John C. Day

Nondestructive DNA extraction from blackflies (Diptera: Simuliidae): retaining voucher specimens for DNA barcoding projects

A nondestructive, chemical‐free method is presented for the extraction of DNA from small insects. Blackflies were submerged in sterile, distilled water and sonicated for varying lengths of time to provide DNA which was assessed in terms of quantity, purity and amplification efficiency. A verified DNA barcode was produced from DNA extracted from blackfly larvae, pupae and adult specimens. A 60‐second sonication period was found to release the highest quality and quantity of DNA although the amplification efficiency was found to be similar regardless of sonication time. Overall, a 66% amplification efficiency was observed. Examination of post‐sonicated material confirmed retention of morphological characters. Sonication was found to be a reliable DNA extraction approach for barcoding, providing sufficient quality template for polymerase chain reaction amplification as well as retaining the voucher specimen for post‐barcoding morphological evaluation.

Cytomegalovirus Viremia as a Risk Factor for Mortality Prior to Antiretroviral Therapy among HIV-Infected Gold Miners in South Africa

Background Cytomegalovirus (CMV) viremia has been shown to be an independent risk factor for increased mortality among HIV-infected individuals in the developing world. While CMV infection is nearly ubiquitous in resource-poor settings, few data are available on the role of subclinical CMV reactivation on HIV. Methods Using a cohort of mineworkers with stored plasma samples, we investigated the association between CMV DNA concentration and mortality prior to antiretroviral therapy availability. Results Among 1341 individuals (median CD4 count 345 cells/µl, 70% WHO stage 1 or 2, median follow-up 0.9 years), 70 (5.2%) had CMV viremia at baseline; 71 deaths occurred. In univariable analysis CMV viremia at baseline was associated with a three-fold increase in mortality (hazard ratio [HR] 3.37; 95% confidence intervals [CI] 1.60, 7.10). After adjustment for CD4 count, WHO stage and HIV viral load (N = 429 with complete data), the association was attenuated (HR 2.27; 95%CI 0.88, 5.83). Mortality increased with higher CMV viremia (≥1,000 copies/ml vs. no viremia, adjusted HR 3.65, 95%CI: 1.29, 10.41). Results were similar using time-updated CMV viremia. Conclusions High copy number, subclinical CMV viremia was an independent risk factor for mortality among male HIV-infected adults in South Africa with relatively early HIV disease. Studies to determine whether anti-CMV therapy to mitigate high copy number viremia would increase lifespan are warranted.

The evolution of the adenylate-forming protein family in beetles: multiple luciferase gene paralogues in fireflies and glow-worms.

Bioluminescence in beetles is dependent upon the enzyme luciferase. It has been hypothesised luciferase evolved from a fatty acyl-CoA synthetase gene deriving a novel bioluminescent function (neofunctionalization) after a gene duplication event. We evaluated this hypothesis within a phylogenetic framework using independent evidence obtained from the genome of Tribolium castaneum, published luciferase genes and novel luciferase and luciferase-like sequences. This phylogenetic study provides evidence for a large gene family of luciferase and luciferase-like paralogues in bioluminescent and non-bioluminescent beetles. All luciferase sequences formed a clade supporting a protoluciferase existing prior to the divergence of the Lampyridae, Elateridae and Phengodidae (Elateroidea). Multiple luciferase genes were identified from members of the Photurinae and the Luciolinae indicating complex gene duplication events within lampyrid genomes. The majority of luciferase residues were identified to be under purifying selection as opposed to positive selection. We conclude that beetle luciferase may have arisen from a process of subfunctionalization as opposed to neofunctionalization early on in the evolution of the Elateroidea.

Confirmation of the species status of the blackfly Simulium galeratum in Britain using molecular taxonomy.

Abstract Since 1920 Simulium reptans (Linnaeus) (Diptera: Simuliidae) has been reported as exhibiting two different larval morphotypes, a typical S. reptans and an atypical S. reptans var. galeratum, which differ in the markings of the larval head capsule. Inconsistent variation in adults and no apparent variation in the pupae have led taxonomists to conclude that these types in Britain are a single species. We investigated populations in Britain where either the typical form or var. galeratum is found, and one population where the two exist sympatrically. A phylogenetic study based upon a region of the mitochondrial cytochrome c oxidase 1 gene (DNA barcoding) produced a tree that delineated the morphotypes into two distinct monophyletic clades. The average Kimura‐2‐parameter distances within each clade (i.e. within each morphotype) were very low (0.67% and 0.78%), with the distances between morphotypes being 9−10‐fold greater (mean 7.06%). This is concordant with differences within and between species in other taxa; based upon the strict correlation between the molecular variation and the morphotypes, we propose the re‐instatement of S. galeratum to species status.

The subgenus Eusimulium (Diptera: Simuliidae: Simulium) in Britain

In northwest and western Europe the blackfly subgenus Eusimulium (Diptera: Simuliidae: Simulium) is represented by four species of which three are widely distributed and one, Simulium petricolum, was previously thought to be largely restricted to the Mediterranean region. Simulium petricolum is reported from southern England, and is shown to be distinct from the other Eusimulium species in terms of mitochondrial DNA sequence (COI DNA barcode), but morphologically and cytotaxonomically indistinguishable from Mediterranean S. petricolum, with the exception of minor differences in larval pigmentation. Simulium petricolum shows a distinctive ecology in Britain and is probably much more widely distributed in Europe than is currently appreciated.

Genomic Structure of the Luciferase Gene from the Bioluminescent Beetle, Nyctophila cf. Caucasica

Abstract The gene coding for beetle luciferase, the enzyme responsible for bioluminescence in over two thousand coleopteran species has, to date, only been characterized from one Palearctic species of Lampyridae. Here we report the characterization of the luciferase gene from a female beetle of an Iranian lampyrid species, Nyctophila cf. caucasica (Coleoptera:Lampyridae). The luciferase gene was composed of seven exons, coding for 547 amino acids, separated by six introns spanning 1976 bp of genomic DNA. The deduced amino acid sequences of the luciferase gene of N. caucasica showed 98.9% homology to that of the Palearctic species Lampyris noctiluca. Analysis of the 810 bp upstream region of the luciferase gene revealed three TATA boxes and several other consensus transcriptional factor recognition sequences presenting evidence for a putative core promoter region conserved in Lampyrinae from -190 through to -155 upstream of the luciferase start codon. Along with the core promoter region the luciferase gene was compared with orthologous sequences from other lampyrid species and found to have greatest identity to Lampyris turkistanicus and Lampyris noctiluca. The significant sequence identity to the former is discussed in relation to taxonomic issues of Iranian lampyrids.

Comparative biometrics of British Marsh Tits Poecile palustris and Willow Tits P. montana

ABSTRACT Biometrics are commonly used to compare bird species. For Marsh Tits and Willow Tits in Britain there are few biometric data from birds of known age and sex, despite their value for population analyses in estimating the proportion of males and females in samples. Comparing measurements between the two species could also aid identification and the monitoring of these declining species in Britain. We present biometrics for a large sample of Marsh Tits of known age and sex, and new data for Willow Tits, which act as reliable reference material. Overall, adults of both species were larger than first-years and males were larger than females, but not among first-year Willow Tits. Marsh Tits were slightly larger and heavier than Willow Tits, but Willow Tits had proportionately longer tails. Discriminant analyses produced new equations for separating the species based on wing length and the measurement between the shortest and longest tail feathers. Probabilities were generated for estimating Marsh Tit population structure from samples of ringing data, but there was a greater overlap between sexes in Willow Tit measurements. We conclude by discussing issues of measurement accuracy and consistency in the collection and analysis of biometric data.

Bioluminescent leakage throughout the body of the glow-worm Lampyris noctiluca (Coleoptera: Lampyridae)

Adult female glow-worms, Lampyris noctiluca, Linnaeus, 1767, emit light from the abdominal ventrites 6 and 7 and a retained larval paired organ on segment 8. Both larvae and adult males exhibit a much smaller paired light organ in the penultimate segment. Using sensitive imaging techniques low level bioluminescence was observed within the light organ even when inactive and was also detected at discrete loci primarily localised at two posterolateral spots on each segment of the larvae. Dispersed light leakage was also detected in the adult male and female glow-worm. These findings indicate the presence of low level bioluminescence present throughout the body in all life stages of the glow-worm L. noctiluca suggesting a secondary role of luciferase within the glow-worm.

How will climate change pathways and mitigation options alter incidence of vector-borne diseases? A framework for leishmaniasis in South and Meso-America

The enormous global burden of vector-borne diseases disproportionately affects poor people in tropical, developing countries. Changes in vector-borne disease impacts are often linked to human modification of ecosystems as well as climate change. For tropical ecosystems, the health impacts of future environmental and developmental policy depend on how vector-borne disease risks trade off against other ecosystem services across heterogeneous landscapes. By linking future socio-economic and climate change pathways to dynamic land use models, this study is amongst the first to analyse and project impacts of both land use and climate change on continental-scale patterns in vector-borne diseases. Models were developed for cutaneous and visceral leishmaniasis in the Americas—ecologically complex sand fly borne infections linked to tropical forests and diverse wild and domestic mammal hosts. Both diseases were hypothesised to increase with available interface habitat between forest and agricultural or domestic habitats and with mammal biodiversity. However, landscape edge metrics were not important as predictors of leishmaniasis. Models including mammal richness were similar in accuracy and predicted disease extent to models containing only climate and land use predictors. Overall, climatic factors explained 80% and land use factors only 20% of the variance in past disease patterns. Both diseases, but especially cutaneous leishmaniasis, were associated with low seasonality in temperature and precipitation. Since such seasonality increases under future climate change, particularly under strong climate forcing, both diseases were predicted to contract in geographical extent to 2050, with cutaneous leishmaniasis contracting by between 35% and 50%. Whilst visceral leishmaniasis contracted slightly more under strong than weak management for carbon, biodiversity and ecosystem services, future cutaneous leishmaniasis extent was relatively insensitive to future alternative socio-economic pathways. Models parameterised at narrower geographical scales may be more sensitive to land use pattern and project more substantial changes in disease extent under future alternative socio-economic pathways.

Offspring sex ratio of a woodland songbird is unrelated to habitat fragmentation

AbstractWe assessed whether the sex ratio of nestling Marsh Tits (Poecile palustris) varied with habitat fragmentation by sampling broods from two British woodland areas with contrasting patch size, and comparing the data with published results from an extensive forest in Poland. We found no evidence for manipulation of offspring sex ratios, supporting findings from Poland, and suggesting that this trait may be typical of the species. The results are important in helping to understand the origin of biased sex ratios observed among adult Marsh Tits in the declining population in Britain.ZusammenfassungDas Geschlechterverhältnis von Nachkommen einer Waldsingvogelart ist unabhängig vom Grad der Fragmentierung des Lebensraums Wir prüften, ob das Geschlechterverhältnis von nestjungen Sumpfmeisen (Poecile palustris) mit dem Grad der Fragmentierung ihres Lebensraumes variiert. Dazu untersuchten wir zwei Populationen in Großbritannien, die sich in der Größe der Waldfragmente unterschieden, und verglichen die Daten mit publizierten Ergebnissen aus einem ausgedehnten Waldgebiet in Polen. Wir fanden keine Hinweise auf einen Einfluss der Fragmentierung auf das Geschlechterverhältnis, was frühere Daten aus Polen unterstützt, dass Daten das Geschlechterverhältnis arttypisch ist. nahmen wir Blutproben (Nachkommen von Waldvogelarten ist unabhängig vom Grad der Fragmentierung des Lebensraums. Dieses Ergebnis ist wichtig, um das ungleiche Geschlechterverhältnis von adulten Sumpfmeisen in abnehmenden Populationen in Großbritannien zu verstehen.