The effect of abrupt salinity change on oxygen consumption in crab megalopae of Chincoteague Bay,USA

Variability in salinity is an environmental stressor that crab megalopae encounter as they are carried by tides and currents throughout Chincoteague Bay. We exposed blue crab (Callinectes sapidus) and fiddler crab (Uca spp.) megalopae to abrupt salinity changes from 10 to 31 ppt and measured their oxygen usage. It was hypothesized that the megalopae would cope with the changes in a manner reflective of the documented abilities and tolerances of adult crabs. It was also hypothesized that lower salinities would have a particularly detrimental effect on the megalopae reflected by both increased oxygen usage and mortality. The megalopae of both species did exhibit an increase in oxygen use at lower salinities, although the effect was more pronounced during the initial transition and decreased during acclimation. The megalopae mirrored the adult responses, with blue crab larvae consuming more oxygen per mg of wet weight at lower salinities, whereas fiddler crab larval oxygen consumption was relatively uniform at all salinities. Mortality of some blue crab postlarvae was observed at 10 ppt while all larval fiddler crabs survived. Coupled with the introduction of additional fresh water into the global water system, these results indicate that further investigation into this subject is necessary.     

Estimating diversity of crabs (Decapoda: Brachyura) in a no-take marine protected area of the SW Atlantic coast through DNA barcoding of larvae

DNA barcoding was used to identify crab larvae from the Marine Biological Reserve of Arvoredo, encompassing a coastal archipelago off the SW Atlantic coast (27°S, 48°W). Partial mitochondrial COI or 16S rRNA gene sequences were obtained for 488 larvae, leading to the identification of 20 species. The COI sequences generated 13 barcode index numbers (BINs) within Barcode of Life Data Systems (BOLD), among which 11 were concordant with single species. DNA from ～ 6% of the larvae did not amplify using the primers tested; based on external morphological characteristics, these larvae represented four possible additional operational taxonomic units (OTUs) at the family level. Intraspecific variation for the COI and 16S rRNA genes was found to be < 2.6% and < 2.1% respectively (Kimura 2-parameter distance), whereas interspecific divergence ranged from 7.9% to 21.5% and 6.4% to 14.5%, respectively. These results imply that both genes are suitable for use in species identification of brachyuran crabs of this area. Molecular identification of this group successfully enabled the diagnosis of larvae of closely related species, including congeners in Mithrax, Achelous and Callinectes. In addition, eight out of 20 species recognized represent new records for the reserve suggesting that the brachyuran fauna in the area has been underestimated based on traditional biodiversity measures. The availability of primers suited to the targeted species, and the development of a taxonomically comprehensive DNA barcoding database are the major recommendations to improve the accuracy and feasibility of using DNA barcoding for species identification of SW Atlantic brachyuran crabs.     

Parasitism,seasonality, and diversity of trombiculid mites (Trombidiformes: Parasitengona,Trombiculidae) infesting bats (Chiroptera) in Poland

The study aims to ascertain the diversity of trombiculid species associated with Chiroptera in Poland, and for the first time in the case of research on Central European Trombiculidae, we use both DNA and morphology in an integrative taxonomic approach to determine species identities of trombiculids. The research was carried out from 2015 to 2019. In total, 2725 larvae were collected from 300 specimens of bats belonging to 11 species. Deutonymphs were obtained through laboratory rearing of larvae; few larvae and deutonymphs were collected also from bats' daily roosts. The presence of trombiculid larvae on hosts was observed between July and April of the following year, with the highest numbers recorded in autumn, during bat swarming. Male bats were infested more often than females (16.4 vs. 6.6%). The highest infestation rate was recorded for Barbastella barbastellus, Myotis nattereri and Plecotus auritus, and the highest prevalence of chiggers (>?30%) for Myotis bechsteinii and P. auritus. The larvae found on bats occupied the areas with free access to the host’s skin: auricles, tragus, and snout. Morphological identification of specimens to the species level was hindered by the mosaic distribution of diagnostic traits. Morphological analyses indicated the presence of Leptotrombidium russicum and Leptotrombidium spp. in the examined material, whereas molecular analyses additionally suggested three other potential species assigned to the same genus based on the assessed scope of intrageneric variation (ASAP method). We argue that the identification of the parasitic larvae (chiggers) using morphological characters does not address the question of actual species boundaries, which, in turn, affects the inferences about host specificity and host range.

     

The adult body size variation of Dryas iulia (Lepidoptera,Nymphalidae, Heliconiinae) in different populations is more influenced by temperature variation than by host‐plant availability during the seasons

The body size of insects is affected by environmental conditions during development and can present considerable intraspecific variations, which can be seen as an ultimate consequence/adaptation to environmental conditions. This paper evaluated whether the body size of the butterfly Dryas iulia from subtropical populations was influenced by changing climate conditions and food source availability during the seasons. The likely reasons behind body size variation were also investigated. First, field data on body size variation, host‐plant availability and climate fluctuation throughout the seasons were recorded. Then, the effects of host‐plant species and temperature on body size were analyzed by controlled experiments. Field data revealed that body size and host‐plant availability varied significantly through the seasons. Populations had the smallest body size during the spring and the biggest size during summer, whereas host‐plant availability was lower during winter and higher during spring. The controlled experiments revealed that both temperature and host‐plant had significant effect on the plasticity of body size. Larvae subjected to winter temperature treatment led to smaller butterflies when compared to immatures reared under summer temperature treatment, and larvae fed with Passiflora misera produced bigger adults when compared to larvae reared on Passiflora suberosa. The combination of data gathered in the field and in the laboratory suggests that seasonal body size variation in D. iulia is related mainly to differences in the temperatures to which larvae are subjected during development, while host‐plant shifts caused by differential availability of food through the seasons had slightly effects on the variation observed.     

On the complementarity of DNA barcoding and morphology to distinguish benign endemic insects from possible pests: the case of Dirioxa pornia and the tribe Acanthonevrini (Diptera: Tephritidae: Phytalmiinae) in Australia

Fruit flies are considered economically important insects due to some species being agricultural pests. However, morphological identification of fruit fly adults and larvae can be difficult requiring a high level of taxonomic expertise, with misidentifications causing problematic false-positive/negative results. While destructive molecular techniques can assist with the identification process, these often cannot be applied where it is mandatory to retain a voucher reference specimen. In this work, we non-destructively (and partial-destructively) processed larvae and adults mostly belonging to the species Dirioxa pornia (Walker, 1849), of the poorly studied nonpest fruit fly tribe Acanthonevrini (Tephritidae) from Australia, to enable molecular identifications whilst retaining morphological vouchers. By retaining the morphological features of specimens, we confirmed useful characters for genus/species-level identification, contributing to improved accuracy for future diagnostics using both molecular and morphological approaches. We provide DNA barcode information for three species of Acanthonevrini known from Australia, which prior to our study was only available for a single species, D. pornia. Our specimen examinations provide new distribution records for three nonpest species: Acanthonevroides variegatus Permkam and Hancock, 1995 in South Australia, Acanthonevroides basalis (Walker, 1853) and D. pornia in Victoria, Australia; as well as new host plant records for D. pornia, from kangaroo apple, apricot and loquat.     

Latitudinal variation of morphological characteristics in the swallowtail Sericinus montelus Gray, 1798 (Lepidoptera: Papilionidae)

Patterns of latitudinal variation in the phenotype or genotype of an organism may provide evidence for natural selection. In this study, we investigated seven populations of swallowtail Sericinus montelus Gray, 1798 (Lepidoptera: Papilionidae), a non‐migratory species, to explore the latitudinal variation of morphological characteristics in adults. The results showed that body size and the development of dark pigmentation on wings in this species responded strongly to latitude. The body size of both male and female adult of S. montelus was negatively correlated with latitude. These findings provided solid evidence to support the converse Bergmann's rule. We considered that the observed variation in morphological characteristics was most likely mediated by the seasonal length and thermoperiod to adapt to different latitudinal environment (e.g. shortened developmental time of immature stages for smaller body size at higher latitude). Moreover, the tendency towards progressively darker colour patterns (only in adult males) at increasingly low latitudes was consistent with Gloger's rule. We suggested that the observed colour variation was most likely associated with thermoregulation. Slight variation in the morphology of the W‐shaped stripe on the forewing of adult females was also found, and we presumed that the functions of sexual preferences, mimicry and thermoregulation might be involved.     

Metamorphosis of mangrove crab megalopae, Ucides cordatus (Ocypodidae): Effects of interspecific versus intraspecific settlement cues

It has recently been shown that metamorphosis of Ucides cordatus megalopae is triggered by substrata from the mangrove forest habitat, and, in particular, adult conspecific odours. Here we demonstrate that the gender of the odour-emitting crabs is insignificant for the metamorphic response in this species. We further investigate whether other estuarine crabs (Goniopsis cruentata, Uca spp., and Callinectes danae) also induce settlement and metamorphosis of U. cordatus megalopae. This is of special interest for population recovery in areas hit by lethargic crab disease (LCD), a fungus that selectively kills U. cordatus but not co-occurring species. Ucides megalopae were reared in four treatments with interspecific-conditioned seawater and tested against the effects of conspecific-conditioned seawater (positive control) and pure seawater (negative control). All megalopae in the positive control metamorphosed successfully, while only one (2%) moulted in the negative control, with a delay of 10 days compared with the latest metamorphosis in the former treatment. In seawater conditioned with U. maracoani and C. danae, which occur on sediment banks and in tidal creeks respectively, all larvae died before reaching the juvenile stage. In the treatments with odours of species that share the same mangrove forest microhabitat as U. cordatus, i.e. G. cruentata and a group of five fiddler crab species (mixed-odour treatment), 20 and 10% respectively of the megalopae moulted with a delay of up to 11 days. No specimens metamorphosed after day 39, but megalopae lived up to 93 days. Since only the conspecific- and coexisting-species treatments stimulated development, we hypothesize that Ucides megalopae are able to precisely identify species-habitat-specific settlement cues. This will be investigated in more detail in future studies, which will also test the effects of the odours of the five forest fiddler crab species separately. The impact of the interspecific odour treatments was much smaller than that of the conspecific odours, nevertheless elevated moulting rates of up to 18% relative to seawater may still significantly accelerate the repopulation of U. cordatus in areas lacking conspecifics, e.g. after massive crab mortalities or at first colonization.     

Thermal attributes of Chrysomya species

The correct identification of forensically important arthropods for post‐mortem interval estimation is crucial, as the rate of larval development can vary substantially between species. The identification of forensically important blowflies of the genus Chrysomya (Diptera: Calliphoridae) may be hampered by their close morphological similarities, especially as immatures. The aim of this study was to establish whether genetically closely related blowfly species would share similar developmental profiles. This could permit the application of developmental data to a number of closely related species, including those for which thermodevelopmental studies are lacking. If Australian Chrysomya were found to share developmental profiles, identification of the blowfly specimen to a level beyond genus may not be necessary, or at least it may not be necessary to distinguish morphologically similar sister species. The three Chrysomya species studied were collected from the same geographical location (Cairns, Australia), reducing the effects of acclimation and population‐level genetic variation. The experimental conditions in this study were virtually identical, which enabled direct comparisons to be made among the species. Blowfly larval lengths were obtained for 24‐hourly intervals at constant temperatures of 25, 30, and 35 °C. The thermal preferences of newly‐hatched feeding larvae were determined by their positions on a temperature gradient apparatus. This study established that all three species investigated differed significantly in their developmental profiles, despite the genetic closeness of the sister species Chrysomya megacephala (Fabricius) and Chrysomya saffranea (Bigot). Because of this, genetic distance was not considered to be a useful factor for predicting thermodevelopment profiles of closely related species within a genus, and highlights the necessity for correct species identification.     

Butterflies of the South Pacific

ABSTRACT

Larvae of the invasive mosquito Culex quinquefasciatus Say are morphologically similar to those of the native Culex pervigilans Bergroth, yet distinguishing these species can be hampered by morphological variations in Cx. quinquefasciatus. We present detail about the extent of these variations in an urban population of Cx. quinquefasciatus in Auckland. To aid in identification of this exotic species, we provide images of key diagnostic characters and some observed exceptions to these. Details regarding variation in diagnostic characters for < 3rd instar and 3rd/4th instar larvae are given. Of the nine characters used for identification, three were highly consistent (dorsal papillae, mantle plate, pecten teeth); each observed in > 90% of larvae, although these characters were not always visible. Other characters were less reliable, for instance, the expected position of seta 1a-S in relation to the pecten teeth was observed in < 10% of larvae. Further exploration of regional morphological variation in both Cx. quinquefasciatus and Cx. pervigilans is recommended, ideally with associated molecular characterisation.     

Evergreen foliage allows early hatching in a pine processionary moth and escape from predation

1. Seasonal variation in leaf quality and climate conditions often imposes constraints on the temporal occurrence of tree‐feeding insect larvae, but the seasonal effects of predation have received limited attention. In temperate climate zones, both the abundance and activity of predators can be expected to vary over time. 2. The study reported herein examined the impact of temporal variation in predator activity levels on the life history of an herbivorous insect feeding on a constant food source: previous‐year needles of Scots pine (Pinus sylvestris L.). In field experiments, the survival and growth rates of colonies of Thaumetopoea pinivora Treitschke larvae that had been manipulated to hatch at three different dates were compared. Eggs of T. pinivora usually hatch by mid‐April in southern Sweden, which is earlier than most other herbivorous insects that overwinter as eggs in this region. 3. Predator exclusion experiments indicated that larvae which hatched later than April experienced a higher level of predator activity, mainly by ants. The final larval size and the timing of pupation were not affected by hatching date. First instar larvae were more extensively preyed on than second instars. 4. The life history of herbivore species can be affected by seasonal variation in predation pressures. This study suggests that early hatching in a lepidopteran species can allow a temporal escape from predation during the vulnerable early life stages.     

Test for latitudinal variation of life history,behavior and mortality in the strictly univoltine damselfly Sympecma fusca (Zygoptera: Lestidae)

A wealth of evidence shows that combinations of ecological stressors interact in shaping life history traits, but little is known about how ecological stressors combine with different seasonal time constraints to shape life history, behavior and mortality across populations. We studied life history, behavior and mortality rate in two latitudinally distant populations of the strictly univoltine, adult‐overwintering damselfly Sympecma fusca. Results from laboratory common‐garden and outdoor experiments indicated countergradient variation of larval development time and growth rate: the more time‐constrained larvae showed faster development and a higher growth rate. This finding led to larger size at emergence in the more time‐constrained individuals. Under conditions of intraspecific interaction (outdoor experiment), northern individuals showed lower survival than southern ones, presumably due to cannibalism. In the absence of intraspecific interactions (laboratory experiment), northern and southern larvae did not differ in survival. Finally, laboratory‐grown northern and southern larvae did not differ in activity level. This is the first time that compensation for seasonal time constraints has been shown in a temperate odonate species that overwinters in the adult stage.     

Complex selection on life-history traits and the maintenance of variation in exaggerated rostrum length in acorn weevils

Trophic interactions can trigger the development of exaggerated specialized characters and promote morphological diversification. For example, acorn weevils (genus Curculio) present strikingly long rostrums, which are used by females to perforate oviposition holes through the seed coat. Species exhibiting longer rostrums are known to exploit larger acorns, and therefore rostrum length is thought to be subject to selection to match the preferred acorn type. However, rostrum length is strongly correlated with body size, and morphological divergence could result from either selection on rostrum length for optimal food exploitation or from other pressures acting on body size. We collected infested acorns at oak forests where the large Curculio elephas and the small-bodied Curculio glandium co-occur. There were no interspecific differences in adult female body size to rostrum length allometric relationships, and rostrum length is equally correlated with body size in either species. MtDNA-based species identification showed that C. glandium larvae were present within acorns of all sizes, whereas C. elephas larvae were restricted to acorns above a minimum size, irrespective of oak species. Hence, exploitation of large acorns can hardly have triggered rostrum enlargement, as the small sized C. glandium adults (with short rostrums) could perforate and oviposit in both small and large acorns. Rather, increased rostrum length is probably a by-product of the larger body sizes of individuals emerging from bigger acorns, which allow increased larval size and enhance larval survival likelihood. Summarizing, when exaggerated feeding traits co-vary with other body features, interspecific morphological variability may result from contrasting selective pressures acting on these correlated characters.     

Seasonal diversity and dynamics of haptophytes in the Skagerrak,Norway, explored by high‐throughput sequencing

Microalgae in the division Haptophyta play key roles in the marine ecosystem and in global biogeochemical processes. Despite their ecological importance, knowledge on seasonal dynamics, community composition and abundance at the species level is limited due to their small cell size and few morphological features visible under the light microscope. Here, we present unique data on haptophyte seasonal diversity and dynamics from two annual cycles, with the taxonomic resolution and sampling depth obtained with high‐throughput sequencing. From outer Oslofjorden, S Norway, nano‐ and picoplanktonic samples were collected monthly for 2 years, and the haptophytes targeted by amplification of RNA/cDNA with Haptophyta‐specific 18S rDNA V4 primers. We obtained 156 operational taxonomic units (OTUs), from c. 400.000 454 pyrosequencing reads, after rigorous bioinformatic filtering and clustering at 99.5%. Most OTUs represented uncultured and/or not yet 18S rDNA‐sequenced species. Haptophyte OTU richness and community composition exhibited high temporal variation and significant yearly periodicity. Richness was highest in September–October (autumn) and lowest in April–May (spring). Some taxa were detected all year, such as Chrysochromulina simplex, Emiliania huxleyi and Phaeocystis cordata, whereas most calcifying coccolithophores only appeared from summer to early winter. We also revealed the seasonal dynamics of OTUs representing putative novel classes (clades HAP‐3–5) or orders (clades D, E, F). Season, light and temperature accounted for 29% of the variation in OTU composition. Residual variation may be related to biotic factors, such as competition and viral infection. This study provides new, in‐depth knowledge on seasonal diversity and dynamics of haptophytes in North Atlantic coastal waters.     

On the Roles of Morphological and Biochemical Criteria in Species Identification: An Example of Dragonfly Larvae of the Genus Aeschna

Dragonflies belong to the group of organisms with numerous well-differentiated species-specific characters at the adult stage, on the one hand, and a significantly smaller number or even the absence of such characters at the early ontogenetic stages. An example of the genus Aeschna is used to show difficulties in revealing morphological and biochemical characters allowing identification of larval dragonflies belonging to closely related species of the family. Distinct morphometric characters can be found only in late-instar larvae. The presence of species-specific proteins in the homogenates of thoracic muscles provides the possibility of using biochemical tests for species identification of larvae. Infestation by parasites has no effects on the biochemical parameters studied. Species identification of the early-instar dragonfly larvae is still problematic.     

Food sources for the early life history stages of the hydrothermal vent crab <Emphasis Type="Italic">Bythograea thermydron</Emphasis>: a stable isotope approach

We used stable isotope analysis to examine food sources for early life-history stages of the vent crab Bythograea thermydron. During two cruises to a hydrothermal vent site along the East Pacific Rise, we collected a variety of endemic organisms, including approximately 300 specimens of B. thermydron. The crab collection consisted mainly of megalopae and early juveniles, but also included adults and a single zoea. As expected, the carbon-isotope composition of newly released zoeae (−12.2%) was similar to the female (−11.1%) and clearly different from megalopae (21.7%). Because we were unsuccessful in culturing the zoea larvae, we were not able to conduct experiments to determine the effect of diet on isotopic composition. The tissue of megalopae was depleted in ¹³C (−21.7%) when compared to indigenous prey species, but was similar to that expected for carnivorous zooplankton dependent on surface primary production. The nitrogen-isotope composition of megalopae was enriched in ¹⁵N (10%) relative to potential prey species, again suggesting a photosynthetic source of primary production. The mean carbon-isotope composition of newly metamorphosed juvenile crabs (−19.9%) resembled megalopae, while tissue from subsequent juvenile stages was enriched in ¹³C (−10.9%) with values similar to those measured in co-occurring prey species. These results imply that megalopae are dependent on a source of primary production exogenous to the vents, but switch to a vent-based food web soon after metamorphosis to the juvenile stage.     

Seasonal variation in larval host plants and consequences for Heliconius erato (Lepidoptera: Nymphalidae) adult body size

Abstract Adult body size, a key life history component, varies strongly within and between Heliconius erato phyllis (Lepidoptera: Nymphalidae) populations. In the present study, we determined whether seasonal variation in adult body size is temperature related and/or determined by seasonal changes of host plants (Passifloraceae) used by the larval stage. A population of H. erato phyllis located in a Eucalyptus plantation (Barba Negra Forest, Barra do Ribeiro County, Rio Grande do Sul State, Brazil) was sampled every 45 days from March 1997 to October 1998 to quantify seasonal variation in adult body size and use of larval host plants. In the laboratory, the effects of the following factors on adult body size were quantified: (i) host plant species (Passiflora misera or Passiflora suberosa); (ii) food quantity consumed by larvae (experimentally manipulated for each passion vine species); (iii) winter and summer temperatures (15 and 25°C, respectively); and (iv) the interaction between host plant species and temperature. Adults emerging during summer were larger than those emerging in other seasons. Female butterflies oviposited selectively on P. misera even when the dominant passion vine was P. suberosa. They only switched from using P. misera to P. suberosa during later autumn and winter, when P. misera vines were completely defoliated. The laboratory feeding trials with both passion vines showed a strong positive association between food quantity consumed by larvae and adult size. They also confirmed that adults are larger when their larvae are reared on P. misera than on P. suberosa. Temperature during larval development had no effect on H. erato phyllis adult size. Thus, seasonal variation of H. erato phyllis adult size in a given place is primarily determined by the availability and quality of host plant species used by the larval stage.     

Insights into species diversity of the genus Hydropsyche Pictet, 1834 (Hydropsychidae,Trichoptera) from the Lake Kinneret catchment (Israel)

Species identification based on larval stages of aquatic insects in Israel is difficult, given the paucity of taxonomic studies. In order to gain deeper understanding of biological processes in Israel's freshwater ecosystems and to develop a bioindication system, species have to be known and characterised at all life stages. Here, we applied an integrative morphological and genetic approach to characterise caddisflies of the genus Hydropsyche Pictet, 1834 from the Lake Kinneret catchment. Seventeen sites in the Lake Kinneret catchment were sampled in spring/summer 2015 and 2016. For 77 hydropsychid larvae and two adult specimens of H. yahfufah Malicky, 2001, we analysed a 658 bp fragment of the mitochondrial COI gene and compared the results of different genetic species delimitation methods to observed morphological variation. Our results suggest the presence of at least five species. This study is an important first step towards assessing stream biodiversity of a yet largely understudied region and highlights advantages of integrative taxonomic approaches.     

Occurrence of an alternative pathway in the larval development of the crab Chasmagnathus granulata Dana, 1851 under laboratory conditions

Larvae of the crab Chasmagnathus granulata were collected in a salt marsh located in the Lagoa dos Patos, Brazil and reared from eclosion to metamorphosis under different dietary regimes. Larvae reared individually in beakers of 40 ml and fed Tetraselmis chuii (zoea III and zoea IV), showed a supplementary stage, here designated as zoea V, with morphological characteristics intermediary between zoea IV and megalopa. No zoeae V molted to megalopa stage. To confirm the occurrence of the supplementary stage, mass cultures of larvae of C. granulata were fed Artemia sp. at high densities, we again detected the fifth zoeal instar. However, when zoeae V were individually placed in beakers and fed Artemia nauplii, they succeeded in molting into megalopae. We observed the occurrence of two types of zoeae IV — a smaller type (from which originated the zoeae V) and a larger type (which directly developed into megalopae). We conclude that stressful nutritional/environmental conditions were responsible for the occurrence of this alternative path of development.     

A Quick Method for Species Identification of Japanese Eel (<Emphasis Type="Italic">Anguilla japonica</Emphasis>) Using Real-Time PCR: An Onboard Application for Use During Sampling Surveys

To compensate for the limited number of morphological characteristics of fish eggs and larvae, we established a convenient and robust method of species identification for eggs of the Japanese eel (Anguilla japonica) using a real-time polymerase chain reaction (PCR) that can be performed onboard research ships at sea. A total of about 1.2 kbp of the mitochondrial 16S ribosomal RNA gene sequences from all species of Anguilla and 3 other anguilliform species were compared to design specific primer pairs and a probe for A. japonica. This real-time PCR amplification was conducted for a total of 44 specimens including A. japonica, A. marmorata, A. bicolor pacifica, and 6 other anguilliform species. Immediate PCR amplification was only observed in A. japonica. We then tested this method under onboard conditions and obtained the same result as had been produced in the laboratory. These results suggest that real-time PCR can be a powerful tool for detecting Japanese eel eggs and newly hatched larvae immediately after onboard sampling during research cruises and will allow targeted sampling efforts to occur rapidly in response to any positive onboard identification of the eggs and larvae of this species.