Cellulosic ethanol holds great potential as biofuel due to its sustainability and renewability, yet recalcitrance of cellulosic feedstocks prevents cost-efficient ethanol production. Enzymatic catalysis of lignocellulosic biomass has the greatest biotechnological potential for cost reductions to the production process. Even though numerous cellulolytic enzymes have been identified in bacteria, plant, and fungi, insects remain as a fairly unexplored prospecting resource. Many insects, either via endogenously or symbiotically derived enzymes, use cellulose as substrate for their energetic needs. Novel cellulases from insects may have the potential to be more efficient than alternative enzymes in the conversion of cellulose to fermentable sugars due to their optimized activity in the highly reducing and extreme pH conditions found in some insect digestive systems. In this work we present data characterizing cellulolytic activity in the grasshopper Dissosteira carolina L. (Orthoptera: Acrididae) and the red flour beetle, Tribolium castaneum Herbst (Coleoptera: Tenebrionidae). After a screening for cellulolytic activity in insect populations from the East Tennessee region, D. carolina was selected due to relatively high cellulolytic activity compared to documented effective insect cellulolytic species. Cellulolytic activity in digestive fluids from gut and head from juvenile and adult stages of D. carolina was measured and an active cellulolytic protein profile demonstrated comparable activities amongst life stages. Partial protein sequences that match those identified from insect and microbial cellulases were obtained from purified 43-kDa and 45-kDa cellulases from D. carolina head digestive fluids. Although unsuccessful, attempts were made to purify and clone these enzymes for recombinant expression. Our research on D. carolina is the first report on the purification of endoglucanase activity in a grasshopper species. Availability of the T. castaneum genome allowed for homology searches using reported insect cellulases to identify a predicted cellulase. We cloned the full-length cDNA for this enzyme and named it TcEG1 (for T. castaneum endo-glucanase-1). TcEG1 was heterologously expressed in bacterial and insect cell culture systems and its activity against cellulose substrates and thermostability measured. Cloning of a cellulase gene from T. castaneum adds to the collection of reported insect cellulases and demonstrates the advantage of using genomic resources for protein discovery.

Hemlock woolly adelgid, Adelges tsugae Annand, is an exotic insect species dramatically reducing populations of eastern hemlock, Tsuga canadensis (L.) Carrieré, throughout the eastern United States. Systemic imidacloprid and horticultural oil are the two primary chemicals used to control infestations of the hemlock woolly adelgid. However, the effect of application timing (fall versus spring) and method on the translocation of imidacloprid throughout the canopy in addition to the quantity of imidacloprid translocated is unknown. Also, the potential effect of both imidacloprid and horticultural oil on non-target canopy insects is unknown. A study was initiated to determine the effect of application timing (fall versus spring) for three imidacloprid application methods (soil drench, soil injection, and tree injection) on the translocation of imidacloprid and concentration levels accumulated in eastern hemlock sap and twig and needle samples, assess the effect of these treatments and horticultural oil on the overall species richness and abundance, guild species richness and abundance, and specific species of non-target phytophagous and transient canopy insects. Eastern hemlocks (n = 30) were selected at Indian Boundary in Cherokee National Forest located in southeast Tennessee on 5 November 2005. This test was arranged in a split plot 2 x 5 factorial complete randomized block design with three replications. Three blocks were established. Each block contained ten trees, arranged in five tree pairs, with one tree in the pair treated in the fall (29-30 November 2005) and the other during the spring (16 April 2006). Five treatments were made; horticultural oil, imidacloprid soil drench, imidacloprid soil injection, imidacloprid tree injection, and the control (no treatment). Enzyme-linked immunosorbant assays were used to determine imidacloprid concentration in sap and combined twig and needle concentrations collected from hemlock branches at three strata (bottom, middle, and top) of the hemlock canopy collected every three months post-treatment. To determine effect on phytophagous and transient canopy insects, monthly sampling consisting of malaise traps, beat-sheets, direct observation/trunk vacuuming/handpicking, and branch pruning was conducted from 16 March 2006 - 18 April 2007. Concentration levels progressively decline from the bottom strata to the top strata of the canopy. This trend was consistent in all chemically treated trees. Tree injections provided the lowest concentration and the most non-uniform distribution of imidacloprid throughout the canopy. Soil drench consistently provided the highest insecticide concentration within the tree across all strata. Species richness and abundance were significantly effected by one or more application methods when compared to the control trees; however, the timing of the applications (fall versus spring) had no significant effect on the insect species. The detritivore and phytophaga guilds were effected by one or more chemical applications. Species richness was significantly lower across all guilds and differed significantly from those species on the control trees. Some 35 insect species were found to be directly effected by these chemical treatments. Of the 35 species, 27 feed directly on eastern hemlock, and as such, ingest the chemical. Eight of the species were psocopterans that feed on decaying organic material (detritivore). The soil drench had the greatest effect on species richness and abundance and guild richness and abundance among non-target phytophagous and transient canopy insects, followed by soil injection, while horticultural oil and tree injections had minimal effect. This data provides more flexibility in the timing and method of application used to have a minimal effect on non-target phytophagous and transient canopy insects.

The hemlock woolly adelgid (HWA), Adelges tsugae Annand, is an invasive pest that is causing declines in populations of eastern hemlock, Tsuga canadensis Carriere, in eastern North American forests. The threat of losing the hemlock as a foundation species in eastern forests prompted reserve managers to devise and implement HWA management strategies integrating cultural, biological, and chemical control tactics. Chemical control methods, systemic imidacloprid applications and horticultural oil foliar sprays, provide the most immediate and effective control of HWA in accessible hemlocks. Non-target impacts of HWA chemical control methods on soil arthropod communities are undocumented. Empirical studies in the field and in the laboratory were performed to determine the extent of effects of the common HWA chemical control treatments to non-target soil arthropods. Treatments were the horticultural oil foliar spray (no imidacloprid), imidacloprid trunk injection, imidacloprid soil injection, imidacloprid soil drench, and untreated controls. Microarthropods in soil drench plots displayed marginally non-significant decreases in abundance and richness. Microarthropod species composition was distinct in all of the imidacloprid treatments when compared to controls. Acari, the mites, consisted of approximately 50% of the observed abundance, and showed no responses to imidacloprid or horticultural oil treatments. Abundance and richness of Collembola, in contrast, were markedly decreased by the soil drench treatments. High performance liquid chromatography (HPLC) was used to quantify concentrations of imidacloprid from soils following imidacloprid treatments. Concentrations of imidacloprid observed in soils from imidacloprid treatment plots exceeded the LD50 and ED50 concentrations for Folsomia candida Willem (Collembola: Isotomidae) observed in the laboratory, especially in the soil drench plots, less frequently so in the soil injection plots and in a few of the trunk injection plots. The springtail Folsomia candida were reared in the laboratory on standard soil substrates containing a series of known imidacloprid concentrations to observe impacts to reproduction and survival. The imidacloprid concentration at which Folsomia candida adults displayed 50% mortality in the laboratory, as inferred from regression analysis of observed dose responses (LD50), was 1.38 mg imidacloprid / kg dry soil. The concentration at which F. candida produced half the number of juveniles observed in control microcosms (ED50) was 0.598 mg imidacloprid / kg dry soil. vi

A molecular phylogenetic investigation was undertaken to identify species within the morphologically homogeneous Simulium jenningsi species group, a pestiferous group of 22 species of black flies restricted to the Nearctic region. Several species in this group have well documented medical and veterinary importance, most notably S. luggeri and S. jenningsi. Unfortunately, females are monomorphic, a conundrum given their pest status. The objective of this study was to examine the utility of molecular data in species identification, with obvious application to identification of pest females. Towards this end, we sequenced approximately 2 kilobases of sequence data from the mitochondrial (Cox I + proximal one-half of Cox II) and nuclear (big zinc finger 2) genomes from positively identified exemplars (pupae, some larvae) and analyzed them phylogenetically using parsimony and Bayesian criteria. Combined analyses were not conducted due to extreme incongruence between the data sets. Mitochondrial and nuclear data sets sufficient for ready identification of approximately one-third and one-half, respectively, of the known species. Both genes recovered S. aranti, S. luggeri, S. ozarkense, S. penobscotense, and portions of the S. fibrinflatum complex (S. fibrinflatum, S. underhilli, S. notiale, and S. snowi). Species positively identified by analyses of independent data sets include S. taxodium and S. chlorum with the mitochondrial data and S. haysi, S. krebsorum, S. dixiense, S. definitum, S. remissum, S. infenestrum, S. podostemi, and S. jenningsi with big zinc finger 2. Future studies to better resolve species identities in this group should focus on additional nuclearly encoded markers or perhaps amplified fragment length polymorphism (AFLP) approaches. However, we suspect that introgression, lineage sorting, and differential sorting of ancestral polymorphisms occurred in various lineages within this group and may make complete phylogenetic reconstruction of all species lineages impossible.

Efficacy of <i>Bt</i> corn hybrids expressing an insecticidal delta-endotoxin through different transformation events was evaluated in Louisiana. Efficacy levels were determined for southwestern corn borer (SWCB), <i>Diatraea grandiosella</i> Dyar; sugarcane borer (SCB), <i>Diatraea saccharalis</i> (F.); corn earworm (CEW), <i>Helicoverpa zea</i> (Boddie); and fall armyworm (FAW), <i>Spodoptera frugiperda</i> (J. E. Smith). Insect (neonates and third instars) susceptibility was evaluated on <i>Bt</i> and non-<i>Bt</i> corn plants (V6 and R1 stages) in field plots and plant tissue in laboratory studies. <i>Bt</i> corn hybrids with MON810 and CBH351 transformation events sustained significantly (<I>P</I> < 0.05) less injury caused by SWCB, SCB, and FAW during infestations at mid-whorl stage. SWCB, SCB, and CEW infestations during silking stage were significantly reduced in <i>Bt</i> hybrids with the MON810 event. However, CEW infestations at silking stage were not significantly (<I>P</I> > 0.05) reduced in the <i>Bt</i> hybrid with the CBH351 event compared to respective non-<i>Bt</i> equivalent hybrid. Performance of BT11 and 176 events against CEW infestations at silking stage was not consistent under observed field conditions. SWCB and SCB were highly susceptible to the MON810 event in leaf tissue experiments. However, consistency in performance of the MON810 event against SWCB and SCB varied among <i>Bt</i> hybrids containing that event. SCB expressed less susceptibility to leaf tissue containing the CBH351 event than to leaf tissue with the MON810 event. Plant resistance to SWCB and SCB attack increased as plants matured, independent of the presence of a <i>Bt</i> construct. Higher CEW survival was observed on <i>Bt</i> silks than on <i>Bt</i> leaves regardless of <i>Bt</i> event. Economic returns above technology fee were consistently observed in central and most areas of northeast Louisiana. Economic returns in northwest Louisiana were highly dependent on corn market prices. Under growing conditions and investment levels observed, soybeans and in some cases grain sorghum production might offer higher economic benefits compared to <i>Bt</i> corn production for late-crop planting in some areas of Louisiana.

Coptotermes formosanus was used as bait to isolate pathogenic fungi from soil. Fifteen pathogenic fungal isolates were obtained, all either Metarhizium anisopliae (six isolates) or Beauveria bassiana (nine isolates). There were no differences in numbers of fungal isolates from the three sampling locations. However, significantly more isolates were found in woodlands (eleven) than in pastures (four). Median lethal doses (LD50s) of these fungal species to C. formosanus were interspersed, indicating that fungal isolates rather than species had the greatest effect on virulence. In vitro growth characteristics were significantly correlated with virulence against termites, suggesting that fungal virulence might be predicted in vitro rather than by bioassay. Conidial production on termite cadavers increased significantly over 11 days post-death. Effects of isolates of M. anisopliae and B. bassiana on in vivo sporulation were significant. B. bassiana isolates could be categorized into a group with high total sporulation (day 11) and low quick sporulation (on days 2 and 3), while M. anisopliae isolates fell into another group with high quick sporulation and low total sporulation. Conidial production was significantly higher in vitro than in vivo. Correlation between in vivo and in vitro conidial production was positive and significant. This may allow preliminary in vitro screening of a large number of isolates for high in vivo sporulation. Fungal species and isolate significantly affected disease prevalence in termite populations. Sporulation of M. anisopliae played a more important role than virulence in producing epizootics in termites, but this was not the case for B. bassiana. Isolates characterized by quick sporulation (day 2 after death) did not produce better epizootics in termites than those with high total sporulation (day 11 after death) in either fungal species. An isolate of M. anisopliae ranked highly in all three categories (virulence, quick sporulation, total sporulation) produced better epizootics than an isolate that was inferior in all three characteristics. High temperature (35ºC) significantly reduced fungal germination rates, leading to significant reduction of epizootics. Thus, fungal characteristics other than virulence must be taken into account for the seasonal colonization approach to termite microbial control.

During Pleistocene glaciation much of the southeastern United States was covered with mixed mesophytic hardwood forest. These forests are composed predominantly of magnolia, holly, and beech with a mixture of other tree species, such as oak and hickory, and a distinct understory. Remnants of mixed mesophytic hardwood forests occur in the southern United States and are important refugia for disjunct and habitat-restricted species. In Louisiana, the mixed mesophytic forest habitat is found mostly in West Feliciana Parish, particularly in the area around St. Francisville. I chose two sites to conduct a study of spider diversity in litter habitats of disturbed and mature forests. Comparing spider species found these two habitats was necessary to understand the effects of disturbance on species richness and abundance. Berlese sampling was used to collect 10-kg samples of forest litter twice monthly from both sites. Collections were made from October 1998 to October 1999. I collected 1725 adult specimens representing 89 species in 14 families. At the mature forest site (Tunica Hills WMA) I collected 909 adult specimens, 58 species in 12 families. At the disturbed forest site (Feliciana Preserve) I collected 816 adult specimens, 73 species in 12 families. Species accumulation and richness estimators indicated the likelihood that additional species were present but not collected during the sampling period. The disturbed site had significantly greater species diversity and more uncommon species, perhaps because of a wider variety of microhabitats and presence of tourist and colonizer species. The mature forest site was less diverse, perhaps as a result of more stable and homogeneous habitat. Multiple disjunct species with northern affinities were found during the course of this study. Twelve species previously unreported for the state were discovered bringing the total to 225 spider species that are known to occur in Louisiana.

Rice is grown over an area of approximately 500,000 acres in Louisiana. The lepidopteran stem borer complex attacking rice in the southern U.S includes stalk borer Chilo plejadellus Zincken, sugarcane borer (SCB) Diatraea saccharalis (F.) and Mexican rice borer, Eoreuma loftini Dyar. With the increasing impact of stem borers in Louisiana, an urgent need exists to develop strategies for management. Currently, no IPM program is in place for stem borers in Louisiana rice and research has been initiated to develop an IPM program for these pests. The first objective of this research was focused on host plant resistance. For this objective, oviposition preference and larval performance of sugarcane borer on commonly grown rice cultivars in Louisiana were investigated. Results from the oviposition preference study revealed significant differences among cultivars. Overall females of D. saccharalis preferred ovipositing on the upper sides of the leaves of rice plants. In the performance study, three different measures of performance were used. Results from these studies revealed significant differences among varieties for these measures of performance. Results also revealed a strong correlation between different measures of performance as well as between performance and preference. Results from the compensation study revealed differences in compensatory response of same eight cultivars. In the silicon soil amendment study, a significant increase in silicon content of rice plants supplemented with calcium silicate was observed compared to the control plants. Soil Si amendment led to lower relative growth rates and reduced boring success of sugarcane borer larvae on. Studies were conducted to evaluate the efficacy of Dermacor seed treatment against sugarcane borer. Dermacor seed treatment was the most effective among different insecticides used in a field study and significantly increased larval mortality in lab and greenhouse experiments. These studies will help facilitate scouting for sugarcane borer in the field and improvement in insecticide timing. Potential exists for current use of these (moderately resistant) cultivars in IPM programs and as sources of resistance in breeding programs for stem borer resistance. Soil Si amendment and Dermacor seed treatments has the potential to fit into the IPM program.

Insect herbivores can induce a range of plant defenses. Signal pathways can be activated that result in the production of secondary metabolites. Many of these compounds can reduce insect fitness, deter feeding, and attract beneficial insects. Additionally, organic and inorganic chemicals applied as a foliar spray or soil drench can activate these plant responses. Azelaic acid, benzothiadiazole (BTH), gibberellic acid (GA3), harpin, and jasmonic acid (JA) are thought to mediate plant response to pathogens and herbivores. The effects of these elicitors on the induction of plant defenses were determined by measuring the weight gain of fall armyworm (FAW), Spodoptera frugiperda (Smith) (Lepidoptera: Noctuidae) on four important crops, cotton, corn, rice, and soybean, treated with elicitors, under greenhouse conditions. JA consistently induced cotton and soybean resistance to FAW. In contrast, azelaic acid, BTH, and harpin treated plant material increased weight gain of FAW, suggesting negative crosstalk between the salicylic acid and JA signaling pathways. No induction of defense was observed in corn and rice, and the lack of spray mixture adhesion to those crops inspired a second experiment in which four adjuvants were co-applied with a reduced rate of JA (0.25X) to corn and cotton. Corn was more responsive to the use of an adjuvant than was cotton. The differential effectiveness of two elicitors, JA and BTH, was investigated on FAW and another noctuid species, the soybean looper (SBL), Chrysodeixis includens (Walker). Weight gain of FAW offered JA-treated soybean was significantly lower than FAW offered non-treated soybean in all trials, whereas growth of SBL was significantly reduced in only one trial. BTH was not effective in reducing weight gain of SBL offered plant material treated at the 1X or 5X rates. BTH reduced weight gain of FAW only in trial 2, at the 5X rate. The findings presented herein provide further support that foliar applications of JA increase resistance to arthropod herbivores, and that this relationship between plant and herbivore could be transitioned to the field, with the ultimate goal of using elicitor-induced defense as a part of an integrated pest management program.

The effects and interactions of chemical, cultural, and biological control of red imported fire ant (RIFA) populations were evaluated in two replicated studies in pastures. The first study was conducted from 2005 through 2007 in Washington parish, Louisiana (WP) where the biological control Psuedacteon tricupsis phorid flies were present and at the LSU AgCenter St. Gabriel Research Station in St. Gabriel, Louisiana (SG) where Psuedacteon tricuspis flies were absent. The second study was conducted from May 2009 to May 2010 at St. Gabriel in the presence of phorid flies. In both studies, RIFA population indices in sites treated with methoprene in the presence of phorid flies were significantly less than at pretreatment after one year compared to six months in the absence of phorid flies. In study one, methoprene treatments did not significantly reduce the abundance of phorid flies or Kneallhazia solenopsae infections. In study two, the effects of plowing and methoprene treatments did not interact, and the height of RIFA mounds in plowed pastures was significantly reduced for nine months while methoprene treatment had no effect on mound height. The results of these two studies suggest that the effects of methoprene treatments on RIFA populations are temporally extended in areas where phorid flies are present and that methoprene treatments do not negatively impact the abundance of phorid flies. Since plowing significantly reduces the height of RIFA mounds for nine months and methoprene does not reduce the height of RIFA mounds, plowing is a more effective tool that producers can use rather than methoprene treatments to negate RIFA related damage to hay equipment. However, treatment of areas to reduce RIFA population density to diminish direct impact of RIFAs on animal and human health can be achieved with methoprene applications, and the temporal and economic benefits of these treatments should be extended in areas where phorid flies are present.