The word entomopathogens was derived from entomology which is the branch of zoology that studies insects and how these insects interact with man, animals and their environment. Entomopathogens are microbes which can be viruses, bacteria, fungi or protozoa. Insects should be studied because they have economic importance that is either detrimental or beneficial to man, plants, animals and the environment. Entomopathogens are microbes including bacteria fungi, and viruses that attack insects. They act as parasites of insects, and thus kill them or stop their reproduction and spread in any environment, especially when such insects cause destruction to plants or the environment. Entomopathogens are a typical example of biological control agents used to control insect infestation in any environment such as the farm land. They are considered natural mortality agents and environmentally safe for controlling insect infestation. Entomopathogens are the natural enemies of insects, and they can be effectively used to control pest or insect infestation in any environment without affecting other non-target organisms in the same environment. The main objectives of the entomopathogens are to prevent, reduce or delay the infestation caused by these insects or pest.
Entomopathogens are microorganisms that are pathogenic to arthropods such as insects, mites, and ticks. They include microbes in the group bacteria (e.g. Bacillus species), viruses (Helicoverpa zea single-enveloped nucleopolyhedrovirus, HzSNVP), nematodes (e.g. Heterorhabditis species) and fungi (e.g. Isaria species). Entomopathogens are better alternatives to the use of insecticides, herbicides or other chemical control agents to control pest or insect in the environment. This is because these chemical control agents are usually poisonous in nature and may affect the ecosystem of where they are used. Chemical control agents or insecticides can also leave some untoward effects on the plant or crop population for which the insecticide was used to protect from insect or pest infestation.
There are many varieties of microbes that infect insects and other arthropods. Also, microbes form symbiotic (mutualistic) association with insects as wells as parasitic relationships. In a mutualistic association, both the insect and the microbe enjoy the company of each other by mutually benefitting from one another’s activities. However, in a parasitic relationship, one member of the association (either the insect or the microbe) suffers from the union. This fact informed the decision to use these insect parasites (entomopathogens) as biological control agents for controlling the nefarious activities of insects or pests on plants and the environment.
Pests include plant pathogens (e.g. fungi, viruses, bacteria), arthropods, weeds, and other small vertebrates that cause damage to plants. The economic cost of pests is enormous, and thus, it is important to control their activities in order to ensure food security across the globe. Pests reduce the yield and quality of crops by:
- Feeding on crops,
- Transmitting infection, diseases or infectious disease agents (pathogens), and
- Competing with crop plants for space and other resources for survival.
Methods of isolation of entomopathogens from natural environments
The natural environments from which insect parasites (entomopathogens) can be isolated from include: the soil, plant parts, dead insects, aquatic forest, agricultural habitat, grazing land (pasture), desert, and urban habitats. However, the soil happens to be the number one site or natural source from which entomopathogens could be isolated from. The soil is a habitat for a wide variety of economically important microbes including antibiotic-producing bacteria. The distribution of these insect parasites in the environment can be affected by both abiotic and biotic factors such as agricultural activities, temperature, humidity, moisture and industrialization.
The methods involved in the isolation of entomopathogens are:
- Soil sampling. Selection of sample (soil) source
- Collection of samples (soil)
- Processing of samples (soil)
- Selection of bait insects or pests
- Preparation of selective culture media for isolation of entomopathogens
- Isolation and identification of entomopathogens
- Bioassay tests. Bioassay is used to quantitatively or qualitatively determine the presence, amount, or functional activity of the entomopathogens of interest isolated. It is used to determine the insecticidal action of the secreted metabolite from the isolated entomopathogens. Bioassay test is used for determining the strength or biological activity of the entomopathogens isolated or produced. This can be carried out using bait insects, mammals, birds or target pests.
- Genomic DNA analysis using PCR and DNA sequencing
Safety tests for microbial control agents against man, animals and other non-target organisms
The safety of microbial control agents is important because these agents can cause several harmful effects to humans and other non-target organisms in the form of inflammation and toxicity. Over a long period, the accumulation of toxins from microbial control agents in humans can cause cancer and other genetic defects. It is therefore important to determine their safety after production before they can be used for any pest control measures. Safety of entomopathogenic microbes used in microbial control practices is a major concern associated with the increased needs for biocontrol agents desired in the exploitation of integrated pest management (IPM) programs for agricultural, veterinary, and medical insect pests.
Before the registration, distribution and usage of microbial control agents, the following aspects of the products are taken into consideration in order to determine their safety to man, plants, non-target organisms and the environment:
- Allergic and toxic properties
- Risks of toxic metabolites
- Genetic recombination and displacement of natural strains of non-target organisms
- Effect on biodiversity, e., on non-target organisms
Microbes used for or as microbial control agents occur naturally in the environment. But the general absence for infection of man with such microbes in the medical literature is important evidence that these agents do not pose a significant human health risk, as well as possible danger to other animals or non-target hosts in the environment. These non-target organisms include: beneficial plants, fish, crustaceans, birds, mammals, and non-target insects and mites. As a precautionary measure, each microbial control agent or microbe used for the production of microbial control agents should be treated individually without generalizing their safety measures since an agent that is safe in one environment may be harmful in another environment.
To test microbial control agents, the entomopathogens are used on non-target organisms including but not limited to fishes, insects, birds, and mammals in the environment. The reason for testing microbial control agents is to ensure that they do not produce or carry toxic substances (inclusive of genes and toxins) that may adversely affect the environment, non-target plants, microbes, animals and humans over their short or prolonged usage.
Bacillus thuringiensis is a Gram-positive bacterium that dwells in the soil, and which is widely used as a biocontrol agent to control pest in the environment (such as in the farm land). B. thuringiensis is unique because it produces exotoxins that have insecticidal activity. The exotoxins that they produce are proteinous in nature, and they have the ability to weaken the activities of pests. The exotoxins target the larvae of insects that are pests.
The major safety studies include:
- Toxicology studies. These are studies designed to study the toxicity or harmful effects of substances or chemicals on living organisms including man. It can be carried out in vitro or in vivo. Toxicological studies are used to find out how poisonous a substance or chemical is to living organisms. Those who carryout such studies are known as toxicologists.
- Pathogenicity tests. Pathogenicity is the ability or capacity of a pathogen to cause disease in a living organism. Pathogenicity tests are studies carried out to check for the possibility of the microbial control agent to cause infection or disease in non-target hosts or organisms. The pathogenicity of the microbial control agent is assessed from the signs and symptoms of disease caused by the entomopathogen used in its production. Pathogenicity test is usually carried out in vivo using laboratory animals such as mice or rats.
- Allergenicity studies. Allergens are antigens or substances that cause allergic (hypersensitivity) reactions in living organisms. Allergenicity studies are tests used to decipher the ability of a microbial control agent to cause allergic reactions in living organisms. It can be carried out in vivo or in vitro by analyzing representative animal, plant, human or environmental samples for the presence of allergens from the microbial control agents.
- Ecological (persistence) risk assessment in environment. This study is aimed at ensuring that there is no unintended impact of the microbial control agent on non-target organisms. It also ensures that there are no contamination of natural resources such as water bodies, faunas, plants and other ecosystems as a result of the use of the microbial control agent. Ecological risk assessment can be carried out by evaluating the dangers of the side effects of these microbial control agents on the general safety of the environment through the assessment of the ecosystems for possible residues or harmful toxins from the microbial control agent.
- Dietary risk assessment. Some microbes used for the production of microbial control agents have been genetically engineered into the genetic makeup of pest resistant crops. For example, Bt corn (insecticide-producing corn) contain proteins (exotoxins) genetically sourced from thuringiensis. BT corn expresses proteins or metabolites that are pest resistant in nature. Such corn becomes toxic to pests that feed on the crop (corn) while remaining harmless to humans/animals. However, the possible toxicity of such corn has been questioned, and this warrants the need to do dietary risk assessment on such products in order to ascertain their safety for consumption. Though the target pests are primarily exposed to the infectious proteins in the corn, small traces or amounts of the protein may find their way to the GIT of consumers.