Sequence and expression analysis of the spermatogenesis-specific gene cognates, wampa and Prosα6T, in Drosophila suzukii.

The sterile insect technique (SIT) is a highly effective biologically-based method for the population suppression of highly invasive insect pests of medical and agricultural importance. The efficacy of SIT could be significantly enhanced, however, by improved methods of male sterilization that avoid the fitness costs of irradiation. An alternative sterilization method is possible by gene-editing that targets genes essential for sperm maturation and motility, rendering them nonfunctional, similar to the CRISPR-Cas9 targeting of ß2-tubulin in the genetic model system, Drosophila melanogaster. However, since genetic strategies for sterility are susceptible to breakdown or resistance in mass-reared populations, alternative targets for sterility are important for redundancy or strain replacement. Here we have identified and characterized the sequence and transcriptional expression of two genes in a Florida strain of Drosophila suzukii, that are cognates of the D. melanogaster spermatocyte-specific genes wampa and Prosalpha6T. Wampa encodes a coiled-coil dynein subunit required for axonemal assembly, and the proteasome subunit gene, Prosalpha6T, is required for spermatid individualization and nuclear maturation. The reading frames of these genes differed from their NCBI database entries derived from a D. suzukii California strain by 44 and 8 nucleotide substitutions/polymorphisms, respectively, though all substitutions were synonymous resulting in identical peptide sequences. Expression of both genes is predominant in the male testis, and they share similar transcriptional profiles in adult males with ß2-tubulin. Their amino acid sequences are highly conserved in dipteran species, including pest species subject to SIT control, supporting their potential use in targeted male sterilization strategies.

Divergence in cell cycle progression is associated with shifted phenology in a multivoltine moth: the European corn borer, Ostrinia nubilalis.

Evolutionary change in diapause timing can be an adaptive response to changing seasonality, and even result in ecological speciation. However, the molecular and cellular mechanisms regulating shifts in diapause timing remain poorly understood. One of the hallmarks of diapause is a massive slowdown in the cell cycle of target organs such as the brain and primordial imaginal structures, and resumption of cell cycle proliferation is an indication of diapause termination and resumption of development. Characterizing cell cycle parameters between lineages differing in diapause life history timing may help identify molecular mechanisms associated with alterations of diapause timing. We tested the extent to which progression of the cell cycle differs across diapause between two genetically distinct European corn borer strains that differ in their seasonal diapause timing. We show the cell cycle slows down during larval diapause with a significant decrease in the proportion of cells in S phase. Brain-subesophageal complex cells slow primarily in G0/G1 phase whereas most wing disc cells are in G2 phase. Diapausing larvae of the earlier emerging, bivoltine E-strain (BE) suppressed cell cycle progression less than the later emerging, univoltine Z-strain (UZ) individuals, with a greater proportion of cells in S phase across both tissues during diapause. Additionally, resumption of cell cycle proliferation occurred earlier in the BE strain than in the UZ strain after exposure to diapause-terminating conditions. We propose that regulation of cell cycle progression rates ultimately drives differences in larval diapause termination, and adult emergence timing, between early- and late-emerging European corn borer strains.

A rapidly evolved shift in life-history timing during ecological speciation is driven by the transition between developmental phases.

For insect species in temperate environments, seasonal timing is often governed by the regulation of diapause, a complex developmental programme that allows insects to weather unfavourable conditions and synchronize their life cycles with available resources. Diapause development consists of a series of distinct phases including initiation, maintenance, termination and post-diapause development. The evolution of insect seasonal timing depends in part on how these phases of diapause development and post-diapause development interact to affect variation in phenology. Here, we dissect the physiological basis of a recently evolved phenological shift in Rhagoletis pomonella (Diptera: Tephritidae), a model system for ecological divergence. A recently derived population of R. pomonella shifted from specializing on native hawthorn fruit to earlier fruiting introduced apples, resulting in a 3-4 week shift in adult emergence timing. We tracked metabolic rates of individual flies across post-winter development to test which phases of development may act either independently or in combination to contribute to this recently evolved divergence in timing. Apple and hawthorn flies differed in a number of facets of their post-winter developmental trajectories. However, divergent adaptation in adult emergence phenology in these flies was due almost entirely to the end of the pupal diapause maintenance phase, with post-diapause development having a very small effect. The relatively simple underpinnings of variation in adult emergence phenology suggest that further adaptation to seasonal change in these flies for this trait might be largely due to the timing of diapause termination unhindered by strong covariance among different components of post-diapause development.

Effect of photoperiod and temperature on the intensity of pupal diapause in the cotton bollworm, Helicoverpa armigera (Lepidoptera: Noctuidae).

The intensity of pupal diapause in the cotton bollworm, Helicoverpa armigera (Hübner) was investigated under both laboratory and natural conditions. By transferring diapausing pupae induced under LD 11:13, LD 12:12 and LD 13:11 at 20, 22 and 25 °C to 25 °C combined with LD 15:9 to terminate diapause the rearing day length of 11 h evoked greater intensity of diapause than did 12 and 13 h at 25 °C; whereas the rearing temperature of 25 °C evoked more intense diapause than did 20 and 22 °C under LD 11:13. By transferring diapausing pupae induced under LD 12:12 at 20 and 22 °C to six temperatures of 18, 20, 22, 25, 28 and 31 °C combined with LD 15:9 to terminate diapause, the duration of diapause was significantly shortened from 146 days at 18 °C to 24 days at 31 °C, showing that high temperatures significantly accelerate diapause development. Furthermore, the duration of diapause was significantly longer at the rearing temperature of 22 °C than that at 20 °C when the diapause-terminating temperatures were 20 and 22 °C. Chilling at 5 °C did not shorten the duration of diapause but lengthened it when chilling period was included. However, chilling plays an important role in synchronizing adult emergence. Rearing temperature of 22 °C also evoked more intense diapause than did 20 °C in most chilling treatments. When the overwintering pupae were transferred at different times from natural temperatures to 25 °C, it was found that the earlier the transfer took place, the earlier the adults emerged when the time spent under natural conditions was included. However, cool temperatures before March showed an enhanced effect on diapause development at 20 °C, suggesting that the high diapause-terminating temperature can offset the effect of chilling on diapause development. The result of diapause termination under natural conditions suggests that the developmental threshold for post-diapause development in H. armigera should be around 17.5 °C.

A comparison of the life-history traits between diapause and direct development individuals in the cotton bollworm, Helicoverpa armigera.

In order to understand the differences of life-history traits between diapause and direct development individuals in the cotton bollworm, Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae), the development time, body size, growth rate, and adult longevity were investigated between the two populations, which were induced under 12:12 L:D and 16:8 L:D photoperiods, respectively, at 20, 22, and 25°C. The results indicated that the larval development time, pupal weight, adult weight, and growth rate were significantly different between diapause and direct developing individuals. The diapause developing individuals had a significantly higher pupal and adult weight and a longer larval time compared with direct developing individuals. However, the growth rate in diapause developing individuals was lower than that in the direct developing individuals. Analysis by GLM showed that larval time, pupal and adult weight, and growth rate were significantly influenced by both temperature and developmental pathway. The pupal and adult weights were greater in males than females in both developmental pathways, exhibiting sexual size dimorphism. The dimorphism in adult weight was more pronounced than in pupal weight because female pupae lost more weight at metamorphosis compared to male pupae. Protogyny was observed in both developmental pathways. However, the protogyny phenomenon was more pronounced at lower temperatures in direct developing individuals, whereas it was more pronounced in diapause developing individuals when they experienced higher temperatures in their larval stage and partial pupal period. The adult longevity of diapause developing individuals was significantly longer than that of direct developing individuals. The results reveal that the life-history strategy was different between diapause and direct developing individuals.

Diapause induction and termination in Hyphantria cunea (Drury) (Lepidoptera: Arctiinae).

The fall webworm, Hyphantria cunea (Drury), enters facultative diapause as a pupa in response to short-day conditions during autumn. Photoperiodic response curves showed that the critical day length for diapause induction was 14 h 30 min, 14 h 25 min and 13 h 30 min at 22, 25 and 28°C, respectively. The photoperiodic responses under non-24 h light-dark cycles demonstrated that night length played an essential role in the determination of diapause. Experiments using a short day length interrupted by a 1-h light pulse exhibited two troughs of diapause inhibition and the effect of diapause inhibition was greater in the early scotophase than in the late scotophase. The diapause-inducing short day lengths of 8, 10 and 12 h evoked greater intensities of diapause than did 13 and 14 h. Diapause can be terminated without exposure to chilling, but chilling at 5°C for 90 and 120 d significantly accelerated diapause development, reduced mortality, and synchronized adult emergence. Additionally, the potential for H. cunea from the temperate region (Qingdao) to emerge and overwinter under field conditions in subtropical regions (Nanchang) of China was evaluated. Pupae that were transferred to Nanchang in early July showed a 60% survival rate and extremely dispersed pupal period (from 12 to 82 days), suggesting that some pupae may undergo summer diapause. Diapausing temperate region pupae that were moved out-of-doors in Nanchang during October showed approximately 20% overwintering survival; moreover, those pupae that overwintered successfully emerged the next spring during a period when their host plants would be available. The results indicate that this moth has the potential to expand its range into subtropical regions of China.

Geographic variation in diapause induction and termination of the cotton bollworm, Helicoverpa armigera Hübner (Lepidoptera: Noctuidae).

Overwintering diapause in Helicoverpa armigera, a multivoltine species, is controlled by response to photoperiod and temperature. Photoperiodic responses from 5 different geographical populations showed that the variation in critical photoperiod for diapause induction was positively related to the latitudinal origin of the populations at 20, 22 and 25°C. Diapause response to photoperiod and temperature was quite different between northern and southern populations, being highly sensitive to photoperiod in northern populations and temperature dependence in southern populations. Diapause pupae from southern population showed a significantly shorter diapause duration than from northern-most populations when they were cultured at 20, 22, 25, 28 and 31°C; by contrast, overwintering pupae from southern populations emerged significantly later than from northern populations when they were maintained in natural conditions, showing a clinal latitudinal variation in diapause termination. Diapause-inducing temperature had a significant effect on diapause duration, but with a significant difference between southern and northern populations. The higher rearing temperature of 22°C evoked a more intense diapause than did 20°C in northern populations; but a less intense diapause in southern population. Cold exposure (chilling) is not necessary to break the pupal diapause. The higher the temperature, the quicker the diapause terminated. Response of diapause termination to chilling showed that northern populations were more sensitive to chilling than southern population. All results demonstrate that H. armigera is not genetically homogeneous throughout its range, but rather is composed of distinct populations genetically adapted to local environmental conditions despite the potential for gene flow via seasonal migration of adults.

Inheritance of photoperiodic control of pupal diapause in the cotton bollworm, Helicoverpa armigera (Hübner).

Pupae of the cotton bollworm, Helicoverpa armigera display a diapause in response to the exposure of their larvae to short photoperiods and relatively low temperatures. Due to geographic variation in photoperiodic response, moths from a northern population, Langfang (39°32'N, 116°41'E), enter diapause in response to short daylengths (D strain) while moths from a southern population, Ledong (18°28'N, 108°53'E), exhibit no diapause under the same conditions (N strain). In the present study, crosses between the two strains are utilized to evaluate the inheritance of diapause under different photoperiods at temperatures 20, 22 and 25°C. The moths in both reciprocal crosses and backcrosses to D strain showed a clear long-day response, similar to that of the D strain, suggesting that the photoperiodic response controlling diapause in this moth is heritable. The incidences of diapause for all F(1) hybrids were intermediate between those of their parents. However, the incidences of diapause at 20°C in F(1) (N×D) strain were significantly higher than those in F(1) (D×N) strain, indicating that the male parent plays a more important role in the determination of diapause. The N strain also showed a short-day photoperiodic response at the lower temperature of 20°C, indicating that the N strain still has the capability to enter a photoperiodically induced diapause, depending on the rearing temperature. Results from all crosses under photoperiods LD 12:12 or LD 13:11 at 22°C showed that inheritance of diapause in H. armigera did not fit an additive hypothesis and that the capacity for diapause was transmitted genetically in the manner of incomplete dominance with non-diapause characteristic partially dominant over the diapausing. Diapause duration in hybrid pupae was also influenced by their inheritance from both parents. Diapause duration in hybrid pupae was intermediate between those of their parents. These results reveal that both diapause induction and duration are under the control of polygene.