Pupal Pigmentation Research Articles

The Pupal Pigmentation Pattern and Pupal Development in the Species of Aphytis Howard (Hymenoptera: Aphelinidae).

Simple SummaryAphytis species (Hymenoptera: Aphelinidae) have been considered as the most important natural enemies in biological control of armoured scale insects (Hemiptera: Diaspididae). However, it is difficult to identify the species of the genus, particularly that of the A. lingnanensis group, based on adult morphological characters. Although the differences in pupal pigmentation of Aphytis species have been documented previously, they have not been much used as taxonomic characters for Aphytis species identification. In this study, we present four characteristic pigmentation patterns of Aphytis pupae, together with photographs, including the categories of: entirely yellow, partly dark brown, entirely or predominantly black, and partly black pupae. No significant intra-specific variation in pupal colour pattern was detected despite relatively high numbers of specimens examined, many from multiple, and different, origins. We summarize the present status of pupal pigmentation in the described species of Aphytis, according to the species groups, which could be used as an important supplementary diagnostic character for distinguishing species of Aphytis, especially closely-related species.Species identification of Aphytis on the basis of adult morphology is extremely difficult, especially in the A. lingnanensis group, with several cryptic species. Pupal pigmentation could be used as one of the taxonomic characters for Aphytis species, and in some instances, pupal pigmentation actually provided the first clue to the distinctness of cryptic Aphytis species. The present study investigated the full-grown larvae or younger pupae of Aphytis species, and pupal pigmentation and pupal development were observed and photographed. Four characteristic pigmentation patterns of Aphytis pupae were summarized including: entirely yellow, partly dark brown, entirely or predominantly black, and partly black. The species in the chilensis and mytilaspidis groups, and some unassigned species, generally have entirely, or predominantly and or partly black pupae. The species in the chrysomphali, funicularis, and proclia groups generally have the pupae entirely yellow. The species of the lingnanensis group have the pupae both entirely yellow, e.g., A. fisheri, and partly dark brown pupae, e.g., A. lingnanensis, A. holoxanthus and A. melinus. The pupae of Aphytis species in this study had a developmental duration of about 5–8 days at 27 ± 1 °C, 70 ± 5% RH and a photoperiod of 10L: 14D. It was found that the pupal skin was always melanized at the beginning stage, generally in the first day, among the pigmented pupae of Aphytis species. As development continued, the pigmentation became darker and the eye colour changed from pale red/brown to green. No significant intra-specific variation in pupal colour pattern was detected despite relatively high numbers of specimens examined, many from multiple, and different, origins. Overall, our study indicates that pupal pigmentation could be more helpful in species identification of Aphytis.

Tyrosine hydroxylase is crucial for pupal pigmentation in Zeugodacus tau (Walker) (Diptera: Tephritidae).

Tyrosine hydroxylase (TH) is the initial enzyme responsible for cuticle sclerotization and pigmentation in many insect species, but to date, no direct functional studies have focused on TH in Zeugodacus tau. Here, the 3336-bp full-length cDNA of TH was isolated from Z. tau, a notorious horticultural pest infesting fruits and vegetables. qRT-polymerase chain reaction revealed that ZtTH transcripts were highly abundant at the time of pupal tanning and during adult emergence and were expressed in the midgut, integument and head of molting larvae. The pupation and eclosion rates gradually decreased when the 1st-instar larvae were fed diets containing higher concentrations of the TH inhibitor 3-iodo-tyrosine (3-IT). Moreover, pupal weights were significantly decreased, and abnormal uncolored phenotypes were observed after 20 mg/g 3-IT was incorporated into the diet. In addition, the suppression of TH function (mediated by RNA interference) led to a decrease in TH mRNAs and eclosion rates, accompanied by less-pigmented phenotypes. There was a severe impairment of larval-pupal cuticle tanning, leading to pupae with less yellowish pigment or that were completely white and transparent, when we injected 2 μL of 24.4 mM or 73.27 mM 3-IT into 3rd-instar larvae or prepupae. These results suggest that TH is an important enzyme for the normal growth and pupal pigmentation of Z. tau and that TH is a potential gene target for use in the control of Z. tau.

Large-scale identification of differentially expressed genes during pupa development reveals solute carrier gene is essential for pupal pigmentation in Chilo suppressalis

Insects undergo metamorphosis, involving an abrupt change in body structure through cell growth and differentiation. Rice stem stripped borer (SSB), Chilo suppressalis, is one of the most destructive rice pests. However, little is known about the regulation mechanism of metamorphosis development in this notorious insect pest. Here, we studied the expression of 22,197 SSB genes at seven time points during pupa development with a customized microarray, identifying 622 differentially expressed genes (DEG) during pupa development. Gene ontology (GO) analysis of these DEGs indicated that the genes related to substance metabolism were highly expressed in the early pupa, which participate in the physiological processes of larval tissue disintegration at these stages. In comparison, highly expressed genes in the late pupal stages were mainly associated with substance biosynthesis, consistent with adult organ formation at these stages. There were 27 solute carrier (SLC) genes that were highly expressed during pupa development. We knocked down SLC22A3 at the prepupal stage, demonstrating that silencing SLC22A3 induced a deficiency in pupa stiffness and pigmentation. The RNAi-treated individuals had white and soft pupa, suggesting that this gene has an essential role in pupal development.

Aspartate Decarboxylase is Required for a Normal Pupa Pigmentation Pattern in the Silkworm, Bombyx mori.

The pigmentation pattern of Lepidoptera varies greatly in different development stages. To date, the effects of key genes in the melanin metabolism pathway on larval and adult body color are distinct, yet the effects on pupal pigmentation remains unclear. In the silkworm, Bombyx mori, the black pupa (bp) mutant is only specifically melanized at the pupal stage. Using positional cloning, we found that a mutation in the Aspartate decarboxylase gene (BmADC) is causative in the bp mutant. In the bp mutant, a SINE-like transposon with a length of 493 bp was detected ~2.2 kb upstream of the transcriptional start site of BmADC. This insertion causes a sharp reduction in BmADC transcript levels in bp mutants, leading to deficiency of β-alanine and N-β-alanyl dopamine (NBAD), but accumulation of dopamine. Following injection of β-alanine into bp mutants, the color pattern was reverted that of the wild-type silkworms. Additionally, melanic pupae resulting from knock-down of BmADC in the wild-type strain were obtained. These findings show that BmADC plays a crucial role in melanin metabolism and in the pigmentation pattern of the silkworm pupal stage. Finally, this study contributes to a better understanding of pupa pigmentation patterns in Lepidoptera.

Involvement of Cyclic Adenosine Monophosphate in the Regulation of Pupal Color Adaptation of the Butterfly, Inachis io

Abstract In the butterfly Inachis io, a pupal melanization reducing factor (PMRF) which is located throughout the entire central nervous system controls the intensity of pigmentation of pupal cuticle depending on the background color of the pupation site. PMRF does not only reduce melanization but, in addition, enhances lutein incorporation in a dose-dependent manner to form pupae with yellow color on bright backgrounds. The present paper reports on the effects on pupal pigmentation caused by cyclic nucleo­ tides and phosphodiesterase (PDE) inhibitors which prevent degradation of cyclic nucleo­ tides. The injection of cAMP did not alter pupal coloration whereas its membrane-permeable analog dibutyryl-cAMP mimicked dose-dependently PMRF activity. Thus, pupae of reduced melanization and, in addition, enhanced yellow coloration were formed. This indicates that an increased intracellular cAMP level is capable of mediating PMRF effect. Also, the injection of the PDE inhibitor isobutylmethylxanthine (IBMX) caused dose-dependently pupae of reduced melanization and enhanced lutein incorporation. Theophylline (another PDE inhibitor) was only slightly effective (23% inhibition of melanization) at the highest dose compared to IBMX. The injection of cGMP and its analog dibutyryl-cGMP exhibited no melanization reducing effect. Extracts of abdominal ganglia (AG) which contained PMRF activity caused significantly brighter pupae when injected in combination with IBMX. However, this stimulation by IBMX became no longer effective at higher AG doses. Therefore, the present results are suggestive of an involvement of cAMP as a second messenger in the action of PMRF on pupal color adaptation.

The control of cuticular melanin and lutein incorporation in the morphological colour adaptation of a nymphalid pupa, Inachis io L.

Pupae of Inachis io show a morphological colour adaptation. Their pigmentation varies, depending on the background colour, between yellow and black. The cuticular pigments involved have been identified as lutein and eumelanin. There are spatial relationships between the locations of both pigments, and the total amounts incorporated in the pupal cuticles are inversely related. During a sensitive period, preceding pupation, the background colour determines the future pupal pigmentation. Ligation experiments show, that in prepupae kept on light background, a factor is released from the anterior part of the body, which reduces the pupal melanization and enhances lutein content. The factor is neither juvenile hormone nor ecdysteroid but rather a water-soluble molecule.

Environmental factors influencing pupal colour determination in Lepidoptera. II. Experiments with Pieris rapae, Pieris napi and Pieris brassicae

The control of the environmentally determined polymorphism for pupal coloration has been investigated in Pieris rapae, Pieris napi and Pieris brassicae . Light stimuli are the most important factors involved in the determination of pupal colour. In particular, yellow and blue spectral wavelengths determine the production of green and brown pupae, respectively, and it is argued that this correlation results from yellow light being closely associated with green foliage and blue light with sites away from green vegetation. Other, non-optical factors, such as relative humidity, may also influence pupal colour determination, but are of less importance than light stimuli. Light as a controlling factor in pupal coloration is considered relevant because the larvae of these species select pupation sites during daylight, when light cues would be of greatest use in distinguishing between green and brown environments. Diapausing pupae of P. rapae have a greater tendency to be brown than green, but the reverse is true in P. brassicae . It is uncertain whether this correlation between pupal colour and diapause is a consequence of behavioural or of physiological differences between larvae destined for pupal diapause and those that develop directly. In P. rapae and P. napi , green pupal pigmentation requires the secretion of a hormonal factor from a centre in the larval head. In the absence of this hormone, brown pupal cuticle is produced.

The effect of juvenile hormone on pupal pigmentation of Pieris brassicae L.

The role of juvenile hormone (JH) in the morphological colour adaptation of pupae of Pieris brassicae controlled by environmental factor was analyzed. First the effects of JH I and its analogue, Farnesyl-Methyl-Ether (FME) were tested. Secondly the JH-titres of the last instar larvae were measured under various light conditions which influence the future pigmentation of the pupae. During the sensitive period, which occurs before pupation, blue light (410 nm) produces the strongest, darkness medium, and yellow light (570 nm) the lightest pigmentation of the pupae. JH I as well as FME has an inhibiting effect on the formation of the black spots in the cuticle. However, this effect only becomes apparent (a) if the insects are kept under blue light during the sensitive period (which normally leads to a strong black pigmentation) and (b) only when these animals were treated with JH I or FME either at the beginning, or 10 hr after the beginning of the sensitive period. In the last larvae instar, JH could be found only during the sensitive period. Fourteen hours after the beginning of the sensitive period the JH concentration reaches a maximum of 30–100 pg JH per insect. The JH-titre resulting from blue light conditions is significantly different from those of the larvae kept under white or yellow light. An additional maximum of 60 pg JH per animal was found 8 hr/after the beginning of the sensitive period. Obviously, JH affects the process of pigmentation of the pupae, but the described results are not sufficient to explain thoroughly the regulation of pigmentation modified by environmental factors. The effects of further factors are discussed.

Biological and systematic studies of developmental stages inAphytis(Hymenoptera: Aphelinidae): III. Meconia as a possible systematic tool inAphytis

Abstract does not appear. First page follows. Introduction This paper is the third in a series of biological and systematic investigations of developmental characteristics in the genus Aphytis Howard (Hymenoptera: Aphelinidae). The systematics of Aphytis has been based almost entirely on imaginal morphology, which often does not show strong interspecific differences. Other than pupal pigmentation, which has proved useful in certain cases (DeBach, 1959), characteristics of developmental stages have not been utilized as systematic tools in this genus. The first paper in this series (Rosen and Eliraz, 1978) presented the developmental history of the generotype of Aphytis, A. chilensis Howard, as a basic background study, and included detailed descriptions of the various developmental stages. The second paper (Eliraz and Rosen, 1978) presented a comparative study of larval characteristics of several species in five species-groups, and the present paper reports the results of a comparative investigation of meconial pellets in several species of Aphytis. In most parasitic Hymenoptera, fecal material accumulates in the midgut throughout the larval feeding period, and is expelled, in the form of a meconium or meconial pellets, only when the blind midgut becomes linked with the proctodaeum prior to pupation (Hagen, 1964). The meconia of numerous parasitic Hymenoptera have been occasionally described, but have not been used much as a diagnostic character for identification or classification. (Flanders (1942)) briefly discussed the meconia of several species, and concluded that “although larval meconium is of little, if any, value in identifying the species of a genus, it may be of value in the recognition of the species which compose the parasitic fauna of a host in the field.” More recently, Viggiani (1969) has shown that the shape of the meconia may be used to separate certain eulophid parasites of leaf-mining insects. All the known species of Aphytis are ectoparasites of armored scale insects, developing externally on the bodies of their hosts beneath the covering scale (Rosen and DeBach, 1976). Their distinctive meconial pellets are very conspicuous under the scale, and have been noted by some of the early workers. (Quayle (1910)), for instance, described the meconia of A. chrysomphali (Mercet) [erroneously recorded by him as A. diaspidis (Howard)] as follows: “Always accompanying the pupa are from six to ten black or dark brown torpedo like bodies .125 mm. long and .055 mm. broad, which are evacuations from the digestive tract and are expelled by the larva preliminary to pupation.” Similarly, (Taylor (1935)), in his description of (presumably) the same species, wrote: “No excretion occurs until the larva is ready to pupate. The excrement is in the form of small, shiny, black or brown bodies, bluntly pointed at each end—and usually 5 or 6 in number, and uniform in shape. They are deposited inside the scale, round the posterior end of the larva, and are so conspicuous in Aspidiotus destructor that they are very useful as a means of detecting the full-grown larvae or pupae in situ.”

Action spectra for light-controlled pupal pigmentation in Pieris brassicae: Melanization and level of bile pigment

Pupal pigmentation in Pieris brassicae is controlled by light during a sensitive period before pupation. Action spectra for melanization and level of biliverdin-IX λ were determined in the range of 383 to 765 nm at equal quantum flux densities. Darkness and light of 500, 728, and 765 nm result in intermediate melanization and a low level of bile pigment. Wavelengths of <500 nm promote, while those of >500 nm inhibit cuticular melanization, but the level of bile pigment is raised in both. Infrared light (748 nm) is shown to have a significant effect, but only the bile pigment is influenced. In both spectra, maximal effects are observed at identical wavelengths: 404, 577, and 661 nm; a minimum occurs at 640 nm. The degree of melanization and the level of biliverdin are correlated positively for λ<500 nm and negatively for λ>500 nm. It is argued that the action spectra may not reflect the absorption of the photoreceptor(s), but rather the action of mechanisms in the central nervous system, which influence both pigments in separate and different ways. Light does not stimulate the synthesis of bile pigment in the larva before pupation, but does inhibit its degradation in the pupa. The results are discussed in the light of the endocrine mechanisms involved, and in respect of the biological significance of colour adaptation in Pieris pupae.

Wavelength dependence of light-controlled pupal pigmentation

Wavelength dependence of light-controlled pupal pigmentation

New Species and Strains of Aphytis (Hymenoptera, Eulophidae) Parasitic on the California Red Scale, Aonidiella Aurantii (Mask.), in the Orient1

This paper deals primarily with the Aphytis species or lesser categories considered to be definite and regular parasites of the California red scale, Aonidiella aurantii (Mask.), in the Orient. The importance of species of Aphytis as parasites of diaspine scale insects is discussed and the potential value of strains or other subspecific categories in biological control is emphasized. Pupal pigmentation is described for four species or groups and is shown to be of taxonomic importance. A key is given to five species (or forms) of Oriental Aphytis parasitic on the California red scale. Two new species, Aphytis melinus and A. fisheri , are described, as are two forms whose status is in doubt, the Assam form of A. hispanicus and the Kagoshima form of A. citrinus. The adult females of A. melinus and A. fisheri are very similar, if not indistinguishable, morphologically, but they do not interbreed and the pupae are quite distinct. The known distribution of these species in the Orient is given.