting this hypothesis are scarce. 2 The flight directions of the pollen beetle Meligethes aeneus to and from a plot of winter oilseed rape and the direction of the wind were recorded concurrently. The beetles were sampled by eight double‐sided window traps encircling the plot. Distal sides of the traps, relative to the plot, sampled the beetles as they flew towards the plot, whereas the proximal sides of the traps sampled them as they flew away from the plot. Paired data on daily catch of beetles in distal or proximal traps and the volume of air impacting each trap were compared. 3 Correlations between daily catch of M. aeneus into distal traps and trap wind volume were negative, indicating that flights by both overwintered‐ and new‐generation insects towards the plot were upwind. 4 Correlations between daily catch of M. aeneus into proximal traps and trap wind volume varied with generation. Catch of overwintered‐generation M. aeneus was negatively correlated with trap wind volume, indicating that flight was upwind. Proximal trap catch of the new‐generation M. aeneus was weakly/modestly positively correlated with trap wind volume, indicating that flights were downwind or crosswind. 5 Understanding the effect of wind direction on flight of M. aeneus holds potential for improving the forecasting of their arrival and spatial distribution on the crop for integrated pest management.     

Development of Odour-Baited Flytraps for Sampling the African Latrine Fly,Chrysomya putoria,a Putative Vector of Enteric Diseases

African pit latrines produce prodigious numbers of the latrine fly, Chrysomya putoria, a putative vector of diarrhoeal pathogens. We set out to develop a simple, low-cost odour-baited trap for collecting C. putoria in the field. A series of field experiments was carried out in The Gambia to assess the catching-efficiency of different trap designs. The basic trap was a transparent 3L polypropylene box baited with 50 g of fish, with a white opaque lid with circular entrance holes. We tested variations of the number, diameter, position and shape of the entrance holes, the height of the trap above ground, degree of transparency of the box, its shape, volume, colour, and the attractiveness of gridded surfaces on or under the trap. Traps were rotated between positions on different sampling occasions using a Latin Square design. The optimal trapping features were incorporated into a final trap that was tested against commercially available traps. Features of the trap that increased the number of flies caught included: larger entrance holes (compared with smaller ones, p<0.001), using conical collars inside the holes (compared with without collars, p = 0.01), entrance holes on the top of the trap (compared with the side or bottom, p<0.001), traps placed on the ground (compared with above ground, p<0.001), the box having transparent sides (compared with being opaque, p<0.001), and with no wire grids nearby (compared with those with grids, p = 0.03). This trap collected similar numbers of C. putoria to other common traps for blow flies. The optimum trap design was a transparent box, with a white plastic lid on top, perforated with 10 conical entrance holes, placed on the ground. Our simple trap provides a cheap, low-maintenance and effective method of sampling C. putoria in the field.     

Diving in head first: trade‐offs between phenotypic traits and sand‐diving predator escape strategy in Meroles desert lizards

Survival, in part, depends on an individual's ability to evade predators. In desert regions some lizard species have evolved head‐first sand‐diving strategies to escape predators. To facilitate this behaviour, a distinctive head morphology that facilitates sand‐diving has evolved. This specialised head morphology may, however, come at a cost to other ecologically relevant functions, particularly bite force. Here, we investigated the relationship between morphology and function in a southern African lacertid lizard genus, Meroles, which consists of eight species that utilise different escape strategies, including sand‐diving and running for cover. It was hypothesized that the specialised head morphology of diving species would negatively affect bite force capacity. We found that species from each escape strategy category differed significantly in head shape, but not bite force performance. A phylogenetic tree of the genus was constructed using two mitochondrial and two nuclear genes, and we conducted phylogenetic comparative analyses. One aspect of the head shape differed between the escape strategies once phylogeny was taken into account. We found that bite force may have co‐evolved with head morphology, but that there was no trade‐off between biting capacity and escape strategy in Meroles.     

Specific detection of the floodwater mosquitoes Aedes sticticus and Aedes vexans DNA in predatory diving beetles

Floodwater mosquitoes (Diptera: Culicidae) are associated with periodically flooded wet meadows, marshes, and swamps in floodplains of major rivers worldwide, and their larvae are abundant in the shallow parts of flooded areas. The nuisance caused by the blood‐seeking adult female mosquitoes motivates mosquito control. Larviciding with Bacillus thuringiensis israelensis is considered the most environmentally safe method. However, some concern has been raised whether aquatic predatory insects could be indirectly affected by this reduction in a potential vital prey. Top predators in the temporary wetlands in the River Dalälven floodplains are diving beetles (Coleoptera: Dytiscidae), and Aedes sticticus and Ae. vexans are the target species for mosquito control. For detailed studies on this aquatic predator–prey system, we developed a polymerase chain reaction (PCR) assay for detection of mosquito DNA in the guts of medium‐sized diving beetles. Primers were designed for amplifying short mitochondrial DNA fragments of the cytochrome C oxidase subunit I (COI) gene in Ae. sticticus and Ae. vexans, respectively. Primer specificity was confirmed and half‐life detectability of Ae. sticticus DNA in diving beetle guts was derived from a feeding and digestion experiment. The Ae. sticticus DNA within diving beetle guts was detected up to 12 h postfeeding, and half‐life detectability was estimated to 5.6 h. In addition, field caught diving beetles were screened for Ae. sticticus and Ae. vexans DNA and in 14% of the diving beetles one or both mosquito species were detected, showing that these mosquito species are utilized as food by the diving beetles.     

Ability of stress‐related volatiles to attract and induce attacks by Xylosandrus germanus and other ambrosia beetles

• 1 Xylosandrus germanus typically colonizes physiologically‐stressed deciduous hosts but it is increasingly being recognized as a key pest of ornamental nursery stock. We tested the attractiveness of common plant stress‐related volatiles to ambrosia beetles occupying the nursery agroecosystem, as well as their ability to induce attacks on selected trees. Experiments were conducted in Ohio, U.S.A.
• 2 Stress volatile attractiveness was first assessed by positioning traps baited with acetaldehyde, acetone, ethanol and methanol in ornamental nurseries. Cumulative trap counts confirmed that ethanol was the most attractive stress‐related volatile to X. germanus. Methanol‐baited traps were slightly attractive to X. germanus, whereas traps baited with acetaldehyde and acetone were not attractive to any ambrosia beetle.
• 3 A series of tree injection experiments were also conducted to determine the ability of these volatiles to induce attacks by ambrosia beetles under field conditions. Injection of ethanol into Magnolia virginiana induced the largest number of attacks, whereas injection of acetaldehyde induced more attacks than methanol or acetone. Xylosandrus germanus was the most predominant species emerging from M. virginiana injected with each of the stress‐related volatiles. No attacks by wood‐boring beetles were observed on water injected or uninjected control trees.
• 4 Solid‐phase microextraction–gas chromatography–mass spectrometry confirmed the emission of acetaldehyde, acetone, ethanol and methanol after their injection into M. virginiana.
• 5 Xylosandrus germanus has an efficient olfactory‐based mechanism for differentiating among host volatile cues. Injecting select trees with stress‐related volatiles, particularly ethanol, shows promise as a trap tree strategy for X. germanus and other ambrosia beetles.

     

Noninvasive sampling of DNA from larval exuvia in diving beetles of the genera Cybister and Dytiscus

Nonlethal DNA sampling is a highly recommendable method in molecular genetic studies of protected and endangered species. To develop a demonstrably nonlethal method of obtaining DNA from larvae of endangered diving beetles (Cybister brevis, C. lewisianus, C. limbatus, C. rugosus, Dytiscus sharpi sharpi and D. sharpi validus), we obtained the larval exuvia (molted skin) of these endangered diving beetles under laboratory conditions. A single exuvia 24 h after molting was sufficient to allow polymerase chain reaction (PCR) detection of a mitochondrial DNA gene, cytochrome‐c oxidase subunit I (COI), and the sequence of the COI gene could be determined directly. Sequences obtained from the exuvial samples were used to further find similarities within DDBJ/EMBL/GenBank. Genomic DNA from the samples was successfully isolated, and we identified the species. This process suggests that exuvia provides a good sample for extracting DNA from endangered diving beetle larvae without killing them.     

Defining the fire trap: Extension of the persistence equilibrium model in mesic savannas

Mesic savannas are dominated by trees that are strong resprouters caught in a frequent fire trap. Persistence within this fire trap has been described by a resprout curve of SizeNext ~ f(Pre‐fire size), defined by the Michaelis‐Menten function. A key feature of this resprout curve is a stable persistence equilibrium that represents the size of individual plants upon which a population will converge over successive inter‐fire time steps under a given fire regime. Here, we contend that such a resprout curve does not adequately describe resprout tree dynamics in frequently burnt mesic savannas because it is constrained to an asymptote. We propose a new framework for modelling the resprout curve, which recognizes that local environmental stochasticity and growth patterns can interact to change the growth response function entirely, and thus more readily reflect the range of feasible resprout responses. Importantly, we define an unstable equilibrium representing the size above which individuals have escaped the fire trap and explore mechanisms that can shift an individual from persistence to escape. Through a case study from northern Australia, we confirm that our framework provides a simple yet practical approach to defining these critical aspects of savanna tree growth dynamics: persistence and escape.     

The Tauber Pollen Trap,a Discussion of Its Usefulness in Pollen Deposition Studies

In connection with pollen deposition studies some discrepancies between Tauber traps and open collectors were discovered. Field observations showed that the aerodynamic lid of the Tauber trap itself acted as a pollen trap by redeposition e.g. because of rainsplash. In an experiment coal dust in the size range of pollen was introduced in a turbulent air stream passing over the trap. Great quantities of coaldust were captured on the aerodynamic lid. By exposure to artificial rain it was observed that the splashing effect of raindrops brought coal dust into the sampler. By application of Corylus pollen on the aerodynamic lid and exposure to rain, it was found that great amount of pollen is transferred by raindrop splash to the trap. This contamination of the pollen contents of the trap is so serious that the usefulness of the Tauber trap for quantitative evaluation of the pollen deposition is doubted unless the trap is protected in some way. Further tests of this effect have been planned.     

Measuring trap efficiency for bark beetles (Col., Scolytidae)

Abstract:  The relative efficiency of cylindrical, linear and cross-barrier traps for trapping bark beetles was investigated based on a theoretical model. Using this model, the effective trap interception area of each trap type was calculated and trap efficiency was defined as the ratio of the effective interception area to the trap surface area. The relative efficiencies of the three trap types were calculated as the ratios of their respective effective interception areas. Based on this approach, assuming random directional movement of dispersing beetles, the order of efficiency of the three trap types, from highest to lowest, was linear, cross-barrier and cylindrical. The expected ratios of trap catches based on the relative efficiencies of the three trap types were fitted to data from trapping experiments with the mountain pine beetle ( Dendroctonus ponderosae Hopkins). In general, there was large variation in trap catches among traps of the same type but the ratios of mean catches per trap conformed to the expected ratios. The results indicate that the model of trap efficiency could be used for designing efficient traps. The methods presented are amenable for assessing the efficiency of other trap designs.     

Cracks or holes in the stems of oviposition plants provide the only exit for hatched larvae of diving beetles of the genera Dytiscus and Cybister

Diving beetles such as Dytiscus and Cybister species (Coleoptera: Dytiscidae) usually oviposit inside an aquatic plant stem beneath the surface of the water. The hatched larvae need to escape from the stem to intake oxygen from the air. To determine where larvae of these diving beetles hatch in the plant stem, the hatchability and escape rates in larvae of Dytiscus sharpi Wehncke, Cybister chinensis Motshulsky, Cybister lewisianus Sharp, and Cybister brevis Aubé were investigated under laboratory conditions. Hatchability of D. sharpi in the stem of Sagittaria trifolia L. (Alismataceae) was extremely low (8.2%). However, it was high (>90%) when late‐stage eggs (2–3 days before hatching) were isolated from the stem and kept in water. On the other hand, the hatchability of Cybister spp. was high (88–95%) in S. trifolia. Usually, Cybister spp. females bite a hole in the plant stem on oviposition. When the oviposition pore in the stem was plugged with glass wool, no larvae could escape from the stem, indicating that the oviposition pore was the only exit for hatched larvae of Cybister spp. In contrast, females of D. sharpi oviposited directly by making a crack in the stem of Oenanthe javanica (Blume) DC. (Apiaceae) without biting. Eggs grew to a length and diameter equal to the stem crack size 2–3 days before hatching. Dytiscus sharpi eggs isolated from O. javanica were artificially inserted into plant stems of O. javanica or S. trifolia (so‐called inserted egg model), and the hatchability and larval escape rates were determined. Larval escape strongly depended on the stem crack width of both O. javanica and S. trifolia, suggesting that the stem crack was an exit for hatched larvae of D. sharpi.     

A portable,remote‐controlled nest‐box trap

Many researchers catch adult birds at nest boxes using a vertical prop that supports a horizontal flap that drops down when an adult enters to feed nestlings. Because the prop is visible, some birds may be too wary to enter nest boxes, or they may dislodge the prop when they lean in; either way, they are not captured. We describe a remote‐controlled nest‐box trap that has the advantages of portability and being invisible to adults delivering food to nestlings. A receiver is installed on the ceiling of a nest box, the receiver is connected to a spring‐loaded flap that is triggered remotely with a transmitter from > 30 m away, and the flap drops down to cover the box entrance. In 2017 and 2018, our remote‐controlled traps did not increase the likelihood of capturing female Tree Swallows (Tachycineta bicolor), but did improve the likelihood of capturing males. We captured 16 male Tree Swallows in 30 attempts (53.3%) using our remote‐controlled trap compared to only 41 captures of males in 139 attempts (29.5%) with prop traps. In addition, whereas prop traps required an average of ~ 50 min to capture adults in successful attempts, our trap required only ~ 25 min. These results suggest that the savings in time for field researchers using our remote‐controlled trap can be substantial, with the added ethical benefit of reducing the amount of time that nestlings are not being fed. Our remote‐controlled trap is also economical to construct, requiring ~ $60 USD for parts and < 3 h to build.     

A carbon dioxide, heat and chemical lure trap for the bedbug, Cimex lectularius

Abstract A trap for the collection of bedbugs, Cimex lectularius Linnaeus (Hemiptera: Cimicidae), is described. The trap was baited with CO₂ (50–400 mL/min), heat (37.2–42.2 °C) and a chemical lure comprised of 33.0 μg proprionic acid, 0.33 μg butyric acid, 0.33 μg valeric acid, 100 μg octenol and 100 μg L‐lactic acid, impregnated into a gel. Laboratory studies, conducted in a square arena measuring 183 cm on each side, showed that traps with and without baits captured adult bedbugs, but traps with CO₂ emissions of 50–400 mL/min caught significantly (P < 0.05) more bedbugs than traps without CO_2. In an infested unoccupied apartment, traps with heat and with or without the chemical lure were tested without CO₂ on 29 trap‐days and with CO₂ on 9 trap‐days. The numbers of bedbugs captured were 656 and 5898 in traps without and with CO₂, respectively. The numbers of bedbugs of all development stages captured were significantly greater in traps with CO₂ (χ² = 15 942, d.f. = 1, P < 10^?9). A non‐parametric two‐way analysis of variance evaluation of six different traps with or without CO₂, heat or a chemical lure monitored over 19 trap‐days in an infested apartment showed that trap type was highly significant (n = 2833 bedbugs collected) (P < 10^?7). The trap with CO₂, heat and a chemical lure captured more bedbugs than the other traps, but only caught significantly more fourth and fifth instar nymphs than all other traps. Otherwise, the catches in this trap did not differ significantly from those caught by traps that contained CO₂ and heat only. The total numbers of bedbugs collected for each trapping date (pooling all six traps) followed an exponential decline over the trapping period. This type of trap, which caught bedbugs in unoccupied apartments with and without furniture, and in an occupied apartment, may have utility in studying the ecology of bedbugs, in detecting bedbug infestations and in reducing numbers of bites by trapping host‐seeking bedbugs.     

Movements and dispersal of farmed Atlantic salmon following a simulated-escape event

One of the major challenges for understanding the ecological impact of escaped farmed Atlantic salmon (Salmo salar L.) is predicting their dispersal patterns after an escape event. Here, we quantify the behaviour of escaped farmed salmon using a simulated-escape experiment within a Norwegian fjord system. Thirty-seven individuals were tagged with acoustic transmitters and their spatial distribution, horizontal movements and diving behaviour was monitored throughout the fjord and accompanying rivers using 29 acoustic receivers. A rapid movement away from the release site occurred. There was no movement into the rivers feeding the fjord and there was no preference for residence at the fish farms: nearly a third of the individuals had a final detection in the outer part of the fjord and no detections occurred from approximately 2 months after release. As it is unlikely that all fish died or remained undetected within the study area it is reasonable to assume that a substantial proportion of the fish moved out of the fjord system. If a recapture program is not implemented immediately after an escape event, we recommend spreading potential recapture efforts over a relatively large area.     

Effect of water depth on predation frequency by diving beetles on mosquito larvae prey

Diving behavior and its frequency may differ among species of mosquito larvae because of differences in predation pressure. The present study aimed to investigate the relationship between water depth and predation frequency on two mosquito species, Culex tritaeniorhynchus (wetland breeder) and Aedes albopictus (container breeder), by the diving beetle Eretes griseus. Culex tritaeniorhynchus spends more time at the surface than A. albopictus, which spends more time thrashing underwater. When intact mosquito larvae of both species were present, the diving beetles consumed almost all A. albopictus larvae (98.3%). After all the A. albopictus larvae had been consumed, the diving beetles began to prey on C. tritaeniorhynchus. In order to compare the effect of position on the predation preference of the diving beetles, equal numbers of both species were heat‐killed and allowed to settle on the bottom of the container. When all the dead mosquito larvae had sunk to the bottom of a plastic container, the diving beetles caught both mosquito species at random. These results indicate that mosquito larvae near the surface were eaten less frequently by diving beetles than those at the bottom. The low diving frequency of C. tritaeniorhynchus is regarded as a form of anti‐predatory behavior.     

Standardized capture rates of the endangered American burying beetle, Nicrophorus americanus Olivier (Coleoptera: Silphidae) using different trap protocols

The U.S. Fish and Wildlife Service has adopted two different pitfall trap protocols to survey the endangered American burying beetle, Nicrophorus americanus. One protocol uses a transect of eight pitfall traps that prohibit bait contact over the course of three trap nights. The other protocol uses buckets that allow for bait contact over a five night trapping period. A trap night is defined as one trap open for one night, and the transect protocol has historically been calculated as eight trap nights per calendar night while; the bucket protocol has been calculated as one trap night per calendar night. This study examined the effectiveness of each protocol based on the number of beetles (Nicrophorus spp.) captured per trap night (BTN) in field and laboratory trials. When each transect was considered as a single trap instead of eight separate traps, no significant difference in BTN was detected between the protocols in any year. Laboratory trials were conducted using Nicrophorus marginatus to determine differences in capture efficiency based on protocol, time after release, and feeding status. The proportion of beetles captured after 5 days was greater when compared to 3 days for either protocol. Our results indicate that cup transects used to sample the American burying beetle should be considered as a single trap when calculating trap night and that the use of five trap nights rather than three would increase the likelihood of capturing beetles.     

Minimizing Leg-Hold Trapping Trauma for Otters With Mobile Phone Technology

ABSTRACT We present and evaluate a protocol for the capture of otters (Lutra lutra) using padded leg-hold traps coupled with Global System for Mobile communication (GSM) trap alarms. The trapping method was highly efficient, capturing 46 otters at 6.9 trap-nights each. Functioning alarms allowed us to remove 36 otters from their traps within 22 (SD = 14) minutes of capture. We caught 10 otters in trap sets with malfunctioning trap alarms and retrieved them the following morning, after ≤24 hours. Functioning alarms reduced the injuries suffered from an average cumulative score of 77.7 to just 5.5 on the International Organization for Standardization 10990-5 trauma scale (Z=-5.074, P ≤ 0.001). As a result, we strongly encourage the use of GSM trap alarms under the principle of refinement in animal experiments.