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Lecanicillium spp. are potential microbial bio-control agent mainly used for the management of sucking insect pests such as aphids, whiteflies, scales, mealy bugs etc. and gaining much importance at present for management of pests. Due to indiscriminate use of chemical pesticides which results in development of resistance, resurgence, outbreak of pests and residue problem, the farmers/growers are forced to use bio-pesticides for sustainable agriculture. Lecanicillium spp. is promising biocontrol agent against sucking insect pests and can be used as one of the components in integrated pest management (IPM). However, optimum temperature and relative humidity are the major environmental factors, for the performance of Lecanicillium spp. under protected/field conditions. The present review is mainly focused on nomenclature of Lecanicillium spp., mode of infection, natural occurrence, influence of temperature and humidity on the growth, factors influencing the efficacy, virulence/pathogenicity to target pests, substrates used for mass production, safety to non-target organisms, compatibility with agrochemicals and commercially available products. This review is mainly useful for the researchers/students to plan their future work on Lecanicillium spp.
chapter and author info
Sajjalavarahalli Gangireddy Eswara Reddy*
Entomology Laboratory, CSIR-Institute of Himalayan Bioresource Technology, Palampur-176061, Himachal Pradesh, India
*Address all correspondence to: firstname.lastname@example.org
The increased use of conventional chemical pesticides over the years has not only contributed to an increase in food production, but also has resulted in adverse effects on the environment and non-target organisms. In view of these side effects, the necessity for sustainable crop production through ecofriendly pest management technique is being largely felt in the recent times. Few biopesticides are available in the market, among them Lecanicilliumspp. based microbial bio-pesticide gaining much importance for sucking pests for organic and sustainable agriculture [1, 2, 3, 4]. Myco pesticides are potential microbial alternative to chemical pesticides and offer a number of benefits such as facility of growth on a variety of substrates, high virulence, trans cuticular penetration, broad host range, less expensive, safe to humans, animals and the environment. Therefore, this review is prepared by compiling the research work done on Lecanicilliumspp. by various research groups on various aspects viz., nomenclature, mode of infection, natural occurrence, effect of temperature and humidity for the growth, factors influencing its efficacy, virulence and pathogenicity against target pests under laboratory/greenhouse/field, substrates used for mass production, safety to non-target organisms, compatibility with agrochemicals and commercially available products were discussed and presented.
The genus Verticilliumcontains diverse host ranges including arthropods, nematodes, plants and fungi . The genus has been redefined using rDNA sequencing, grouping insect pathogens into the new genus Lecanicilliumwhich includes L. attenuatum, L. lecanii, L. longisporum, L. muscariumand L. nodulosum, which were all formerly classified as V. lecanii[5, 6, 7].
When L. lecaniiconidia comes in contact with the host integument, it gets adhere to the epicuticle and germinate. Germinated conidia form germ tubes that penetrate cuticle directly or grow over the surface of the epicuticle. The germ tube penetrates by lysing both the epicuticle and the procuticle [8, 9]. This is accomplished by the mechanical pressure exerted by appresorium (penetration peg) and secretion of enzymes viz., proteases, chitinases and esterase’s which plays an important role during cuticle penetration of insect host and also serve as cuticle degrading enzymes. The fungus proliferates throughout the insect’s body, draining the insect of nutrients, and eventually killing it in around 48–72 hours. The mycotoxins produced by L. lecaniiare bassianolide [10, 11], vertilecanin-A1, decenedioic acid and 10-hydroxy-8-decenoic acid) [12, 13, 14]. As the host nutrients are depleted, the blastopores’ differentiate into elongated hyphae which extend outward from the body forming a mycelial mat of conidiophores over the surface of the integument resulting in mummification. Under favourable environmental condition, conidiophores mature giving rise to conidia which continues the disease cycle further.
4. R & D publications on different aspects of Lecanicilliumspp.
The number of publications related to Lecanicilliumspp. from 1971 to 2020 was presented in the Figures 1 and 2. The data clearly indicated that, during 1971–80sthe publications were completely nil, but during 1981–91s, the R&D work has been initiated in the entire world and the publications were increased gradually reaching 58% during 2001–2020 (Figure 2). While, considering the number publications on various aspects of Lecanicilliumspp., more research work has been done on virulence and pathogenicity (Figure 3) followed by biotechnology and biochemistry as compared to morphology, diversity, ecology, mass production. The number of publications was meagre on effect of environmental factors (temperature and humidity), safety to natural enemies and compatibility with pesticides .
Lecanicilliumspp. is the most widely distributed and generally found on infected insects both in temperate and tropical areas throughout the world. There are number of reports on natural infection of Lecanicilliumspp. on different insect pests but out of the reported insects and pests, maximum are sucking pests belonging to Hemiptera, Thysanoptera and Acarina which indicates its possible spectrum for use as a biocontrol agent for pest management. Reports of natural occurrence of Lecanicilliumspp. on sucking insects presented in the Table 1.
6. Effect of temperature and humidity on the growth of Lecanicilliumspp.
Temperature and humidity are the main factors influencing the growth of the fungus. Effect of different temperature on conidial germination, growth rate, colony size and mycelial growth was discussed and presented in Table 2.
Effect of temperature on growth of Lecanicilliumspp.
Temperature affects the Lecanicilliumspp. in different ways by influencing the germination, growth and viability of the fungus in the host insect and environment. High temperature inactivates the fungus before contact with the pest insect or may reduce or accelerate the growth within an insect depending on the temperature requirements of the fungus and the host insect. In contrast, low temperatures reduce or stop the germination and growth. Optimal germination and growth rates of Lecanicilliumspp. range between 23°C and 28°C, growth rapidly slows >30°C and ceases at 34 to 37°C. Similarly, conidial germination is adversely affected by temperatures above 30°C. Temperature below 16°C increasingly slows germination and growth and thus affects efficacy in terms of a longer survival of the target population. Lecanicilliumstrains showed optimum growth at 25°C; the aerial conidia of Lecanicilliumstrains germinate in a broad temperature range (15–30°C) and L. lecanii41,185 was the only strain with conidial germination at 35°C .
Effect of different temperature on conidial germination, growth rate, colony size and mycelial growth of L lecaniiwas discussed and presented in Table 2. At 25°C and 0.975 aw (water activity) conidial germination occurred in all the isolates ranging from 28.7 to 98% whereas isolate PFC 10 no conidial germination had. Per cent germination decreased from highest values at 25°C to the lowest trend at 10°C in Mycotol (20.6°C). Maximum germination of conidia was observed between 15 and 25°C . Most of the isolates showed growth at 5 and 30°C and mean growth rate increased as temperature increased. Optimum growth rate occurred at 25°C (1.64 to 2.07 mm) for all isolates) . Colony size of the fungus was influenced by the temperature, the colony growth is maximum at 25°C (47.3 to 53.6 mm) as compared to the temperature between 5 to 20°C . The optimum temperature for the mycelial growth of L. lacaniiCA-1-G was 23°C (37.57 mg/cm2) and 26°C (39.43 mg/cm2) as compared to 20°C (29.43 mg/cm2), 29°C (20.7 mg/cm2) and 32°C (20.63 mg/cm2). Similarly, L. lecaniigrew and sporulated over a wide range of temperatures (20–32°C). The optimum temperature for growth was 23°C (46.45 x105 conidia cm−2) or 26°C (33.76 x105 conidia cm−2) for L. lecaniiCA-1-G . Virulence of Lecanicilliumspp. isolates was evaluated against third instar T. vaporariorumon tomato plants at 23°C. Colony radial growth, conidial production and germination decreased with the reduction in water activity, while 32°C was extremely detrimental for all fungal isolates. However, some isolates were able to grow and produce conidia at low water activity and high temperature . L. muscariumcan multiplied in temperature range of 15–30°C but optimum temperature against M. persicaebetween 20 to 30°C .
Humidity is another important environmental factor affecting the efficacy and survival of Lecanicillium. Spore germination on the insect cuticle and sporulation after outgrowth of the dead host insect require high moisture. Generally high humidity is required for germination of spores under in vitro, insects can become infected at much lower humidity. Under fluctuating humidity, daily saturated humidity requirement of at least 16 h for causing death in Trialeurodes vaporariorum(Westwood) infected with L. lecanii. Several previous studies provided evidence that a threshold time period at high humidity was required for infection. Conidia of L. lecaniirequired at least 72 h at 100% RH and 20°C before removal to 70% RH to reach >90% infectivity of Myzus persicae(Sulzer) . Similarly, at 25°C temperature and 75% relative humidity (RH), L. lecanii41,185 showed highly virulent pathogenicity (100%) against M. persicaeand Aphis gossypiiGlover . Application of L. longisporumagainst A. gossypiion cucumber in controlled environment (Temperature; 19–26°C and humidity; 80–98%) resulted in 100% mortality [32, 33]. L. muscariumgrow at optimum temperature but higher mortality observed against M. persicaebetween 55 and 90% humidity .
7. Factors influencing the efficacy of Lecanicilliumspp. against sucking insect pests
The virulence and pathogenicity of Lecanicilliumspp. vary with strain, stage of the insect and dose of the fungus.
Virulence of the Lecanicilliumspp. varies with strain to strain or isolate to isolate. The isolate ICAL6 was more virulent (LC50 = 1.05 x 107 conidia mL−1) to nymphs of M. persicaethan Macrosiphum euphorbiae(Thomas) (LC50 = 1.26 x 107 conidia mL−1)) and Nasonovia ribisingri(Mosley) (LC50 = 2.78 x 107 conidia mL−1) . The strain Vl 6063 imported from Canada was more virulent to Bemisia tabaci(Gennadius) (2.57 x 105 conidia mL−1)) than the domestic strains V3450 and Vp28 (LC50 = 6.03 x 105 conidia mL−1) . L. lecanii@ 1x107 conidia mL−1 is more effective against nymphs of Plannococcus citri(84% mortality) after six days of treatment as compared to L. longisporum(59% mortality) . L. muscariumisolate FI 2481 @ 1x107 conidia mL−1 was more effective against Thaumastocoris peregrinus(72% mortality) as compared to L. lecaniiisolate FI 2482 which reported 50% mortality . Similarly, L. lecaniihybrid strain 2aF4 was more promising (LC50 = 5.3x104 conidia mL−1) for the management of Trialeurodes vaporariorumthan L. lecanii2aF4 (LC50 = 7.8x104 conidia mL−1) .
7.2 Stage of the insect
Stage of host plays important role in the success of Lecanicilliumspp. and not all stages of insect life cycle are equally susceptible to fungal infection. So, the fungal application can be successful against the particular pest when it can be done at the condition where the susceptible stage or weaker stages of the particular pest become dominant among population.
First and third instar nymphs of B. tabaci(38 and 65% mortality) were significantly more susceptible to L. muscariumthan the fourth instar (15%) in verbena plants. Similarly, first and second instars B. tabaciwas more susceptible (50 and 55% mortality) than the third and fourth instar (25 and 20% mortality) on tomato foliage . L. lecanii(ARSEF 7460) showed higher mortality against nymphs of T. vaporariorumfollowed by L. longisporum(ARSEF 7207) and L. muscarium(ARSEF 74601) @ 1 x 107 conidia mL−1) . The pathogenicity of L. lecaniistrains was more in pupae (59–72.5%) than adults (34–52.6%) after 6 days of inoculation . L. lecanii(2.8 x 107 conidia/ml) isolated from Scirtothrips bispinosus(Bagnall) in tea showed higher mortality against larvae (60%) than adults (30%) of S. bispinosusunder laboratory at same dose . Mortality of nymphs of Plannococcus citriwere more susceptible (84% mortality) after six days of treatment to L. lecanii@ 1x107 conidia mL−1 as compared to adults which showed 40% mortality . L. lecaniihybrid strain 2aF43 @ 1x107 conidia mL−1 showed more efficacy against first instar nymphs of T. vaporariorum(68% mortality) as compared to 4th instar nymphs (30% mortality) and adults (60% mortality). Similarly, L. lecaniihybrid strain 2aF4 is more effective against first instar nymphs (46% mortality) as compared to 4th instar nymphs (30% mortality) .
7.3 Dose/inoculums level
Fungal inoculum level is the important factor which affects the performance. It is general trend that the higher fungal inoculum level gives higher insect mortality. However, sufficient inoculum level should be worked out for the particular pest to prevent the over inoculum wastage and to achieve higher mortality. Higher dose of L. lecanii(1.2 x 109 conidia ha−1) was caused 92.30 and 80.93% mortality of Brevicoryne brassicaeLinnaeus and Aleurodicus disperses(Russell) respectively at 10 days after treatment in the laboratory, whereas in field conditions L. lecanii(Vl3) at 2 x 1012 conidia ha −1) causing 61.16% and 66.50% mortality of B. brassicaeand A. craccivorarespectively .
8. Efficacy of Lecanicilliumspp. against sucking pests under laboratory/greenhouse/field
Efficacy of Lecanicilliumspp., against aphids, whiteflies, thrips, scales and mealy bugs in the laboratory/greenhouse/field conditions w.r.to its mortality, LC50 and LT50 values were presented in the Table 3.
9. Substrates used for mass production of Lecanicilliumspp.
Lecanicilliumspp. can be mass multiplied by solid state fermentation (SSF) and liquid state fermentation (LSF) using different growth media. In SSF, different grains, agars and non-synthetic solid media were used for mass production of Lecanicilliumspp. (Table 4).
Substrates (media, grains, agro wastes) used for mass production of Lecanicilliumspp.
Among grains, rice is most suitable substrates for mass production (1.97 x 109 spores g−1) followed by sorghum (1.90 x 109 spores g−1) as compared to finger millet, wheat and corn (1.6–1.80 x 109 spores g−1) . Similarly, crushed rice +1% yeast extract recorded higher spore yield (24.59 x 108 conidia g−1) followed by crushed bajra +1% yeast extract (17.49 x 108 spores g−1) as compared to crushed sorghum, maize and wheat .
Among different agro wastes used for multiplication of L. lecanii, thegrowth and sporulation were found to be better on rice bran +10% molasses (30.86 x 104 conidia g −1) followed by wheat bran +10% molasses (18.76 x 104 conidia g−1) and rice bran (15.98 x 104 conidia g−1). Complete inhibition of growth and reproduction of the fungus was noticed on bagasse and pressmud with 1 per cent yeast extract alone. However, growth was recorded when bagasse and press mud was supplemented with 10% molasses (10.88 and 7.90 conidia g−1 respectively) . Among agars, malt extract agar (MEA) yields high conidia production (5.23 x 107 conidia cm−2) followed yeast peptone dextrose agar (4.58 x107 conidia cm−2) as compared to potato dextrose agar and sabaroud dextrose agar (2.91 and 2.87 x 107 conidia cm−2 respectively) . In non-synthetic media, jack seeds produced high spore yield (4.11 x 108 spores g−1) followed by ladies’ finger (3.12 x 108 spores g−1), carrot (2.17 x 108 spores g−1) and rice husk (1.27 x 108 spores g−1) .
In LSF, molasses yeast broth (MYB) supported maximum spore production of L. lecanii(8.33 x 107 spores ml−1) followed by potato carrot broth (6.5 x 107 spores/ml) and potato sucrose broth (6.3 x 107 spores ml−1) as compared to Sucrose yeast broth, Jaggery yeast broth and Potato dextrose broth (2.45–3.95 x 107 spores mL−1). Among non-synthetic liquid media, coconut water produced higher spores (5.27 x 107 spore’s g−1) and biomass production than rice wash water (3.12 x 107 spore’s g−1) as compared to rice cooked water and wheat wash water (1.21–2.11 x 107 spores g−1) . The growth of L. longisporumconidial spores are higher in rice bran [24 x 107 conidia g−1] as compared to beet pulp [23 x 107 conidia g−1] .
10. Safety of Lecanicilliumspp. to parasitoids/predators/pollinators
The safety of any bio control agent to parasitoids/predators/pollinators is the important aspect which should be studied thoroughly before its commercialization to avoid the hazards and disturbance of ecological balance. Effect of L. lecaniion aphid parasitoid Aphidius colemani(Viereck) which showed the normal development (approximately 90% adult emergence) when its cotton aphid, A. gossypiihost was treated with L. lecaniiconidia 5 or 7 days after parasitization. Fungus exposure 1 day before or up to 3 days after parasitization, however, reduced Aphidius colemani(Viereck) emergence from 0 to 10%. They suggested that the parasitoid and fungus may be used together for aphid bio control . L. lecaniishowed pathogenicity against predatory mite, Phytoseiulus persimilisAthias-Henriot but its effect was lower than that of spider mite, Tetranychus urticae(Koch) . L. lecaniiis safer to predatory coccinellid, Coccinella septempunctataLinnaeus and predatory mites, Amblyseius ovalis(Evans) and Amblyseius longispinosus(Evans) under field conditions . The fungus L. lecaniiwas not pathogenic to Chrysoperla carnea(Stephens), Coccinella septempunctata(Linnaeus), Episyrphus balteatus(De Geer) and Samia cynthia ricini(Boisduval), but was found to be pathogenic to Bombyx mori(Linnaeus). Parasitism, adult emergence and adult longevity of Trichogramma chilonis(Ishii) were affected by fungal treatments. Aphid mummification and Diaeretiella rapaeadult emergence were affected by the fungus. Results suggest that L. lecaniiis compatible with natural enemies of cabbage aphid, T. chilonisand is harmless to silk worm . L. muscariumat 106 and 107 spores mL−1 was safer to predatory mite P. persimilis. Number of parasitized larvae of Eretmocerus sp. nr. furuhashiisurvival decreased with increasing concentrations of L. muscariumand only 29% emergence of pupae was observed at a conidial concentration of 1 × 108 conidia mL−1. Similarly, 67% emergence of adult E. sp. nr. Furuhashiiwas observed . Parasitoid (Diaeretiella rapae) emergence was affected by application of L. longisporumbefore or after parasitism and longevity decreased in female F1 populations . In the laboratory conditions, application of L. muscarium(1 x 108 conidia/ml) against A. colemanihad not affected longevity and fertility of the female A. colemani. The combination of Aphidius colemaniwith L. muscariumreduced the aphid infestation in the semi field conditions as compared to A. colemanialone .
The Lecanicilliumspp. is not harmful to humans during handling in the laboratory and field for the control of pests.
11. Compatibility of Lecanicilliumspp. with agro chemicals
Chemical pesticides may have antagonistic or synergistic effect on the potentiality of Lecanicilliumspp. and may disrupt natural epizootic. Under such epizootic condition, it is expected to enhance effectiveness through joint action of pathogen and compatible insecticides, which would reduce not only the cost of protection but also reduce the contamination of the environment. The literature on compatibility of Lecanicilliumspp. with agrochemicals is lacking.
Among different insecticides studied for their effect on L. lecaniiunder in-vitro, malathion was significantly detrimental (69.18% inhibition) than all other insecticides except quinalphos (66.76%). Conversely, endosulfon and chlorpyriphos were significantly safer (37.31 to 44.37%), followed by oxydemeton methyl and dimethoate (45.33 to 48.27% inhibition) . Similarly, endosulfan completely inhibited the germination of conidia and hyphal growth. In contrast, diafenthiuron, thiamethoxam, imidacloprid, thiodicarb, primicarb, omethoate, acetamiprid, and pymetrozine were compatible with L. lecanii in planta. Imidacloprid and cyromazine were compatible with L. lecaniiin terms of vegetative growth, sporulation, conidial viability and pathogenicity against T. urticae.At the recommended concentration, the fungicides carbendazim, chlorothalonil, propiconazole, mancozeb and wettable sulphur completely inhibited the germination of candida (100%) except iprodione and triadimefom allowed 37.38 and 41.62% conidia to germinate respectively .
12. Commercial formulations
The commercial formulations based on Lecaniilliumspp. are available in India and other countries are presented in Table 5. Number of manufacturers based on Lecanicilliumspp. products is more in India however; the production is very low and not available to the farmers/stakeholders/growers on time as compared to synthetics due to dominant in pesticides market and lack of awareness to farmers/growers about biopesticides. In India, the efficacy of Lecanicilliumspp., based products was less due to high temperature and low humidity as compared to temperate countries, even though in India, these products were used as one of the components in IPM and also used for the management of sucking pests of flowers and vegetables in greenhouse.
Commercially available products based on Lecanicilliumspp. [58, 59].
Lecanicilliumspp. is promising biocontrol agent and can be used as one of the components of integrated pest management under green house and field conditions against sucking insect pests. Lecanicilliumis multiplying on commercially available media (potato dextrose agar and broth etc.) till date but it can be mass multiplied at cheaper rate on solid grain media of sorghum and rice; liquid media of sugar cane molasses. It can be used effectively in conjunction with other natural enemies and compatible pesticides.
Author is grateful to Director, Institute of Himalayan Bioresource Technology (Council of Scientific and Industrial Research, New Delhi), Ministry of Science and Technology, Government of India, Palampur, Himachal Pradesh, India for encouragement and support.
Conflict of interest
Author declares that no conflict of interest is reported.
Sajjalavarahalli Gangireddy Eswara Reddy (October 22nd 2020). <em>Lecanicillium</em> spp. for the Management of Aphids, Whiteflies, Thrips, Scales and Mealy Bugs: Review, Arthropods - Are They Beneficial for Mankind?, Ramón Eduardo Rebolledo Ranz, IntechOpen, DOI: 10.5772/intechopen.94020. Available from:
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