Advantages to the Use of Insect-Resistant

Crop Varieties
• Economic benefits occur because crop yields are saved from loss to
insect pests and money is saved by not applying insecticides that would
have been applied to susceptible varieties.
• Ecological and environmental benefits arise from increases in species
diversity in the agroecosystem, in part because of reduced use of
insecticides. Increases in species diversity increase ecosystem stability
which promotes a more sustainable system far less polluted and
detrimental to natural resources.
• Use of insect-resistant crop varieties as a component of IPM
• Insect-resistant cultivars synergize the effects of natural, biological, and
cultural insect pest-suppression tactics. The “built-in” protection of
resistant plants from insect pests functions at very basic level, disrupting
the normal association of the insect pest with its host plant.
• Plant resistance to insect pest have advantages over other direct control
tactics. For example, plant resistance to insects is compatible with
insecticides use, while biological control is not. Plant resistance is specific,
only affecting the target pest. Often effects of use of insect-resistant
cultivars are cumulative over time. Usually the effectiveness of resistant
cultivars is long-lasting.
 Chapter 3
NATURE OF BIOLOGICAL CONTROL
AGENTS
Parasitoid
• The term parasitoid was coined in 1913 by the German
writer O.M. Reuter (and adopted in English by his
reviewer, William Morton Wheeler) to describe the
strategy in which, during its development the parasite
lives in or on the body of a single host individual,
eventually killing that host, the adult parasitoid being
free-living.
• About 10% of described insect species are
entomophagous parasitoids.
Two other orders with parasitoidal members:

• The “twisted-wing parasites” (order Strepsiptera), which is a small group

consisting entirely of parasitoids.
• The beetles (order Coleoptera), which includes at least two families,
Ripiphordae and Rhipiceridae, tha are largely parasitoids, and rove beetles
(family Staphylinidae) of the genus Aleochara.
• Occasional members of others can be parasitoids; one of the remarkable is
the moth family Epipyropidae, which are ectoparasitoids of planthoppers
and Cicadas. The genus Cyclotorna has even more elaborate habits,
beginning its growth period parasitizing plant bugs, and concluding by
feeding on ant larvae in their colonies.
Types of parasitoids
Ectoparasitoid – it is the one which develops externally
on its host.
Example : Bracon brevicornis on coconut black headed
caterpillars.

Endoparasitoid – it is the one which develops within

the body of its host.
Example : Eriborius trochanteratus on coconut black
headed caterpillars.
Parasites
a. Lady Beetles
 Most adult lady Beetles are round to oval, brightly
colored and often spotted.
 Lady beetle larvae are elongated, usually dark
Adult lady beetle Larvae lady beetle
colored, and flecked with orange or yellow.
 Adult and larvae feed on large numbers of small,
soft-bodied insects such as aphids.
 One group of small, black lady beetles (Stethorus)
is important in controlling spider mites and others
specialize in scale insects.
 One species of lady beetle, however, the Mexican
bean beetle, is a plant pest. This common Colorado
insect is found feeding on bean leaves. It is
distinguished from other lady beetles by spotting
and color in the adult stage. Larvae of Mexican black lady beetles (Stethorus) Mexican lady beetles
bean beetle are yellow and spiny.
 b. Green Lacewings
• adult stage: a pale green insect with
large, clear, highly-veined wings that are
held over the body when at rest.
• Adult green lacewings primarily feed on
nectar and other fluids, but some species
also consume a few small insects.
• Green lacewings lay a distinctive stalked
egg. Lacewing larvae emerge in four to 10
days.
• These larvae, sometimes called aphid
lions, are voracious predators capable of
feeding on small caterpillars and beetles,
as well as aphids and other insects.
 c. Syrphid Flies
• Several names : flower flies or hover flies.
• Most are brightly colored, yellow or orange and black,
and may resemble bees or yellow jacket wasps.
• It is the larval stage of the sryphid fly that preys on
insects.
• Variously colored, the tapered maggots crawl over foliage
and can eat dozens of small, soft-bodied insects each day.
• Sryphid flies are particularly important in controlling
aphid infestations early in the season, when cooler
temperatures may inhibit other predators. Similar in
appearance to sryphid fly larvae is a small, bright orange
predatory midge (Aphidoletes).
 d. Predatory Bugs
• True bugs (Order: Hemiptera) are predators of
insects and mites. All feed by piercing the prey
with their narrow mouthparts and sucking out
body fluids. A red and black species of
predatory stink bug, capable of feeding on
fairly large insects such as caterpillars and
potato beetle larvae, is most conspicuous.
• Most common of all the predatory bugs are
the small (less than 1/8 inch) minute pirate
bugs. Minute pirate bugs are most frequently
seen in flower or in crevices of a green plant,
where they feed on thrips, spider mites and
insect eggs. Other predatory bugs common in
yards and gardens include ambush bugs and
assassin bugs.
e. Ground Beetles
• Immature stages are
distinctly different from
adults and more often are
found within the top few
inches of soil.
• Ground beetles are general
feeders with powerful jaws.
f. Mantids
• Mantids are general predators that feed
on almost any insects of the right size.
• They have one generations per year
with winter spent as eggs within a pod.
• One species of mantids, the Chinese
mantid, is sometimes available for sale.
 g. Hunting wasps
• A large number of wasps from several
families prey on insect pest.
• Many take their prey, whole or in
pieces, back to their mud, soil or
paper nests to feed to the immature
wasps.
• Example, the common Polistes paper
wasps, when hunting, may thoroughly
search plants and feed on caterpillars,
often providing substantial control of
these insects.
h. Predatory Mites
• Several mite species are predators of plant-
feeding spider mites. Typically, these
predatory mites are a little larger than spider
mites but are more rounded in shape and
faster moving than their prey.
• Predatory mites often can provide good
control of spider mites. Low humidity can
restrict their activity. They are also more
susceptible to insecticides than are plant-
feeding species.
Predatory mite used to control spider mite in
tomatoes
i. Spiders
• All spiders feed on insects or other small
arthropods.
• Most people are familiar with many
common web-making species. However,
there are many others spiders – wolf Black widow spider
spiders, crab spiders, jumping spiders –
that do not build webs but instead move
about and hunt their prey on soil or plants.
• These less conspicuous spiders can be
important in controlling insect pests such
as beetles, caterpillars, leafhoppers and
aphids.
3. Pathogens of Insects
• Pathogen - an entity that can incite disease.
• Pathogenicity – the capability of a pathogen to cause
• Disease- any malfunctioning of host cells and tissues that results from
continuous irritation by a pathogenic agent or environmental factor and leads
to development of symptoms.
• Pathogens are viruses or microorganisms that cause disease.
• Insects are susceptible to a variety of diseases caused by pathogens. Insect
pathogens have a relatively narrow host range and thus are considered to be
more environmentally friendly than synthetic chemical insecticides.
• The pathogens that cause disease in insects fall into four main groups: viruses,
bacteria, fungi, and protozoa.
a. Viruses
• Viruses are obligate intracellular parasites, meaning that they can reproduce
only in living cells and are composed in the simplest form of a nucleic acid,
either DNA or RNA, and a protein shell referred to as the capsid.
• Insect viruses can be cultured in living hosts (in vivo) or in cultures insect
cells (in vitro).
• Insect viruses are divided into two broad nontaxonomic categories:
• Occluded viruses are so named because after formation in infected cells, the
mature virus particles (virions) are occluded within a protein matrix, forming a
paracrystalline bodies that are generically referred to as either inclusion or
occlusion bodies.
• Nonoccluded viruses, the virions occur freely or occasionally form
paracrytalline arrays of virioins that are also known as inclusion bodies. These,
however, have no occlusion body protein interspersed among the virioins.
 Types of insect viruses

Iridoviridaeviridae: Iridovirus
 The viral structure is non-enveloped, non-
occluded, isocahedral viral particles.
 Viral particles are organized into crystalline
arrays. Light reflected from such arrays
interferes with incident light, resulting in the
characteristic iridescent colors that are the
most obvious sign of patent infection. Iridovirus infected (blue) larva of Aedes aegypti
next to a healthy larva.
 The small iridovirus tend to display colors
from violet to turquoise.
 Iridoviridaeviridae: Iridovirus
 Although some iridoviruses infect frogs and fishes, those infecting insects
belong to two genera: Iridovirus, whose viral particles fluctuate between 120
to 130 nm in size.
 They mostly infect arthropods, particularly insects, in damp or aquatic
habitats worldwide. Most commonly reported from larval stages of Diptera
larvae, such as mosquito larvae, as well as from larvae of Coleoptera and
Lepidoptera.
 They are highly infectious by injection but have low infectivity by ingestion.
 Horizontal transmission can occur by cannibalism or predation of patently
infected individuals, or the virus may even be vectored by nematodes and
parasitoid wasps that introduce viral particles into the host insect during the
act of penetration or oviposition.
 Cytoplasmic Polyhedrosis Viruses
• The cytoplasmic polyhydrosis viruses (family Reoviridae) are occluded double-stranded RNA viruses with
a genome divided into 9 or 10 segments of RNA.
• These viruses, commonly referred to as CPVs, cause a chronic disease and reproduce only in the stomach
of insects, where typically they from large (ca. 0.5-2 im) polyhedral to spherical occlusion bodies in the
cytoplasm of midgut epithelial cells.
• Infection in early instars retards growth and development, extending the larval phase by weeks.
• The disease is often fatal. In advance stages of disease, the infected midgut is white rather than
translucent brown, because of large numbers of accumulated polyhydra.
• This virus type is relatively common among lepidopterous insects and among dipterous insects of the
suborder Nematocera (e.g., mosquitoes, blackflies, midges).
• CPVs are typically easy to transmit by feeding to species that belong to the same family of the host from
which they were isolated, and thus the host range of this virus type is quite broad.
 Poxviridae:Entomopox Viruses

GV NPV

GV NPV
 Poxviridae:Entomopoxvirus
 Member of the family of Poxviridae has a wide host, including
vertebrates and invertebrates.
 Chicken pox and Small pox virus belong to this family.
 The show allantoid – to brick-shaped virions, occluded within
ovoid OBs called Spheroids.
 Entomopoxvirus has been isolated from 27 orthopterans,
lepidopterans, dipterans and coleopterans.
 The subfamily Poxvirinae includes three genera, i.e.
Entomopoxvirus A, Entomopoxvirus B, and Entomopoxvirus Anomala cuprea (Coleoptera) larvae infected with
an entomopoxvirus show the symptoms of the
C infection such as a whitish appearance and
 Entomopoxvirus A infects only coleopteran species; underdevelopment (left, infected larva; right,
Entomopoxvirus B infects lepidopteran and coleopteran healthy one).

species; Entomopoxvirus C infects only dipteran species.

 The fourth group, group D, has been proposed by ICTV
which only attacks hymenopterans.
 Ascoviridae: Ascovirus
 A few species of Ascovirus has been isolated only from insects,
specifically from Lepidopterans (Noctuidae).
 Enveloped virions of ascoviruses are bacilliform, ovoid or allantoid in
shape, and occluded within vesicle-like OBs.

An ascovirus-infected caterpillar
Structure of Ascovirus virions
 Ascoviridae: Ascovirus
Ascovirus Lifecycle
 Ascovirus establishes in the Helicoverpa population through spring-
summer.
 The disease is transmitted from caterpillar to caterpillar by wasps (such
as Microplitis).
 Ascovirus could be transmitted directly from one caterpillar to another by
spitting - for example, when caterpillars encounter each other on the plant.
 When ascovirus particles enter the caterpillar´s body they multiply in
tissue cells, eventually infecting the haemolymph (blood). This causes the
haemolymph to change from clear to milky. The caterpillar stops eating,
but may not die for several days or weeks, surviving in a lethargic state.
 Ascoviridae: Ascovirus

Ascovirus symptoms
 In cases where a Microplitis wasp has both parasitised a caterpillar and
infected it with ascovirus, the symptoms seen are those of the disease rather
than of the parasitoid. When ascovirus kills the caterpillar, it also kills the
developing Microplitis larva.
 Caterpillars infected with ascovirus will generally stop eating within two
days. They stop growing, but can live for weeks in a lethargic state before
they die.
 The blood of an ascovirus-infected caterpillar is white and creamy, whereas
the blood of a healthy caterpillar is clear. Blood colour gives the best
diagnosis in the laboratory and can be tested by splitting or pricking the
caterpillar.
Baculoviridae: Granulosis viruses (GV)

 Baculoviruses (family Baculoviridae) are large, enveloped,

double-stranded, occluded DNA viruses.
 About 65 species oval or ovoid granules.
 These viruses are divided into two main types, commonly
known as the nuclear polyhedrosis viruses (NPVs) and the
granulosis viruses (GVs).
 Infections: Granulosis viruses attack the adipose tissue.
 Host: Lepidoptera larvae
 Survival: granulosis viruses form particles inside a crystalline
protein structure (occlusion body). This allows the virus to
survive outside the host. Can survive for year out of sunlight.
 Biocontrol agents: Cydia pomonella GV (codling moth) ,
Phythorimaca operculelle GV (potato tuber moth)
Use of Viruses as Insect Control Agents
NPV of the European spruce sawfly, Gilpinia hercyniae, as a classical
biological control agents.

Nuclear polyhedrosis virus (NPV) - part of the family of baculoviruses,

is a virus affecting insects, predominantly moths and butterflies. It has
been used as a pesticide.

The European spruce sawfly was introduced into eastern Canada from
northern Europe around the turn of the last century and had become a
severe forest pest by the 1930s. Hymenopteran parasitoids were
introduced from Europe in the mid-1930s as part of a biological control
effort, and inadvertently along with these came the NPV, which was first
detected in 1936.

Most sawfly populations were reduced to below economic threshold

levels by 1943 and remain under natural control today, the control being
affected by a combination of the NPV, which accounts for more than 90%
of the control, and the wasp parasitoids.
cytoplasmic polyhydosis viruses (CPVs) - for suppression of the
pine caterpillar, Dendrolimus spectabilis in Japan.
Ascoviruses and ento-mopoxviruses have not been developed as
control agents for any insect owing to lack of efficacy.
 one baculovirus, the Autographa
californica nucleopolyhedrovirus (AcMNPV) noted earlier, has
been developed as an expression vector for producing a large
number of foreign proteins in vitro.
 This expression system takes advantage of the strong
polyhedron promoter system, which in the wild-type viruses
produces large amounts of the polyhedria used to occlude
virions.
 By substituting foreign genes for the polyhedrin gene, it is
possible to synthesize in insect cell cultures large quantities of
foreign proteins, such as the capsid proteins of viruses that
attack the vertebrates used for vaccine development and basic
biochemical research.
 B. Bacteria
• Bacteria ate relatively simple unicellular microorganisms that lack internal
organelles such as a nucleus and mitochondria, and reproduce by binary
fission.
• A variety of bacteria are capable of causing diseases in insects.
• Most study are spore-forming bacilli (family Bacillaceae), especially B.
thuringiensis.
• Many subspecies of Bt are used as bacterial insecticides and as a source of
genes for insecticidal proteins added to make transgenic plant resistant to insect
attack, especially attack by caterpillars and beetles.
• The other bacterial insect pathogens that have received various degrees of
study are B. sphaericus, Paenibacillus popilliae, and P. larvae, the latter being
the etiological agent of foulbrood, an important disease of honey bee larvae,
Serratia entomophila and S. marcescens.
Bacillus thuringiensis
 What are some products that contain Bacillus thuringiensis
(Bt)?
• Currently, Bt strains are found in over 180 registered
pesticide products. Bt products are used on crops and
ornamental plants. Others are used in and around
buildings, in aquatic settings, and in aerial applications.
These products are commonly sprays, dusts, granules,
and pellets. Some of these products are approved for use
in organic agriculture.
• Some crops have been engineered to make the Bt toxin.
These plant-incorporated protectants include corn,
cotton, and soybeans.
 How does Bacillus thuringiensis (Bt) work?
• Bt makes toxins that target insect larvae when eaten. In their gut, the toxins
are activated. The activated toxin breaks down their gut, and the insects die
of infection and starvation. Death can occur within a few hours or weeks.
• The different types of Bt create toxins that can only be activated by the target
insect larvae. In contrast, when people eat the same toxins, the toxins are not
activated and no harm occurs.
• Each type of Bt toxin is highly specific to the target insect. For example, the
‘kurstaki’ type targets caterpillars. The ‘isrealensis’ type targets immature flies
and mosquitoes. Little to no direct toxicity to non-target insects has been
observed.
 What happens to Bacillus thuringiensis
(Bt) in the environment?
• Toxins created by Bt are rapidly broken down by sunlight and in acidic soil.
Other microbes in soil can also break it down. Bt does not readily leach in
soil. It typically remains in the top several inches of soil. Bt remains dormant
in most natural soil conditions. However, there has been some reproduction
in nutrient rich soils. On the soil surface, dormant Bt cells last only a few
days. However, below the soil surface, they can last for months or years.
The half-life in unfavorable soil is about 4 months. Bt toxins break down
much faster. In one study, 12% remained after 15 days.
• In water, Bt does not readily reproduce. A study found Bt toxins in the air
were broken down rapidly by sunlight. Forty-one percent (41%) of the toxin
remained after 24 hours. On plant surfaces, sunlight breaks down Bt; the
half-life of Bt toxins is 1-4 days.
Bacillus sphaericus
• Many isolates of B. sphaericus (Bs) are toxic to certain mosquito species.
• Over the past three decades, three isolates have been evaluated for their mosquito control
potential,
– 1593 from Indonesia and 2297 from Sri Lanka - isolates were obtained from soil and
water samples at mosquito breeding sites.
– 2362 from Nigeria - isolated from a dead adult blackfly.
• Bs acquires its toxicity as the result of protein endotox-ins that are produced during
sporulation and assembled into par-asporal bodies.
• Bs is unusual in that the main toxin is a binary toxin (i.e., composed of two protein subunits).
These are proteolytically activated in the mosquito midgut to release peptides having
molecular masses of, respectively, 43 and 39 kDa, that associate to form the binary toxin,
with the former protein constituting the binding domain, and the latter the toxin domain.
• The toxins bind to micro-villi of the midgut epithelium, causing hypertrophy and lysis of cells,
destroying the midgut and killing the mosquito larva.
 c. Paenibacillus popilliae
 P. popilliae is a highly fastidious bacterium that is the primary eti-ologial agent of the so-
called milky diseases of scarab larvae.
 The term “milky disease” is derived from the opaque white color that characterizes
diseased larvae and results from the accumulation of sporulating bacteria in larval
hemolymph (blood).
 These insects are the immature stages of beetles, such as the Japanese beetle, Popillia
japonica, that are important grass and plant pests belonging to the coleopteran family
Scarabaeidae.
 The disease is initiated when grubs feeding on the roots and vegetative cells invade the
midgut epithelium, where they grow and reproduce, changing in form as they progress
toward invasion of the homocoel (body cavity).
 After passing through the basement membrane of the midgut, the bacteria colonize the
blood over a period of several weeks and sporulate, reaching populations of 100,000,000
cells ml-1.
d. Serratia entomophila
 A novel bacterium named. S. entomophila cause amber disease in the grass grub,
Costelystra zealandica an important pest of pastures in New Zealand, and has been
developed as a biological control agent for this pest.
 This bacterium adheres to the chitinous intima of the foregut, where it grows extensively,
eventually causing the larvae to develop an amber color; the result of infection is death.
 The bacterium is easily grown and mass-produced in vitro and can now be grown to
densities as hogh as 4 X 1010 cells ml_1.
 Successful mass production of S. entomophila led to its rapid commercialization. It is now
used to treat infested pastures in New Zealand at a rate of one liter of product per hectare.
 Liquid formulations of this living, nonspore-forming bacterium are applied with subsurface
application equipment.
 The rapid development and commercialization of the bacterium, even though the use is
rather restricted, shows how microbials can be successful in niche markets, where there are
few alternatives, and mass production methods, the most critical factor, are available.
Vegetables - animal science

Rice - entomology

Fruit and Plantation Crops - plant breeding

Life cycle or mode of action of entomopathogenic fungi against the insect pests
 a. Aquatic Fungi
• Aquatic fungi of two types that attack mosquito larvae have received considerable study:
species of Coelomomyces (class Chytridiomucetes: order Blastocladiales) and Lagenidium
giganteum (class Oomycetes: order Lagenidium).
• The genuse Coelomomyces comprises over 80 species of obligately parasitic fungi that have a
complex life cycle involving an illustration of sexual (gametophytic) and asexual (sporophytic)
generations.
• The sexual phase parasitizes a microcrustacean host, typically a cope-pod, whereas the
asexual generations develops, with rare exception, in mosquito larvae.
• In the life cycle, a biflagellate zygospore invades the homocoel of a mosquito larva, where it
produces a sporophyte that colonizes the body and forms resistant sporangia.
• The larva dies and subsequently the sporangia undergo meiosis, producing uni-flagellate
meiospores that invade the homocoel of a copepod host, where a gametophyte develops. At
maturation, the gametophyte cleaves, forming thousands of uniflagellate gametes.
b. Terrestrial Fungi
• The fungi that have received the most attention for use in
biological control are terrestrial fungi, with most emphasis placed
on the development of selected species of hyphomycetes such as
M. anisopliae and B. bassiana for use as microbial insecticides. In
addition, the more specific and nutritionally fastidious
entomophthoraceous fungi continue to receive attention, but for
their potential use as classical biological control agents rather
than as microbial insecticides.
 ENTOMOPHTHORALES
• The Entomophthorales are an order of mainly entomogenous fungi. They are
usually associated with Diptera, although some are pathogens of other
arthropods, nematodes and tardigrades, amongst others.
• Entomophthorales comprises a large order of zygomycete fungi that contains
numerous genera, many species of which are commonly found parasitizing
insects and other arthropods.
• These fungi routinely cause localized and in some cases widespread
epizootics in populations of hemipterous and homopterous insects,
particularly aphids and leafhoppers, but also in other types of insects such as
grasshoppers, flies, beetle larvae, and caterpillars.
 ENTOMOPHTHORALES
• The typical life cycle of the entomopathogenic species of Entomophthorales
involves the invasion of the host by germ hyphae produced by adhesive
spores (termed conidia) which are airborne.
• The fungus invades the abdomen of the host and, following its death,
sporophores are produced, typically between the individual segments of the
abdomen, where a new crop of forcibly discharged spores are produced.
• Culture media : two recipes have proven successful in many cases: SDA
supplemented with egg yolk and milk, and EYSMA.
• A simple type of medium consisting of SDA supplemented with sesame oil
and sucrose fatty acid esters was found to be appropriate for maintenance
and preservation of several species of Entomophthorales (Feng & Xu, 2001),
although adding egg yolk resulted in faster growth.
 PROTOZOA
• Protos – first; Zoion- animal
• Protozoa is a diverse group of eukaryotic unicellular motile microorganisms
that belong to what is now known as the kingdom Protista.
• The word “protozoa” was by coined by GEORG AUGUST GOLDFUSS in
1818.
• The cell contains a variety of organelles, but no cell wall, and cells vary greatly
in size and shape among different species.
• Cannot make their own food; they ingest algae, yeasts, bacteria and smaller
protozoa for nutrients.
• Feeding is by ingestion or more typically by adsorption, and vegetative
reproduction is by binary or multiple fission.
• Potential for use as fast-acting microbial insecticides because of the chronic
nature of the diseases.
MICROSPORIDIA
• The microsporidia (phylum Microspora) are
the most common and best studied of the
protozoans that cause important diseases of
insects.
• over a 1000 species are known, and most of
these have been described from insects
• Microsporidia have been most commonly
described from insects of the orders
Coleoptera, Lepidoptera, Diptera, and
Orthoptera
• The epizootics in insect populations caused
by protozoa are usually due to
microsporidia.
• All microsporidia are obligate intracellular parasites and
are unusual in that they lack mitochondria.
• They produce spores that are distinguished from the
spores of organisms of all other known types by the
presence of a polar filament along coiled tube inside the
spore used to infect hosts with the sporoplasm.
• The typical microsporidian life cycle begins with the
ingestion of the spore by a susceptible insect. Once inside
the midgut, the polar filament everts, rapidly injecting the
sporoplasm into host tissue.
• The disease often lasts for several weeks during which
billions of spores may accumulate in the tissues of a
single infected host.
• Many microsporidia are transmitted vertically from adult
females to larvae via the egg (transovarially).