Michael I. Haverty

Intercaste, intercolony, and temporal variation in cuticular hydrocarbons ofCopotermes formosanus shiraki (Isoptera: Rhinotermitidae)

We characterized the variation in cuticular hydrocarbon mixtures between seven colonies of the Formosan subterranean termite,Coptotermes formosanus Shiraki, from the same population. We report differences between castes, between colonies, and within the population over time to assess seasonality. Colonies ofC. formosanus from Oahu, Hawaii, were sampled for 25 months. Each month, one sample each of 200 workers, 50 soldiers, nymphs, or alates from each colony was subjected to gas chromatography-mass spectrometry (GC-MS) analysis of the cuticular hydrocarbons. We resolved 39 individual peaks and identified 52 individual or isomeric mixtures of hydrocarbons. Onlyn-alkanes and methyl-branched alkanes occur; no olefins were found. Internally branched monomethylalkanes were the most abundant class of hydrocarbons, representing 45% to 50% of the total 9-;11-;13-Methyl-heptacosane accounted for over 30% of the total hydrocarbon for all castes. 2-Methyl- and 3-methylalkanes comprise approximately 30% of the total. Internally branched dimethylalkanes constitute 15% to 20% of the total cuticular hydrocarbon. Only one trimethylalkane, 13,15,17-trimethylnonacosane, was found in small amounts. The hydrocarbon mixtures of all four castes were similar. Quantitative differences in hydrocarbon mixtures among the castes were easily displayed using canonical discriminant analysis. Soldiers and workers are significantly different from one another and from nymphs and alates. Nineteen peaks are statistically significant between workers and soldiers. Nymphs and alates were not statistically different. We detected statistically significant quantitative differences between colonies in 18 peaks for workers and 12 peaks for soldiers. Each of the colonies ofC. formosanus can be separated from the others by the proportions of their hydrocarbon components. We detected statistically significant differences between months of the year for 12 peaks for workers and four peaks for soldiers; two peaks each for workers and soldiers showed distinct, seasonal trends. This seasonal shift in proportions of hydrocarbons correlates with the production of alates.

Correlation of mitochondrial haplotypes with cuticular hydrocarbon phenotypes of sympatric Reticulitermes species from the southeastern United States.

Three sympatric Reticulitermes species have been identified in Georgia, USA, based on morphological characters from alates and soldiers: R. flavipes, R. virginicus, and R. hageni, but species identification at individual collection sites is often difficult because alate production is seasonal and soldiers comprise 1–3% of the colony. We therefore set up an experiment to determine if chemical phenotypes and mtDNA haplotypes can be used together to separate species of subterranean termites. Subterranean termites of the order Reticulitermes (Isoptera: Rhinotermitidae) were collected from 20 inspection ports across four soil provinces in Georgia. Each collection was identified to species using dichotomous keys. Two collections, HH11 and BH25, however, could not be unequivocally keyed to species and were classified as unknown. The mitochondrial cytochrome oxidase II (COII) gene was sequenced from individual members of each collection and the variation in cuticular hydrocarbon phenotypes from these same collections was characterized. The cuticular hydrocarbon and mtDNA phylogenetic analyses show agreement with both unknown collections falling out in a separate clade. Specimens from HH11 nad BH25 are different morphologically, chemically, and genetically from the three known sympatric species in Georgia. Our results suggest that these two collections may represent at least one new taxon in Reticulitermes. Furthermore, the association of cuticular hydrocarbon phenotypes and mtDNA haplotypes demonstrates that, when combined with morphological characters, they are useful in separating known species, determining new species, and understanding termite evolution.

Correspondence of soldier defense secretion mixtures with cuticular hydrocarbon phenotypes for chemotaxonomy of the termite genus Reticulitermes in North America

Soldier defense secretions from samples of Reticulitermes collected in California, Nevada, Arizona, New Mexico, and Georgia were characterized and correlated with cuticular hydrocarbon phenotypes. Twenty-seven cuticular hydrocarbon phenotypes have been defined, and soldier defense secretion (SDS) phenotypes have been described for 25 of these. Forty-five terpenoid compounds were found, including monoterpenes, sesquiterpenes, and a few diterpenes. The monoterpenes include (−)-α-pinene, (−)-β-pinene, (−)-camphene, myrcene, (Z)- and (E)-ocimene, and (−)-limonene. The major sesquiterpenes produced are (+)-γ-cadinene, (+)-γ-cadinene aldehyde, (−)-germacrene A, germacrene B, γ-himachalene, and β-bisabolene. Some SDS phenotypes pair with more than one cuticular hydrocarbon phenotype; however, with two exceptions, each hydrocarbon phenotype is associated with only one SDS phenotype. These chemical characterizations lend support to the conclusion that there are numerous undescribed species of Reticulitermes in North America.

Response of Reticulitermes spp. (Isoptera: Rhinotermitidae) in Northern California to Baiting with Hexaflumuron with Sentricon Termite Colony Elimination System

Abstract Colonies of Reticulitermes spp. were baited with prototype and commercial Sentricon stations (Dow AgroSciences LLC, Indianapolis, IN) to test the efficacy of hexaflumuron in different concentrations and bait matrices and to document reinvasion of the foraging territories vacated by eliminated colonies. Seven colonies of Reticulitermes spp. from two sites were characterized with cuticular hydrocarbon analyses and mark-release-recapture and agonistic behavioral studies. Three colonies were observed as controls and four colonies were baited. When a connection between the bait station and the monitoring station could not be confirmed by mark-release-recapture studies, the results of the baiting were equivocal. The monitoring stations of a colony at our wildland site were devoid of termites 406 d after baiting with one Sentricon station, but became reoccupied with the same species of termites ≈6 mo after baiting. A colony at the residential site was baited with 0.5% hexaflumuron in the Recurit II bait matrix; 60 d later termites were absent from all monitoring stations. These monitoring stations remained unoccupied for ≥18 mo. Foraging Reticulitermes spp. appeared in three of the seven monitoring stations 18, 24, and 36 mo after baiting, respectively. Using cuticular hydrocarbon analyses and agonistic behavior studies, we determined that the Reticulitermes spp. occupying these monitoring stations were from three different colonies; none were members of the original colony destroyed by baiting. Another colony at the residential site was baited using a noncommercial, experimental bait; 52 d later termites were absent from all monitoring stations. The monitoring stations remained unoccupied for ≥9 mo. A different Reticulitermes sp. colony invaded one monitoring station 9 mo after baiting.

Nest Growth and Survivorship in Three Species of Neotropical Nasutitermes (Isoptera: Termitidae)

Abstract Long-term growth and survivorship of individual arboreal nests were studied in three species of Neotropical termites in the genus Nasutitermes. Of the 29 N. corniger (Motschulsky) and seven N. ephratae (Holmgren) nests monitored in an area of young second-growth in Panama, 12 (41%) N. corniger and four (57%) N. ephratae nests remained active throughout the 9- to 11-mo study. There was no significant difference in survivorship between small and large nests of either species. In surviving N. corniger nests with a single queen, the net increase in volume was highly correlated (r = 0.87, n = 9) with queen wet weight. There was a marked seasonality to nest expansion in both N. corniger and N. ephratae, with growth occurring almost exclusively during the wet season. Seventeen N. acajutlae (Holmgren) nests were monitored for 4–9 yr on Guana Island, British Virgin Islands. Four of the 17 (23.5%) N. acajutlae nests survived the study period, and two more abandoned their original nest and relocated. Within this limited sample of colonies, N. acajutlae nests that were large (>150,000 cm3) at the beginning of the study had a higher probability of survival than did small (<100,000 cm3) nests. Nest budding, relocation, and resprouting are mechanisms that Nasutitermes may use to create a new nest for all or a portion of an established colony. The ontogeny of incipient Nasutitermes colonies is discussed as a sequence in which a young colony remains cryptic within wood, building its population size to a point where the colony can maintain and defend a nest. Early in a wet season, termites then venture from within wood to build and occupy a small arboreal nest.

Modification of cuticular hydrocarbons of Cryptotermes brevis (Isoptera: Kalotermitidae) in response to temperature and relative humidity.

Abstract To assess their ability to modify cuticular hydrocarbon (CHC) composition and survive adverse conditions, Cryptotermes brevis (Walker) nymphs were subjected to various combinations of temperature and relative humidity. Cuticular hydrocarbon profiles of C. brevis were consistent with previous studies. Alkenes were the most prevalent in the CHC mixture, comprising 54.5% of the total hydrocarbon ( n = 12), whereas n-alkanes and branched alkanes comprised 24.8 and 6.3%, respectively. Sixteen compounds yielded >2% of the total hydrocarbon and were subsequently tested for temperature and humidity effects in two successive experiments. In both experiments, temperature effects were found: n-C29 increased and n-C25 decreased with increasing temperature. Similarly, five compounds ( n-C25, n-C27, n-C29, C39:2, and C41:2) were analyzed for relative humidity effects in experiment 2, based on the results of experiment 1; only C41:2 indicated a significant positive relative humidity effect. The remaining 11 compounds comprising >2% of the total hydrocarbon were tested, and a single statistically significant increase was found with C45:3 with increasing temperature. Significant positive effects were found with total alkenes, dienes, and trienes; relative humidity had the opposite effect on total n-alkanes. In both experiments warm, damp conditions were detrimental to survival. Overall, CHC modification was minor; and given that C. brevis has numerous adaptations for dealing with desiccation and an inability to tolerate high relative humidity it suggests that this species may not vary widely from a highly desiccation-tolerant state.

Laboratory Evaluation of Within-Species, Between-Species, and Parthenogenetic Reproduction in Reticulitermes Flavipes and Reticulitermes Virginicus

Considerable interest currently exists regarding the reproductive strategies of social insects (Blum and Blum, 1979; Crozier, 1979). Among termites (Order Isoptera) colony foundation by alate pairs, fusion of existing colonies, splitting of existing colonies, and parthenogenesis have all been reported (Nutting, 1969). Little information is available regarding the relative importance of each of these strategies. The genus Reticulitermes (Rhinotermitidae) contains six Nearctic and twelve Palearctic species, three of which have been critically examined for reproductive modes. Pickens (1932) and Weesner (1956) studied colony foundation of R. hesperus Banks by male + female dealate pairs, as well as by parthenogenesis. Buchli (1950) studied similar strategies for R. lucifugus Rossi. Clement (1979) studied interspecific hybridizationof R. santonensis Feytaud and R. iucifugus. More limited studies on colony foundation by male + female dealate pairs of R.flavipes (Kollar) were conducted by Beard (1974). Field studies with R. flavipes (Howard and Haverty, 1980) suggest that an important reproductive strategy for this species is colony splitting with subsequent production of numerous (several hundred) neotenic reproductives. However, sizeable alate flights are also a prominent feature of the biology of Reticulitermes spp. and

ODORANT-BINDING PROTEINS FROM A PRIMITIVE TERMITE

Hitherto, odorant-binding proteins (OBPs) have been identified from insects belonging to more highly evolved insect orders (Lepidoptera, Coleoptera, Diptera, Hymenoptera, and Hemiptera), whereas only chemosensory proteins have been identified from more primitive species, such as orthopteran and phasmid species. Here, we report for the first time the isolation and cloning of odorant-binding proteins from a primitive termite species, the dampwood termite, Zootermopsis nevadensis nevadensis (Isoptera: Termopsidae). A major antennae-specific protein was detected by native PAGE along with four other minor proteins, which were also absent in the extract from control tissues (hindlegs). Multiple cDNA cloning led to the full characterization of the major antennae-specific protein (ZnevOBP1) and to the identification of two other antennae-specific cDNAs, encoding putative odorant-binding proteins (ZnevOBP2 and ZnevOBP3). N-terminal amino acid sequencing of the minor antennal bands and cDNA cloning showed that olfaction in Z. n. nevadensis may involve multiple odorant-binding proteins. Database searches suggest that the OBPs from this primitive termite are homologues of the pheromone-binding proteins from scarab beetles and antennal-binding proteins from moths.

Agonistic Behavior Between Individual Worker Termites from Three Cuticular Hydrocarbon Phenotypes of Reticulitermes (Isoptera: Rhinotermitidae) from Northern California

Abstract Bioassays examining aggression in termites have typically been performed by pairing groups of workers from different colonies. We examined whether similar results would be observed using bioassays with individual workers. We report the results of pairings of individual workers of Reticulitermes spp. of three different hydrocarbon phenotypes (CA-A, CA-B, and CA-C). Resulting agonistic behavior between individuals can be used to infer relatedness of workers from different foraging groups because identification of species of Reticulitermes is often difficult. Pairings consisted of two termites from the same colony or from different colonies with the same or different phenotypes. We recorded avoidance behavior and immediate aggression during 5-min observations and mortality at 24 h. Intercolonial/intraphenotype pairings paralleled group bioassays with low levels of immediate aggression; however, individual pairings had lower levels of mortality at 24 h. Bioassays using individual Reticulitermes spp. workers in interphenotype pairings yielded results comparable with previously reported group bioassays and could be used in lieu of group bioassays. However, if immediate aggression is lacking, it would be advisable to use group bioassays or increase the number of replications to differentiate intracolonial and intercolonial pairings. Survival rates were the same for all phenotypes when paired with either of the other two phenotypes. There were, however, significant differences in the number of avoidance responses; pairings with CA-C had the highest number of avoidance responses followed by pairings with CA-B and CA-A. There were significant differences in immediate aggression; CA-C was the most aggressive, followed by CA-B and CA-A. Different levels of aggression among phenotypes provide additional evidence that these phenotypes represent distinct taxa or species.

Flight Phenology of Sympatric Populations of Reticulitermes (Isoptera: Rhinotermitidae) in Northern California: Disparate Flight Intervals Indicate Reproductive Isolation among Cuticular Hydrocarbon Phenotypes

Abstract The diurnal flights of Reticulitermes spp. have been reported to occur in the spring and early fall in northern California. From the spring of 1994 until the fall of 2001, we collected hundreds of alates of Reticulitermes spp. cuticular hydrocarbon phenotypes CA-A/A′ and CA-D during flights from multiple locations in the San Francisco Bay Area. As predicted, there were two flight seasons: one in the spring from February to May and one in the fall from October to December. The flights in the spring were made only by phenotypes CA-A/A′; those in the fall exclusively included phenotype CA-D. These disparate flight times indicate that reproductive isolation is complete and provide further biological evidence that phenotypes CA-A and CA-A′ are variants of one taxon and are distinct from CA-D.