Integrated pest management strategies targeting the Florida kissing bug, Triatoma sanguisuga: Preventing this vector of Chagas disease from invading your home.

Triatomines (Hemiptera: Reduviidae: Triatominae), commonly called "kissing bugs", are blood-sucking pests and vectors of the protozoan parasite Trypanosoma cruzi, the causative agent of Chagas disease (CD). Eleven species of kissing bugs occur throughout the southern half of the USA, four of which are well known to invade human dwellings. Certain kissing bugs in the USA are known to transmit T. cruzi to humans and other animals and their bites can also lead to serious allergic reactions, including anaphylaxis. In Florida, the kissing bug Triatoma sanguisuga frequently invades homes, bites residents, and has been found infected with T. cruzi, placing humans and companion animals at risk for CD. This review outlines integrated pest management (IPM) strategies for minimizing human exposure to T. sanguisuga and CD. A comprehensive IPM plan for kissing bugs includes detailed inspections, removal of vertebrate host nesting areas, and kissing bug harborage, home improvements to exclude kissing bugs from entering structures, pest removal, and judicious use of pesticides. This approach can limit or eliminate kissing bug entry into residential structures, thereby preventing kissing bug bites, and CD infections in humans and companion animals.

Tunneling Performance Increases at Lower Temperatures for Solenopsis invicta (Buren) but not for Nylanderia fulva (Mayr).

Nylanderia fulva (Mayr), the tawny crazy ant, is an invasive pest established in Florida and several other Gulf Coast states. In their invasive ranges in the Southeastern USA, large N. fulva populations have reduced species abundance, even displacing another invasive ant, Solenopsis invicta (Buren). In North Florida, N. fulva populations survive winter temperatures that reach below freezing for extended periods. However, the shallow littoral debris used by N. fulva for nest construction offers little insulation to brood and reproductives when exposed to freezing temperatures. Field populations of N. fulva in North Florida were observed tunneling below ground, a previously undescribed behavior. Other invasive ants exhibit similar subterranean tunneling behavior as a means of thermoregulation. To test the hypothesis that N. fulva has the capacity to construct subterranean tunnels across a range of ecologically relevant temperatures, tunneling performance for N. fulva and S. invicta, another invasive ant that tunnels extensively, were compared at four temperatures (15.0, 18.0, 20.0, and 22.0 °C). Overall, N. fulva tunneled significantly less than S. invicta. Nylanderia fulva tunneled furthest at warmer temperatures whereas S. invicta tunneled furthest at cooler temperatures. However, N. fulva constructed subterranean tunnels at all temperatures evaluated. These data support the hypothesis that N. fulva is capable of tunneling in temperatures as low as 15.0 °C, confirming that this ant can also perform a behavior that is used by other ants for cold avoidance.

Detection of Permethrin Resistance and Fipronil Tolerance in Rhipicephalus sanguineus (Acari: Ixodidae) in the United States.

Permethrin is a commonly used acaricide for tick control on domestic animals and in residential environments, while fipronil use is restricted to on-animal treatment. Following widespread reports of permethrin and fipronil application failures to control indoor infestations of Rhipicephalus sanguineus (Latreille), the brown dog tick, 31 tick populations were obtained from Florida and Texas for acaricide resistance screening. These field-collected ticks from kennels and residential facilities were challenged with technical grade permethrin and fipronil to create dose response curves that were compared with an acaricide-susceptible strain. Permethrin resistance was successfully screened in nine populations, all of which were resistant or highly resistant. Tick susceptibility to fipronil was conducted on four populations, which were found to be tolerant, with resistance ratios below 10. This is the first documentation of R. sanguineus permethrin resistance and fipronil tolerance in the United States. Potential causes of resistance development and recommendations on future brown dog tick control management plans are discussed.

RNA interference in the termite Reticulitermes flavipes through ingestion of double-stranded RNA.

RNA interference (RNAi) represents a breakthrough technology for conducting functional genomics research in non-model organisms and for the highly targeted control of insect pests. This study investigated RNAi via voluntary feeding in the economically important pest termite, Reticulitermes flavipes. We used a high-dose double-stranded (ds) RNA feeding approach to silence two termite genes: one encoding an endogenous digestive cellulase enzyme and the other a caste-regulatory hexamerin storage protein. Contrary to results from previous low-dose studies that examined injection-based RNAi, high-dose silencing of either gene through dsRNA feeding led to significantly reduced group fitness and mortality. Hexamerin silencing in combination with ectopic juvenile hormone treatments additionally led to lethal molting impacts and increased differentiation of presoldier caste phenotypes (a phenotype that is not capable of feeding). These results provide the first examples of insecticidal effects from dsRNA feeding in a termite. Additionally, these results validate a high-throughput bioassay approach for use in (i) termite functional genomics research, and (ii) characterizing target sites of conventional and novel RNAi-based termiticides.

Molecular and biochemical markers for monitoring dynamic shifts of cellulolytic protozoa in Reticulitermes flavipes.

Lower termites rely on cellulolytic protozoa to aid in the digestion of their wood-based diet. However, despite the major contribution of protozoa to the lower termite digestive system, few techniques have been developed to monitor shifts in protozoan populations. This study investigated whether quantitative real-time PCR (qRT-PCR) and/or cellulase enzyme assays can be used to monitor changes of cellulolytic protozoan populations in the lower termite, Reticulitermes flavipes (Kollar). Previously developed cellulase primer sets were used to test for changes in cellulase gene expression, while three different cellulase enzyme assays were used to assess changes in cellulase enzyme activity. The results from this study indicate that qRT-PCR is a reliable method to monitor shifts in cellulolytic protozoan populations. Specifically, qRT-PCR can serve as a useful monitoring technique during high-throughput screening of novel termite control agents such as cellulase inhibitors, and help to answer questions relating to whether or not such control agents impact cellulolytic protozoan populations.

Correlation of cellulase gene expression and cellulolytic activity throughout the gut of the termite Reticulitermes flavipes.

Termites have developed cellulose digestion capabilities that allow them to obtain energy and nutrition from nutritionally poor food sources, such as lignocellulosic plant material and residues derived from it (e.g., wood and humus). Lower termites, which are equipped with both endogenous (i.e., of termite origin) and symbiotic cellulases, feed primarily on wood and wood-related materials. This study investigated cellulase gene diversity, structure, and activity in the lower termite, Reticulitermes flavipes (Kollar). We initially used a metagenomics approach to identify four genes encoding one endogenous and three symbiotic cellulases, which we refer to as Cell-1, -2, -3 and -4. These four genes encode proteins that share significant sequence similarity with known endoglucanases, exoglucanases and xylanases. Phylogenetic analyses further supported these inferred relationships by showing that each of the four cellulase proteins clusters tightly with respective termite, protozoan or fungal cellulases. Gene structure studies revealed that Cell-1, -3 and -4 are intron-free, while Cell-2 contains the first intron sequence to be identified from a termite symbiont cellulase. Quantitative real-time PCR (qRT-PCR) revealed that the endogenous Cell-1 gene is expressed exclusively in the salivary gland/foregut, whereas symbiotic Cell-2, -3, and -4 are highly expressed in the hindgut (where cellulolytic protists are harbored). Cellulase activity assays mapped the distribution pattern of endoglucanase, exoglucanase and xylanase activity throughout the R. flavipes digestive tract. Cellulase gene expression correlated well with the specific types of cellulolytic activities observed in each gut region (foregut+salivary gland, midgut and hindgut). These results suggest the presence of a single unified cellulose digestion system, whereby endogenous and symbiotic cellulases work sequentially and collaboratively across the entire digestive tract of R. flavipes.

Speed of efficacy and delayed toxicity characteristics of fast-acting fire ant (Hymenoptera: Formicidae) baits.

Efficacy and speed of action of fire ant (Hymenoptera: Formicidae) baits that claim fast control of colonies were compared with a standard bait. More than 85% of red imported fire ant, Solenopsis invicta Buren, laboratory colonies provided bait containing the active ingredient indoxacarb died within 3 d, and all colonies were dead in 6 d. Standard bait containing hydramethylnon resulted in death of 60% of the colonies in 9 d. Bait containing spinosad did not cause colony death. Under field conditions, one-half of the areas treated with the indoxacarb bait did not have any active fire ant nests within 3 d, whereas 11 d was needed to reach the same level of control with the hydramethylnon bait. Spinosad had a maximum of 17% of the treated areas without nests after 3 d. The delay in death of S. invicta adults treated in the laboratory with the indoxacarb and spinosad baits was shorter than the standard hydramethylnon bait, which had mortality similar to the traditional delayed toxicity criterion of < 15% mortality after 24 h and > 89% mortality over the test period. Indoxacarb caused mortality of 57% at 24 h and 100% at 48 h; however, visual symptoms of toxicity were not readily observed for at least 8 h before the abrupt increase in death. Spinosad caused 96% mortality by 24 h, and initial mortality became apparent at 4 h. Time required for death of 15% of a treated population (LT15) of spinosad, indoxacarb, and hydramethylnon was 3, 9, and 16 h, respectively. Delayed toxicity characteristics of the fast-acting indoxacarb bait may be useful for the development of other fast-acting ant baits.

Social exploitation of hexamerin: RNAi reveals a major caste-regulatory factor in termites.

Lower termites express a unique form of eusocial polyphenism in that totipotent workers can differentiate into either soldier or reproductive caste phenotypes. In this initial effort using RNA interference in termites, we found that two hexamerin genes, Hex-1 and Hex-2, participate in the regulation of caste polyphenism. Our methodology involved a dual gene-silencing approach that used a single short-interfering RNA fragment to silence the two homologous hexamerin genes. We performed validation studies that evaluated effects on nontarget housekeeping genes, silencing of a nonhousekeeping control gene, and effects at the protein level. We found that the two hexamerin proteins, which are inducible by the morphogenetic juvenile hormone and which constitute a significant proportion of total termite protein, suppress juvenile-hormone-dependent worker differentiation to the soldier caste phenotype. This mechanism allows termite colonies to retain high proportions of altruistic worker caste members, thus apparently enhancing colony-inclusive fitness. These findings demonstrate a unique status quo regulatory mechanism for termite worker caste retention and provide an example of previously undescribed preadult developmental/caste-regulatory genes from any social insect.

Influence of soil compaction on tunnel network construction by the eastern subterranean termite (Isoptera: Rhinotermitidae).

Eastern subterranean termites, Reticulitermes flavipes (Kollar), workers were introduced into arenas containing low, moderate, and high compaction builder's sand (1.05 g/cm3, 1.18 g/cm3 or 1.35 g/cm3 bulk densities, respectively), and they immediately began tunneling. Termites built the tunnel network significantly fastest in soil of low compaction compared with moderately or highly compacted soil. In soil of low compaction, 221.67 +/- 4.73 cm of total tunnel distance was constructed in 1 d compared with only 96 cm of tunneling in highly compacted soil. At 14 d, total tunnel distance averaged 216.83 +/- 4.56 cm in soil of low compaction compared with 169.70 +/- 4.10 and 181.18 +/- 6.13 cm in moderately and highly compacted soil, respectively. Decreases in total tunnel distance between 1 and 14 d were caused by backfilling of seldom-used tunnels. Termites did the majority of tunneling during the first day of introduction into arenas. In soil of low and moderate compaction, termites essentially constructed the entire tunnel network within the first day, only modifying it by backfilling or maintaining tunnels. In highly compacted soil, 53% of the final tunnel network was constructed during the first day, 87% was constructed by the third day, and 97% was constructed by the seventh day. Soil compaction did not affect the number of primary tunnels or the number and diameter of secondary tunnels. The angle between the secondary tunnel and primary tunnel also was not significantly affected by soil compaction. However, the number of secondary tunnels in soil of low compaction (5.89 +/- 0.51) was significantly greater than in moderately (2.74 +/- 0.36) and highly (3.58 +/- 0.59) compacted soils.

Ability of canine termite detectors to locate live termites and discriminate them from non-termite material.

Dogs were trained to detect Eastern subterranean termites, Reticulitermes flavipes (Kollar), using the United States Customs method of scent detection dog training modified with a food reward. Dogs were tested with various numbers of Eastern subterranean termites placed in vented PVC containers. Trained dogs were 95.93% accurate in finding > or = 40 Eastern subterranean termite workers (positive indications) and incorrectly indicated the presence of termites in 2.69% of the containers without termites. Multiple species of termites [dark southern subterranean, R. virginicus (Banks); Formosan subterranean, Coptotermes formosanus Shiraki; powderpost, Cryptotermes cavifrons Banks; and southeastern drywood termites, Incisitermes snyderi (Light)], were similarly evaluated. Dogs trained to locate Eastern subterranean termites were also 100% accurate in finding dark southern subterranean termites, 98.89% accurate in finding Formosan subterranean termites, 97.33% accurate in finding powderpost termites, and 88.89% accurate in finding southeastern drywood termites. Dogs were able to discriminate live termites from non-termite material. Trained dogs' false response rate was 25.33% to Eastern subterranean termite-damaged wood, 6.67% to American cockroaches, Periplaneta americana (L.), and 2.67% to Florida carpenter ants, Camponotus floridanus Buckley.