Contents lists available at ScienceDirect Journal of Invertebrate Pathology journal homepage: www.elsevier.com/locate/jip Evaluation of different sponge types on the survival and infectivity of stored entomopathogenic nematodes Mustapha Touray a , Baris Gulcu b , Derya Ulug a , Sebnem H. Gulsen a , Harun Cimen a , Harry K. Kaya c , Ibrahim Cakmak d , Selcuk Hazir a, ⁎ a Department of Biology, Faculty of Arts and Science, Aydin Adnan Menderes University, Turkey b Department of Biology, Faculty of Arts and Science, Duzce University, Turkey c Department of Entomology and Nematology, University of California, One Shields Ave., Davis, CA, United States d Department of Plant Protection, Faculty of Agriculture, Aydin Adnan Menderes University, Turkey ARTICLE INFO Keywords: Entomopathogenic nematodes Heterorhabditis Steinernema Formulation Biological control Storage ABSTRACT Sponges are one of the cheapest and most suitable substrates used to formulate and/or store the infective ju- veniles (IJs) of entomopathogenic nematodes (EPNs). Our study investigated the survival and infectivity of the IJs on five different sponges compared to that in an aqueous suspension (control). The sponges were Oasis® floral, Nanosponge, Scotchbrite TM , or Lysol® and natural sea sponge. EPN species tested were Heterorhabditis bacteriophora, Steinernema carpocapsae and S. feltiae. The recovery efficiency of the IJs from sponges was initially assessed. Subsequently, IJs were stored in the sponges and placed in plastic bags or Falcon tubes and incubated at 10° or 27 °C for 8 months or 11 weeks, respectively. IJ survival and infectivity were monitored monthly for the 10 °C and weekly for 27 °C in these sponge types. The IJs were recovered from the sponges, and their survival was based on observing their movement under a dissecting microscope, and infectivity was based on larval mortality in Galleria mellonella. Recovery efficiency of IJs was best for the Oasis floral sponge for all nematode species ranging between 83 and 91%. The survival and infectivity of stored IJs in all sponge types and control for both 10° and 27 °C gradually decreased over time. IJs stored in Scotchbrite, Lysol, and Nanosponge had the best survival and infectivity, whereas Oasis floral and natural sea sponges showed the poorest results. After 8 months at 10 °C in plastic bags, the survival ratio of all IJs in these three sponges (Scotchbrite, Lysol, and Nanosponge) was approximately 55%. IJs in Scotchbrite and Nanosponge were also able to survive and retain their infectivity at 27 °C for 3 months. IJs stored in Falcon tubes had survival that ranged from 26 to 53% at 27 °C and 55 to 77% at 10 °C. H. bacteriophora IJs lost their infectivity when stored at 27 °C after 10 weeks. However, S. carpocapsae and S. feltiae exhibited 85% infectivity when stored in Scotchbrite and 50% in Nanosponge, respectively. Overall, sponges made from polyurethane (Scotchbrite) followed by melamine (Nanosponge) and cellulose (Lysol) are recommended for long-term nematode storage and transportation of nematode samples. However, Oasis floral sponge may be preferred for short-term IJ formulation for field applications because of easier recovery of IJs. 1. Introduction The genera, Steinernema (Family: Steinernematidae) and Heterorhabditis (Family: Heterorhabditidae), are soil-dwelling nema- todes with significant biological applications in pest management, especially of soil insects and those that occur in cryptic habitats (Hazir et al., 2003; Gulcu et al., 2017). These nematodes are mutualistically associated with bacteria in the genera Xenorhabdus (Steinernema) and Photorhabdus (Heterorhabditis). Entomopathogenic nematodes (EPNs) have a ubiquitous presence in soils around the world, and a variety of attributes as biocontrol agents including a durable non-feeding, infective juvenile (IJ) stage, a broad host range against many important soil insect pests, the ability to seek out hosts in soil or in cryptic ha- bitats, safety to non-target organisms and amenability to mass pro- duction and formulation. These qualities are some of the reasons for their growing use as commercialized biocontrol agents. The develop- ment of commercialized EPNs is mainly due to growing concerns raised by insect resistance to and unwanted side effects of chemical pesticides on beneficial organisms, the environment, and human health (Gaugler and Kaya, 1990; Ehlers, 2002; Shapiro-Ilan et al., 2012, 2014). Mass production and formulation are key issues to EPN commer- cialization and improving these factors will enhance the efficacy of the https://doi.org/10.1016/j.jip.2020.107332 Received 29 November 2019; Received in revised form 13 January 2020; Accepted 19 January 2020 ⁎ Corresponding author. E-mail address: selcuk.hazir@gmail.com (S. Hazir). Journal of Invertebrate Pathology 171 (2020) 107332 Available online 03 February 2020 0022-2011/ © 2020 Elsevier Inc. All rights reserved. T