Using the Traditional Ex Vivo Whole Blood Model to Discriminate Bacteria by Their Inducible Host Responses.

Whole blood models are rapid and versatile for determining immune responses to inflammatory and infectious stimuli, but they have not been used for bacterial discrimination. Staphylococcus aureus, S. epidermidis and Escherichia coli are the most common causes of invasive disease, and rapid testing strategies utilising host responses remain elusive. Currently, immune responses can only discriminate between bacterial 'domains' (fungi, bacteria and viruses), and very few studies can use immune responses to discriminate bacteria at the species and strain level. Here, whole blood was used to investigate the relationship between host responses and bacterial strains. Results confirmed unique temporal profiles for the 10 parameters studied: IL-6, MIP-1α, MIP-3α, IL-10, resistin, phagocytosis, S100A8, S100A8/A9, C5a and TF3. Pairwise analysis confirmed that IL-6, resistin, phagocytosis, C5a and S100A8/A9 could be used in a discrimination scheme to identify to the strain level. Linear discriminant analysis (LDA) confirmed that (i) IL-6, MIP-3α and TF3 could predict genera with 95% accuracy; (ii) IL-6, phagocytosis, resistin and TF3 could predict species at 90% accuracy and (iii) phagocytosis, S100A8 and IL-10 predicted strain at 40% accuracy. These data are important because they confirm the proof of concept that host biomarker panels could be used to identify bacterial pathogens.

Anatomy of a Neotropical insect radiation.

BACKGROUND: Much evolutionary theory predicts that diversity arises via both adaptive radiation (diversification driven by selection against niche-overlap within communities) and divergence of geographically isolated populations. We focus on tropical fruit flies (Blepharoneura, Tephritidae) that reveal unexpected patterns of niche-overlap within local communities. Throughout the Neotropics, multiple sympatric non-interbreeding populations often share the same highly specialized patterns of host use (e.g., flies are specialists on flowers of a single gender of a single species of host plants). Lineage through time (LTT) plots can help distinguish patterns of diversification consistent with ecologically limited adaptive radiation from those predicted by ecologically neutral theories. Here, we use a time-calibrated phylogeny of Blepharoneura to test the hypothesis that patterns of Blepharoneura diversification are consistent with an "ecologically neutral" model of diversification that predicts that diversification is primarily a function of time and space. RESULTS: The Blepharoneura phylogeny showed more cladogenic divergence associated with geography than with shifts in host-use. Shifts in host-use were associated with ~ 20% of recent splits (< 3 Ma), but > 60% of older splits (> 3 Ma). In the overall tree, gamma statistic and maximum likelihood model fitting showed no evidence of diversification rate changes though there was a weak signature of slowing diversification rate in one of the component clades. CONCLUSIONS: Overall patterns of Blepharoneura diversity are inconsistent with a traditional explanation of adaptive radiation involving decreases in diversification rates associated with niche-overlap. Sister lineages usually use the same host-species and host-parts, and multiple non-interbreeding sympatric populations regularly co-occur on the same hosts. We suggest that most lineage origins (phylogenetic splits) occur in allopatry, usually without shifts in host-use, and that subsequent dispersal results in assembly of communities composed of multiple sympatric non-interbreeding populations of flies that share the same hosts.

Phylogenetic relationships of the tribe Toxotrypanini (Diptera: Tephritidae) based on molecular characters.

Current hypotheses of relationship among the species of the fruit fly genera Anastrepha and Toxotrypana are tested using sequence data from six DNA regions: the mitochondrial regions 16S, CAD, and COI, and the nuclear regions EF1a, PER, and PGD. DNA sequences were obtained from 146 species of Anastrepha, representing 19 of the 21 species groups as well as five of the six clades of the robusta group, and four species of Toxotrypana in addition to species of Hexachaeta, Pseudophorellia, Alujamyia, and 13 other tephritid genera used as outgroups. The results indicate that Hexachaeta is more closely related to the Molynocoelia group than to Toxotrypana and Anastrepha, and it is removed from the tribe Toxotrypanini. The group Anastrepha+Toxotrypana and the genus Toxotrypana are strongly supported as monophyletic, consistent with previous studies, but Toxotrypana arises within Anastrepha, confirming that Anastrepha as currently defined is paraphyletic. The placement of Toxotrypana within Anastrepha is clearly defined for the first time with high support, as the sister group to the cryptostrepha clade of the robusta group of Anastrepha. Within Anastrepha, the daciformis, dentata, leptozona, raveni, and striata species groups are highly supported clades. The serpentina group is recognized with lower support, and the fraterculus and pseudoparallela groups are supported with minor alterations. The robusta group is resolved as polyphyletic, but four of the six species clades within it are recovered monophyletic (one clade is not represented and another is represented by one species). The punctata and panamensis groups are resolved together in a clade. At least some species of the mucronota group are related, however this group requires further study. The benjamini, grandis, and spatulata groups appear to be polyphyletic. Relationships among the species groups are generally poorly resolved, with the following exceptions: (1) the lineage including Toxotrypana, the cryptostrepha clade, and the tripunctata group; (2) the sister group relationship of the daciformis+dentata groups; (3) a clade comprising the punctata and panamensis groups; and (4) the large clade comprising the pseudoparallela+spatulata+ramosa+grandis+serpentina+striata+fraterculus groups.

Geographic population structure of the sugarcane borer, Diatraea saccharalis (F.) (Lepidoptera: Crambidae), in the southern United States.

The sugarcane borer moth, Diatraea saccharalis, is widespread throughout the Western Hemisphere, and is considered an introduced species in the southern United States. Although this moth has a wide distribution and is a pest of many crop plants including sugarcane, corn, sorghum and rice, it is considered one species. The objective was to investigate whether more than one introduction of D. saccharalis had occurred in the southern United States and whether any cryptic species were present. We field collected D. saccharalis in Texas, Louisiana and Florida in the southern United States. Two molecular markers, AFLPs and mitochondrial COI, were used to examine genetic variation among these regional populations and to compare the sequences with those available in GenBank and BOLD. We found geographic population structure in the southern United States which suggests two introductions and the presence of a previously unknown cryptic species. Management of D. saccharalis would likely benefit from further investigation of population genetics throughout the range of this species.

Lethal interactions between parasites and prey increase niche diversity in a tropical community.

Ecological specialization should minimize niche overlap, yet herbivorous neotropical flies (Blepharoneura) and their lethal parasitic wasps (parasitoids) exhibit both extreme specialization and apparent niche overlap in host plants. From just two plant species at one site in Peru, we collected 3636 flowers yielding 1478 fly pupae representing 14 Blepharoneura fly species, 18 parasitoid species (14 Bellopius species), and parasitoid-host associations, all discovered through analysis of molecular data. Multiple sympatric species specialize on the same sex flowers of the same fly host-plant species-which suggests extreme niche overlap; however, niche partitioning was exposed by interactions between wasps and flies. Most Bellopius species emerged as adults from only one fly species, yet evidence from pupae (preadult emergence samples) show that most Bellopius also attacked additional fly species but never emerged as adults from those flies.

Molecular Survey for the Invasive Leafminer Pest Liriomyza huidobrensis (Diptera: Agromyzidae) in California Uncovers Only the Native Pest Liriomyza langei.

Liriomyza huidobrensis (Blanchard) is a highly destructive invasive leafminer pest currently causing extensive damage to vegetable and horticultural crops around the world. Liriomyza langei Frick is a leafminer pest native to California that cannot currently be morphologically distinguished from L. huidobrensis. We used a DNA-barcoding approach, a published PCR-RFLP method, and a new multiplex PCR method to analyze 664 flies matching the morphological description of huidobrensis-langei. We found no evidence for the presence of L. huidobrensis in our extensive samples from California. In addition to the new molecular method, this work is important because it provides definitive data that the California "pea leafminer" is currently, and has probably always been, L. langei. These data will also be important in the event that the highly invasive L. huidobrensis ever becomes established.

The relationship between variable host grouping and functional responses among parasitoids of Antispila nysaefoliella (Lepidoptera: Heliozelidae).

Our study investigated the importance of variability in the parasitoid community as a source of selection on host group size using a field population of the tupelo leafminer, Antispila nysaefoliella Clemens, which specializes on tupelo, Nyssa sylvatica Marsh. Larvae were collected from leaves with variable numbers of larvae and screened for parasitism using polymerase chain reaction of mitochondrial cytochrome oxidase I using markers designed specifically for amplifying parasitoid DNA while excluding host DNA. This method of selective PCR was effective for detecting the presence and identifying species of immature stages of three hymenopteran superfamilies: Chalcidoidea, Ichneumonoidea and Platygastroidea, which represented 83.4%, 16.0% and 0.6% of the total detectable parasitism, respectively. Our resulting sequences were then calibrated with sequences from identified adult parasitoids that had been either reared or field-captured. A cluster analysis revealed 10 distinct clades that showed differences in attack patterns with respect to host traits and season. Total parasitism followed an inverse density-dependent or density-independent pattern with respect to host density (number per leaf). However, when parasitoid taxa were considered separately, one clade, which could be a cryptic species of Pnigalio maculipes Crawford (Chalcidoidea: Eulophidae), was found to increase its per leaf attack rate with host density. Our results suggest that parasitoid community composition and differences among species in their attack strategies can play a large role in determining the adaptive advantage of host grouping.

Hidden neotropical diversity: greater than the sum of its parts.

The diversity of tropical herbivorous insects has been explained as a direct function of plant species diversity. Testing that explanation, we reared 2857 flies from flowers and seeds of 24 species of plants from 34 neotropical sites. Samples yielded 52 morphologically similar species of flies and documented highly conserved patterns of specificity to host taxa and host parts. Widespread species of plants can support 13 species of flies. Within single populations of plants, we typically found one or more fly species specific to female flowers and multiple specialists on male flowers. We suggest that neotropical herbivorous insect diversity is not simply a function of plant taxonomic and architectural diversity, but also reflects the geographic distribution of hosts and the age and area of the neotropics.