Remnant populations of the regal fritillary (Speyeria idalia) in Pennsylvania: Local genetic structure in a high gene flow species

The Regal Fritillary butterfly, Speyeria idalia (Drury) (Lepidoptera: Nymphalidae), has been described as a high gene flow species. Supporting this assertion, previous studies in the Great Plains, where it is still relatively widespread, have found evidence of gene flow across hundreds of kilometers. Using mitochondrial and microsatellite loci, we examined the spatial genetic structure of a very isolated Pennsylvania population of these butterflies that occupies three separate meadows located within ten kilometers of each other. We found restricted gene flow and a distinct structure, with each meadow having a unique genetic signature. Our findings indicate that even a species that normally exhibits high gene flow may show fine-scale genetic subdivision in areas where populations have been largely extirpated.Authors contributed equally.     

The Cryptococcus neoformans catalase gene family and its role in antioxidant defense

In the present study, we sought to elucidate the contribution of the Cryptococcus neoformans catalase gene family to antioxidant defense. We employed bioinformatics techniques to identify four members of the C. neoformans catalase gene family and created mutants lacking single or multiple catalase genes. Based on a phylogenetic analysis, CAT1 and CAT3 encode putative spore-specific catalases, CAT2 encodes a putative peroxisomal catalase, and CAT4 encodes a putative cytosolic catalase. Only Cat1 exhibited detectable biochemical activity in vitro, and Cat1 activity was constitutive in the yeast form of this organism. Although they were predicted to be important in spores, neither CAT1 nor CAT3 was essential for mating or spore viability. Consistent with previous studies of Saccharomyces cerevisiae, the single (cat1, cat2, cat3, and cat4) and quadruple (cat1 cat2 cat3 cat4) catalase mutant strains exhibited no oxidative-stress phenotypes under conditions in which either exogenous or endogenous levels of reactive oxygen species were elevated. In addition, there were no significant differences in the mean times to mortality between groups of mice infected with C. neoformans catalase mutant strains (the cat1 and cat1 cat2 cat3 cat4 mutants) and those infected with wild-type strain H99. We conclude from the results of this study that C. neoformans possesses a robust antioxidant system, composed of functionally overlapping and compensatory components that provide protection against endogenous and exogenous oxidative stresses.     

Feasibility of using the national hospital discharge survey to estimate the prevalence of selected birth defects

BACKGROUND: Nationally representative data on the prevalence of certain birth defects are largely unavailable. We evaluated the feasibility of using data from the National Hospital Discharge Survey (NHDS) to describe the prevalence of selected birth defects. METHODS: All live births recorded in the NHDS during 1999-2001 were included. The prevalence for selected birth defects was calculated using weighted ratio estimators. Prevalence ratios comparing the NHDS estimates to published national estimates from the National Birth Defects Prevention Network (NBDPN) were calculated. RESULTS: With the exception of common truncus, the NHDS prevalence for the selected defects was consistently lower than the NBDPN estimates. The prevalence ratios ranged from 0.38 for trisomy 18 and anopthalmia/micropthalmia to 1.16 for common truncus. The NHDS prevalence estimates for spina bifida without anencephaly (PR 0.89, 95% CI: 0.57-1.22) and gastroschisis/omphalocele (PR 0.94, 95% CF: 0.48-1.40) most closely approximated the NBDPN estimates. CONCLUSIONS: NHDS data underestimate the prevalence of most birth defects. Additional research is needed to determine whether NHDS estimates may be useful for evaluating trends in certain conditions. Surveillance systems employing active case-finding continue to provide more accurate estimates of birth defects prevalence.     

Temporal gene induction patterns in sheepshead minnows exposed to 17beta-estradiol

Gene arrays provide a powerful method to examine changes in gene expression in fish due to chemical exposures in the environment. In this study, we expanded an existing gene array for sheepshead minnows (Cyprinodon variegatus) (SHM) and used it to examine temporal changes in gene expression for male SHM exposed to 100 ng 17beta-estradiol (E(2))/L for five time points between 0 and 48 hr. We found that in addition to the induction of genes involved in oocyte development (vitellogenin [VTG], zona radiata [ZRP]), other genes involved in metabolism and the inflammatory response are also affected. We identified five patterns of temporal induction in genes whose expression was modified due to E(2) exposure. We validated the gene array data for the expression of VTG 1, VTG 2, ZRP 2 and ZRP 3 and found that with low levels of exogenous E(2) (100 ng E(2)/L) exposure, ZRP expression precedes VTG expression. However, at higher concentrations of E(2) (500 ng E(2)/L), the difference in temporal expression appears to be lost. Exposure to high levels of environmental contaminants may affect the normal ordered expression of genes required for reproduction. Gene expression profiling using arrays promises to be a valuable tool in the field of environmental toxicology. As more genes are identified for species used in toxicological testing, researchers will be better able to predict adverse effects to chemical exposures and to understand the relationships between changes in gene expression and changes in phenotype.     

A visual thalamocortical slice

We describe a thalamocortical slice preparation in which connectivity between the mouse lateral geniculate nucleus (LGN) and primary visual cortex (V1) is preserved. Through DiI injections in fixed brains we traced and created a three-dimensional model of the mouse visual pathways. From this computer model we designed a slice preparation that contains a projection from LGN to V1. We prepared brain slices with these predicted coordinates and demonstrated anatomical LGN-V1 connectivity in these slices after LGN tracer injections. We also revealed functional LGN-V1 connectivity by stimulating LGN electrically and detecting responses in layer 4 of V1 using calcium imaging, field potential recordings and whole-cell recordings. We also identified layer-4 neurons that receive direct thalamocortical input. Finally, we compared cortical activity after LGN stimulation with spontaneous cortical activity and found significant overlap of the spatiotemporal dynamics generated by both types of events.     

Two-stage case–control designs for rare genetic variants

The search for the association of rare genetic variants with common diseases is of high interest, yet challenging because of cost considerations. We present an efficient two-stage design that uses diseased cases to first screen for rare variants at stage-1. If too few cases are found to carry any variants, the study stops. Otherwise, the selected variants are screened at stage-2 in a larger set of cases and controls, and the frequency of variants is compared between cases and controls by an exact test that corrects for the stage-1 ascertainment. Simulations show that our new method provides conservative Type-I error rates, similar to the conservative aspect of Fisher’s exact test. We show that the probability of stopping at stage-1 increases with a smaller number of cases screened at stage-1, a larger stage-1 continuation threshold, or a smaller carrier probability. Our simulations also show how these factors impact the power at stage-2. To balance stopping early when there are few variant carriers versus continuation to stage-2 when the variants have a reasonable effect size on the phenotype, we provide guidance on designing an optimal study that minimizes the expected sample size when the null hypothesis is true, yet achieves the desired power.     

Potential for the Anopheles gambiae Densonucleosis Virus To Act as an “Evolution-Proof” Biopesticide

“Evolution-proof” or “late-life-acting” insecticides (LLAIs) preferentially kill older adult mosquitoes and are of extreme interest to control vector-borne diseases such as malaria. We used quantitative PCR to assess whether the Anopheles gambiae densonucleosis virus (AgDNV) had potential as an LLAI. After infection, AgDNV titers increased modestly during larval development but replicated slower than the host cells, resulting in a significant decrease in the normalized virus titer during larval and pupal development. Normalized virus titers dramatically increased after adult emergence, peaking in 7- to 10-day-old adults. Unlike other DNVs, AgDNV does not significantly replicate in preadult mosquitoes but rather preferentially replicates in older adults. The natural dynamics of AgDNV make it ideal for expression of insect-specific toxin genes as a biological LLAI.Malaria infects several hundred million people and results in over one million deaths annually. Vector control is a major component of current malaria control strategies. However, the evolution of insecticide resistance by Anopheles mosquito vectors has severely hampered control efforts (5, 13, 14). Although novel chemistries are being explored, new insecticides will face similar problems with resistance evolution. Recently, late-life-acting insecticides (LLAIs) have been proposed as novel agents to control vector-borne diseases such as malaria (1, 10). LLAIs selectively kill the older mosquitoes responsible for the bulk of parasite transmission while allowing for reproduction of the younger age classes that contribute to the bulk of evolutionary fitness, i.e., there is an optimal window of time wherein mosquitoes live long enough to reproduce but not long enough to transmit pathogens (8, 9). Reproduction allows for relaxation of evolutionary pressures that select for resistance to the agent. If resistance alleles exert fitness costs, there are theoretical scenarios under which resistance is not expected to evolve, leading some to provocatively term LLAIs as “evolution-proof” (1, 10).LLAI do not have to be conventional chemical pesticides. Like all organisms, mosquitoes can be infected with pathogens of their own (including fungi, bacteria, or viruses) that can be exploited to shorten mosquito life span to control disease (6, 9-12). Some pathogens can be vertically transmitted (9, 11), which not only results in relaxed selection pressure but also allows for their transmission and spread into the vector population.Densonucleosis viruses (or densoviruses [DNVs]) are icosahedral, nonenveloped parvoviruses that have been identified from many invertebrate taxa, including multiple mosquito species (2, 11). Naturally occurring DNVs typically infect mosquitoes during the aquatic larval phase. Infection of young larvae (first or second instar) is generally lethal, resulting in virus amplification and release into the larval environment. Larvae infected at later time points (third or fourth instar) develop into infected adults that inoculate virus vertically and horizontally into the larval environment during oviposition, completing the virus life cycle (2, 11).The Aedes aegypti densovirus (AeDNV) is generally lethal to Ae. aegypti larvae in a dose-dependent manner, and high virus titers in larvae are observed both by quantitative PCR (qPCR) and by expression of foreign transgenes such as green fluorescent protein (GFP) (2, 8, 15). In contrast, the Anopheles gambiae densovirus (AgDNV) is not lethal to An. gambiae larvae (11). Using GFP-transducing virus and epifluorescence microscopy, we have also never seen GFP expression in larvae, pupae, or young adults; GFP is not observable until the adults are ∼1 week postemergence (J. L. Rasgon and X. Ren, unpublished observations) (Fig. ​(Fig.1).1). Based on these observations, we hypothesized that AgDNV has different replication kinetics in An. gambiae than AeDNV has in Ae. aegypti, remaining at relatively low titers in the immature life stages but replicating to high titers after adult emergence.Open in a separate windowFIG. 1.Epifluorescent image (dorsal view) of a 10-day-old adult An. gambiae female infected with GFP-expressing AgDNV (as described in reference 11). GFP is not visible in infected mosquitoes until 7 to 10 days postemergence.