John D. Pinto

The utility of internally transcribed spacer 2 DNA sequences of the nuclear ribosomal gene for distinguishing sibling species of Trichogramma

The usefulness of the internally transcribed spacer 2 (ITS2) of the nuclear ribosomal gene complex is tested for providing taxonomic characters to identify Trichogramma species. The ITS2 sequences of a group of sibling species of the T. deion/T. pretiosum complexes were determined. A simple and precise identification key to the species of these assemblages was constructed using as taxonomic characters the size of the ITS2 and the difference in restriction length polymorphism of species with similarly sized ITS2. Individual wasps can be identified by amplification of their ITS2 with general primers, determining the size of the PCR product using standard agarose electrophoresis, followed in some species by a DNA-digestion with a restriction enzyme. Because this system works well for a number of closely related species we are hopeful that similar PCR-based identification can be extended to all species of the genus once their ITS2 sequences have been determined. The advantage of this identification system over the morphology-based system is that non-specialists are able to quickly and cheaply identify individual specimens. In addition, species specific primers were tested for the two most common species of these groups (i.e. T. pretiosum and T. deion). These primers can be used either as a direct identification tool or as a method to confirm the identification using the general key. The phylogeny of this group of wasps was also analyzed based on the ITS2 sequence.

Phylogenetic studies of Meloidae (Coleoptera), with emphasis on the evolution of phoresy

One of the most recent classifications of Meloidae is based on the assumption that phoretic first‐instar larvae evolved twice in the family, once in Meloinae and again in Nemognathinae. Within Meloinae, this scheme places all presumed phoretic taxa in Meloini regardless of other characteristics. This paper challenges this classification with a cladistic analysis of all meloid genera whose first‐instar larvae were available for study. It concludes that phoresy evolved several times in Meloinae alone and that Meloini, when defined to include all phoretic genera, is polyphyletic. Cladistic support also is presented for four subfamilies of Meloidae and for several of the traditional tribes recognized in recent classifications.

Crossing Incompatibility Between Trichogramma minutum and T. platneri (Hymenoptera: Trichogrammatidae): Implications for Application in Biological Control

Abstract The two egg parasitoid species Trichogramma minutum Riley and T. platneri Nagarkatti are closely related. No morphological or molecular characters are known to distinguish them. Their treatment as distinct species rests on the absence of female offspring in between-species crosses. The current species definitions, which must be considered tentative, assume their natural ranges to differ, with T. minutum occurring east of the Rocky Mountains and T. platneri occurring to the west. Both species are used throughout North America for the biological control of moths in orchards and forests. They are available to growers and researchers through a large number of biological control suppliers. Because the species can only be identified after crosses with known cultures, it is likely that producers often cannot be sure of the identity of their wasps. Here we studied the causes of the lack of female offspring in the between-species crosses and what happened when one species is released for biological control in the native area of the other. Our results show that females in between-species crosses are inseminated and use the sperm to fertilize their eggs, but that these fertilized eggs die. In addition, females do not preferentially mate with males of their own species when exposed to conspecific and nonconspecific males. These results are used in a model to predict the effect of releasing the non-native species in the native area of the other species. This model shows that such introductions can result in a prolonged and substantial reduction of intended biological control.

The systematics of the Trichogramma minutum species complex (Hymenoptera: Trichogrammatidae), a group of important North American biological control agents: the evidence from reproductive compatibility and allozymes

Abstract Two North American Trichogramma , Trichogramma minutum Riley, and Trichogramma platneri Nagarkatti (the T. minutum complex), cannot be distinguished morphologically and their species status has been questioned. Both are commercially available for biological control and are important parasitoids of the eggs of Lepidoptera in agricultural ecosystems. Results are reported of an extensive survey of reproductive compatibility and allozymes at the phosphoglucomutase ( Pgm ) locus in collections representing 85 localities throughout much of the geographical range of this species complex. In total, 103 reproductive crosses were performed and 161 collections were analyzed electrophoretically. Results support the recognition of T. minutum and T. platneri as distinct species. The two are distinguishable electrophoretically at the Pgm locus. High levels of reproductive incompatibility are correlated with the allozymic differences. Although T. platneri is western in distribution (W of 115° longitude) and T. minutum is primarily eastern, the two are sympatric in the Pacific Northwest. A low level of interspecific hybridization in laboratory studies notwithstanding, there is no evidence that introgression occurs in the zone of sympatry. This study continues to stress the importance of species recognition and the inadvisability of using T. minutum and T. platneri interchangeably in biological control.

Precise sex allocation, local mate competition, and sex ratio shifts in the parasitoid wasp Trichogramma pretiosum

Abstract We determined the sex, order, and clutch size of eggs laid by the parasitoid wasp, Trichogramma pretiosum Riley, in the eggs of one of its natural hosts, Trichoplusia ni (Hübner). The parasitoid allocated sex non-randomly to hosts in the laboratory with a variance significantly less than that of a binomial (random) distribution, our null model. More clutches of two or more eggs contained a single male egg as the second or third egg laid than would be expected by chance and none contained two or more male eggs. T. pretiosum also increased the sex ratio (% male) of its offspring with increasing foundress numbers by increasing the frequency of male offspring as the second egg in a two-egg clutch allocated to unparasitized hosts and as the single egg allocated to previously parasitized hosts. These results indicate that T. pretiosum allocates the sex of its offspring precisely. Precise sex allocation is favored under local mate competition because it reduces variation in the number of sons per patch thus maximizing the number of inseminated daughters emigrating from the patch. Similar combinations of female and male offspring emerged from T. ni eggs parasitized by T. pretiosum in the field, again with a sex ratio variance less than that expected for a binomial distribution. These results strongly suggest that this parasitoid species manifests local mate competition.

Effects of water stress and rehydration on hemolymph volume and amino acid content in the blister beetle, Cysteodemus armatus

1. 1. Hemolymph volume, osmolality, amino acid concentrations and fecal amino acid concentrations were measured in a desert blister beetle, Cysteodemus armatus LeConte, which were not desiccated, 10% desiccated (% of body mass), or rehydrated. 2. 2. Desiccation and rehydration had significant effects on the hemolymph volume in this species but not hemolymph osmolality. 3. 3. Total amino acid concentrations were significantly affected by hydration state. 4. 4. Hemolymph urea, asparagine, methionine and histidine were significantly affected by hydration state, but other ninhydrin positive substances (NFS) were not affected. 5. 5. While production of fecal material (frass) decreased with desiccation, the concentration of most amino acids increased, indicating excretion as an avenue of hemolymph osmoregulation.

A molecular phylogeny of the Trichogrammatidae (Hymenoptera: Chalcidoidea), with an evaluation of the utility of their male genitalia for higher level classification

Abstract A molecular phylogeny of the Trichogrammatidae (Hymenoptera: Chalcidoidea) is presented. This group of minute egg parasitoids is known from a broad range of host insects. The phylogeny produced, the first of any kind for the family, utilizes 121 taxa in fifty‐two of the eighty‐four recognized genera. Results were inferred from the ribosomal RNA regions 18S, 28S‐D2 and 28S‐D3, all aligned according to secondary structure models. Parsimony analysis was performed on both a complete and reduced dataset, in which ambiguous regions as defined by secondary structure were eliminated. The reduced dataset produced a much less resolved phylogenetic hypothesis. Only the complete dataset was utilized for Bayesian and maximum likelihood analyses. A robust‐choice Bayesian hypothesis stemmed from the concatenation of five distinct character set parameters. The results are compared with the current classification based primarily on male genitalia. Although our conclusions are partially congruent with the accepted hypothesis of trichogrammatid relationships, none of the currently adopted assemblages was recovered as monophyletic. Nevertheless, the structure of the male genitalia does correspond with relative taxon position in the molecular hypotheses. In general, the greatest genitalic simplification and fusion characterizes taxa treated as the most derived in the molecular hypotheses. Several groups are consistently recovered, but relationships between these groups and other genera vary with the analytical method. A new classification of the Trichogrammatidae is proposed. It includes one tribe, Trichogrammatini, within the subfamily Trichogrammatinae, and three tribes, Paracentrobiini, Chaetostrichini and Oligositini, within the Oligositinae. All tribes are more narrowly defined than previously and many genera are treated as incertae sedis within each subfamily. The results are interpreted in the light of morphological evidence, and the placement of genera not represented in molecular analysis is inferred by morphology alone. Host associations throughout the family are varied, although the molecular results suggest that Trichogrammatidae primitively parasitized Coleoptera, with more recent radiations onto other insect orders, such as Lepidoptera and Hemiptera.

Non-reciprocal cross-incompatibility in Trichogramma deion

In non‐reciprocal cross‐incompatibility (NRCI), the crossing of a female of a strain A with a male of a strain B results in hybrid offspring, whereas the reciprocal cross produces few or no hybrids. Only females are of hybrid origin in Hymenoptera because they arise from fertilized eggs; males arise from unfertilized (haploid) eggs. Crosses between many strains of Trichogramma deion showed some degree of NRCI. Crosses between a T. deion culture collected in Seven Pines, California (SVP) with one from Marysville, California (MRY) showed an extreme form of NRCI in which practically no female offspring was produced when MRY females were crossed with SVP males. The reciprocal cross produced a close to normal proportion of female and male offspring. Detailed studied of this cross indicated that 1) the female offspring produced in the compatible interstrain cross were not the result of parthenogenesis but were true hybrids, 2) the incompatible interstrain cross did not produce female offspring because fertilized eggs died during development, 3) the death of these eggs could not be prevented by either antibiotic or temperature treatment, 4) cytoplasmically inherited factors causing NRCI could be discounted because backcrossed females with the genome of MRY and the cytoplasm of SVP, exhibit the NRCI relationship characteristic of their genome. Therefore the NRCI between these strains appears to be caused by a modification coded for by the nuclear genes of MRY that results in incompatibility when SVP sperm fertilizes MRY eggs. In addition the level of incompatibility in crosses between the SVP females and MRY males is temperature sensitive, the higher the rearing temperature the lower the level of compatibility.

First-instar larvae, courtship and oviposition in Eletica: amending the definition of the Meloidae (Coleoptera: Tenebrionoidea)

Abstract. The first data are presented on larvae, oviposition, and mating behaviour in the Eleticinae, the most primitive subfamily of Meloidae. Taxa studied include two African species of Electica. Larval descriptions of E.rubripennis and E.wahlbergi are presented, as well as observations of oviposition in E.rubripennis and of courtship leading to copulation in E.wahlbergi. The placement of eggs under bark and the absence of a triungulin larva in both species are unique for the family. They represent plesiotypic traits found in other groups of Tenebrionoidea.

Water Relations of the Desert Blister Beetle Cysteodemus armatus (Leconte) (Coleoptera: Meloidae)

Several parameters of the xeric adaptiveness of the desert blister beetle Cysteodemus armatus were studied in the field and laboratory. Field measurements demonstrated that these beetles were most active during warmer parts of the day with exposure to high air (>35 C) and soil (>55 C) temperatures. Beetles had body temperatures consistently higher than ambient air temperatures but less than substrate temperatures. Rates of water loss (173 mg day⁻¹, including fecal losses), gravimetrically determined in the laboratory at 30 C, were higher than field-determined tritiated water turnover (125 mg day⁻¹). The Q10s for water loss were highest (5.4) at 25-30 C and lowest at 30-35 C (1.6). Cysteodemus armatus is subjected to stressful conditions and compensates for water losses with water-rich food (creosote flowers with 85% water content). However, the voracious feeding habits of these beetles are most likely devoted to energy accommodation rather than the water balance. Females ingest a mean of 185 cal day⁻¹, with a fecal output of 41 cal day⁻¹ and a metabolic output of 71 cal day⁻¹, providing about 73 cal day⁻¹ for egg production, with a batch of 100 eggs produced every 3.2 days. Under stress with no food available the beetles have higher rates of water loss than any other desert species studied. The major water saving displayed by stressed animals is through reduction of fecal water output. A total water budget showed a gain of 200 mg day⁻¹ and losses of 184 mg day⁻¹, excluding reproductive losses.