Wing pattern evolution in Heliconius butterflies provides some of the most striking examples of adaptation by natural selection. The genes controlling pattern variation are classic examples of Mendelian loci of large effect, where allelic variation causes large and discrete phenotypic changes and is responsible for both convergent and highly divergent wing pattern evolution across the genus. We characterize nucleotide variation, genotype-by-phenotype associations, linkage disequilibrium (LD), and candidate gene expression patterns across two unlinked genomic intervals that control yellow and red wing pattern variation among mimetic forms of Heliconius erato. Despite very strong natural selection on color pattern, we see neither a strong reduction in genetic diversity nor evidence for extended LD across either patterning interval. This observation highlights the extent that recombination can erase the signature of selection in natural populations and is consistent with the hypothesis that either the adaptive radiation or the alleles controlling it are quite old. However, across both patterning intervals we identified SNPs clustered in several coding regions that were strongly associated with color pattern phenotype. Interestingly, coding regions with associated SNPs were widely separated, suggesting that color pattern alleles may be composed of multiple functional sites, conforming to previous descriptions of these loci as “supergenes.” Examination of gene expression levels of genes flanking these regions in both H. erato and its co-mimic, H. melpomene, implicate a gene with high sequence similarity to a kinesin as playing a key role in modulating pattern and provides convincing evidence for parallel changes in gene regulation across co-mimetic lineages. The complex genetic architecture at these color pattern loci stands in marked contrast to the single casual mutations often identified in genetic studies of adaptation, but may be more indicative of the type of genetic changes responsible for much of the adaptive variation found in natural populations.

Recent studies indicate that relatively few genomic regions are repeatedly involved in the evolution of Heliconius butterfly wing patterns. Although this work demonstrates a number of cases where homologous loci underlie both convergent and divergent wing pattern change among different Heliconius species, it is still unclear exactly how many loci underlie pattern variation across the genus. To address this question for Heliconius erato, we created fifteen independent crosses utilizing the four most distinct color pattern races and analyzed color pattern segregation across a total of 1271 F2 and backcross offspring. Additionally, we used the most variable brood, an F2 cross between H. himera and the east Ecuadorian H. erato notabilis, to perform a quantitative genetic analysis of color pattern variation and produce a detailed map of the loci likely involved in the H. erato color pattern radiation. Using AFLP and gene based markers, we show that fewer major genes than previously envisioned control the color pattern variation in H. erato. We describe for the first time the genetic architecture of H. erato wing color pattern by assessing quantitative variation in addition to traditional linkage mapping. In particular, our data suggest three genomic intervals modulate the bulk of the observed variation in color. Furthermore, we also identify several modifier loci of moderate effect size that contribute to the quantitative wing pattern variation. Our results are consistent with the two-step model for the evolution of mimetic wing patterns in Heliconius and support a growing body of empirical data demonstrating the importance of major effect loci in adaptive change.

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Encephalomyocarditis virus (EMCV) is a picornavirus that produces lytic infections in murine and human cells. Employing a genome-wide CRISPR-Cas9 knockout screen to find host factors required for EMCV infection, we identified a role for ADAM9 in EMCV infection. CRISPR-mediated deletion of ADAM9 in multiple human cell lines rendered the cells highly resistant to EMCV infection and cell death. Primary fibroblasts from ADAM9 KO mice were also strongly resistant to EMCV infection and cell death. In contrast, ADAM9 KO and WT cells were equally susceptible to infection with other viruses, including the picornavirus Coxsackie virus B. ADAM9 KO cells failed to produce viral progeny when incubated with EMCV. However, bypassing EMCV entry into cells through delivery of viral RNA directly to the cytosol yielded infectious EMCV virions from ADAM9 KO cells, suggesting that ADAM9 is not required for EMCV replication post-entry. These findings establish that ADAM9 is required for the early stage of EMCV infection, likely for virus entry or viral genome delivery to the cytosol.IMPORTANCEViral myocarditis is a leading cause of death in the United States, contributing to numerous unexplained deaths in people ≤35 years old. Enteroviruses contribute to many cases of human myocarditis. Encephalomyocarditis virus (EMCV) infection causes viral myocarditis in rodent models, but its receptor requirements have not been fully identified. CRISPR-Cas9 screens can identify host dependency factors essential for EMCV infection and enhance our understanding of key events that follow viral infection, potentially leading to new strategies for preventing viral myocarditis. Using a CRISPR-Cas9 screen, we identifiedadisintegrinandmetalloproteinase 9 domain (ADAM9) as a major factor required for the early stages of EMCV infection in both human and murine infection.

: This report describes a method for the determination of species identity of newly hatched larvae of five sympatric Hawaiian amphidromous gobioids (Lentipes concolor, Sicyopterus stimpsoni, Awaous guamensis, Stenogobius hawaiiensis, and Eleotris sandwichensis). Polymerase chain reaction (PCR) was used to amplify a homologous section of the cytochrome b (Cyt b) region of the mitochondrial genome (mtDNA) from adults of all five species. The resulting PCR-amplified DNA was subjected to restriction fragment length polymorphism (RFLP) analysis producing species-specific restriction patterns. PCR products from the five species were sequenced to substantiate correct amplification, restriction site locations, and fragment sizes. The sequence data were also used to construct a phylogenetic tree. Individual, newly hatched, wild-caught larvae of amphidromous gobioids of unknown species affinity were sorted into six morphotypes based on physical characteristics. These typed larvae and those from two species that spawned in captivity were subjected to the same molecular analysis as the adults. The RFLP results from adults and larvae were compared, allowing larval morphotypes to be assigned to the appropriate species. These comparisons permitted construction of an identification key to the newly hatched larvae of these species based solely on physical characteristics for use in future field studies.Key Words: fish larvae, species identification, Hawaii, gobioidea, PCR, RFLP.http://link.springer-ny.com/link/service/journals/10126/bibs/1n2p167.html

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ABSTRACT:Animal studies have shown that induction of cytochrome P450 (CYP) in the lung by oxygen exposure may result in the release of free radical oxidants and arachidonic acid metabolites, which can cause lung injury that is reduced by treatment with cimetidine, a CYP inhibitor. To determine whether cimetidine would reduce lung injury in human infants at risk for chronic lung disease, we conducted a randomized clinical trial in which we administered either cimetidine or a placebo for 10 d beginning Ͻ24 h after birth to 84 newborn infants weighing Յ1250 g who were receiving O 2 and mechanical ventilation. Cimetidine had no significant effect on severity of respiratory insufficiency assessed at 10 d postnatal age. F 2 -isoprostane levels (a marker of oxidant injury) in tracheal aspirates were significantly higher in the cimetidine group at 4 d and at 10 d. There were no significant differences between the groups in tracheal aspirate levels of inflammatory markers (leukotriene B 4 , IL-8, and nucleated cell count) or arachidonic acid metabolites. We conclude that cimetidine does not reduce lung injury in newborn premature infants receiving O 2 and mechanical ventilation. It is possible that cimetidine was not an adequate CYP inhibitor in this context. (Pediatr Res 59: 795-800, 2006) C imetidine, a cytochrome P450 (CYP) inhibitor as well as an H 2 blocker, prevents the severe pulmonary gas exchange failure that occurs in newborn lambs after 72 h of breathing 95% oxygen (1). It is postulated that this beneficial effect occurs as a result of the inhibitory action of this drug on CYP metabolism in the lung, since this enzyme system, which is induced by oxygen exposure, can be a source of free radical oxidants and specific metabolites of arachidonic acid that have the potential to cause lung injury (2).There is a variety of in vivo evidence to support the concept of CYP mediated oxidant injury. For example, survival of newborn rats in 100% O 2 is significantly decreased by treatment with 3-methylcholanthrene, an inducer of the 1A1 isoform of CYP in the lung (3). Treatment with IL-1, which prolongs the survival of rats exposed to lethal concentrations of O 2 , also results in a significant decrease in pulmonary CYP, a decrease of CYP 2B1 isoform mRNA, and a significant decrease in superoxide anion generation from isolated pulmonary microsomes (4). In a rabbit model of lung reperfusion injury, CYP inhibitors (including cimetidine) markedly decrease lung edema and prevent the reperfusion-related increase in lung microvascular permeability (5). In an isolated perfused rat heart model, cimetidine and other CYP inhibitors confer a dose-dependent reduction in myocardial damage following ischemia and reperfusion, an effect attributed to suppression of reactive oxygen species production (6). Carbon tetrachloride liver injury is known to be mediated by a CYPcatalyzed reaction that produces free radicals. In rats exposed to carbon tetrachloride, inhibition of CYP has been shown to decrease hepatic lipid peroxidation (7). There a...