Pigment-dispersing factor is present in circadian clock neurons of pea aphids and may mediate photoperiodic signalling to insulin-producing cells.

The neuropeptide pigment-dispersing factor (PDF) plays a pivotal role in the circadian clock of most Ecdysozoa and is additionally involved in the timing of seasonal responses of several photoperiodic species. The pea aphid, Acyrthosiphon pisum, is a paradigmatic photoperiodic species with an annual life cycle tightly coupled to the seasonal changes in day length. Nevertheless, PDF could not be identified in A. pisum so far. In the present study, we identified a PDF-coding gene that has undergone significant changes in the otherwise highly conserved insect C-terminal amino acid sequence. A newly generated aphid-specific PDF antibody stained four neurons in each hemisphere of the aphid brain that co-express the clock protein Period and have projections to the pars lateralis that are highly plastic and change their appearance in a daily and seasonal manner, resembling those of the fruit fly PDF neurons. Most intriguingly, the PDF terminals overlap with dendrites of the insulin-like peptide (ILP) positive neurosecretory cells in the pars intercerebralis and with putative terminals of Cryptochrome (CRY) positive clock neurons. Since ILP has been previously shown to be crucial for seasonal adaptations and CRY might serve as a circadian photoreceptor vital for measuring day length, our results suggest that PDF plays a critical role in aphid seasonal timing.

Progress in the characterization of insulin-like peptides in aphids: Immunohistochemical mapping of ILP4.

Aphids were the first animals described as photoperiodic due to their seasonal switch from viviparous parthenogenesis to sexual reproduction (cyclical parthenogenesis) caused by the shortening of the photoperiod in autumn. This switch produces a single sexual generation of oviparous females and males that mate and lay diapausing cold-resistant eggs that can overcome the unfavourable environmental conditions typical of winter in temperate regions. Previous studies have hinted at a possible implication of two insulin-like peptides (ILP1 and ILP4) in the aphid seasonal response, changing their expression levels between different photoperiodic conditions. Moreover, in situ localization of their transcripts in particular neurosecretory cells (NSCs) in the aphid brain supported the idea that these neuropeptides could correspond to the formerly called virginoparin, an uncharacterized factor originally proposed to be transported directly to the aphid embryos to promote their development as parthenogenetic individuals. To further investigate the fate of these ILPs, we raised a specific antiserum against one of them (ILP4) and mapped this neuropeptide by immunohistochemistry (IHC) in Acyrthosiphon pisum and Megoura viciae aphids. Coincident with in situ localization, our results show that ILP4 is synthesized in two groups (one in each brain hemisphere) of four neurosecretory cells in the pars intercerebralis (NSC group I) and then it is transported outside the brain to the corpora cardiaca. From there, three nerves (two laterals and one medial) transport it to the abdomen. Although no precise site of release has been found, the terminations of these nerves near the germaria would be compatible with the proposal of a direct connection between group I of NSCs and the reproductive system by localized release. In addition, we detected some collateral arborizations originating from the eight NSCs going to the pars lateralis, where clock neurons and some photoreceptors have been previously localized, suggesting a possible communication between the circadian and photoperiodic systems.

Insulin-like peptides involved in photoperiodism in the aphid Acyrthosiphon pisum.

Aphids were the first animals reported as photoperiodic as their life cycles are strongly determined by the photoperiod. During the favourable seasons (characterised by long days) aphid populations consist exclusively of viviparous parthenogenetic females (known as virginoparae). Shortening of the photoperiod in autumn is perceived by aphids as the signal that anticipates the harsh season, leading to a switch in the reproductive mode giving place to the sexual morphs (oviparae females and males) that mate and lay winter-resistant (diapause-like) eggs. The molecular and cellular basis governing the switch between the two reproductive modes are far from being understood. Classical experiments identified a group of neurosecretory cells in the pars intercerebralis of the aphid brain (the so called group I of neurosecretory cells) that were essential for the development of embryos as parthenogenetic females and were thus proposed to synthesise a parthenogenesis promoting substance that was termed "virginoparin". Since insulin-like peptides (ILPs) have been implicated in the control of diapause in other insects, we investigated their involvement in aphid photoperiodism. We compared the expression of two ILPs (ILP1 and ILP4) and an Insulin receptor coding genes in A. pisum aphids reared under long- and short-day conditions. The three genes showed higher expression in long-day reared aphids. In addition, we localised the site of expression of the two ILP genes in the aphid brain. Both genes were found to be expressed in the group I of neurosecretory cells. Altogether, our results suggest that ILP1 and ILP4 play an important role in the control of the aphid life-cycle by promoting the parthenogenetic development during long-day seasons while their repression by short days would activate the sexual development. Thus we propose these ILPs correspond to the so called "virginoparin" by early bibliography. A possible connection with the circadian system is also discussed.

Aggressive mimicry coexists with mutualism in an aphid.

Understanding the evolutionary transition from interspecific exploitation to cooperation is a major challenge in evolutionary biology. Ant-aphid relationships represent an ideal system to this end because they encompass a coevolutionary continuum of interactions ranging from mutualism to antagonism. In this study, we report an unprecedented interaction along this continuum: aggressive mimicry in aphids. We show that two morphs clonally produced by the aphid Paracletus cimiciformis during its root-dwelling phase establish relationships with ants at opposite sides of the mutualism-antagonism continuum. Although one of these morphs exhibits the conventional trophobiotic (mutualistic) relationship with ants of the genus Tetramorium, aphids of the alternative morph are transported by the ants to their brood chamber and cared for as if they were true ant larvae. Gas chromatography-mass spectrometry analyses reveal that the innate cuticular hydrocarbon profile of the mimic morph resembles the profile of ant larvae more than that of the alternative, genetically identical nonmimic morph. Furthermore, we show that, once in the brood chamber, mimic aphids suck on ant larva hemolymph. These results not only add aphids to the limited list of arthropods known to biosynthesize the cuticular chemicals of their deceived hosts to exploit their resources but describe a remarkable case of plastic aggressive mimicry. The present work adds a previously unidentified dimension to the classical textbook paradigm of aphid-ant relationships by showcasing a complex system at the evolutionary interface between cooperation and exploitation.

Molecular systematics of aphids (Homoptera: Aphididae): new insights from the long-wavelength opsin gene.

Viviparous aphids (Aphididae) constitute a monophyletic group within the Homoptera with more than 4000 extant species worldwide but higher diversity in temperate regions. Several aspects of their biology account for attention paid to this group of insects. Their plant-sap-sucking way of feeding with many species transmitting viruses to crop plants has important implications on crop management strategies. Cyclical parthenogenesis associated in many groups to host alternation and elaborate polyphenisms is of special interests for evolutionists. Finally, the ancient association of most aphid species with intracellular endosymbiotic bacteria (Buchnera sp.) has also received much attention from evolutionists interested in mechanisms involved in the symbiotic process. Knowing the phylogenetic relationships among major aphid taxa is of special interest to evolutionists interested in the above issues. However, until recently, molecular approaches to aphid phylogeny were absent and discussions on the evolution of aphid life-cycles and on evolutionary aspects of their symbiotic association with Buchnera were framed by morphology-based phylogenies. Recently, two reports using molecular approaches attempted to address the yet unresolved phylogeny of Aphididae with limited although somehow different conclusions. In the present report we study the utility of the long-wave opsin gene in resolving phylogenetic relationships among seven subfamilies within the Aphididae. Our results corroborate some previously proposed relationships and suggest a revision of some others. In particular, our data support grouping the analysed aphid species into three main clades, being the subfamily Lachninae one of them, which contradicts its generally accepted sistership relationship with the subfamily Aphidinae. Moreover, our data also suggest a basal position of Lachninae which has implications on current discussions about the ancestrality of conifer-feeding in modern aphids.

Molecular evolution of aphids and their primary (buchnera sp) and secondary endosymbionts: implications for the role of symbiosis in insect evolution

Los pulgones mantienen una asociación endosimbiótica obligada con Buchnera sp., una bacteria estrechamente relacionada con Escherichia coli. La bacteria se encuentra alojada en estructuras celulares llamadas bacteriomas en el hemocele de los pulgones, y son maternalmente transmitidas. Los estudios filogenéticos indican que la asociación tuvo un único origen hace aproximadamente 200-250 millones de años, y desde entonces hospedero y endosimbionte han evolucionado en paralelo. Sin embargo, el patrón de ramificación en la familia de los áfidos no ha sido resuelto, lo cual impide la evaluación de, por ejemplo, el papel de la transferencia horizontal de genes en la evolución temprana de Buchnera. El papel principal de Buchnera en esta asociación es la biosíntesis y aprovisionamiento de aminoácidos esenciales a su áfido hospedero. Esta afirmación está apoyada por estudios fisiológicos, metabólicos y recientemente por estudios genéticos. Las transformaciones genéticas experimentadas por Buchnera en su adaptación a la vida endosimbiótica son variadas. Entre ellas, la reducción de su genoma comparando con bacterias de vida libre, el incremento en A+T, tasas de evolución diferentes, incremento relativo de las tasas de substitución no sinónimas y la amplificación génica mediada por plásmidos. Pero la endosimbiosis es un proceso activo en los insectos, como se puede comprobar por el estado intermedio de los endosimbiontes secundarios, los cuales presentan valores intermedios de las características anteriormente citadas en comparación con las bacterias de vida libre y Buchnera