Bacillus thuringiensis strains with high insecticidal activity against insect larvae of the orders Coleoptera and Lepidoptera

Valeri Bagiyan; Marina Kinosyan; Narine Ghazanchyan; Nune Khachaturyan; Sona Gevorgyan; Karine Chitchyan; Sona Barseghyan; Arshaluys Verdyan; Vigen Goginyan

doi:10.1515/opag-2022-0384

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Bacillus thuringiensis strains with high insecticidal activity against insect larvae of the orders Coleoptera and Lepidoptera

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Published/Copyright: October 30, 2024

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From the journal Open Agriculture Volume 9 Issue 1

Abstract

The concept of sustainable agricultural development is aimed at reducing the use of chemical pest control agents. Since the use of chemical pesticides poses a significant environmental threat, one of the possible solutions to overcome this problem is the study of biological approaches, in particular, the use of entomopathogenic microorganisms as biological pest control agents. Insecticidal activity of over 220 strains of Bacillus thuringiensis (Bt) and Lysinibacillus sphaericus from the collection of the Laboratory of Spore-Forming Microorganisms of the Microbial Depository Center of the Scientific and Production Center “Armbiotechnology” and 15 strains isolated from samples of dead insects (imago) of representatives of various genera of the Coleoptera order was studied. The pathogenic effect of the strains was assessed in relation to the insect species Tenebrio molitor, Zophobas morio, Leptinotarsa decemlineata Say and Pieris brassicae, belonging to the Coleoptera and Lepidoptera orders. Among the test objects, the representative of Lepidoptera Pieris brassicae demonstrated the highest degree of sensitivity to B. thuringiensis strains up to 100%, with some Bt strains causing 60–100% mortality of Tenebrio molitor and Zophobas morio larvae of different ages. The larvae of the beetle Leptinotarsa decemlineata Say were also partially susceptible, presenting mortality levels of between 31 and 60%. The article is devoted to the study of Bt bacterial strains that have pronounced insecticidal activity against insect larvae of the orders Coleoptera and Lepidoptera.

Keywords: entomopathogenic bacteria; Bacillus thuringiensis ; toxins; insecticidal activity; coleopteran pests

1 Introduction

Nowadays, synthetic pesticides are still widely used to control insect pests of agricultural crops, although their negative impact on natural biocenoses is well known. In this regard, the use of biological control methods that can minimize the impact of chemical pesticides on the environment is an alternative for sustainable agricultural development. Similarly, a number of authors, e.g., Baker et al. [1] and Singh et al. [2], emphasize that the implementation of biological approaches in integrated pest management is a priority of sustainable agricultural development. As noted by Heckel [3], the use of entomopathogenic strains of Bacillus thuringiensis (Bt) as a biological method of insect control has a number of advantages over chemical ones, primarily in terms of environmental safety.

Bt, a member of the Bacillus cereus group, is ubiquitous in the environment, predominantly in soil [4,5]. Lacey et al. [6] reported that Bt has found wide application as a microbial insecticide in agriculture due to its insecticidal activity based on the combined action of its spores and crystals. The ability to form parasporal crystalline inclusions is a characteristic feature of Bt strains, which distinguishes them from other members of the Bacillus group [7,8]. Rosas-García [9] and Berini et al. [10] also reported the safety of insecticidal products based on Bt strains for humans and the environment as an important factor.

According to Güneş et al. [11] and Bravo et al. [12], the pathogenic effect of Bt is due to the obligatory presence of crystals of the delta-endotoxin protein (Cry toxin). At the same time, as observed by Roh et al. [13], the presence of crystals does not always ensure the obligatory effect of the Bt strains against insect pests.

Perhaps, as Letowski et al. [14] and Noguera and Ibarra [15] believe, this is explained by the presence of more than one crystal protein gene in pathogenic Bt strains. Entomocidal crystals are a type of protein toxins that is only harmful to insects when administered orally to insects [16]. Afrikian [17] describes that when silkworm (Bombyx mori) caterpillars were fed mulberry leaves treated with Bt crystals, it was found that after 1 h, the caterpillars stopped consuming food due to paralysis of the midgut, after which general paralysis and death of the insects are observed within 1–2 days. According to Bravo et al. [18], the mechanism of action of the endotoxin protein described for Lepidoptera is most likely identical for other orders of insects.

However, Federici [19] pointed out a difference between the suppressive toxicity of most of the Cry toxin proteins studied against lepidopteran pests and a small number of known delta-endotoxin crystal proteins that are toxic to Coleoptera. Nester et al. [20] and Portela-Dussán et al. [21] reported that Bt, in addition to Cry toxin proteins, also synthesizes a biodegradable vegetative insecticidal protein (Vip) in the vegetative growth phase. However, Palma et al. [22] emphasized that among the identified families of Vip proteins, only Vip1/Vip2 proteins exhibit toxicity against Coleoptera pests. According to Chakroun et al. [23], a fundamentally important fact regarding the pathogenic action of vegetative-insecticidal proteins has been established. High insect mortality is achieved only with the combined action of Vip1 and Vip2 proteins.

It should also be noted that the development of new technologies has opened up prospects for the production of more effective Bt formulations that will overcome some of the problems associated with limited field stability and loss of insecticidal activity, in particular, due to degradation of the pesticidal protein under the influence of ultraviolet radiation [24,25].

Although the toxicity of Bt proteins against target organisms from various insect orders has been studied for many years, research on the interactions between this bacterium and insect species classified in the order Coleoptera is more limited compared to those in the orders Lepidoptera and Diptera [7,26,27,28,29]. The work is devoted to the study of Bt bacterial strains with pronounced insecticidal activity against insect larvae from the order Coleoptera.

2 Materials and methods

The development of microbiological plant protection products with pronounced insecticidal activity against agricultural pests from the order Coleoptera was carried out using strains of entomopathogenic bacteria, including Bt, Brevibacillus laterosporus, and Lysinibacillus sphaericus (more than 3,000 strains) from the Culture Collection of the Microbial Depository Center (MDC) of the Scientific and Production Center “Armbiotechnology” of the National Academy of Sciences of Armenia, potentially possessing entomopathogenic action against insects of the order Coleoptera.

2.1 Strain maintenance and isolation

The collection strains and 15 new isolates used in this study are stored in the MDC Spore Forming Bacteria Laboratory. Strains were stored at 20°C as spores on meat-peptone agar slants (MPA – in g/100 ml: 1.0 peptone, 0.5 NaCl, 2.0 agar and 100 ml meat broth, pH 7.2) and activated in fish-yeast extract medium (FYE – in g/100 ml: 2.0 fish extract paste, 0.5 yeast extract, 2.0 agar, 100 ml water, pH 7.2) for 3–5 days. New isolates, designated NI 1 to NI 15, were recovered from dead insect samples on FYE agar with incubation at 30°C. The identity of each isolate was confirmed by a series of morphological and physiological tests. The type of strains of Bacillus cereus (MDC-2185/ATCC-25621/DSM-609), Bt (MDC-735/ATCC-10792/DSM-2046), Brevibacillus laterosporus (MDC-2176/ATCC-6456/DSM-335) and Lysinibacillus sphaericus (MDC-2177/ATCC-19394/DSM-488) from the Spore-Forming Bacteria Laboratory, and MDC Culture Collection were used as controls.

2.2 Preparation of material for bioassays

To study the insecticidal activity, the test strains were initially grown on FYE agar medium with incubation at 30°C for 48 h. Standardized suspensions of 2-day cultures (with a cell count of 4–6 × 10⁹ CFU) were used for the experiment. The inoculum included resuspended biomass (viable vegetative bacterial cells with abundant spore formation and endotoxin inclusions). Direct observations of the sporulation process and endotoxin crystal formation, determination of the cell count (in a Goryayev chamber), and the study of the growth phase of each culture were carried out using light microscopy.

2.3 Bioassays

In all bioassays using Tenebrio molitor and Zophobas morio, larvae were fed an artificial diet based on cabbage, wheat germ, and egg white. Colorado potato beetle (Leptinotarsa decemlineata Say) larvae were fed potato haulms. For the representative of the Lepidoptera order Pieris brassicae, common cress (Barbarea vulgaris) was used as a supplementary diet. The criteria used to define the association between mortality and toxicity were those established by us: (+) – low toxicity (1–30% mortality); (++) – medium toxicity (31–60% mortality); and high toxicity (+++) – high toxicity (61–100% mortality).

Mealworm and Zophobas (family Tenebrionidae) bioassays. Ten 2-week-old Tenebrio molitor and 4-week-old Zophobas morio larvae were placed in an 18 cm × 12 cm × 6 cm container filled with a feeding substrate treated with a bacterial suspension of 2-day-old cultures of the selected strains with a cell count of 10⁹ CFU/ml. Each analysis consisted of five repetitions of each test strain. The mortality rate was determined after 48 and 72 h.
Colorado potato beetle bioassay (family Chrysomelidae). The 10 s-instar larvae of Leptinotarsa decemlineata Say were placed in an 18 cm × 12 cm × 6 cm container filled with a feeding substrate treated with a bacterial suspension of 2-day-old cultures of the selected strains (which showed insecticidal properties against the beetle larvae of the pilot test objects Zophobas morio and Tenebrio molitor with a cell count of 10⁹ CFU/ml). Each analysis consisted of three repetitions of each test strain. After 48 h, dead larvae were removed, and the remaining larvae were transferred to a new container containing the control diet. Mortality levels were monitored for 5 days.
Pieris brassicae bioassay (family Pieridae, Lepidoptera). The 10 s-instar caterpillars were placed in a container measuring 18 cm × 12 cm × 6 cm, containing a substrate for feeding treated with a bacterial suspension of 2-day cultures of selected strains (which showed insecticidal properties against the beetle larvae of the pilot test objects Zophobas morio and Tenebrio molitor with a cell count of 10⁹ CFU/ml). Each assay consisted of three replicates of each test strain. Mortality rates were determined after 24 and 48 h.

The insect larvae for bioanalysis were provided by the Laboratory of Entomology and Soil Zoology of the Scientific Center of Zoology and Hydroecology of the National Academy of Sciences of Armenia.

2.4 Molecular genetic analysis

The bacterial genomic DNA was extracted using the FastPure^® Blood/Cell/Tissue/Bacteria DNA Isolation Mini Kit (Vazyme Biotech Co., Ltd, Nanjing, China) in accordance with the manufacturer’s instructions To perform bacterial 16S rRNA gene amplification, 27F (AGAGTTTGATCMTGGCTCAG) and 1492R (TACGGYTACCTTGTTACGACTT) universal primers were used. Sequencing was carried out at Novogene company (UK). For comparative analysis of nucleotide sequences, the BLAST program was used.

3 Results and discussion

3.1 Selection of strains of entomopathogenic bacteria with potential insecticidal activity according to the database of the Collection of Microorganism Cultures

The MDC contains more than 3,000 strains of entomopathogenic bacteria belonging to the species Bt (different serotypes), Lysinibacillus sphaericus, Brevibacillus laterosporus, and Bacillus popilliae. Given the large number and diversity of strains, the criterion for selecting priority strains for the study was their origin and synthesis of bacterial insecticides: dead insects of the orders Coleoptera and Lepidoptera (larvae or adults of the Colorado potato beetle, weevils, bark beetles, flour beetles, fruit borers, mulberry moths and other insect pests). As a result of monitoring the database of the Collection of Microorganism Cultures, over 220 cultures of Bt and Lysinibacillus sphaericus were selected:

172 strains isolated from dead insects of the order Coleoptera;
29 strains isolated from dead insects of the order Lepidoptera;
23 strains isolated from soil.

In addition, 15 new strains of entomopathogenic bacteria were isolated from dead insects (imago) belonging to various genera within the order Coleoptera, samples of which were collected during expeditionary work.

3.2 Selection of strains of entomopathogenic bacteria by cultural and morphological characteristics

By reviving museum cultures of the selected entomopathogens from dry shoals on the corresponding nutrient media (MPA, FYE), their viability and reproductive capacity were assessed, after which the second stage of selection was carried out based on the micromorphology of the strains. Microscopy was carried out after 24 and 48 h of incubation at 30°C, which is the optimal temperature for cultivating Bt bacteria. The main requirement for Bt strains used in industry is high entomocidal activity, along with other production indicators, such as phage resistance and high yield of the product. The Bt culture should ensure the highest possible percentage of spore and entomocidal inclusion formation since asporogenous variants and strains that do not form entomocidal inclusions do not have insecticidal activity. Since the pathogenic action of Bt is due to the presence of the delta-endotoxin protein, subsequent screening by phase-contrast microscopy showed that only actively growing 2-day cultures of 58 strains abundantly produced crystalline endospores during the sporulation phase. The form of the synthesized endotoxin was also taken into account. According to Afrikian [17], insecticidal action on representatives of the order Coleoptera is mainly possessed by strains of entomopathogenic bacteria with a quadrate and diamond crystal (endotoxin) shape. Further studies were conducted on the indicated strains with entomocidal inclusions to determine their insecticidal activity.

3.3 Testing the effect of entomopathogenic bacteria strains on test object larvae

By the nature of their entomopathogenic action, Bt crop insecticides can be classified as intestinal poisons. This is true primarily for toxins of a crystalline nature. In all cases, for the Bt culture to manifest its effect, endotoxin crystals must enter the insect’s intestinal tract, i.e., the spectrum of entomocidal activity is based on oral administration of native Bt culture liquid together with the food consumed and refers to the spore–cell–toxin complex. The nature of the entomopathogenic action of Bt cultures is highly dependent on the insect larvae and the age of the larval stage, as well as on the influence of other factors – temperature, food composition, intestinal microflora, etc. Based on this, various substrates were studied, which, after treatment with a suspension of 48 h, cultures of selected strains with a titer of 10⁹ CFU/ml were used to feed the larvae. Cabbage, wheat germ, and egg white were used as substrates for feeding. The feeding substrate without treatment with culture suspension was used as a control. The larvae of different ages of Zophobas morio (4–6 weeks) and Tenebrio molitor (2–4 weeks) were used as test objects. As shown in Table 1, when using a suspension of the same strain to treat different substrates, the variant with the egg demonstrates higher insecticidal activity. It can be stated that it is possible to use animal protein as a carrier to enhance the insecticidal effect of endotoxin, compared to other substrates.

Table 1

Influence of feed composition on the level of insecticidal activity of Bt strains towards the larvae of Tenebrio molitor and Zophobas morio

Strain, MDC	Mortality (%)
	Tenebrio molitor, age 2 weeks			Zophobas morio, age 4 weeks
	Egg white	Wheat germ	Cabbage	Egg white	Wheat germ	Cabbage
837	++	+	−	+	−	ND
1008	++	+	−	+	+	−
1063	+	−	ND	−	−	ND
1115	−	−	ND	−	−	ND
2267	++	−	−	−	−	−
2299	+	−	−	−	−	ND
2374	++	+	−	+	+	−
2393	+	−	−	+	−	−
2493	+	−	−	−	−	ND
2661	++	+	−	+	+	−
2661+1008	+++	++	+	+++	+	+
2670	+	+	−	+	−	−
2670+2661	++	+	+	+++	+	+
2673	+	+	−	+	−	ND
2673+1008	++	++	−	+++	+	+
2697	+	−	−	−	−	−
2700	+	−	−	−	ND	ND
2701	+	−	−	−	−	−
2702	+	+	−	−	−	−
2703	++	+	+	+	+	−
2706	+	−	−	−	−	ND
2728	+	−	−	−	−	−
2730	−	−	−	−	−	−
2767	+	−	−	−	−	−
2858	+	−	−	−	−	−
2867	+	−	−	−	−	−
2867	++	−	−	−	−	−
2936	+	−	−	−	ND	ND
14041	+++	+	+	++	−	−
14955	+++	−		++	−	−
15258	+++	++	−	++	+	−
15282	+	−	+	−	−	−
15282+1008	++	+	−	+++	+	−
15336	+++	+	−	++	+	−
15374	+	−	+	+	−	ND
15875	+	−	−	−	−	−
15891	+++	−	−	++	+	=
15891	+++	+	+	++	−	−
15892	+	−	−	−	−	−
15929	−	−	−	+	−	−
16084	++	−	−	−	−	−
16311	+++	++	+	+++	+	−
NI 2	+	−	−	_	−	ND
NI 3	+	−	−	+	−	ND
NI 5	+	−	+	+	−	−
NI 7	+	+	−	+	−	ND
NI 8	+	_	−	_	−	ND
NI 13	+	+	−	_	−	ND

* (−) – without toxicity, (+) – low toxicity; (++) – medium toxicity, (+++) – high toxicity, NI – newly isolated strain, ND – not done.

The tests were carried out both with individual Bt strains and with a combination consisting of different strains. The counting was carried out after 48 and 72 h. As a result of testing the effects of the selected strains of entomopathogenic bacteria and their combinations, their full or partial insecticidal activity was established in relation to the larvae of test objects of representatives of the Coleoptera order. It should also be noted that some strains demonstrated a higher mortality rate of test objects in the combined test than when used separately. It is possible that in these variants with strains 1008 or 2661, there is a combination of vegetative insecticidal proteins Vip1 and Vip2, the toxicity of which increases with combined action, which is consistent with known studies [22,23].

It was found that with the increasing age of the test object larvae, the insecticidal activity of entomopathogenic bacteria against larvae in terms of percentage decreases. It was also found that for the correct interpretation of the test results of the insecticidal activity, Bt strains demonstrated against test object larvae, the temperature in the laboratory should not exceed 25°C. The appearance of dead test object larvae after exposure to Bt strains is shown in Figures 1 and 2.

Figure 1

Insecticidal effect of the strain Bt 15891 on 4-week-old mealworm larvae.

Figure 2

Insecticidal effect of the strain Bt 16311 on Zophobas larvae (6 weeks old).

The initiation of sporulation apparently coincides with the beginning of crystal biosynthesis. The formation of these inclusions in the sporulating cell is an irreversible process, and its completion does not require additional exotrophic factors. When sporulating cells are suspended in distilled water, the formation of free crystals, as well as spores, is completed without any additional nutrient sources. The correlation of the quantitative content of spores and endotoxin crystals of Bt strains with insecticidal activity is presented in Table 2.

Table 2

Microscopic characteristics of selected strains of Bt and their toxic levels towards coleopteran test objects

Strain (MDC)	Mortality of larvae (%)		Microscopy, after 48 h	Isolation source and area
Strain (MDC)	Tenebrio molitor	Zophobas morio	Microscopy, after 48 h	Isolation source and area
837	++	+	80% free spores, 5% sporulating cells, crystals in large quantities	Silkworm, Lepidoptera, Bombyx mori L., Georgia, Kakheti region
1008	++	+	70% free spores, sporulating cells contain crystals	Colorado potato beetle (Leptinotarsa decemlineata Say), Armenia, Gegharkunik province
2374	++	+	70% free spores, sporulating cells contain crystals	Rat flea, (Xenopsylla cheopis), Russia, Krasnodar Krai
2661	++	+	80% free spores, 5% sporulating cells, crystals in large quantities	Small flour beetle (Tribolium confusum Duv.), Armenia, Kotayk province
2703	++	+	80% free spores, there are cells with crystals	Alfalfa leaf weevil, Coleoptera (Phytonomus variabilis Hbst.), Armenia, Armavir province
14041	+++	++	90% free spores, 10% sporulating cells, large amounts of diamond-shaped crystals	May beetle, Coleoptera, Melolontha pectoralis Germ., Armenia, Syunik province
14955	+++	++	90% free spores, sporulating cells with crystals	Cereal leaf beetle, Coleoptera (Lema melanopus L.,) Russia, Krasnodar Krai
15258	+++	++	80% free spores with remnants of sporangium, 10% vegetative cells, diamond-shaped crystals in large quantities	Colorado potato beetle (Leptinotarsa decemlineata Say), Armenia, Gegharkunik province
15336	+++	++	90% free spores, oval, large, single sporulating cells with crystals	Pit oak splendour beetle, Coleoptera (Chrysobothris affinis L., Armenia, Kotayk province
15891	+++	++	80% free spores, 20% sporulating cells, crystals in large quantities	Silkworm pupa Lepidoptera (Bombyx mori L.), Iran, Mazandaran province
16311	+++	+++	80% free spores, there are cells with a crystal	Silkworm farm, soil, Iran, Mazandaran province

(++) – medium toxicity and (+++) – high toxicity.

3.4 Insecticidal activity of Bt strains against lepidopteran pests

It is well known that Bt strains exhibiting insecticidal activity against Coleoptera due to the presence of the delta-endotoxin protein (Cry toxin) are also toxic to Lepidoptera [19]. As a test object, we chose the cabbage white butterfly, the caterpillars that cause the most severe damage in white cabbage and cauliflower. According to Carter and Hargreaves [30], the caterpillars of the second generation are the most harmful, which appear in late July–early August.

Studies of the insecticidal activity of selected Bt strains against lepidopteran caterpillars showed that almost all strains have a lethal effect on Pieris brassicae (L.) Moreover, in variants with strains MDC 2661, MDC 15258, and MDC 15336, 100% death of caterpillars is observed after 24 h, in contrast to strain MDC 837 (40% mortality), known for its activity against lepidopterans (Table 3).

Table 3

Insecticidal activity of strain Bt against cabbage white caterpillar lepidopteran Pieris brassicae (L.) (second-instar larvae)

Strain	Mortality of caterpillars (%)		Eating of leaf (%)
Strain	24 h	48 h	Eating of leaf (%)
MDC 837	++	+++	20
MDC 1008	+++	+++	10
MDC 2661	+++	+++	10
MDC 14041	+	+++	20
MDC 15258	+++	+++	10
MDC 15336	+++	+++	20
MDC 15891	++	+++	40
MDC 16311	++	+++	40
Control	0	0	50

(++) – medium toxicity and (+++) – high toxicity.

3.5 Insecticidal activity of Bt strains against coleopteran pests Colorado potato beetle (Leptinotarsa decemlineata Say)

The larvae and adults of beetles cause serious damage to agricultural crops due to their feeding habits, resulting in significant economic losses [31]. The Colorado potato beetle, Leptinotarsa decemlineata Say (Coleoptera, Chrysomelidae), is considered to be one of the most destructive insect pests of cultivated potatoes globally [32]. Alyokhin et al. [33] contend that pest control is difficult due to their natural tendency to adapt to toxic substances and due to their high resistance to pesticides, to which they quickly develop immunity. This has led to the emergence of a large number of Colorado potato beetle populations that are resistant to insecticides. Insecticide resistance in this insect remains a serious problem for pest management specialists.

The sublethal effect of the Bt delta-endotoxin protein on the larvae of Leptinotarsa decemlineata [34] and the pathogenic effect of the Bt Cry3A toxin on the development of the Colorado potato beetle [35] is known from the literature data. According to Hussein et al. [36], it is possible that the Cry3A toxin accumulates in the tissues of insects before the imago stage and negatively affects the reproduction of the Colorado potato beetle In this regard, a biological method of control against Leptinotarsa decemlineata Say is promising, especially since strains of Bt bacteria are known to exhibit activity against the Colorado potato beetle, provided that they are used at the first two larval stages [37].

We studied the effect of Bt strains with high insecticidal activity against Colorado potato beetle larvae. As shown in Table 4, strains MDC 15258, 15891, and 16311, and a combination of strains MDC 1008 and MDC 2661 showed toxicity against Leptinotarsa decemlineata Say larvae. However, it should be noted that to achieve the lethal concentration LC50 against second-instar larvae Colorado potato beetle, the exposure to strain MDC 12258 was 72 h, unlike other strains. It can also be stated that at a suspension cell count of the test strains of 4 × 10⁹ CFU/ml, the LC50 concentration of the toxin is achieved, which is necessary to destroy 50% of the exposed population of Colorado potato beetle larvae.

Table 4

Insecticidal activity of Bt strains against Colorado potato beetle (second-instar larvae)

Strain, MDC	Mortality (%)		Eating of leaf (%)
Strain, MDC	72 h	120 h	Eating of leaf (%)
1008+2661	+	++	80
14041	—	—	90
15258	++	++	60
15336	—	—	90
15891	+	++	60
16311	+	++	80
Control	−	−	100

(+) – low toxicity and (++) – medium toxicity.

The appearance of dead Colorado potato beetle larvae after exposure to Bt strain MDC 15258 is shown in Figure 3.

Figure 3

Insecticidal effect of Bt 15258 strain on second-instar larvae Colorado potato beetle.

3.6 Phylogenetic sequencing of the B. thuringiensis strains

The study of the genetic profile of strains of entomopathogenic bacteria with high insecticidal activity was carried out at Novogene Company (UK). As expected, the results of comparative analysis of the 16S rRNA sequence confirmed the phylogenetic relationship of the selected strains with Bt. Figure 4 shows the phylogenetic tree of the Bt strain MDC 15258 constructed using the neighbor joining method.

Figure 4

Phylogenetic tree of strain Bt MDC-15258.

4 Conclusion

In summary, the bioassay data indicate a wide activity spectrum of the Bt strains from the MDC Spore Forming Bacteria Laboratory collection against species of the insect orders Coleoptera and Lepidoptera, stimulating further experiments to evaluate the use of the Bt strains MDC 15258, MDC 15891, and MDC 16311 as a bioinsecticide, particularly against another agricultural pest, the click beetle. Another potential area of research is to elucidate the molecular basis for the increased toxicity of the combined action of Bt strains MDC 1008 and MDC 2661.

Acknowledgments

This work was supported by the Laboratory of Entomology and Soil Zoology of the Scientific Center of Zoology and Hydroecology of the National Academy of Sciences of Armenia. The authors express their gratitude to Dr. Mark Kalashyan for assistance in providing insect larvae of test objects.

Funding information: This research was funded by the Science Committee of the Ministry of ESCS of Armenia (grant number 23EDP-4D003).
Author contributions: All authors have accepted responsibility for the entire content of this manuscript and consented to its submission to the journal, reviewed all the results and approved the final version of the manuscript. VB – supervision, conceptualization, and writing – original draft; MK – supervision and investigation; NG – formal analysis and investigation; NK – methodology and investigation; SG – data curation and investigation; KC – investigation and validation; SB – writing – review and editing and software; AV – resources; VG – conceptualization, project administration, and writing – review and editing.
Conflict of interest: Authors state no conflict of interest.
Data availability statement: All data generated or analyzed during this study is included in this published article.

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Received: 2024-09-02

Revised: 2024-10-04

Accepted: 2024-10-09

Published Online: 2024-10-30

This work is licensed under the Creative Commons Attribution 4.0 International License.

Articles in the same Issue

https://doi.org/10.1515/opag-2022-0384

Keywords for this article

entomopathogenic bacteria; Bacillus thuringiensis; toxins; insecticidal activity; coleopteran pests

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