Self-Recognition and Allorecognition Mechanisms Exert a Significant Influence on the Sex Allocation Patterns of the Pea Aphid.

The mechanism controlling sex allocation in the pea aphid, Acyrthosiphon pisum (Harris), remains a crucial yet unresolved issue in the field of evolutionary ecology. This study aims to assess the influence of the presence of both self and non-self clones, along with juvenile hormone III (JH III) titer, on the sex allocation of aphid offspring. To this end, red and green clones were utilized as experimental subjects, and the agar method was employed. Initially, three distinct experimental treatments were established using sexuparae, and the daily offspring count and sex allocation in each treatment zone were recorded. Subsequently, an additional experimental condition involving mixed-clone treatments was introduced. This procedure entailed the transfer of a single sexupara and 20 oviparous females from either the red (1G + 20Rov) or green clone (1G + 20Gov) onto a leaf on agar medium. Simultaneously, a control setup with a new sexupara (1G) was established. Three days following sexupara production, a dose of 0, 25, or 50 ng of JH III was applied to the aphids' abdomens. Subsequently, the titers of JH III in the sexuparae across each treatment group were quantified, and the extent of sex allocation was tallied. The findings demonstrated pronounced disparities in sex allocation among the various treatments and, notably, a substantial increase in the total offspring and oviparous number in the mixed-clone treatment group. The effects of mixed-clone treatment on the sex allocation patterns of the sexupara progeny could be determined by the application of exogenous JH III, indicating that JH may mediate the effects of mixed-clone treatment on sex allocation. Consequently, it can be concluded that A. pisum sexuparae possess the capability to modulate their sex allocation in response to the nature of adjacent competitor clones, thereby demonstrating a variety of sex allocation patterns. Throughout this process, JH III plays a pivotal role.

Interspecific recognition based on cuticular hydrocarbons mediates reproduction control in aphids.

The preset study tested whether an aphid species can control its reproduction by recognizing the presence and density of a rival species. Acyrthosiphon pisum and Megoura crassicauda often coexist on the same leguminous plant. We established clonal colonies from each species and mixed colonies with one A. pisum and one M. crassicauda adult. There were no significant differences in the population growth patterns of the two species at 20 °C. However, mixed colonies increased faster and attained larger colony sizes than the clonal colonies. Thus, positive interspecific interactions were confirmed. A mixed colony was dominated by the members of a clone that produced a greater number of newborns in the initial stage, irrespective of the species. Thus, we confirmed the priority effect in the interspecific competition. To simulate the priority effect, 15 glass beads coated with the hexane extract of M. crassicauda aphids were attached to a cut leaf, to which one A. pisum adult was transferred. The presence of the hexane extract of M. crassicauda greatly reduced the reproductive rate of A. pisum adults. We conclude that aphids can control their reproduction by evaluating the relative density of rivals to fellow aphids based on the cuticular hydrocarbons.

Taxonomic revision of the Tetraneura akinire species group (Insecta, Aphididae, Eriosomatinae), with description of a new species and a correction of a nomenclatural confusion.

Gall-forming aphid species Tetraneura nigriabdominalis and T. fusiformis and its closely related species are taxonomically revised. By referring to the original descriptions, the name T. nigriabdominalis (Sasaki, 1899) is discarded as an erroneous combination, and T. akinire Sasaki, 1904 is adopted as a valid name. The T. akinire species group is defined as having long claws in the first instar nymphs of the root generation. Of the T. akinire species group distributed in Korea and Japan, T. ovaliformis sp. nov., which induces globular galls on the leaves of Ulmus davidiana var. japonica, is described, and T. akinire and T. sorini Hille Ris Lambers, 1970 are redescribed. A molecular phylogeny based on partial sequences of mitochondrial cytochrome c oxidase subunit I (COI) indicates that T. akinire is composed of two clusters, one (type A) which is distributed widely from Europe to East Asia on Ulmus spp., and the other (type B) which is found in Hokkaido, northern Japan on U. davidiana var. japonica and in tropical regions as anholocyclic lineages. Tetraneura fusiformis Matsumura, 1917, which has often been treated as a junior synonym of T. nigriabdominalis, likely corresponds to type B. The taxonomic status of T. fusiformis is discussed and this species is tentatively considered as a junior synonym of T. akinire sensu novo.

Self and non-self recognition affects clonal reproduction and competition in the pea aphid.

The spatial interaction of clonal organisms is an unsolved but crucial topic in evolutionary biology. We evaluated the interactions between aphid clones using a colour mutant (yellow) and an original (green) clone. Colonies founded by two aphids of the same clone and mixed colonies, founded by a green aphid and a yellow aphid, were set up to observe population growth for 15 days. We confirmed positive competition effects, with mixed colonies increasing in size more rapidly than clonal colonies. In mixed colonies where reproduction started simultaneously, green aphids overwhelmed yellow aphids in number, and yellow aphids restrained reproduction. However, when yellow aphids started to reproduce earlier, they outnumbered the green aphids. To test whether aphids have the ability to control reproduction according to the densities of self and non-self clones, one yellow aphid or one antennae-excised yellow aphid was transferred into a highly dense green clone colony. Intact yellow aphids produced fewer nymphs in crowded green colonies, whereas the fecundity of antennae-excised aphids did not change. Thus, we conclude that aphid clones can discriminate between self and non-self clones, and can regulate their reproduction, depending on whether they are superior or inferior in number to their competitors.

Liquid Marbles in Nature: Craft of Aphids for Survival.

Some aphids that live in the leaf galls of the host plant are known to fabricate liquid marbles consisting of honeydew and wax particles as an inner liquid and a stabilizer, respectively. In this study, the liquid marbles fabricated by the galling aphids, Eriosoma moriokense, were extensively characterized with respect to size and size distribution, shape, nanomorphology, liquid/solid weight ratio, and chemical compositions. The stereo microscopy studies confirmed that the liquid marbles have a near-spherical morphology and that the number-average diameter was 368 ± 152 µm, which is 1 order of magnitude smaller than the capillary length of the honeydew. The field emission scanning electron microscopy studies indicated that micrometer-sized wax particles with fiber- and dumpling-like shapes coated the honeydew droplets, which rendered the liquid marbles hydrophobic and nonwetting. Furthermore, the highly magnified scanning electron microscopy images confirmed that the wax particles were formed with assemblage of submicrometer-sized daughter fibers. The contact angle measurements indicated that the wax was intrinsically hydrophobic and that the liquid marbles were stabilized by the wax particles in the Cassie-Baxter model. The weight ratio of the honeydew and the wax particles was determined to be 96/4, and the honeydew consisted of 19 wt % nonvolatile components and 81 wt % water. The 1H nuclear magnetic resonance, Fourier transform infrared spectroscopy, and mass spectroscopy studies confirmed that the wax mainly consisted of triglycerides and that the honeydew mainly consisted of saccharides (glucose and fructose) and ribitol. The atomic force microscopy studies confirmed that honeydew is sticky in nature.

Effects of Radiation From Contaminated Soil and Moss in Fukushima on Embryogenesis and Egg Hatching of the Aphid Prociphilus oriens.

Radiation-contaminated soils are widespread around the Fukushima Daiichi Nuclear Power Plant, and such soils raise concerns over its harmful effect on soil-dwelling organisms. We evaluated the effects of contaminated soil and moss sampled in Fukushima on the embryogenesis and hatching of aphid eggs, along with the measurement of the egg exposure dose. Cs-137 concentration in soil and moss from Fukushima ranged from 2200 to 3300 Bq/g and from 64 to 105 Bq/g, respectively. Eggs of the eriosomatine aphid Prociphilus oriens that were collected from a non-contaminated area were directly placed on the soil and moss for 4 or 3 months during diapause and then incubated until hatching. The total exposure dose to the eggs was estimated as ca. 100-200 mGy in the 4-month soil experiment and 4-10 mGy in the 4-month moss experiment. There was no significant difference in egg hatchability between the contaminated soil treatment and the control. No morphological abnormalities were detected in the first instars that hatched from the contaminated soil treatment. However, we found weak effects of radiation on egg hatching; eggs placed on the contaminated moss hatched earlier than did the control eggs. On the contaminated soil, the effects of radiation on egg hatching were not obvious because of uncontrolled environmental differences among containers. The effects of radiation on egg hatching were detected only in containers where high hatchability was recorded. Through the experiments, we concluded that the aphid eggs responded to ultra-low-dose radiation by advancing embryogenesis.

Transgenerational seasonal timer for suppression of sexual morph production in the pea aphid, Acyrthosiphon pisum.

Many aphid species switch reproductive modes seasonally, with the sexual generations appearing in autumn. Sexual generations are induced by short days. It has been reported that the appearance of sexual morphs is suppressed by a transgenerational factor (a seasonal timer) over several generations after hatching from overwintered eggs. The present study examined whether the seasonal timer measures the number of days from hatching or the number of generations from hatching using the pea aphid, Acyrthosiphon pisum Harris (Homoptera: Aphididae). Effects of temperature and photoperiod on the seasonal timer were also examined by successive rearing. The ability to produce sexual morphs was strongly suppressed in stem mothers (the foundress generation), and gradually recovered over successive generations produced during a few months. The duration for which the seasonal timer could function depended on the number of days from hatching and temperature, but not on photoperiod or the number of generations from hatching. We thus showed in a single study that the seasonal timer of the pea aphid has all the physiological characteristics shown in separate studies in different aphid species.

Cryptic diversity of the subfamily Calaphidinae (Hemiptera: Aphididae) revealed by comprehensive DNA barcoding.

Aphids are a species rich group comprising many important pests. However, species identification can be very difficult for aphids due to their morphological ambiguity. DNA barcoding has been widely adopted for rapid and reliable species identification as well as cryptic species detection. In this study, we investigated cryptic diversity in the subfamily Calaphidinae (Hemiptera: Aphididae) based on 899 sequences of cytochrome c oxidase I (COI) for 115 morphospecies (78 species collected in this study and sequences of 73 species downloaded from Genbank). Among these 115 morphospecies, DNA barcoding results of 90 (78.3%) species were identical to results of morphological identification. However, 25 (21.7%) morphospecies showed discrepancies between DNA barcoding and traditional taxonomy. Among these 25 discordances, a total of 15 cryptic species were identified from 12 morphospecies. We also found three morphologically distinct species pairs that sharing DNA barcoding. Based on molecular operational taxonomic unit (MOTU) estimation, we discussed on species delimitation threshold value for these taxa. Our findings confirm that Calaphidinae has high cryptic diversity even though aphids are relatively well-studied.

Genetic Variability of the Invasive Species Metcalfa pruinosa (Hemiptera: Flatidae) in the Republic of Korea.

Metcalfa pruinosa (Say, 1830) (Hemiptera: Flatidae) has caused substantial agricultural damage since its recent introduction to the Republic of Korea; however, the source of this introduction is still unclear. To examine the genetic divergence and phylogenetic relationships among several populations of M. pruinosa from Korea and foreign countries, 251 COI sequences from 251 samples collected from Korea, France, Italy, Spain, Slovenia, and the United States were newly analyzed, together with seven published COI sequences from Canada. In total, 19 haplotypes were detected from the 258 COI sequences, and three haplotypes, H1, H3, and H9, were detected from samples in Korea. The MJ network and Bayesian inference revealed that the three haplotypes of Korea were closely connected with samples of Italy, Spain, Slovenia, France, and the United States. Our study revealed the possibility of multiple invasions of M. pruinosa from Europe and/or North America into Korea.

A new species of Megoura (Hemiptera, Aphididae) from Japan.

A new species of the genus Megoura, M. lathyricola sp. n., was collected from Lathyrus japonicus subsp. japonicus (Leguminosae) in seashore areas of northern and southern Japan. This species is described and illustrated, and a revised key to the identification of the world species of Megoura is presented.

Morphological abnormalities in gall-forming aphids in a radiation-contaminated area near Fukushima Daiichi: selective impact of fallout?

To evaluate the impact of fallout from the Fukushima Daiichi Nuclear Power Plant accident on organisms, this study compared the morphology and viability of gall-forming aphids between the Fukushima population and control populations from noncontaminated areas. This study, in particular, focused on the morphology of first-instar gall formers derived from the first sexual reproduction after the accident. Of 164 first instars from Tetraneura sorini galls collected 32 km from Fukushima Daiichi in spring 2012, 13.2% exhibited morphological abnormalities, including four conspicuously malformed individuals (2.4%). In contrast, in seven control areas, first instars with abnormal morphology accounted for 0.0-5.1% (on average, 3.8%). The proportions of abnormalities and mortality were significantly higher in Fukushima than in the control areas. Similarly, of 134 first instars from T. nigriabdominalis galls, 5.9% exhibited morphological abnormalities, with one highly malformed individual. However, of 543 second-generation larvae produced in T. sorini galls, only 0.37% had abnormalities, suggesting that abnormalities found in the first generation were not inherited by the next generation. Although investigation is limited to one study site, this result suggests that radioactive contamination had deleterious effects on embryogenesis in eggs deposited on the bark surface, but a negligible influence on the second generation produced in closed galls. Furthermore, analysis of both species samples collected in spring 2013 indicated that the viability and healthiness of the aphids were significantly improved compared to those in the 2012 samples. Thus, the results of this study suggest the possibility that a reduced level of radiation and/or selection for radiation tolerance may have led to the improved viability and healthiness of the Fukushima population.

Taxonomic status of the Bemisia tabaci complex (Hemiptera: Aleyrodidae) and reassessment of the number of its constituent species.

Bemisia tabaci (Hemiptera: Aleyrodidae) is one of the most important insect pests in the world. In the present study, the taxonomic status of B. tabaci and the number of species composing the B. tabaci complex were determined based on 1059 COI sequences of B. tabaci and 509 COI sequences of 153 hemipteran species. The genetic divergence within B. tabaci was conspicuously higher (on average, 11.1%) than interspecific genetic divergence within the respective genera of the 153 species (on average, 6.5%). This result indicates that B. tabaci is composed of multiple species that may belong to different genera or subfamilies. A phylogenetic tree constructed based on 212 COI sequences without duplications revealed that the B. tabaci complex is composed of a total of 31 putative species, including a new species, JpL. However, genetic divergence within six species (Asia II 1, Asia II 7, Australia, Mediterranean, New World, and Sub Saharan Africa 1) was higher than 3.5%, which has been used as a threshold of species boundaries within the B. tabaci complex. These results suggest that it is necessary to increase the threshold for species boundaries up to 4% to distinguish the constituent species in the B. tabaci complex.

Seasonal changes in the genetic structure of an aphid-ant mutualism as revealed using microsatellite analysis of the aphid Tuberculatus quercicola and the ant Formica yessensis.

The present study examined whether the mutualistic relationship between the aphid Tuberculatus quercicola (Matsumura) (Homoptera: Aphididae) and the attending ant Formica yessensis Forel (Hymenoptera: Formicidae) has had any mutual effects on the microgeographical genetic population structure of both partner species. The aphids and the attending ants were collected in June, August, and October 2004 from six trees of the Daimyo oak Quercus dentata Thunberg (Fagales: Fagaceae) and were genotyped using microsatellite loci. Significant genetic differentiation was detected among T. quercicola populations on the respective trees across seasons (an average of pairwise F(ST) = 0.183). Similarly, significant genetic differentiation was found among populations of F. yessensis that attended aphid colonies on the respective host trees, though the averages of pairwise F(ST) were lower (an average of pairwise F(ST) = 0.070). An analysis of molecular variance and two-way ANOVA detected a significantly large genetic difference between spring and summer samples in F. yessensis but not in T. quercicola, indicating that changes in genetic composition occurred in the F. yessensis colony. In spite of a drastic seasonal change in the genetic difference in F. yessensis, principle coordinate analysis showed that the relative position among the six populations was maintained from spring to summer, suggesting that the tree where honeydew was available for a long time was occupied by F. yessensis over the same period and that the honeydew sources were inherited at the level of the ant colony. It is hypothesized that the suitability of host trees for the aphid T. quercicola may have an affect on the genetic structure of the attending ant F. yessensis. Within a colony of aphids, clonal diversity decreased significantly as the season progressed. The reduction in clonal diversity may be due to an increase in identical genotypes by parthenogenesis or selective pressure from host plant deterioration.

Allelic and genotypic diversity in long-term asexual populations of the pea aphid, Acyrthosiphon pisum in comparison with sexual populations.

Many aphid species exhibit geographical variation in the mode of reproduction that ranges from cyclical parthenogenesis with a sexual phase to obligate parthenogenesis (asexual reproduction). Theoretical studies predict that organisms reproducing asexually should maintain higher allelic diversity per locus but lower genotypic diversity than organisms reproducing sexually. To corroborate this hypothesis, we evaluated genotypic and allelic diversities in the sexual and asexual populations of the pea aphid, Acyrthosiphon pisum (Harris). Microsatellite analysis revealed that populations in central Japan are asexual, whereas populations in northern Japan are obligatorily sexual. No mixed populations were detected in our study sites. Phylogenetic analysis using microsatellite data and mitochondrial cytochrome oxidase subunit I (COI) gene sequences revealed a long history of asexuality in central Japan and negated the possibility of the recent origin of the asexual populations from the sexual populations. Asexual populations exhibited much lower genotypic diversity but higher allelic richness per locus than did sexual populations. Asexual populations consisted of a few predominant clones that were considerably differentiated from one another. Sexual populations on alfalfa, an exotic plant in Japan, were most closely related to asexual populations associated with Vicia sativa L. The alfalfa-associated sexual populations harboured one COI haplotype that was included in the haplotype clade of the asexual populations. Available evidence suggests that the sexuality of the alfalfa-associated populations has recently been restored through the northward migration and colonization of alfalfa by V. sativa-associated lineages. Therefore, our results support the theoretical predictions and provide a new perspective on the origin of sexual populations.

Inbreeding depression, increased phenotypic variance, and a trade-off between gonads and appendages in selfed progeny of the aphid Prociphilus oriens.

Trade-offs are potentially common among two or more traits whose development is dependent on the same resources. To detect genetic trade-offs, the techniques of quantitative genetics, pedigree analyses, and selection experiments have been used. This study demonstrates genetically based trade-offs between gonads and appendages in hatched larvae of the aphid Prociphilus oriens by focusing on enlarged variance among the families of selfed progeny. The selfed and outbred families were compared in respect to the size of morphological traits, gonad volume, and hatch dates as well as egg volume. Selfing not only increased the among-family variance component in all larval traits examined, but it also increased the mean size of all the morphological traits significantly. In contrast, gonad volume, a fitness component, was reduced with selfing. Calculation of the allometry (log-transformed regression) of larval traits to egg volume indicated that in the outbred group, morphological traits grew slowly relative to egg volume with slopes below 0.25, whereas gonads exhibited isometric growth. With selfing, most morphological traits had significantly steeper slopes, whereas the slope for gonads was greatly decreased. When the effect of egg volume was statistically removed from the means of selfed families, significant negative correlation was detected between the adjusted means of gonad volume and those of tibia length. This result suggests genetic trade-offs between gonad volume and tibia length. Thus, the evidence implies that at the loci governing the development of appendages, the dominant alleles function to canalize the development of tibiae into an optimal size, irrespective of egg volume. It is hypothesized that increased homozygosity of the deleterious recessive alleles reduced gonad volume through increasing the resource allocation to tibiae. The hypothesis of the gonad-appendage trade-off could be applied to explain the phenotypic evolution in some aphid species.

Experimental hybridisation between X0 and XY chromosome races in the grasshopper Podisma sapporensis Shir. (Orthoptera, Acrididae). I. Cytological analysis of embryos and F1 hybrids.

The results of experimental hybridisation between some chromosome subraces belonging to the X0 and XY chromosome races of the brachypterous grasshopper P. sapporensis are presented. Pre-zygotic reproductive isolation mechanisms in experimental pairs were not confirmed. In crossings of XY-standard x X0-standard and XY-standard x X0-Naganuma chromosome subraces, a zygotic barrier has been found. All embryos of XY-standard x X0-standard crosses and the vast majority of embryos of XY-standard x X0-Naganuma crosses were obtained from female diploid or haploid/diploid cells as a result of parthenogenesis. In very rare cases, when the zygotic barriers had been surmounted, normal embryo heterozygotes and a F1 hybrid generation were obtained in XY-standard x X0-Naganuma crosses. On the contrary, crosses between the XY-Tanno and X0-standard subraces gave viable offspring in spite of many chromosome differences such as a X-A translocation and fixed pericentric inversions in four pairs of autosomes. The results obtained do not support the hypothesis that chromosomal differences play a key role in restricting gene flow between X0 and XY races of P. sapporensis. The presence of crossing barriers explains the phenomena of the purity of the X0 and XY chromosomes races.

Ant attendance changes the sugar composition of the honeydew of the drepanosiphid aphid Tuberculatus quercicola.

Mutualistic interactions between aphids and ants are mediated by the honeydew produced by aphids. Previous work showed that when attended by the ant Formica yessensis, nymphs of the aphid Tuberculatus quercicola developed into significantly smaller adults with lower fecundity than when not ant-attended. This study tested the hypothesis that this cost of ant attendance arises through changes in the quality and quantity of honeydew. Ant-attended and ant-excluded aphid colonies were prepared in the field, and the sugar concentration and sugar composition of the honeydew of ant-attended colonies were compared with those of ant-excluded colonies. The frequency and amount of honeydew excretion were also quantified in the two types of colonies. The aphids excreted smaller droplets of honeydew more frequently in ant-attended colonies than in ant-excluded colonies. There was no significant difference in total sugar concentration between the honeydew of ant-attended aphids and ant-excluded aphids. However, ant-attended aphids produced honeydew containing a significantly lower proportion of glucose and higher proportions of sucrose and trehalose than did ant-excluded aphids. These results suggest that the enhanced rate of honeydew-excretion behavior under ant attendance led to changes in the aphid's physiological status. We suggest that the increase in the proportions of sucrose and trehalose in honeydew leads to a shortage of carbohydrates available for energy metabolism, resulting in lower performance of the aphids under ant attendance.