Natural Occurrence, Distribution and Isolation of Native Strains of Entomopathogenic Nematodes (Steinernematidae and Heterorhabditidae) in Vegetable Crops in Haryana, India

Author: Deepak Kumar*, Anil Kumar, Prakash Banakar and Vinod Kumar

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Abstract

Entomopathogenic nematodes (EPNs), Steinernematidae and Heterorhabditidae have a potential bio-control agents since these organisms have a wide range of geographic areas, soil types and are adapted to several hosts. They showed better performances over chemical and microbial insecticides. But for the better performances of EPNs, should be adapted to the local environmental conditions. The search for local EPN isolates though systematic survey is the first critical step in building an effective biological management program for insect pests. The populations of EPNs were isolated by ‘Galleria trap’ method from the soils around the roots of various crops in Haryana (Hisar, Jind, Panipat, Sonipat, Karnal, Mewat, Palwal and Fatehabad districts). A total of 160 soil samples were collected from the rhizosphere of different crops and were processed for the detection of EPNs by insect bait method, 40 samples (25%) were found to be positive for EPNs. Based on samples collected from different vegetable crops, the frequency of EPNs observed were chilli (40.0%), cucurbitaceae (31.1%), capsicum (33.3%) and tomato (27.6%). Out of 40 samples found positive for EPNs collected from various vegetable crops, the frequency of occurrence of Steinernematid and Heterorhabditid nematodes were 80 and 20 percent, respectively. Identification of EPNs was done at generic level (Heterorhabditis and Steinernema) based upon the colour of host cadaver. Galleria mellonella larvae infected with EPN of the genus Heterorhabditis may impart reddish-orange colouring while those infected with Steinernema, imparts grey-brown colouring. So, the bio-efficacy of EPNs proved better of native strains which already compatible to the same environment.

Keywords

Distribution, Frequency, Isolation, Heterorhabditidae, Steinernematidae

Conclusion

The safest biocontrol agents, and a potential alternative to insecticides, are entomopathogenic nematodes. EPNs can be utilised on a number of crops because of their broad host range. However, only 40 of the 160 soil samples taken during the survey from various vegetable crops in Haryana detected entomopathogenic nematodes. The survival of Steinernema species and Heterorhabditis species emphasises the significance of carrying out more thorough surveys in Haryana.

References

INTRODUCTION Commercial production of entomopathogenic nematodes (EPNs) from the families Steinernematidae and Heterorhabditidae is used to manage insect pests biologically, especially for those that live in soil and cryptic environments. From various nations throughout the world, 21 species of Heterorhabditis and about 100 genuine species of Steinernema have been identified. But till now, only fifteen species of EPNs (12 for Steinernema and 3 for Heterorhabditis species) are recorded from different parts of India (Kumar et al., 2022). Identification of naturally adapted species in a particular location is crucial for EPN to be a successful biological pesticide (Stock et al., 1999). Surveys showed that, with the exception of Antarctica, these nematodes are present throughout the earth (Campos-Herrera et al., 2012). As a result of many surveys, numerous new species have been described, as well as numerous new isolates of previously described species. Entomopathogenic nematode research on Steinernema carpocapsae, S. glaseri, S. feltiae, and Heterorhabditis bacteriophora was started in India utilizing alien species and strains. However, the nematodes' weak capacity to adapt in a strange environment is what caused the results to be so uneven (Kaya et al., 2006). Since then, surveys have been concentrated on identifying native species, leading to the recovery of a number of species and strains (Lalramliana and Yadav, 2010; Hussaini et al., 2001; Kulkarni et al., 2012; Bhat et al., 2017). In the genera Xenorhabdus for steinernematids and Photorhabdus for heterorhabditids, these nematodes are linked to symbiotic bacteria. Once it has located an appropriate host, the infectious juvenile stage of the nematode enters it through a natural entrance and moves into the hemocoel. Then the associated bacteria quickly grow in insect hemocoel and cause septicemia, which kills the host within 24 to 72 hours. The nematodes start the development process, then eat the bacterial cells and tissues of the host until there are no more food supplies left in the host carcass. Finally, they appear in the soil as a fresh batch of infectious juveniles (IJs) looking for fresh hosts (Hazir et al., 2003). After two or three generations in the cadaver, the nematodes feed on the bacterial cells and host tissues before emerging as infectious juveniles into the soil environment. When they infect a new insect host, the infectious juveniles start the life cycle all over again (Kaya et al., 2006). The main objective of this work is to isolate native EPN from the district of Haryana, India, a biodiversity hotspot location, in addition to analyzing the occurrence and their potential use as a biological control agent moving forward. MATERIALS AND METHODS District surveyed: Hisar, Jind, Panipat, Sonipat, Karnal, Mewat, Palwal and Fatehabad districts of Haryana were surveyed for EPNs (Fig. 1). Soil sampling: From 2019 to 2022, soil samples were taken at random from a diversity of vegetable-growing regions in the districts of Haryana, India. A total of 160 samples, spanning around 8 districts, were taken from different sampling sites (Plate 1). Using a hand shovel, 5 subsamples from each site were obtained from a 20–25 cm depth and combined to obtain roughly 1 kg of composite samples (Orozco et al., 2014). The appropriate elevations and geographic coordinates were noted. For further processing, all soil samples were collected in polyethylene bags to avoid dehydration, proper tagged, sealed, and placed in boxes before being transported to the laboratory, Department of Nematology, College of Agriculture, CCS Haryana Agricultural University, Hisar. Culture of insects (Galleria mellonella): Entomopathogenic nematodes were isolated on greater wax moth, G. mellonella (Woodring and Kaya 1988). In a lab conditions, the larger wax moth culture was kept alive on an artificial diet. Insect-baiting and nematode culture: According to Bedding and Akhurst's original report (1975), insect-baiting was undertaken. Each glass bottles containing soil samples received ten G. mellonella larvae in their last instar, which were subsequently inserted and incubated at 25+2°C. Regular inspections were conducted on the samples to assess for successful insect infection and soil moisture. Until infective juveniles (IJs) emerged, all collected cadavers were cleaned with distilled water and placed in a White trap (White, 1927). Next, IJs were collected, cleaned, and kept in storage at 16+2°C. Re-inoculation of G. mellonella larvae was performed three times in order to get pure culture of the nematodes retrieved from the soil (Hoy et al., 2008). RESULTS AND DISCUSSION The main objective of the current investigation was to identify native EPN species with excellent strains so that they may be used for successful bio-management of various insect pests in different parts of the state of Haryana. Eight districts were intended to obtain a total of 160 soil samples that represented various soil types and irrigation systems. These samples were then processed to the soil baiting technique with G. mellonella, a widely preferred host for isolating EPNs from soil. Out of 160 soil samples, 40 were found to have G. mellonella infestations with EPNs, according to the data. Out of 40 positive samples for EPNs, the eight soil samples taken from vegetable fields contained Heterorhabditis spp. which were identified based on the colour of the afflicted corpses, that turned brick red. Remaining 25 soil samples with positive EPNs results were determined to include Steinernema spp. based on the cadaver Galleria's grayish-white colour (Plate 3). According to data on the prevalence of EPNs acquired from eight districts in the state of Haryana (Table 1), the highest frequency of EPNs was found in Panipat (40.9%) followed by Hisar (37.5%) and Sonipat district (36.4%), while Fatehabad district had the lowest incidence of EPNs (20%). Mewat and Palwal districts were found to have no EPN prevalence. Following Jind, whose soil samples tested positive for EPNs in 30.4 percent of cases, was Karnal district, which had a 19.0 percent EPN presence. Six (Hisar, Jind, Panipat, Sonipat, Karnal, and Fatehabad) of the eight districts in Haryana state that were surveyed for the presence of EPNs were found to be positive for Steinernema spp., while five (Hisar, Jind, Panipat, Sonipat, and Fatehabad) were found to be positive for Heterorhabditis spp. Out of 160 soil samples, 45 came from cucurbitaceae crops, including 29 tomato, 23 brinjal, 18 okra, 11 potato, 7 carrot, 5 capsicum, 5 chilli, 5 onion, 4 cruciferous, 4 coriander and 3 garlic crop. Based on samples collected from different vegetable crops, the frequency of occurrence of EPNs were observed maximum in chilli (40.0%) followed by capsicum (33.3%) cucurbitaceae (31.1%), tomato (27.6%), brinjal (26.1%), cruciferous (25.0%), onion (20.0%), potato (18.2%), okra (16.7%) and carrot (14.3). Coriander and garlic crops were found with none EPNs frequency. Out of 40 samples found positive for EPNs collected from various vegetable crops, the frequency of occurrence of Steinernematid and Heterorhabditid nematodes were 80 and 20 percent, respectively (Table 2). The aforementioned findings are consistent with those made by Uribe-Lorio et al. (2005). According to their findings, 20.50 percent of all soil samples tested positive for the presence of EPNs. In contrast, Barbosa-Negrisoli et al. (2010) and Myers et al. (2015) found entomopathogenic nematodes in 15.7 percent and 21 percent, respectively, of the soil samples. The conclusions of Hussaini et al. (2000) are likewise supported by the current findings. They noted that Andhra Pradesh has a diverse range of EPN species. The distribution and incidence of EPNs were reported by Sunanda et al. (2016) in 1.38 percent of all soil samples taken from the state of Telangana. Lalramliana and Yadav (2010), Singh et al. (2015), and Josephrajkumar and Sivakumar (1997) have also reported the prevalence and distribution of EPNs from various sources. Gowda et al., 2020 also recovered 3 soil samples containing EPNs out of 130 samples during a survey of Uttar Pradesh, India from 2016-2017 with frequency of 2.3 percent. Out of 200 soil samples collected from 40 soil sites in Thialand and found the prevalence of EPNs was 8.0 percent (Ardpairin et al., 2020). Out of 313 soil samples collected from different districts of Haryana during 2018-2021, 99 samples (31.6%) were found to be positive for the EPNs. Maximum frequencies of occurrence of EPNs was found in ber orchards (Steinernematid 65.6% and Heterorhabditid 34.4%), followed by sugarcane, wheat and cluster been, as only 71.1, 37.5, 35.7 and 35.3 percent, respectively (Kumar et al., 2021).

How to cite this article

How to cite this article: Deepak Kumar, Anil Kumar, Prakash Banakar and Vinod Kumar (2022). Natural Occurrence, Distribution and Isolation of Native Strains of Entomopathogenic Nematodes (Steinernematidae and Heterorhabditidae) in Vegetable Crops in Haryana, India. Biological Forum – An International Journal, 14(2a): 354-359.