Bacteria from the Midgut of Common Cockchafer (Melolontha melolontha L.) Larvae Exhibiting Antagonistic Activity Against Bacterial Symbionts of Entomopathogenic Nematodes: Isolation and Molecular Identification

The mechanisms of action of the complex including entomopathogenic nematodes of the genera Steinernema and Heterorhabditis and their mutualistic partners, i.e., bacteria Xenorhabdus and Photorhabdus, have been well explained, and the nematodes have been commercialized as biological control agents against many soil insect pests. However, little is known regarding the nature of the relationships between these bacteria and the gut microbiota of infected insects. In the present study, 900 bacterial isolates that were obtained from the midgut samples of Melolontha melolontha larvae were screened for their antagonistic activity against the selected species of the genera Xenorhabdus and Photorhabdus. Twelve strains exhibited significant antibacterial activity in the applied tests. They were identified based on 16S rRNA and rpoB, rpoD, or recA gene sequences as Pseudomonas chlororaphis, Citrobacter murliniae, Acinetobacter calcoaceticus, Chryseobacterium lathyri, Chryseobacterium sp., Serratia liquefaciens, and Serratia sp. The culture filtrate of the isolate P. chlororaphis MMC3 L3 04 exerted the strongest inhibitory effect on the tested bacteria. The results of the preliminary study that are presented here, which focused on interactions between the insect gut microbiota and mutualistic bacteria of entomopathogenic nematodes, show that bacteria inhabiting the gut of insects might play a key role in insect resistance to entomopathogenic nematode pressure.

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Nanopore-Sequencing Characterization of the Gut Microbiota of Melolontha melolontha Larvae: Contribution to Protection against Entomopathogenic Nematodes?

Pathogens ◽

10.3390/pathogens10040396 ◽

2021 ◽

Vol 10 (4) ◽

pp. 396

Author(s):

Ewa Sajnaga ◽

Marcin Skowronek ◽

Agnieszka Kalwasińska ◽

Waldemar Kazimierczak ◽

Karolina Ferenc ◽

...

Keyword(s):

Gut Microbiota ◽

Entomopathogenic Nematodes ◽

Bacterial Species ◽

Antagonistic Activity ◽

Rrna Gene ◽

Nanopore Sequencing ◽

Bacterial Phyla ◽

Melolontha Melolontha

This study focused on the potential relationships between midgut microbiota of the common cockchafer Melolontha melolontha larvae and their resistance to entomopathogenic nematodes (EPN) infection. We investigated the bacterial community associated with control and unsusceptible EPN-exposed insects through nanopore sequencing of the 16S rRNA gene. Firmicutes, Proteobacteria, Actinobacteria, and Bacteroidetes were the most abundant bacterial phyla within the complex and variable midgut microbiota of the wild M. melolontha larvae. The core microbiota was found to include 82 genera, which accounted for 3.4% of the total number of identified genera. The EPN-resistant larvae differed significantly from the control ones in the abundance of many genera belonging to the Actinomycetales, Rhizobiales, and Clostridiales orders. Additionally, the analysis of the microbiome networks revealed different sets of keystone midgut bacterial genera between these two groups of insects, indicating differences in the mutual interactions between bacteria. Finally, we detected Xenorhabdus and Photorhabdus as gut residents and various bacterial species exhibiting antagonistic activity against these entomopathogens. This study paves the way to further research aimed at unravelling the role of the host gut microbiota on the output of EPN infection, which may contribute to enhancement of the efficiency of nematodes used in eco-friendly pest management.

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Synergism of gut microbiota to double-stranded RNAs in RNA interference of a leaf beetle

10.1101/824581 ◽

2019 ◽

Cited By ~ 1

Author(s):

Letian Xu ◽

Shijing Xu ◽

Liuwei Sun ◽

Yiqiu Zhang ◽

Jing Luo ◽

...

Keyword(s):

Rna Interference ◽

Gut Microbiota ◽

Insect Pests ◽

Leaf Beetle ◽

Gut Bacteria ◽

Biotic And Abiotic Stress ◽

Epithelial Surface ◽

Pass Through ◽

Insect Gut ◽

Insect Gut Bacteria

AbstractRNA interference (RNAi) has emerged as an efficient tool to control insect pests. When lethal double-stranded RNAs (dsRNAs) were ingested by the insects, strong gene silencing and mortality can be induced. To exert their function, dsRNA molecules must pass through insect’s gut and enter epithelial cells and/or the hemolymph. Gut bacteria are known to inhabit on the epithelial surface to confer host new capabilities to counter both biotic and abiotic stress. Whether there is a crosstalk between gut bacteria and dsRNAs and the effects of the microbiome on RNAi efficiency remains unknown. Here, using a leaf beetle-gut microbiota system, we investigated whether and how gut bacteria interact with dsRNA molecules and its effects on host insects. We firstly showed that the leaf beetle Plagiodera versicolora (Coleoptera) is highly susceptible to RNAi. Then, we found that ingestion of dsRNAs by non-axenic P. versicolora larvae results in (i) significantly accelerated mortality compared to axenic larvae, and (ii) over-growth and dysbiosis of the gut microbiota. The latter is mainly caused by the bacterial utilization of the dsRNA degraded products initiated by the host insect. Furthermore, we found that Pseudomonas putida, a gut bacterium of P. versicolora, was a main commensal-to-pathogen strain that accelerated the death of P. versicolora larvae. Taken together, our findings reveal a synergistic role of gut microbiota to dsRNA-induced mortality of pest insects, which provides new insights in the ecological functions of insect gut bacteria, and also contributes to a better understanding of the RNAi mechanisms in insects.

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Non-lethal effects of entomopathogenic nematode infection

Scientific Reports ◽

10.1038/s41598-021-96270-2 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Camila C. Filgueiras ◽

Denis S. Willett

Keyword(s):

Biological Control ◽

Entomopathogenic Nematodes ◽

Insect Pests ◽

Entomopathogenic Nematode ◽

Pupal Stage ◽

Nematode Infection ◽

Biological Control Agents ◽

Lethal Effects ◽

Risk Of Death ◽

Strain Dependent

AbstractEntomopathogenic nematodes are typically considered lethal parasites of insect hosts. Indeed they are employed as such for biological control of insect pests. The effects of exposure to entomopathogenic nematodes are not strictly limited to mortality, however. Here we explore non-lethal effects of exposure to entomopathogenic nematodes by introducing the relatively non-susceptible pupal stage of Delia antiqua to thirteen different strains. We specifically chose to inoculate the pupal stage because it tends to be more resistant to infection, yet resides in the soil where it could come into contact with EPN biological control agents. We find that there is no significant mortality at the pupal stage, but that there are a host of strain-dependent non-lethal effects during and after the transition to adulthood including altered developmental times and changes in risk of death compared to controls. We also find that exposure to specific strains can reduce risk of mortality. These results emphasize the strain-dependent nature of entomopathogenic nematode infection and highlight the positive and negative ramifications for non-lethal effects for biological control of insect pests. Our work emphasizes the need for strain-specific screening of biological control agents before wide-spread adoption.

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The Invasion and Encapsulation of the Entomopathogenic Nematode, Steinernema abbasi, in Aedes albopictus (Diptera: Culicidae) Larvae

Insects ◽

10.3390/insects11120832 ◽

2020 ◽

Vol 11 (12) ◽

pp. 832

Author(s):

Wei-Ting Liu ◽

Tien-Lai Chen ◽

Roger F. Hou ◽

Cheng-Chen Chen ◽

Wu-Chun Tu

Keyword(s):

Aedes Albopictus ◽

Entomopathogenic Nematodes ◽

Biocontrol Agent ◽

Insect Pests ◽

Entomopathogenic Nematode ◽

Asian Tiger Mosquito ◽

The Public ◽

The Past ◽

Asian Tiger ◽

Vector Mosquito

The Asian tiger mosquito, Aedes albopictus, is of crucial concern to the public and veterinary health because of its vector role in transmission of several mosquito-borne diseases. Over the past decades, entomopathogenic nematodes (EPNs) have been used to control important agricultural insect pests and are considered to be effective against mosquitoes as well. The objectives of this study were to investigate the mosquitocidal effects of Steinernema abbasi to Ae. albopictus and the encapsulation processes of invading nematodes in the mosquito host. In this study, we found that S. abbasi was pathogenic to 3rd and 4th instar larvae of Ae. albopictus by entering the hemocoel of the 3rd and 4th instar larvae mainly through mouth and gastric caecum or by penetrating pupae through the intersegmental membrane or trumpet. The mosquito larvae infected with a single nematode caused a high mortality. Although EPNs in the hemocoel of mosquitoes were melanized and encapsulated, most Ae. albopictus larvae failed to survive after infection with S. abbasi. Overall, we demonstrated that S. abbasi is pathogenic to Ae. albopictus larvae, suggesting that this S. abbasi isolate has potential as a biocontrol agent for managing this vector mosquito.

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Efficacy of Entomopathogenic Nematode, Steinernema dharanaii (TFRIEPN-15) against Termite, Odontotermes obesus (Isoptera: Termitidae)

Indian Journal of Forestry ◽

10.54207/bsmps1000-2019-p4t27o ◽

2019 ◽

Vol 42 (4) ◽

pp. 392-397

Author(s):

Sanjay Paunikar ◽

◽

N. Kulkarni ◽

Keyword(s):

Native Species ◽

Entomopathogenic Nematodes ◽

Insect Pests ◽

Entomopathogenic Nematode ◽

Effective Field ◽

Field Application ◽

Control Treatment ◽

Application Techniques ◽

Odontotermes Obesus ◽

Plantation Crops

Entomopathogenic nematodes (EPNs) in the genera Steinernema and Heterorhabditis and their associated bacteria such as Xenorhabdus spp. and Photorhabdus spp. are lethal parasites of soil dwelling and cryptic habit insects. Termites are one of the most severe soil insect pests of number of forestry, agricultural, households items and plantation crops in India and abroad. The efficacy of new native species of entomopathogenic nematode (TFRIEPN-15), Steinernema dharanaii against termites, Odontotermes obesus was tested under laboratory conditions. The termite, Odontotermes obesus was exposed to Infective Juveniles (IJs) from numbers 3 to 40 IJs termite-1. The dose- dependent mortality was noted of the termites. The lowest dose of 3IJs termites-1 caused 26.67% mortality which was significantly superior control treatment. The highest dose of 40IJs termites-1 caused 72.00% mortality was recorded. These experiments have proved that native isolated entomopathogenic nematodes can be used successfully to control termites and other soil insect pests with simple and effective field application techniques achieving 100% mortality of the treated insects.

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Keefektifan Patogenesitas Steirmema Carpocapsae (all strain) terhadap Hama Plutella xylostella L.

Jurnal Agrinika : Jurnal Agroteknologi dan Agribisnis ◽

10.30737/agrinika.v5i1.1555 ◽

2021 ◽

Vol 5 (1) ◽

pp. 51

Author(s):

Mariyono Mariyono ◽

Eko Yuliarsha Sidhi ◽

Nugraheni Hadiyanti

Keyword(s):

Plutella Xylostella ◽

Entomopathogenic Nematodes ◽

Insect Pests ◽

Entomopathogenic Nematode ◽

The Body ◽

Steinernema Carpocapsae ◽

Infective Juvenile ◽

Lc50 Value ◽

Steinernema Glaseri ◽

Significant Difference

The utilization of entomopathogenic nematodes is an example of the uses of an environmentally friendly biological agent. Entomopathogenic nematodes of the family Steinernematidae and Hetrorhabditidae are very potential to control insect pests. The purpose of this study was to study the pathogenicity of the entomopathogenic nematode Steinernema carpocapsae (all strains) as a biological control against Plutella xylostella. This research includes 2 stages, namely the preparation stage and laboratory experiments. The experimental design in this study was a completely randomized design (CRD) consisting of six treatments and three replications. The treatments tested were entomopathogenic nematode concentrations consisting of six levels: 0, 8, 16, 32, 64 and 128 infective juvenile per ml. The LC50 and LT50 values were calculated using Probit analysis. The results of observations of nematodes that enter the insect body and pest mortality were analyzed using analysis of variance (ANOVA), once showing a significant difference, it was then continued to the DMRT test at 5% level. Based on the results of the study, the highest pathogenicity against Plutella xylostella was Steinernema carpocapsae (all strains) when compared to Steinernema glaseri (NC) and Steinernema sp. local isolates. Resistance to Steinernema carpocapsae (all strains) and the LC50 value were determined by the age of Plutella xylostella larvae. The bigger and older the larvae, the more its resistance to Steinernema carpocapsae (All strains) and the LC50 value. The number of entomopathogenic nematodes that enter the body of Plutella xylostella increased with increasing length of contact time.Penggunaan nematoda entomopatogen merupakan salah satu pemanfaatan agensia hayati yang ramah lingkungan. Nematoda entomopatogen dari famili Steinernematidae dan Hetrorhabditidae sangat potensial untuk mengendalikan serangga hama. Tujuan dari penelitian ini adalah mempelajari patogenisitas nematoda entomopatogen Steinernema carpocapsae (all strain) sebagai pengendali hayati terhadap hama Plutella xylostella. Penelitian ini meliputi 2 tahap yaitu tahap persiapan dan percobaan laboratorium. Rancangan percobaan pada penelitian ini adalah Rancangan Acak Lengkap (RAL) yang terdiri enam perlakuan dan tiga ulangan. Perlakuan yang diujikan adalah konsentrasi nematoda entomopatogen yang terdiri enam taraf: 0, 8, 16, 32, 64 dan 128 infektif juvenile/ml. Nilai LC50 dan LT50 dihitung menggunakan analisis probit. Hasil pengamatan nematoda yang masuk dalam tubuh serangga dan mortalitas hama dianalisis menggunakan analisis sidik ragam (ANOVA), apabila menunjukkan beda nyata dilanjutkan uji DMRT taraf 5%. Berdasarkan hasil penelitian, patogenisitas tertinggi terhadap Plutella xylostella adalah Steinernema carpocapsae (all strain) bila dibandingkan dengan Steinernema glaseri (NC) dan Steinernema sp. isolat lokal. Ketahanan terhadap Steinernema carpocapsae (All strain) dan nilai LC50 ditentukan umur larva Plutella xylostella. Semakin besar dan tua umur larva, ketahanan terhadap Steinernema carpocapsae (All strain) dan nilai LC50 juga semakin meningkat. Jumlah nematoda entomopatogen yang masuk kedalam tubuh Plutella xylostella semakin banyak seiring dengan bertambah lamanya waktu kontak.

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Entomopathogenic nematodes: A tool in biocontrol of insect pests of vegetables-A review

Agricultural Reviews ◽

10.18805/ag.v38i02.7945 ◽

2017 ◽

Vol 38 (02) ◽

Cited By ~ 3

Author(s):

Gitanjali Devi ◽

Dhrubajyoti Nath

Keyword(s):

Biological Control ◽

Entomopathogenic Nematodes ◽

Insect Pests ◽

Entomopathogenic Nematode ◽

Insect Pest ◽

Nematode Species ◽

Vegetable Crops ◽

Biological Control Agents ◽

Isolation And Identification ◽

Identification Technique

Biological control agents have become increasingly important component in integrated pest management programme. Entomopathogenic nematodes are effective biological control agents for many important insect pests of vegetable crops. Therefore entomopathogenic nematodes are gaining attention in the field of biocontrol research worldwide. With the development and improvement of isolation and identification technique many novel species and strains have been utilized for management of several insect pests. This review aims to explore the potentiality of entomopathogenic nematode species against economically important insect pest of vegetables in India as well as in other countries.

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Characterization of symbiotic and associated bacteria from entomopathogenic nematode Heterorhabditis sp. (nematode: Heterorhabditidae) isolated from India

Egyptian Journal of Biological Pest Control ◽

10.1186/s41938-020-00343-9 ◽

2020 ◽

Vol 30 (1) ◽

Author(s):

Rashid Pervez ◽

Showkat Ahmad Lone ◽

Sasmita Pattnaik

Keyword(s):

Insect Pests ◽

Entomopathogenic Nematode ◽

Alcaligenes Faecalis ◽

Symbiotic Bacteria ◽

Rrna Gene ◽

Main Body ◽

Associated Bacteria ◽

Molecular Approaches

Abstract Background Entomopathogenic nematodes (EPNs) harboring symbiotic bacteria are one of the safest alternatives to the chemical insecticides for the control of various insect pests. Infective juveniles of EPNs locate a target insect, enter through the openings, and reach the hemocoel, where they release the symbiotic bacteria and the target gets killed by the virulence factors of the bacteria. Photorhabdus with Heterorhabditis spp. are well documented; little is known about the associated bacteria. Main body In this study, we explored the presence of symbiotic and associated bacteria from Heterorhabditis sp. (IISR-EPN 09) and characterized by phenotypic, biochemical, and molecular approaches. Six bacterial isolates, belonging to four different genera, were recovered and identified as follows: Photorhabdus luminescens, one each strain of Providencia vermicola, Pseudomonas entomophila, Alcaligenes aquatilis, and two strains of Alcaligenes faecalis based on the phenotypic, biochemical criteria and the sequencing of 16S rRNA gene. Conclusion P. luminescens is symbiotically associated with Heterorhabditis sp. (IISR-EPN 09), whereas P. vermicola, P. entomophila, A. aquatilis, and A. faecalis are the associated bacteria. Further studies are needed to determine the exact role of the bacterial associates with the Heterorhabditis sp.

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Virulence of entomopathogenic nematodes (Rhabditida: Steinernematidae and Heterorhabditidae) for the control of Diabrotica speciosa germar (coleoptera: chrysomelidae)

Ciência e Agrotecnologia ◽

10.1590/s1413-70542011000600015 ◽

2011 ◽

Vol 35 (6) ◽

pp. 1149-1156 ◽

Cited By ~ 6

Author(s):

Viviane Santos ◽

Alcides Moino Junior ◽

Vanessa Andaló ◽

Camila Costa Moreira ◽

Ricardo Alves de Olinda

Keyword(s):

Biological Control ◽

Entomopathogenic Nematodes ◽

Entomopathogenic Nematode ◽

Larval Mortality ◽

Steinernema Glaseri ◽

Soil Insects ◽

Diabrotica Speciosa ◽

Pupal Mortality

Entomopathogenic nematodes (EPNs) are used in biological control of soil insects and show promise in the control of D. speciosa. The objective of this work was to evaluate the potential of native and exotic entomopathogenic nematode isolates in the control of D. speciosa under laboratory and greenhouse conditions. Results showed that all of EPNs caused larval mortality. The most virulent were Heterorhabditis sp. RSC01 (94%), Steinernema glaseri (84%), Heterorhabditis sp. JPM04 (82%) and Heterorhabditis amazonensis RSC05 (78%). There was no effect of the Heterorhabditis sp. RSC01 and S. glaseri isolates on eggs. The maximum mortality of D. speciosa larvae by Heterorhabditis sp. RSC01 was observed at a concentration of 300 IJ/ insect, while by S. glaseri observed the highest mortality at the concentration of 200 IJ/ insect. The Heterorhabditis sp. RSC01 isolate caused over 80% pupal mortality at a concentration of 250 IJ/insect. The virulence of Heterorhabditis sp. RSC01 and S. glaseri was affected by temperature. The Heterorhabditis sp. RSC01 isolate caused reduction in larva survival under greenhouse conditions at all of the tested concentrations and there was no difference in mortality among different concentrations of infectid juveniles.

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First Glimpse of Gut Microbiota of Quarantine Insects in China

10.1101/2020.12.10.420612 ◽

2020 ◽

Author(s):

Yanxue Yu ◽

Qi Wang ◽

Ping Zhou ◽

Na Lv ◽

Wei Li ◽

...

Keyword(s):

Environmental Factors ◽

Gut Microbiota ◽

Geographical Origin ◽

Geographic Origin ◽

Insect Species ◽

Monitoring Method ◽

Geographical Source ◽

Insect Gut ◽

Tracing Method ◽

New Habitats

AbstractQuarantine insects are economically important pests that frequently invade new habitats. A rapid and accurate monitoring method to trace the geographical source of invaders is therefore needed for prevention, detection, and eradication. Current methods based on insect genetics are often too slow. We developed a novel tracing method based on insect gut microbiota. The source location of microbiota of insects can be used to rapidly determine the insects’ geographic origin. We analyzed 179 gut microbiota samples belonging to 591 individuals of 22 quarantine insect species collected from 36 regions in China and abroad. The gut microbiotas of these insects mainly included Actinobacteria, Bacteroidetes, Cyanobacteria, Firmicutes, Proteobacteria, and Tenericutes. The diversity of the insect gut microbiota was closely related to geographic and environmental factors. Different insect species could be distinguished at the phylum level of microbiota. Populations of individual insect species from different regions could be distinguished at the genus level of microbiota. A method for determining the geographical origin of invasive insect species was tentatively established, but its practical applicability requires further study.

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