ASU Scholarship Showcase

Introduction: Urbanization can considerably impact animal ecology, evolution, and behavior. Among the new conditions that animals experience in cities is anthropogenic noise, which can limit the sound space available for animals to communicate using acoustic signals. Some urban bird species increase their song frequencies so that they can be heard above low-frequency background city noise. However, the ability to make such song modifications may be constrained by several morphological factors, including bill gape, size, and shape, thereby limiting the degree to which certain species can vocally adapt to urban settings. We examined the relationship between song characteristics and bill morphology in a species (the house finch, Haemorhous mexicanus) where both vocal performance and bill size are known to differ between city and rural animals.

Results: We found that bills were longer and narrower in more disturbed, urban areas. We observed an increase in minimum song frequency of urban birds, and we also found that the upper frequency limit of songs decreased in direct relation to bill morphology.

Conclusions: These findings are consistent with the hypothesis that birds with longer beaks and therefore longer vocal tracts sing songs with lower maximum frequencies because longer tubes have lower-frequency resonances. Thus, for the first time, we reveal dual constraints (one biotic, one abiotic) on the song frequency range of urban animals. Urban foraging pressures may additionally interact with the acoustic environment to shape bill traits and vocal performance.

Although insulin resistance in skeletal muscle is well-characterized, the role of circulating whole blood in the metabolic syndrome phenotype is not well understood. We set out to test the hypothesis that genes involved in inflammation, insulin signaling and mitochondrial function would be altered in expression in the whole blood of individuals with metabolic syndrome. We further wanted to examine whether similar relationships that we have found previously in skeletal muscle exist in peripheral whole blood cells. All subjects (n=184) were Latino descent from the Arizona Insulin Resistance registry. Subjects were classified based on the metabolic syndrome phenotype according to the National Cholesterol Education Program’s Adult Treatment Panel III. Of the 184 Latino subjects in the study, 74 were classified with the metabolic syndrome and 110 were without. Whole blood gene expression profiling was performed using the Agilent 4x44K Whole Human Genome Microarray. Whole blood microarray analysis identified 1,432 probes that were altered in expression ≥1.2 fold and P<0.05 after Benjamini-Hochberg in the metabolic syndrome subjects. KEGG pathway analysis revealed significant enrichment for pathways including ribosome, oxidative phosphorylation and MAPK signaling (all Benjamini-Hochberg P<0.05). Whole blood mRNA expression changes observed in the microarray data were confirmed by quantitative RT-PCR. Transcription factor binding motif enrichment analysis revealed E2F1, ELK1, NF-kappaB, STAT1 and STAT3 significantly enriched after Bonferroni correction (all P<0.05). The results of the present study demonstrate that whole blood is a useful tissue for studying the metabolic syndrome and its underlying insulin resistance although the relationship between blood and skeletal muscle differs.

The probiotic effects of Lactobacillus reuteri have been speculated to partly depend on its capacity to produce the antimicrobial substance reuterin during the reduction of glycerol in the gut. In this study, the potential of this process to protect human intestinal epithelial cells against infection with Salmonella enterica serovar Typhimurium was investigated. We used a three-dimensional (3-D) organotypic model of human colonic epithelium that was previously validated and applied to study interactions between S. Typhimurium and the intestinal epithelium that lead to enteric salmonellosis. Using this model system, we show that L. reuteri protects the intestinal cells against the early stages of Salmonella infection and that this effect is significantly increased when L. reuteri is stimulated to produce reuterin from glycerol. More specifically, the reuterin-containing ferment of L. reuteri caused a reduction in Salmonella adherence and invasion (1 log unit), and intracellular survival (2 log units). In contrast, the L. reuteri ferment without reuterin stimulated growth of the intracellular Salmonella population with 1 log unit. The short-term exposure to reuterin or the reuterin-containing ferment had no observed negative impact on intestinal epithelial cell health. However, long-term exposure (24 h) induced a complete loss of cell-cell contact within the epithelial aggregates and compromised cell viability. Collectively, these results shed light on a potential role for reuterin in inhibiting Salmonella-induced intestinal infections and may support the combined application of glycerol and L. reuteri. While future in vitro and in vivo studies of reuterin on intestinal health should fine-tune our understanding of the mechanistic effects, in particular in the presence of a complex gut microbiota, this the first report of a reuterin effect on the enteric infection process in any mammalian cell type.

Extra-intestinal pathogenic E. coli (ExPEC), including avian pathogenic E. coli (APEC), pose a considerable threat to both human and animal health, with illness causing substantial economic loss. APEC strain χ7122 (O78∶K80∶H9), containing three large plasmids [pChi7122-1 (IncFIB/FIIA-FIC), pChi7122-2 (IncFII), and pChi7122-3 (IncI₂)]; and a small plasmid pChi7122-4 (ColE2-like), has been used for many years as a model strain to study the molecular mechanisms of ExPEC pathogenicity and zoonotic potential. We previously sequenced and characterized the plasmid pChi7122-1 and determined its importance in systemic APEC infection; however the roles of the other pChi7122 plasmids were still ambiguous. Herein we present the sequence of the remaining pChi7122 plasmids, confirming that pChi7122-2 and pChi7122-3 encode an ABC iron transport system (eitABCD) and a putative type IV fimbriae respectively, whereas pChi7122-4 is a cryptic plasmid. New features were also identified, including a gene cluster on pChi7122-2 that is not present in other E. coli strains but is found in Salmonella serovars and is predicted to encode the sugars catabolic pathways. In vitro evaluation of the APEC χ7122 derivative strains with the three large plasmids, either individually or in combinations, provided new insights into the role of plasmids in biofilm formation, bile and acid tolerance, and the interaction of E. coli strains with 3-D cultures of intestinal epithelial cells. In this study, we show that the nature and combinations of plasmids, as well as the background of the host strains, have an effect on these phenomena. Our data reveal new insights into the role of extra-chromosomal sequences in fitness and diversity of ExPEC in their phenotypes.

Cancer therapy selects for cancer cells resistant to treatment, a process that is fundamentally evolutionary. To what extent, however, is the evolutionary perspective employed in research on therapeutic resistance and relapse? We analyzed 6,228 papers on therapeutic resistance and/or relapse in cancers and found that the use of evolution terms in abstracts has remained at about 1% since the 1980s. However, detailed coding of 22 recent papers revealed a higher proportion of papers using evolutionary methods or evolutionary theory, although this number is still less than 10%. Despite the fact that relapse and therapeutic resistance is essentially an evolutionary process, it appears that this framework has not permeated research. This represents an unrealized opportunity for advances in research on therapeutic resistance.

Background: Urbanization can strongly impact the physiology, behavior, and fitness of animals. Conditions in cities may also promote the transmission and success of animal parasites and pathogens. However, to date, no studies have examined variation in the prevalence or severity of several distinct pathogens/parasites along a gradient of urbanization in animals or if these infections increase physiological stress in urban populations.

Methodology/Principal Findings: Here, we measured the prevalence and severity of infection with intestinal coccidians (Isospora sp.) and the canarypox virus (Avipoxvirus) along an urban-to-rural gradient in wild male house finches (Haemorhous mexicanus). In addition, we quantified an important stress indicator in animals (oxidative stress) and several axes of urbanization, including human population density and land-use patterns within a 1 km radius of each trapping site. Prevalence of poxvirus infection and severity of coccidial infection were significantly associated with the degree of urbanization, with an increase of infection in more urban areas. The degrees of infection by the two parasites were not correlated along the urban-rural gradient. Finally, levels of oxidative damage in plasma were not associated with infection or with urbanization metrics.

Conclusion/Significance: These results indicate that the physical presence of humans in cities and the associated altered urban landscape characteristics are associated with increased infections with both a virus and a gastrointestinal parasite in this common songbird resident of North American cities. Though we failed to find elevations in urban- or parasite/pathogen-mediated oxidative stress, humans may facilitate infections in these birds via bird feeders (i.e. horizontal disease transmission due to unsanitary surfaces and/or elevations in host population densities) and/or via elevations in other forms of physiological stress (e.g. corticosterone, nutritional).

Our previous studies show reduced abundance of the β-subunit of mitochondrial H+-ATP synthase (β-F1-ATPase) in skeletal muscle of obese individuals. The β-F1-ATPase forms the catalytic core of the ATP synthase, and it is critical for ATP production in muscle. The mechanism(s) impairing β-F1-ATPase metabolism in obesity, however, are not completely understood. First, we studied total muscle protein synthesis and the translation efficiency of β-F1-ATPase in obese (BMI, 36±1 kg/m²) and lean (BMI, 22±1 kg/m²) subjects. Both total protein synthesis (0.044±0.006 vs 0.066±0.006%·h^-1) and translation efficiency of β-F1-ATPase (0.0031±0.0007 vs 0.0073±0.0004) were lower in muscle from the obese subjects when compared to the lean controls (P<0.05). We then evaluated these same responses in a primary cell culture model, and tested the specific hypothesis that circulating non-esterified fatty acids (NEFA) in obesity play a role in the responses observed in humans. The findings on total protein synthesis and translation efficiency of β-F1-ATPase in primary myotubes cultured from a lean subject, and after exposure to NEFA extracted from serum of an obese subject, were similar to those obtained in humans. Among candidate microRNAs (i.e., non-coding RNAs regulating gene expression), we identified miR-127-5p in preventing the production of β-F1-ATPase. Muscle expression of miR-127-5p negatively correlated with β-F1-ATPase protein translation efficiency in humans (r = – 0.6744; P<0.01), and could be modeled in vitro by prolonged exposure of primary myotubes derived from the lean subject to NEFA extracted from the obese subject. On the other hand, locked nucleic acid inhibitor synthesized to target miR-127-5p significantly increased β-F1-ATPase translation efficiency in myotubes (0.6±0.1 vs 1.3±0.3, in control vs exposure to 50 nM inhibitor; P<0.05). Our experiments implicate circulating NEFA in obesity in suppressing muscle protein metabolism, and establish impaired β-F1-ATPase translation as an important consequence of obesity.

Strategies are needed to improve repopulation of decellularized lung scaffolds with stromal and functional epithelial cells. We demonstrate that decellularized mouse lungs recellularized in a dynamic low fluid shear suspension bioreactor, termed the rotating wall vessel (RWV), contained more cells with decreased apoptosis, increased proliferation and enhanced levels of total RNA compared to static recellularization conditions. These results were observed with two relevant mouse cell types: bone marrow-derived mesenchymal stromal (stem) cells (MSCs) and alveolar type II cells (C10). In addition, MSCs cultured in decellularized lungs under static but not bioreactor conditions formed multilayered aggregates. Gene expression and immunohistochemical analyses suggested differentiation of MSCs into collagen I-producing fibroblast-like cells in the bioreactor, indicating enhanced potential for remodeling of the decellularized scaffold matrix. In conclusion, dynamic suspension culture is promising for enhancing repopulation of decellularized lungs, and could contribute to remodeling the extracellular matrix of the scaffolds with subsequent effects on differentiation and functionality of inoculated cells.

Background: The coevolution of male traits and female mate preferences has led to the elaboration and diversification of sexually selected traits; however the mechanisms that mediate trait-preference coevolution are largely unknown. Carotenoid acquisition and accumulation are key determinants of the expression of male sexually selected carotenoid-based coloration and a primary mechanism maintaining the honest information content of these signals. Carotenoids also influence female health and reproduction in ways that may alter the costs and benefits of mate choice behaviors and thus provide a potential biochemical link between the expression of male traits and female preferences. To test this hypothesis, we manipulated the dietary carotenoid levels of captive female house finches (Carpodacus mexicanus) and assessed their mate choice behavior in response to color-manipulated male finches.

Results: Females preferred to associate with red males, but carotenoid supplementation did not influence the direction or strength of this preference. Females receiving a low-carotenoid diet were less responsive to males in general, and discrimination among the colorful males was positively linked to female plasma carotenoid levels at the beginning of the study when the diet of all birds was carotenoid-limited.

Conclusions: Although female preference for red males was not influenced by carotenoid intake, changes in mating responsiveness and discrimination linked to female carotenoid status may alter how this preference is translated into choice. The reddest males, with the most carotenoid rich plumage, tend to pair early in the breeding season. If carotenoid-related variations in female choice behavior shift the timing of pairing, then they have the potential to promote assortative mating by carotenoid status and drive the evolution of carotenoid-based male plumage coloration.

Background: Obesity is a metabolic disease caused by environmental and genetic factors. However, the epigenetic mechanisms of obesity are incompletely understood. The aim of our study was to investigate the role of skeletal muscle DNA methylation in combination with transcriptomic changes in obesity.

Results: Muscle biopsies were obtained basally from lean (n = 12; BMI = 23.4 ± 0.7 kg/m[superscript 2]) and obese (n = 10; BMI = 32.9 ± 0.7 kg/m[superscript 2]) participants in combination with euglycemic-hyperinsulinemic clamps to assess insulin sensitivity. We performed reduced representation bisulfite sequencing (RRBS) next-generation methylation and microarray analyses on DNA and RNA isolated from vastus lateralis muscle biopsies. There were 13,130 differentially methylated cytosines (DMC; uncorrected P < 0.05) that were altered in the promoter and untranslated (5' and 3'UTR) regions in the obese versus lean analysis. Microarray analysis revealed 99 probes that were significantly (corrected P < 0.05) altered. Of these, 12 genes (encompassing 22 methylation sites) demonstrated a negative relationship between gene expression and DNA methylation. Specifically, sorbin and SH3 domain containing 3 (SORBS3) which codes for the adapter protein vinexin was significantly decreased in gene expression (fold change −1.9) and had nine DMCs that were significantly increased in methylation in obesity (methylation differences ranged from 5.0 to 24.4 %). Moreover, differentially methylated region (DMR) analysis identified a region in the 5'UTR (Chr.8:22,423,530–22,423,569) of SORBS3 that was increased in methylation by 11.2 % in the obese group. The negative relationship observed between DNA methylation and gene expression for SORBS3 was validated by a site-specific sequencing approach, pyrosequencing, and qRT-PCR. Additionally, we performed transcription factor binding analysis and identified a number of transcription factors whose binding to the differentially methylated sites or region may contribute to obesity.

Conclusions: These results demonstrate that obesity alters the epigenome through DNA methylation and highlights novel transcriptomic changes in SORBS3 in skeletal muscle.