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Laboratory Report About Isolation Of Entomopathogenic Nematodes (Lab Report Sample)


Laboratory report about Isolation of Entomopathogenic Nematodes

Exercise IX: Isolation of Entomopathogenic Nematodes
Telan, Jose Angel Jude B 5/28/18 ENT 275
-17145209423000Entomopathogenic nematodes or EPNs are soil-living organisms and are usually attributed to the families Heterorhabditidae and Steinernamatidae. Members of these families usually harbor symbiotic bacteria specific to each family which causes lethality to the potential hosts of these entomopathogenic nematodes. In this experiment, isolation process of these EPNs will be conducted along with the observation of its infection process on insect larvae. Baiting setups containing soil mixed with known EPN species were prepared and provided to the students. Exposure of the insect larvae were done by placing the larvae on the baiting setups. The cadavers were isolated in white traps after achieving mortality in the baiting setup. Observations on the pathology is mainly on the color of the cadavers since Heterorhabdus-infected larvae shows reddish to brown color while Steirnema-infected larvae have cadavers with dark color. IJs emerged and were isolated on the white traps. These methods allowed the students to be familiarized with the EPN isolation and infection process.
Nematodes are worm-like organisms that are widely available in the soil. Entomopathogenic nematodes or EPNs are represented by the families Steinernematidae and Heterorhabditidae which are related to their respective symbiotic bacteria Xenorhabdus spp. and Photorhabdus spp. respectively. These symbiotic bacteria are the main cause of the lethality of these entomopathogenic nematodes. Once the juveniles of these EPNs infect and penetrate potential hosts, the bacteria is released inside the host and able to cause septicemia in the insect. These juveniles are labeled as IJs or infective juveniles and have potential in biological control applications especially against specific insect targets. (Atwa, 2011; Kaya & Gaugler, 1993) In this experiment, students will be familiarized with the process of nematode isolation from the soil and recognize basic characterization of nematode infection in insects.
Materials and methods
Baiting setups were initially prepared for this experiment. Soils treated with entomopathogenic nematodes (EPNs) were mixed with soil packed in the baiting setup composed of circular microwaveable containers with the soil inside. Two known genera of EPNs were used for the setups, thus six of these baiting setups were prepared and labeled MBLB and BSDS, with three different larvae (Asiatic Corn Borer, Lesser Waxmoth, and mealworm larvae) subjected on each genus of EPN. Changes were observed after 48 hours post infection and once mortality was achieved, the insect cadavers were transferred on a white trap. A white trap could be described as a 35mm petri plate lined with filter paper inside a larger petri dish with distilled water enough just to fill the surface. Observations were further noted since IJs were to be observed after white trapping.
Results and Discussion
Some cadavers were immediately isolated after 24 hrs while the remaining insect cadavers which achieved mortality later was isolated and transferred on individual white traps after 24hrs. The larvae that achieved later deaths were precisely the mealworms. The MBLB labeled traps showed infected cadavers that were beginning to darken in color after 48hrs (figure 1) while the BSDS labeled specimens showed that somewhat brown to reddish cadaver color (Hu & Webster, 2000). Seven days after isolation, IJs were observed on the white traps. Also, some MBLB labeled specimens appeared to have fungal growth on the cadavers which could indicate contamination of fungus in the setups. BSDS labeled cadavers could be Heterorhabditis spp. of nematodes while MBLB labeled cadavers could be Steinernema spp. because of the gross pathologies observed during the course of nematode infection.
4829175153035Figure 1. Heterorhabditis spp. larvae 4 days PI. Left: ACB, Right: LWM.00Figure 1. Heterorhabditis spp. larvae 4...
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