HORMONAL PRODUCTS IN COCKROACH EMBRYOS; HOST SPECIFICITY OF ENTOMOPATHOGENIC NEMATODES AND THE EFFECT OF SURFACE COAT PROTEINS FROM NEMATODES ON INSECT IMMUNITY

I Juvenile hormone (JH), produced by the corpora allata (CA), regulates molting and reproduction in many insects, including cockroaches. It is known, however, that JH is produced only after dorsal closure, a conspicuous event in embryogenesis. Dorsal closure is an important physiological change in embryos. I found that the ratio of dorsal closure to embryo development time was consistent (about 45% of total embryo development) across most cockroach species. This conservation was linked to reproductive biology of the cockroaches. The only viviparous cockroach, Diploptera punctata, completed dorsal closure at 20.8 % of embryo development time. Blattella germanica, whose reproductive mode is different from other oviparity cockroaches, finished its dorsal closure at 38.5 % of embryo development time. Other oviparous and ovoviviparous cockroaches completed dorsal closure at similar percentages of the embryo development time. It is reported that embryonic CA produce both JH and its immediate precursor methyl farnesoate (MF) in Nauphoeta cinerea. Using a radiochemical assay, the present iv research found that cockroach embryos produced and released both JH and MF across all three reproductive modes. These include Periplaneta americana, Eurycotis floridana, Blaberus discoidalis, Byrsotria fumigata, Rhyparobia maderae, Nauphoeta cinerea, and Diploptera punctata. I also found that the control of conversion of MF into JH by epoxidase, the last step of biosynthesis of JH, is species dependent. These results suggest that the conversion of MF into JH is a rate-limiting step and was species specific. Abstract II Entomopathogenic nematodes (EPNs) are good candidates for biological-control agents for soil-dwelling insects. Infective juveniles (IJ) of EPNs enter insect hosts and release symbiotic bacteria that kill the hosts. Insects defend against EPNs by a rapid cellular immune response that includes encapsulation and melanization, which kills EPNs. EPNs have to overcome insects’ innate immunity to survive and reproduce. This study was designed to understand host immune responses to two species of nematodes, Heterorhabditis bacteriophora and Steinernema glaseri, and the relationship of immunity to host specificity. I hypothesized that EPNs induce or suppress the immuneII Entomopathogenic nematodes (EPNs) are good candidates for biological-control agents for soil-dwelling insects. Infective juveniles (IJ) of EPNs enter insect hosts and release symbiotic bacteria that kill the hosts. Insects defend against EPNs by a rapid cellular immune response that includes encapsulation and melanization, which kills EPNs. EPNs have to overcome insects’ innate immunity to survive and reproduce. This study was designed to understand host immune responses to two species of nematodes, Heterorhabditis bacteriophora and Steinernema glaseri, and the relationship of immunity to host specificity. I hypothesized that EPNs induce or suppress the immune responses in their host based on their surface coat proteins (SCPs). The insect hosts I tested were immature stages of wax worm Galleria mellonella, oriental beetle larvae Exomala orientalis, Japanese beetle larvae Popillia japonica, tobacco horn worm larvae Manduca sexta, northern masked chafer larvae Cyclocephala borealis, and adult house cricket Acheta domesticus. I found that H. bacteriophora and S. glaseri infected and reproduced in their susceptible hosts well. In these hosts, EPNs were melanized and encapsulated at low v percentages and a high percentage of EPNs were free-moving. In the resistant hosts, most EPNs were melanized and encapsulated and few EPNs were free moving. S. glaseri NC strain was more successful compared to the S. glaseri FL strain in the same hosts. These results suggest the nematodes elicited immune responses in hosts that correlated with their infectivity. I also found that hemocytes from M. sexta, a susceptible host, recognized S. glaseri at a low percentage during the first hour post nematode introduction. After 24 hours, H. bacteriophora escaped recognition of hemocytes from G. mellonella, a susceptible host. I demonstrated that different species and strains of EPNs had different SCPs. I isolated and characterized the SCPs from S. glaseri NC strain. These SCPs suppressed immune responses in the oriental beetle larva, a susceptible host for S. glaseri, thus protecting H. bacteriophora from being killed in the same host, as it normally would be. Immuno-suppression was dose-dependent. Also, multiple injections of the SCPs protected H. bacteriophora better in Oriental beetle larvae. In a nondenatured state, two isolated SCPs in the SCPs of S. glaseri each conveyed this immuno-suppressive effect. The two SCPs were composed of smaller proteins when separated on two dimensional PAGE. Hemocytes of oriental beetle larvae started degrading after exposure to the proteins for 3 hours. Some of the SCPs from S. glaseri NC strain were sequenced and one of them was enolase, which is also secreted by other parasites. vi TABLE OF CONTENTS List of figures . ix List of tables . xi Acknowledgements . xii Chapter 1: Overview of hormonal products in cockroach embryos . 1 1.