Several Manduca sexta eggs; one can see the outline of a developing larva in the foremost egg on the left, and the right-most larva has already hatched, leaving behind an empty egg-case.
I have recently had the opportunity to rear one of the most prolific insects around, known not only for its role as a model organism in many biological studies but also for its reputation as a major agricultural pest. The tobacco hornworm (Manduca sexta) can defoliate entire plants and is commonly found on many hosts from the Solanaceae family, most importantly tomato and tobacco.
Tobacco hornworms have a widespread distribution throughout the eastern United States, occurring year-round in southern states and seasonally in its northern range. Growing to more than 80 mm in length during the final larval instar (meaning molting stage for my non-entomologically inclined readers), these caterpillars are one of the largest pest insects around.
Newly-hatched, first-instar Manduca sexta larva on standard insect diet
As part of the Insect Physiology course I am enrolled in at the Univ. of Illinois, I am documenting the growth of two tobacco hornworms from egg to adult. In the relatively controlled conditions of my apartment, I expect my larvae to pupate within 2 weeks, though this rate of development varies drastically in nature according to season, temperature, and numerous other environmental stimuli. In transitioning through the typical 5 larval stages that most hornworms undergo, these larvae grow more than 1000 times their original size in just under two weeks! Imagine if these life history traits characterized human development: assuming a newborn baby is 50 cm in height, that baby would grow to be 500 m tall in only two weeks!
I am rearing Manduca sexta caterpillars on pre-made caterpillar diet for the Insect Physiology course I am taking, IB427, at the Univ. of Illinois at Urbana-Champaign. Here is the first larval instar, just recently hatched from its egg, taking its first nibbles of food.
Please also check out this separate blogpost on rearing Manduca larvae, complete with high-quality images of the adult moth and interesting facts about hornworm life history:
http://normalbiology.blogspot.com/2010/12/its-hard-to-be-hornworm.html
The rate of growth is not the only fascinating aspect of insect development, however. Each progression from instar to instar involves shedding the insect's cuticle (ie. skin), which has become too small, in a process known as molting. Since most insects are passive breathers, depending on simple gas exchange to keep oxygen flowing into their spiracles, and since the tracheal tubes leading air from the external environment into the insect are lined with cuticle, the molting process can be quite tricky and involves completely removing the lining of all tracheae during each molt. Clearly, this is a very vulnerable period in an insects' life, one in which the insect is unable to breath and defend itself from predators or inhospitable conditions. Check out the following video to see the tracheal linings being removed along with the rest of the old larval cuticle:
Note the long, thin, white tracheal linings seen emerging from the new larval instar's spiracles visible near the anterior ventral side of the body. I claim no ownership of this video.
Before a hornworm transitions from the pupal stage to the adult stage and becomes a moth, a broad spectrum of behavioral, physiological, and developmental processes take place, including several molts from the first larval instar to its last. The majority of these processes are induced and regulated via cascades of biochemical reactions within the insect's body.
The major actors in these biochemical cascades include hormones (intercellular messaging proteins derived from the endocrine system - think slower), neurohormones (intercellular messaging proteins derived from neurosecretory cells - think faster), enzymes, and the tissues/organs these molecules interact with. Among the myriad biochemical pathways that have been described and the countless processes yet to be discovered, one of the most well-understood biochemical mechanisms is that which controls molting in insects and crustaceans (Covi et al. 2012).
First-instar Manduca sexta larva in defensive position
Aside from gaining valuable parenting experience caring for my Manduca larvae, another goal of this project is to examine the hormonal control of molting behavior through a classic ligation experiment. Early insect physiologists ingeniously devised a simple experiment to examine the effects of limiting hormone circulation throughout a developing larva. By ligating, or tying off, one end of a final-instar larva with a fine string, hemolymph circulation was restricted, though the larva continued to survive. The idea here was to inhibit circulation of specific molecules (now recognized as prothoracicotropic hormone [PTTH], juvenile hormone [JH], and ecdysteroids) produced in the insect brain and its accessory organs. If caterpillars were ligated 5 or more hours post-molting to the fourth instar, researchers found that the entire ligated caterpillar showed signs of molting; however, larvae that were ligated within 5 hours of emerging as fourth instars only displayed pupal characteristics on the posterior section of the body (Gibbs & Riddford 1977). This tipped researchers off to the fact that some molecule from the insect's brain must be initiating ecdysis and further molting processes, and that there exists a critical period in Manduca during which this molecule is released.
Second-instar Manduca sexta larva; note the drastic change in coloration, the typical stripes on the sides of the larva that are faintly visible, and the small black spiracles on the larva's sides which it uses to breath
My classmates and I aim to recreate these experiments with our own Manduca sexta larvae and document the results here.
In a healthy hornworm larva, molting is regulated both by environmental and physiological conditions. Both temperature and photoperiod (the relative length of exposure to light and dark within a day) have significant impacts on the timing of tobacco hornworm molting. Additionally, a hornworm will only molt once it has fed sufficiently and grown an appropriate size. Once these so-called "gates" are passed, a hormone known as prothoracicotropic hormone (PTTH) is released from one of two neurohemal organs known as the corpus allatum and the corpus cardiacum, which are located just behind the brain in most insects. This hormone travels to the prothoracic gland where it then triggers the release of molting hormones (ecdysteroids) into the hemolymph. These molting hormones then trigger many cellular structures throughout the insect's body to prepare for molting and to begin the ecdysial process.
For more details on ecdysis and to learn exactly how the process occurs, check out the following: http://nelson.beckman.illinois.edu/courses/neuroethol/models/manduca_ecdysis/ecdysis.html
Our group's first ligated Manduca sexta larva, freshly molted to the 4th instar. Over the next several days, we will be monitoring the posterior and anterior sections of this and our other ligated larvae to see whether or not each side shows signs of pupation.
My classmates and I are watching for signs of pupal characteristics in caterpillars that we either ligated soon after emerging as a fourth instar or waited to ligate at least 24 hours post-emergence. Unfortunately, my fellow classmates' and my own larvae have not yet pupated so the results of our ligation experiment are forthcoming.
Finally, watch this healthy, non-ligated hornworm larvae emerge as a pupae!
Check out this awesome video (with questionable music) showing an M. sexta caterpillar molting to the pupal stage! I claim no ownership of this video and thank user "ustc07207" for uploading it to YouTube.com
Literature Cited:
Covi, J.A., E.S. Chang and D.L. Mykles. 2012. Neuropeptide signaling mechanisms in crustacean and insect molting glands. Invertebrate Reproduction & Development 56(1): 33-49. DOI: 10.1080/07924259.2011.588009 permalink: http://dx.doi.org/10.1080/07924259.2011.588009
Gibbs, D., and L.M. Riddiford. 1977. Prothoraciotropic hormone in Manduca sexta: localization by a larval assay. Journal of Experimental Biology 66: 255-266.
permalink: http://jeb.biologists.org/content/66/1/255