Insect Biodiversity

T.Y.B.Sc Zoology Pune University, Semester II Paper V

ZY-345: Paper V
Total Lectures: 45.
Introduction: (2)
1.1 Definition & scope.
1.2 Types of diversities among insects.
2. Insect & it’s environment: (4)
2.1 Insect diversity & adaptations with reference to terrestrial habitats: forest, agriculture, subterranean, cave, glacier, mountain & desert.
2.2 Insect diversity & adaptations with reference to aquatic habitats : river, stream, lake, pond, torrents, marine, estuarine & ephemeral water bodies.
3. Population dynamics of Insects: (3)
3.1 Concept of population dynamics.
3.2 Factors affecting population dynamics in insects.
3.3 Seasonal variations in insect populations.
4. Insect taxonomy: (9)
4.1 Outline of scheme of classification of insects as given by Richards & Davis.
4.2 Distinguishing features of Apterygotan insects.
4.3 Distinguishing features of Pterygotan insects : Exopterygota & Endopterygota.
4.4 Distinguishing taxonomic features & significance of following major insect orders:
Orthoptera, Diptera, Hemiptera, Lepidoptera & Coleoptera.
4.5 Useful contribution in molecular phylogenetic studies.
5. Insects in social groups: (5)
Definition, intraspecific & interspecific relationships among insects.
5.2 Social organizations in ants, wasps & termites.
5.3 Significance of social organizations.
6. Food & feeding behavior in insects: (4)
6.1 Selection of food by insects.
6.2 Food diversity among insects.
6.3 Significance of diversity in food & feeding habits.
7. Breeding behavior in insects: (6)
7.1 Diversity in courtship & oviposition behavior in insects.
7.2 Diversity in oviposition sites among insects.
7.3 Parental care & nest building diversity in insects.
7.4 Diapause behavior in insects.
8. Diversity in insect relationships:
8.1 Diversity in mutualistic associations: ant-aphids, ant-coccids, ant-bug, ant-butterfly &
ant- membracids.
8.2 Insects as predators, parasites & parasitoids.
8.3 Insect plant interaction: Role of insects as plant bodygaurds.
9. Survival strategies in insects: (3)
Escape, flight, sting, poison, mimicry, hide, camouflage & migration.
10. Effect of changing climate & human interference on insect diversity: (4)
10.1 Impact of global changes on diversity of insects at various levels such as local, regional, national & global.
10.2 Important steps essential for conversation & management of insect diversity.
Reference books:
1.Imm’s General Text book of Entomology Vol. I & 11(1993), Richards O.W. &Davis R.F.., B.I. Pul (Indian edition) New Delhi.
2. Principals of insect morphology, Snodgrass R.E. (1994) Indian Reprint, SBS Pub. New Delhi.
3. Structure & functions of Insects. Chapman R.F. (1983), edition, ELBS, London.
4. Entomology, Gillott Cedric (1980), Plenum Press, New York.
5.The Science of Entomology, Romoser W.S. (1981) 2’’ edition, Mac millon Co., New York.
6. General Entomology, Mani M.S. (1998) Reprint Oxford- IBH, India.
7 An Introduction to Entomology, Srivastav R.D. & Singh R.P. (1997), Concept Pub. New Delhi.
8.General & Applied Entomology, Nayar K.K., T.N. anantkrishanan & B.V. David, (1983), tata McGrow Hill, Pub. New Delhi.
9. Insects, Mani M.S. (2006) Reprint NBT Pub. New Delhi.

Population Dynamics
A population describes a group of individuals of the same species occupying a specific area at a specific time. Some characteristics of populations are the population density, the birthrate, and the death rate. And immigration (Foreigners coming to settle in our country) into the population, or emigration ( leaving own country to settle elsewhere) out of it.
Together, these population parameters, or characteristics, describe how the population density changes over time. The ways in which population densities fluctuateincreasing, decreasing, or both over timeis the subject of population dynamics.
Population density measures the number of individuals per unit area, for example, the number of deer per square kilometer. Although this is straightforward in theory, determining population densities for many species can be challenging in practice.
I. Insect Populations and Population Dynamics.
A. Definition of Population: a collective group of individuals of the same species occupying a particular space.
B. Populations may be divided into demes. A deme is a smaller unit of a population found in a localized area. Demography is the study of the vital statistics of a population.
C. Definition of Population Dynamics: that aspect of population ecology dealing with forces affecting changes in population densities or affecting the form of population growth.
D. Populations have 2 basic requirements:
1. They have a minimum size; and
2. They occupy an area containing all needed resources.
Measuring Population Density
One way to measure population density is simply to count all the individuals. Alternatively, good estimates of population density can often be obtained via the quadrat method. In the quadrat method, all the individuals of a given species are counted in some subplot of the total area. Then that data is used to figure out what the total number of individuals across the entire habitat should be.
The quadrat method is particularly suited to measuring the population densities of species that are fairly uniformly distributed over the habitat. For example, it has been used to determine the population density of soil species such as insects. It is also commonly used to measure the population density of plants.
The birthrate of a population describes the number of new individuals produced in that population per unit time.
The death rate, also called mortality rate, describes the number of individuals who die in a population per unit time.
The immigration rate is the number of individuals who move into a population from a different area per unit time.
The emigration rates describe the numbers of individuals who migrate out of the population per unit time.
The values of these four population parameters allow us to determine whether a population will increase or decrease in size. The "intrinsic rate of increase r " of a population is defined as r = (birth rate immigration rate +)-(death rate + emigration rate ).
If r is positive, then more individuals will be added to the population than lost from it. Consequently, the population will increase in size. If r is negative, more individuals will be lost from the population than are being added to it, so the population will decrease in size. If r is exactly zero, then the population size is stable and does not change. A population whose density is not changing is said to be at equilibrium .
 A population of organisms may display:
1. Growth
2. Dispersion
3. Genetic variability
4. Continuity in time.
G. Populations are not isolated entities, but exist in "communities" with other populations of species.

II. Communities
A. Communities are associations of different species which are relatively consistent associations and are classified based upon the major plant species within the community (i.e., Oak-Hickory forest, Tropical Rain forest).
B. Trophic levels or nutritional associations can be distinquished between interacting species. These different levels are referred to as:
1. Primary producers (green plants)
2. Primary consumers (herbivores)
3. Secondary consumers (carnivores)
4. Decomposers
5. Scavengers

C. Definite food chains can be distinquished in the community. Branching food chains make up food webs.
1. Food chain: a trophic path or succession of populations through which energy flows in an ecosystem as a result of consumer consumed relationships.
2. Food web: a complex of branching, joining, or diverging food chains that connect together the various populations in an ecosystem.
D. Communities exist within "ecosystems".
III. Ecosystems
A. An "ecosystem" is an interacting, self-sustaining, natural system of living organisms (the community) and of a chemical-physical component (the abiotic environment). This is the basic functional unit in ecology.
B. In biological control we deal with "agroecosystems" which are ecosystems composed of cultivated land, the plants contained or grown thereon, and the animals associated with these plants. These are extremely limited systems and they can be very unstable.
C. Within ecosystems, populations are thought to exist in a state of "homeostasis". This idea was suggested by Herbert Spencer (1897). Homeostasis is the tendency of a system to maintain a dynamic equilibrium, and if disturbed, to restore that equilibrium. The more complex a biological system (community) the more stable it usually is to disturbances. The more complicated the system the more reliable is the system of checks and balances against rise and fall.
IV. Arthropod Pest Populations Within Agroecosystems.
A. In agroecosystems one strives to maintain arthropod pests at noneconomic levels (not a "pest free" situation).
B. Economic Injury Level: the level at which insect induced damage can no longer be tolerated and therefore the level at or before which it is desirable to initiate deliberate control activities.
C. Economic Threshold: the pest density at which control measures should be applied to prevent an increasing pest population from reaching the economic injury level.
D. Economic threshold may also be referred to as the "action threshold", "density treatment level" or "treatment threshold". These terms usually do not include the influence of market values on the system and mainly refer to the impact of the pest upon the plant's productivity.
E. Most insects are maintained at very low population levels by "natural control" and are not viewed as pest species. Only 10,000 to 30,000 of the one million insect species known are pests. These pests feed on crops, forests, pasture lands, or livestock or menance our health, comfort or possessions.
F. Natural control is the maintenance of a more or less fluctuating population density within certain definable upper and lower limits over a period of time by the combined actions of the whole environment.
V. Population Growth and Regulation
A. Population ecology was originally confined to demography of human populations as was practiced by Chinese and Egyptian census takers.
B. Malthus (1803) published "Essay on the Principles of Population" which considered overpopulation of mankind. His ideas were similar to those of Giovanni Botero (1588) who proposed the same concept  of population regulation.
C. Malthus' idea was that "Man" increases at a geometric rate but his food supply does not. Soon became known as the "Malthusian Principle". Malthus was criticized because "Man" was considered to be capable of controlling his own birthrate. Another problem was that he considered the increase in subsistence (food & housing) was in a arithmetic ratio which was a major error. If this was so populations would increase without limit.
D. Charles Darwin (1859) accepted Malthus' principle. He placed much stress on:
1. Biotic mortality factors in the environment;
2. Competition as a major regulation factor; and
3. Climate as a factor in limiting population growth.
E. Herbert Spencer (1897) emphasized the concept of stability of the heterogeneous state and the instability of the homogeneous state.
F. Harry Scott Smith (1929) argued that the introduction of a complex of natural enemies (rather than a single species) should be employed in classical BC programs. He emphasized that the different kinds of natural
enemies available vary as to the particular habitats where they excel and that the biological control to be expected from their combined actions, as conditions vary from place to place and time to time, would be greater than that to be expected from the action of any one of them alone.


A specialized segment of the population of social insects, castes have different functions within the society and sometimes different morphologies. Castes have distinct divisions of labor
Social systems characterized by parental care of young, overlap of generations, and reproductive division of labor. True sociality.
The maintenance of a functional steady state in an organism or superorganism
Behavioral differences among castes
Caste members are radically different in appearance, us. results from environmental (food) differences
Social Insects
Insects that live cooperatively in colonies and exhibit a division of labor among distinct castes. ex. termites, ants, bees, some wasps.
A social insect colony described as a multicellular animal, individual members of the colony are similar to individual cells in an animal

Traits of Eusocial Insects

1) Parental care of young (young couldn't survive without parental care)
2) Overlap of generations (essential for 1)
3) Reproductive division of labor, i.e.. there are egg-laying females and other females, may be other castes
example: Honey Bees
  • Reproductive Castes - queen and drone
    • Queen - produces eggs to maintain the colony.
    • Drones - mate with new queens.
  • Worker Caste - sisters, all daughters of the queen
    • Care for the eggs, larvae, queen and drones.
    • Maintain and defend the hive, and forage for food.


Most insects are not social, some aggregate or contact other members of their species for short periods to mate or for other functions. Some even dispense with mating and reproduce asexually.
Only a few groups are truly social.
All termites (Isoptera), some Hymenoptera (all ants, honey bees, stingless bees, bumble bees, and some members of other bee groups, and at least one wasp sp.).
True social insects, esp. the ants and termites, are dominant ecological groups.


Solitary Insects
Social Insects
Hide from predators
Colony productivity increased

No competition with others of your species
Group defense and alarm

Live in small spaces
Food gathering

Exploit small food resources
Nest building

Care of young
Lack of social benefits
Intense predation, parasitism, disease
For societies to persist, they must survive and reproduce more successfully than solitary individuals.


  • Reproductives - queen and drones
  • Workers
  • Soldiers
    • May be distinct morphological types, esp. in ants.
    • Lacking in wasps and bees.

Three Castes of Termites

A Winged Adult Reproductive Termite

A Wingless Termite Worker

A Wingless Termite Soldier

Winged Adult Termite
A Termite Worker
A Termite Soldier

Comparison of Isoptera and Hymenoptera Caste Systems

Workers and Soldiers

Male Reproductives (Drones, Kings)
Immature or adult
Permanent attendant of the queen
Die after mating

Unlike bees, in termites the male is not a drone. Workers may be males and the king has functions other than mating. The queen may live for more than 10 years. But if she is killed or her egg production declines, secondary queens replace her.

Caste members may be radically different in appearance from one another or polymorphic and castes may have subcastes that differ in appearance and function. This usually results from environmental (food) differences not genetic differences. Behavioral differences among castes are called polyethism.


The Queen Bee
  •  Lays all the eggs and regulates sex of offspring (parthenogenesis).
    • Unfertilized eggs -> males
    • Fertilized eggs -> females
  • All members of the hive are the queen's progeny.
  • The queen's pheromones identify hive members.
The Worker Bees
  •  Workers determine type of egg laid by queen.
    • Large cells receive unfertilized eggs that develop into males -- males haploid.
    • Smaller cells receive fertilized eggs that develop into females -- females diploid.
  • Workers determine whether a female egg develops into a reproductive or worker.
    • Workers receive royal jelly only their first three days.
    • Queens receive royal jelly throughout the larval stage.


  • Wolves, killer whales, lions, man
  • SW African naked mole rats


Steps in evolution of animal size and complexity
  •  Single celled animals
  • Colonial animals
    • Clusters of cells capable of independent life
  • Multicellular animals
    • Groups of interdependent cells
      • Cellular division of labor
      • Cells not capable of independent life
      • Cellular communication and coordination essential
      • Most cells no longer reproducing new individuals
    • Superorganisms
      • Groups of interdependent, individual insects
        • Individual division of labor - castes
        • Individuals not capable of independent life
        • Colony individual to individual communication essential
        • Most individuals are not reproductive.

Peculiarities of Insect Societies


A biological, not a cultural trait, that is wide-spread among ants. Most ant battles you see are actually slave raids. Ant slavery is unique because ant slavery is usually between species, unlike human slavery.
Slave making ants
  • Capture larvae and pupae of another species.
  • Carry them back to there own nest where:
    • They acquire the nest odor.
    • Develop into adults and act as workers for their new colony.
Some slave making ant species are incapable of surviving without slave workers. They are no longer able to collect food or feed their immatures or themselves.


Embodies restless aggression, territorial conquest, and genocidal annihilation of neighboring colonies. Ants war with their own and other species and use a variety of tactics.
Imported Fire Ant, Solenopsis invicta vs. the Woodland Ant, Pheidole dentata
The fire ants have colonies hundred times larger than the woodland ant and whenever they discover a woodland ant colony they completely destroy it. Yet woodland ant colonies are abundant around fire ants. Whenever, a woodland worker discovers a fire ant scout soldiers are so rapidly deployed that the scout rarely makes it back to its colony. The soldiers do not sting or spray poisons like many ants but rely on large mandibles to cut their opponents into pieces. If despite this the woodland nest is discovered the soldiers fall back to form a short perimeter around the nest which keeps the invading fire ants at bay temporarily. The colony evacuates the nest and after the battle and the fire ants have departed, they will return and reclaim their nest.


Many ants keep insect livestock in the order Homoptera. Commonly seen in our area are ants tending aphids. The ants herd the aphids and protect them from predators and parasites, in turn, the aphids reward the ants by providing with droplets of sweet and nourishing honeydew. Besides aphids, scale insects, other Homoptera, are farmed and some insects in other orders.
Other ants and some termites are gardeners. They collect plant material, bring it into their nests, compost it, and use it to grow fungus which they feed on. Leaf cutter and parasol ants are examples.


Some social insects are able to maintain steady state conditions in their colonies or nests, e.g. in temperature and humidity. This is called homeostasis and is essential for colony health.
Examples: Honey bees
  • Ventilate their hives - if too hot, wax melts.
  • Cluster to stay warm in the winter - if too cold, individuals die.
  •  Soft bodied, very susceptible to desiccation.
  • "Air conditioned" termite mounds - vent heat and retain humidity.

  ~ Disappearing Act

Habitat: deserts Size: 2.5cm long Adaptation: can dig a hole in the sand using its stout, spiny legs & disappear into it within a few seconds; drink moisture that condenses on plants early in the morning Diet: aphids Predator(s):   

                                                  WEEVIL ~ Long Snout Beetle
Habitat: deserts Size: less than 6mm Adaptation: lay its eggs in the stalk, seed or fruit of plants & the young grubs eat their way out Diet: Predator(s):       

                                                  DARKLING BEETLE ~ Stinker
Habitat: deserts Size: 2-35mm Adaptation: stands with some of its feet lifted off the hot ground; can emit a foul-smelling black fluid, driving away its adversaries Diet: dry, decomposing plant or animal tissue Predator(s): birds, reptiles & amphibians                                 

                                                  ANTLION aka Doodlebug
  ~ Spiky Jaws

Habitat: deserts Size: ≈ 1cm long Adaptation: digs funnel-shaped pits near cacti to trap ants & insects Diet: ants & insects Predator(s): Extra: Adult ant lions look like dragonflies           

                                                  BUTTERFLY ~ Quick Breeder
Habitat: deserts Size: 0.8-4cm from tip to tip of its spread wings Adaptation: breed quickly Diet: nectar of flowers & other plant liquids Predator(s): birds; flies & wasps lay their eggs on or in the bodies of the butterfly caterpillars           

                                                  JEWEL WASP
  ~ Paralyzing Sting

Habitat: deserts Size: Adaptation: has a sting to paralyse other insects Diet: larva feeds on flesh of paralysed insects; adult feeds on nectar Predator(s):       
 This very unique desert animal is found in the deserts of Western North America. The female black widow is pitch black in colour except for an ominous looking red hour glass shaped mark on its abdomen while the male is roughly half the size of a female. It is a highly venomous spider that is unique in that the larger females devour the males after a courtship.

Sand grasshopper
Urnisa guttulosa

Desert grasshoppers such as the sand grasshopper can only feed and reproduce following rains that lead to the germination of green plants.
The rest of the time, and during the heat of the day, the sand grasshopper must shelter. Some desert grasshoppers shelter on trees and shrubs, but the sand grasshopper covers itself with sand. The sand grasshopper has special long middle legs to scoop out sand and eyes located at the very top of its head so it can sit buried with just its eyes and antennae exposed.
Desert grasshoppers do not breed regularly. Whenever it rains they feed, reproduce and lay eggs in the soil. Eggs can remain dormant for years if necessary, until the next heavy rain.
The sand grasshopper and many others such as the gibber grasshopper use the "stay-put" strategy of desert survival, and never reach plague numbers or damage crops like some locusts. 

Ants and Aphids association

Ants and aphids share a well-documented relationship of mutualism. Ants feed on the sugary honeydew left behind by aphids. In exchange, the ants protect the aphids from predators and parasites. In fact, honey ants will go to unusual lengths to ensure the health of the aphids in their care.
Aphids suck the sugar-rich fluids from their host plants. Because these liquids are low in nitrogen, the aphids must consume large quantities of them to gain adequate nutrition. The aphids then excrete equally large quantities of waste, called honeydew, which is high in sugar content.
Where there's sugar, there's bound to be ants. Some ants are so hungry for the honeydew, they'll actually "milk" the aphids to make them excrete it. The ants use their antennae to stroke the aphids, stimulating them to release the honeydew. Some aphid species have lost the ability to poop on their own, and now depend on their caretaker ants to milk them.
Aphid-herding ants make sure their "cattle" stay well-fed and safe. When the host plant is depleted of nutrients, the ants carry their aphids to a new food source. If predatory insects or parasites attempt to harm their wards, the ants will defend them aggressively. Some honey ants even go so far as to destroy the eggs of known aphid predators like lady beetles.
Some species of honey ants continue to care for their aphids during winter. The ants carry the aphid eggs home, and tuck them away in their nests for the winter months. They store the precious aphids where temperatures and humidity are optimal, and move them as needed when conditions in the nest change. In spring, when the aphids hatch, the ants carry them to a host plant to feed.
While it appears the ants are generous caretakers of their aphid charges, they've really got their own interests in mind. Aphids are almost always wingless, but certain environmental conditions will trigger them to develop wings. If the aphid population becomes too dense, or food sources decline, the winged aphids can fly to a new location. Not wanting to lose their food source, honey ants may prevent aphids from dispersing.
Ants have been observed tearing the wings from aphids before they can become airborne. A recent study has also shown that ants can use semiochemicals to stop the aphids from developing wings, and to impede their ability to walk away.

Study of soil, aquatic, scavenging, arborial & cave dwelling insects with respect to their body part adaptations & ecological significance.

Ecological significance.
Terrestrial Insects. The vast majority of insect species are adapted to a terrestrial life style. A rigid exoskeleton is often accompanied by a body surface that is resistant to water loss. The evolution of wings and the elaborate development of legs has afforded numerous options for locomotion on land.
Aquatic Insects. Although only about 5% of the insect species are aquatic, those that are represent an array of taxonomic groups. Aquatic insects may be predators, herbivores, or scavengers. Adaptations of aquatic insects are also varied, enhancing the likelihood of survival in various aquatic habitats. Various structures, such as suckers, claws and hooks, have been modified from legs and body parts to allow insects to cling to an underwater substrate in moving streams. Insects that inhabit still water are often physiologically adapted to an environment with reduced oxygen. Some insects have their legs adapted to swimming organs.
2) Soil Insects. There are numerous species of insects that have become specialized to inhabit soil. Springtails, beetles, termites, ants, and fly larvae are notable groups with soil-inhabiting members. Insects may ingest organic material near the soil surface and defecate organic remains deeper into the soil, thereby contributing to the process by which litter is transformed. Excavation by soil insects also alters the soil, mixing layers and creating pores where oxygen can be present.
Structural features of soil insects include reduction of wings and presence of a body that is round in cross section. Sometimes the forelegs are modified for digging. Mating often occurs above the ground during brief periods. Eyes and antennae may be reduced or absent.
Insects inhabit almost every imaginable habitat, including plants, wood, stored food products, and living animal tissue (parasitic insects).
Examine the material on display to become familiar with examples of adaptation and radiation of insects into different habitats.

Insect Adaptations to Low Temperatures

The ability of insects to adapt to diverse ecological conditions is legendary. This tremendous diversity is justly illustrated by their ability to withstand the intense cold of arctic and alpine environments. Indeed, the Arctic spring is accompanied by a veritable deluge of biting insects; a grim but unmistakable testament to their overwintering capabilities.
The adaptations that have evolved to allow insects to survive low temperatures are legion, but they can be classified along two general lines, freeze tolerance (the ability to survive following ice formation within the body cavity) and freeze avoidance (the prevention of ice formation within the body cavity at temperatures where such freezing would normall occur).
In addition to the problems posed by ice formation, there are also significant problems that must be solved for normal metabolism to occur at low temperatures. The maintenance of neural function, fluidity of cell membranes, pH control, activity of enzymes, adaptation to hypoxia, dehydration of body fluids, etc. all present obstacles to low temperature survival. Although these difficulties are formidable, they will not be discussed further here, as the adaptations to freezing temperatures are the focus of this chapter.
Very few insect species are actually exposed to the full rigors of winter temperatures as most choose an overwintering microhabitat that provides a buffered temperature. The habitats provided inside vegetation (logs, stumps, etc.) or under the soil provide thermal buffering, especially when covered with snow. In many climates, however, the organisms are still exposed to potentially lethal conditions throughout the winter. The particular adaptations associated with freeze tolerance and freeze avoidance allow these organisms to survive in such harsh environments.

·  Freeze Tolerance

Many species of insects have developed a tolerance for ice formation within their body fluids. The degree to which these species withstand freezing varies widely, from just a few degrees below freezing to -87°C for an Alaskan beetle. The strategies employed are legion, although the principle means for minimizing injury from ice formation is the use of cryoprotectants to reduce the amount of ice formed and the salt concentration at a given temperature.

·         Cryoprotectants

The cryoprotectants used by freeze tolerant species are similar (almost exactly so) to the compounds used by freeze avoidant insects to colligatively depress the freezing point. Glycerol is the most common cryoprotectant, followed by sorbitol and erythritol, ribitol, threitol, and sucrose. A multicomponent cryoprotection scheme is common, to reduce the concentrations of any given cryoprotectant to sub-toxic levels. The mechanism of cryoprotection appears to be simple colligative action, as in cryopreservation, with the additional benefit of stabilization of protein structure against low temperature denaturation.

·         Ice Nucleating Proteins

One strategy for mitigating the damaging effects of ice formation is to nucleate ice at a high sub-freezing temperature to avoid the high osmotic stresses associated with rapid freezing. Ice nucleators that have been found in insects are generally not too efficient, initiating ice growth at supercooling points between -7°C and -10°C, thus they are probably involved in avoiding intracellular ice growth rather than the directed ice growth seen in freeze-tolerant amphibians.
Some freeze tolerant insects have neither specialized ice nucleating proteins, nor an absence of ice nucleation sites (as in some freeze- avoidant insects). In a dry environment, they will supercool to near -20°C and then freeze and die. If they are inoculated with environmental ice at higher temperatures, however, they can survive. It seems, though, that most freeze tolerant insects produce ice nucleating proteins as part of their strategy for survival.

·         Stabilization of Bound Water

The water associated with macromolecules (bound water) is important for maintaining the tertiary structure of some molecules as well as the structure of membranes. It has been found that freeze tolerant organisms increase the amount of bound water in their systems during cold acclimation. That this confers additional tolerance to freezing is still a point of speculation.

·         Antifreeze Proteins

In many cases, damage from freezing has been linked to the rate of thawing, implicating the degree to which the ice undergoes recrystallization as a damaging mechanism. Since a few freeze tolerant insects produce antifreeze proteins, it has been speculated that these proteins minimize the injury associated with recrystallization of extracellular ice (AFP's are potent inhibitors of recrystallization). In fact, many of the freeze tolerant species that produce AFP's contain them in too low a concentration to produce thermal hysteresis; they are concentrated enough to inhibit recrystallization, however, since the thermodynamic driving force is much lower than for crystal growth in supercooled water.

·         Vitrification

It has been found that the hemolymph of a particular insect can be partially vitrified at cooling rates that are likely to occur in nature. Furthermore, the very low temperatures that some insects survive in the absence of ice indicate that vitrification could easily be achieved if the temperature went below the glass transition temperature. Thus it has been speculated that vitrification, or at least partial vitrification, may well be a strategy for freeze tolerance although direct evidence is thus far lacking.

·  Freeze Avoidance

Insects that are freeze-susceptible (ice formation in their body fluids is lethal) need to avoid freezing during the winter months. There are three basic strategies that insects employ to avoid ice formation within their body cavity: 1. Colligative depression of the freezing point through the concentration of a low molecular weight solute; 2. Production of an antifreeze protein (AFP) to lower the crystal-growth temperature non-colligatively; 3. Lowering of the nucleation temperature by removal of ice nucleation sites.

·         Colligative Freezing Point Depression

Colligative freezing point depression is simply the addition of low molecular weight solutes to the body fluids, exactly as occurs in normal cryoprotectant use. The solutes must be non-toxic in the concentrations required (molar), excluding many salts and small organic molecules. The polyhydroxy alcohols are the most common antifreeze solutes. Glycerol is undoubtedly the most prevalent polyol found in insects, but other compounds, such as ethylene glycol, sorbitol, and mannitol are also found in some species. There are other polyols that are found in elevated concentrations during the winter, but not in the molar quantities of an antifreeze solute (the combined effect, however, is to further reduce the freezing point of the solution); these include inositol, fucitol, arabitol, zylitol, rhamnitol, and ribitol. In conjunction with the polyols, elevated levels of the sugars trehalose, glucose and fructose are often found during the winter, as are elevated levels of the amino acid alanine. Such a multicomponent approach to freezing point depression allows a significant colligative action without bringing any one solute to the point of chemical toxicity.

·         Non-Colligative Freezing Point Depression

Thermal hysteresis producing antifreeze proteins (AFP's) have been found in many species of insects. The AFP's found in insects have, in some cases, been found to have a much higher activity than the AFP's isolated from polar fishes, primarily due to the increased concentration found in insects. Insect AFP activity has been found with a depression of the crystal growth temperature by as much as 6°C to 9°C below the melting point. None of the insect AFP's have been found to have carbohydrate moieties, in contrast to the antifreeze glycoproteins commonly found in antarctic fish.
The advantage of AFP's over colligative freezing point depression is in the much lower concentrations required and the ability to concentrate these molecules in the gut, where ice contamination is likely.

·         Supercooling

Pure water has a homogeneous nucleation temperature of -40°C, thus in the absence of any nucleation sites, it should be possible for an insect to survive very cold temperatures in a supercooled state if it could rid itself of all ice nucleators and prevent external ice from contacting its body fluids. It has been shown that some insects become much more susceptible to freeze injury if they are fed ice nucleating bacteria before exposure to cold temperatures, indicating that supercooling is a naturally occuring strategy for freeze-avoidant insects. In addition, insects have been found to have significantly lower supercooling points in winter, as compared with summer, without any lowering of their melting points or crystal growth temperatures. Some species remove ice nucleation sites seasonally whereas others have removed them permanently, over evolutionary time (a strategy that is evidently not compatible with all lifestyles).
There are some insect species found in the Canadian Rockies that have supercooling points of -60°C, combining colligative freezing point depression with an absence of ice nucleation sites to avoid ice formation at any terrestrial temperature (although these species are also freeze tolerant, there is simply no chance for them to experience temperatures of -60°C except in the laboratory).
Many aquatic insects live in cold regions where most aquatic habitats are ice-covered or frozen during winter. Insects survive the cold and icy conditions by a variety of adaptations: different species may move seasonally between habitats, choose particular overwintering sites, modify local conditions by constructing shelters, and withstand the effects of ice break-up and flooding during the spring thaw. In particular, although data are limited, several kinds of aquatic insects withstand subfreezing temperatures while surrounded by ice. A review of the available information suggests fruitful avenues for research, including detailed study of the cold hardiness of individual species in relation to habitat conditions, mechanical protection, dehydration, time, production of different types of cryoprotectants, and seasonal movements.