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Distance-based tests for homogeneity of multivariate dispersions.

The traditional likelihood-based test for differences in multivariate dispersions is known to be sensitive to nonnormality. It is also impossible to use when the number of variables exceeds the number of observations. Many biological and ecological data sets have many variables, are highly skewed, and are zero-inflated. The traditional test and even some more robust alternatives are also unreasonable in many contexts where measures of dispersion based on a non-Euclidean dissimilarity would be more appropriate. Distance-based tests of homogeneity of multivariate dispersions, which can be based on any dissimilarity measure of choice, are proposed here. They rely on the rotational invariance of either the multivariate centroid or the spatial median to obtain measures of spread using principal coordinate axes. The tests are straightforward multivariate extensions of Levene's test, with P-values obtained either using the traditional F-distribution or using permutation of either least-squares or LAD residuals. Examples illustrate the utility of the approach, including the analysis of stabilizing selection in sparrows, biodiversity of New Zealand fish assemblages, and the response of Indonesian reef corals to an El Niño. Monte Carlo simulations from the real data sets show that the distance-based tests are robust and powerful for relevant alternative hypotheses of real differences in spread.

Animals↗

Polarized light cues underlie compass calibration in migratory songbirds.

Migratory songbirds use the geomagnetic field, stars, the Sun, and polarized light patterns to determine their migratory direction. To prevent navigational errors, it is necessary to calibrate all of these compass systems to a common reference. We show that migratory Savannah sparrows use polarized light cues from the region of sky near the horizon to recalibrate the magnetic compass at both sunrise and sunset. We suggest that skylight polarization patterns are used to derive an absolute (i.e., geographic) directional system that provides the primary calibration reference for all of the compasses of migratory songbirds.

Alaska↗

Phylogenetic analysis of Buggy Creek virus: evidence for multiple clades in the Western Great Plains, United States of America.

We present the first detailed phylogenetic analysis of Buggy Creek virus (BCRV), a poorly known alphavirus with transmission cycles involving a cimicid swallow bug (Oeciacus vicarius) vector and cliff swallows (Petrochelidon pyrrhonota) and house sparrows (Passer domesticus) as the principal avian hosts. Nucleotide sequences of a 2,075-bp viral envelope glycoprotein-coding region, covering the entire PE2 gene, were determined for 33 BCRV isolates taken from swallow bugs at cliff swallow colonies in Nebraska and Colorado in the summer of 2001 and were compared with the corresponding region of BCRV isolates collected from Oklahoma in the 1980s. We also analyzed isolates of the closely related Fort Morgan virus (FMV) collected from Colorado in the 1970s. Phylogenetic analysis indicated that BCRV falls into the western equine encephalomyelitis complex of alphaviruses, in agreement with antigenic results and a previous alphavirus phylogeny based on the E1 coding region. We found four distinct BCRV/FMV clades, one each unique to Nebraska, Colorado, and Oklahoma and one containing isolates from both Nebraska and Colorado. BCRV isolates within the two clades from Nebraska showed 5.7 to 6.2% nucleotide divergence and 0.7 to 1.9% amino acid divergence, and within these clades, we found multiple subclades. Nebraska subclades tended to be confined to one or a few cliff swallow colonies that were close to each other in space, although in some cases, near-identical isolates were detected at sites up to 123 km apart. Viral gene flow occurs when cliff swallows move (bugs) between colony sites, and the genetic structure of BCRV may reflect the limited dispersal abilities of its insect vector.

Alphavirus↗

Hypothalamic expression of thyroid hormone-activating and -inactivating enzyme genes in relation to photorefractoriness in birds and mammals.

Photorefractoriness is the insensitivity of gonadal development to the stimulatory effects of long photoperiods in birds and to the inhibitory effects of short photoperiods in small mammals. Its molecular mechanism remains unknown. Recently, it has been shown that reciprocal expression of thyroid hormone-activating enzyme [type 2 deiodinase (Dio2)] and -inactivating enzyme [type 3 deiodinase (Dio3)] genes in the mediobasal hypothalamus is critical for photoperiodically induced gonadal growth. Since thyroid hormones are required not only for photoinduction, but also for the induction of photorefractoriness, we examined the expression of these genes in relation to photorefractoriness in birds and mammals. Transfer of birds to long photoperiods induced strong expression of Dio2. This was maintained in tree sparrow when they later became photorefractory, but decreased somewhat in quail. In hamsters, transfer to long photoperiods also induced strong expression of Dio2. High values were not maintained under long photoperiods, and, indeed, expression decreased at the same rate as in animals transferred to short photoperiods. There was no renewed expression of Dio2 associated with testicular growth as animals became refractory to short photoperiods. Expression of Dio3 was high under short photoperiods and low under long photoperiods in all the animals examined, except for the short photoperiod-refractory hamsters. Our present study revealed complex regulation of deiodinase genes in the photoinduction and photorefractory processes in birds and mammals. These gene changes may be involved in the regulation of photorefractoriness, as well as photoinduction.

Animals↗

Distributed and selective auditory representation of song repertoires in the avian song system.

For many songbirds, the vocal repertoire constitutes acoustically distinct songs that are flexibly used in various behavioral contexts. To investigate how these different vocalizations are represented in the song neural system, we presented multiple song stimuli while performing extracellular recording in nucleus HVC in adult male song sparrows Melospiza melodia, a species known for its complex vocal repertoire and territorial use of song. We observed robust auditory responses to natural song stimuli in both awake and anesthetized animals. Auditory responses were selective for multiple songs of the bird's own repertoire (BOR) over acoustically modified versions of these stimuli. Selectivity was evident in both awake and anesthetized HVC, in contrast to auditory selectivity in zebra finch HVC, which is apparent only under anesthesia. Presentation of multiple song stimuli at different recording locations demonstrated that stimulus acoustic features and local neuronal tuning both contribute to auditory responsiveness. HVC auditory responsiveness was broadly distributed and nontopographic. Variance in auditory responsiveness was greater among than within HVC recording locations in both anesthetized and awake birds, in contrast to the global nature of auditory representation within zebra finch HVC. To assess the spatial consistency of auditory representation within HVC, we measured the repeatability with which ensembles of BOR songs were represented across the nucleus. Auditory response ranks to different songs were more consistent across recording locations in awake than in anesthetized animals. This spatial reliability of auditory responsiveness suggests that sound stimulus acoustic features contribute relatively more to auditory responsiveness in awake than in anesthetized animals.

Acoustic Stimulation↗

Photoperiod-independent changes in immunoreactive brain gonadotropin-releasing hormone (GnRH) in a free-living, tropical bird.

Timing of seasonal reproduction in high latitude vertebrates is generally regulated by photoperiodic cues. Increasing day length in the spring is associated with changes in the brain that are responsible for mediating reproductive activities. A primary example of this is the increased content of gonadotropin-releasing hormone (GnRH) in the preoptic area of the hypothalamus in birds as they enter the spring breeding season. Increased GnRH activity stimulates the release of luteinizing hormone and follicle-stimulating hormone from the anterior pituitary. These gonadotropins induce growth of the gonads and release of sex steroids which act on the brain to mediate reproductive behaviors. By contrast, seasonal breeding in the tropics can occur in the absence of significant changes in photoperiod. To our knowledge, no studies have investigated whether seasonal breeding in free-living tropical vertebrates is associated with seasonal changes in the GnRH system. We studied two populations of rufous-collared sparrows (Zonotrichia capensis) at the equator, separated by only 25 km, but with asynchronous reproductive phenologies associated with local climate and independent of photoperiodic cues. We collected brains and measured GnRH immunoreactivity (GnRH-ir) during each population's breeding and non-breeding periods. Breeding males had larger, but not more, GnRH-ir cells than non-breeding birds. The plasticity of the GnRH system was associated with local climate, such that the two populations exhibited asynchronous changes in GnRH-ir despite experiencing identical photoperiod conditions. Our results demonstrate that tropical birds can exhibit neural changes similar to those exhibited in higher latitude birds. However, these tropical populations appear to be using supplementary cues (e.g., rainfall, temperature, food availability) in a similar way to higher latitude species using an initial predictive cue (photoperiod). These results raise questions about the evolution of reliance upon photoperiodism and the strength of reproductive responses to other environmental cues in congeners from higher latitudes. The ability to respond to a multitude of environmental cues might be part of the ancestral condition, and the subsequent obligate photoperiodism in high-latitude congeners could reflect a loss of flexibility in response to environmental signals.

Analysis of Variance↗

Plasticity of the avian song control system in response to localized environmental cues in an equatorial songbird.

A striking feature of the vertebrate brain is its plasticity. In high-latitude vertebrates, seasonal plasticity of the brain is driven by ubiquitous photoperiod cues and therefore is highly predictable and synchronous across extensive geographic ranges. A pronounced example of seasonal brain plasticity occurs in the nuclei that regulate song behavior in songbirds. These nuclei are larger in breeding than in nonbreeding birds. In the tropics, photoperiod varies little annually, and other environmental cues important for breeding can show considerable local geographic variability. We investigated whether localized patterns of seasonal breeding in tropical birds are associated with brain plasticity. We studied two populations of rufous-collared sparrows (Zonotrichia capensis) that breed, only 25 km apart, on the equator but out of phase with each other. We measured gonadal activity and the size of song nuclei (high vocal center, robust nucleus of the arcopallium, and area X) during each population's breeding and nonbreeding periods. Breeding males had larger song nuclei and greater gonadal activity than did nonbreeding birds. This plasticity was associated with local environmental cues, such that the two populations exhibit asynchronous changes in brain structure. These results demonstrate the sensitivity of the brain and its ability to use a variety of environmental cues to coordinate seasonal plasticity and reproduction.

Animals↗

Temporal estimates of effective population size in species with overlapping generations.

The standard temporal method for estimating effective population size (N(e)) assumes that generations are discrete, but it is routinely applied to species with overlapping generations. We evaluated bias in the estimates N(e) caused by violation of this assumption, using simulated data for three model species: humans (type I survival), sparrow (type II), and barnacle (type III). We verify a previous proposal by Felsenstein that weighting individuals by reproductive value is the correct way to calculate parametric population allele frequencies, in which case the rate of change in age-structured populations conforms to that predicted by discrete-generation models. When the standard temporal method is applied to age-structured species, typical sampling regimes (sampling only newborns or adults; randomly sampling the entire population) do not yield properly weighted allele frequencies and result in biased N(e). The direction and magnitude of the bias are shown to depend on the sampling method and the species' life history. Results for populations that grow (or decline) at a constant rate paralleled those for populations of constant size. If sufficient demographic data are available and certain sampling restrictions are met, the Jorde-Ryman modification of the temporal method can be applied to any species with overlapping generations. Alternatively, spacing the temporal samples many generations apart maximizes the drift signal compared to sampling biases associated with age structure.

Adult↗

Rapid West Nile virus antigen detection.

We compared the VecTest WNV antigen assay with standard methods of West Nile virus (WNV) detection in swabs from American Crows (Corvus brachyrhynchos) and House Sparrows (Passer domesticus). The VecTest detected WNV more frequently than the plaque assay and was comparable to a TaqMan reverse transcription-polymerase chain reaction.

Animals↗

Prevalence of Bordetella bronchiseptica in certain central Iowa.

Bordetella bronchiseptica was isolated from 6 of 13 short-tailed shrews (Blarina brevicauda) and 1 of 47 house sparrows (Passer domesticus) trapped in the vicinity of a swine Bordetella rhinitis experimental area. The organism was found in four of 50 foxes (Vulpes fulva), 2 of 36 opossums (Didelphis marsupialis) and 1 of 37 raccoons (Procyon lotor) trapped in the Ames, Iowa area. This bacterium was not culturally isolated from 14 deer mice (Peromyscus maniculatus), 64 house mice (Mus Musculus), 10 masked shrews (Sorex cinereus) and 54 starlings (Sturnus vulgaris).

Animals↗

Wild bird mortality and West Nile virus surveillance: biases associated with detection, reporting, and carcass persistence.

Surveillance targeting dead wild birds, in particular American crows (Corvus brachyrhynchos), plays a critical role in West Nile virus (WNV) surveillance in the United States. Using crow decoy surrogates, detection and reporting of crow carcasses within urban and rural environments of DeKalb County, Georgia were assessed for potential biases that might occur in the county's WNV surveillance program. In each of two replicated trials, during July and September 2003, 400 decoys were labeled with reporting instructions and distributed along randomly chosen routes throughout designated urban and rural areas within DeKalb County. Information-theoretic methods were used to compare alternative models incorporating the effects of area and trial on probabilities of detection and reporting. The model with the best empirical support included the effects of area on both detection and reporting of decoys. The proportion of decoys detected in the urban area (0.605, SE=0.024) was approximately twice that of the rural area (0.293, SE=0.023), and the proportion of decoys reported in the urban area (0.273, SE=0.023) was approximately three times that of the rural area (0.103, SE=0.028). These results suggest that human density and associated factors can substantially influence dead crow detection and reporting and, thus, the perceived distribution of WNV. In a second and separate study, the persistence and fate of American crow and house sparrow (Passer domesticus) carcasses were assessed in urban and rural environments in Athens-Clarke, Madison, and Oconee counties, Georgia. Two replicated trials using 96 carcasses of each species were conducted during July and September 2004. For a portion of the carcasses, motion sensitive cameras were used to monitor scavenging species visits. Most carcasses (82%) disappeared or were decayed by the end of the 6-day study. Carcass persistence averaged 1.6 days in rural areas and 2.1 days in urban areas. We analyzed carcass persistence rates using a known-fate model framework in program MARK. Model selection based on Akaike's Information Criteria (AIC) indicated that the best model explaining carcass persistence rates included species and number of days of exposure; however, the model including area and number of days of exposure received approximately equal support. Model-averaged carcass persistence rates were higher for urban areas and for crow carcasses. Six mammalian and one avian species were documented scavenging upon carcasses. Dead wild birds could represent potential sources of oral WNV exposure to these scavenging species. Species composition of the scavenger assemblage was similar in urban and rural areas but "scavenging pressure" was greater in rural areas.

Animals↗

Mechanism of spring relapse in avian malaria: effect of gonadotropin and corticosterone.

Previous work demonstrated that corticosterone induces relapse of avian malaria, and that this effect varies markedly from winter to spring. In the present study, English sparrows with latent Plasmodium relictum infections were treated in winter with corticosterone, gonadotropin, corticosterone + gonadotropin, or a control regimen consisting of the oil vehicle. Gonadotropin neither induced relapse nor potentiated the induction of relapse by corticosterone. These data cast doubt on the hypothesis that spring relapse in malarial infections is mediated by seasonal changes in reproductive hormone levels.

Animals↗

Characterization of avipoxviruses from wild birds in Norway.

Avipoxviruses from different geographic regions of the world have been characterized to study their genetic and biological properties, but so far, no such work has been performed on Norwegian isolates. Lesions suggestive of avian pox, found on a Norwegian wild sparrow (Passer domesticus) and wood pigeon (Palumbus palumbus), were obtained in 1972 and 1996, respectively. Histologically, these lesions were demonstrated to be characteristic of poxvirus infections and the poxvirus was observed using an electron microscope. The resulting viruses were propagated in chicken embryo fibroblast cells. Restriction fragment length polymorphism of genomes from 2 Norwegian isolates and fowl pox vaccine strain, generated by BamHI, revealed a high degree of heterogeneity among the isolates. The profiles of avipoxviruses isolated from wild birds were clearly distinct from each other and also to the fowl poxvirus strain. Furthermore, chickens experimentally infected with pigeon poxvirus had higher antibody titers and extensive lesions compared to other isolates. This may suggest that pigeon poxvirus is more virulent than the other isolates.

Animals↗

Major histocompatibility alleles associated with local resistance to malaria in a passerine.

Malaria parasites are a major cause of human mortality in tropical countries and a potential threat for wildlife, as witnessed by the malaria-induced extinction of naive Hawaiian avifauna. Identifying resistance mechanisms is therefore crucial both for human health and wildlife conservation. Patterns of malaria resistance are known to be highly polygenic in both humans and mice, with marked contributions attributed to major histocompatibility (Mhc) genes. Here we show that specific Mhc variants are linked to both increased resistance and susceptibility to malaria infection in a wild passerine species, the house sparrow (Passer domesticus). In addition, links between host immunogenetics and resistance to malaria involved population-specific alleles, suggesting local adaptation in this host-parasite interaction. This is the first evidence for a population-specific genetic control of resistance to malaria in a wild species.

Alleles↗

Molecular cloning and sequencing of the gene for CDP-diglyceride synthetase of Escherichia coli.

The cds gene of Escherichia coli codes for the enzyme CDP-diglyceride synthetase. We now report the construction of plasmids which carry cds. Using these plasmids, we have sequenced 1274 base pairs of DNA, including a 750-base pair open reading frame which is the coding region of the cds gene. This DNA sequence allows the deduction of the primary peptide sequence for CDP-diglyceride synthetase. The protein is very hydrophobic, and, assuming no processing or modification, has a molecular weight of 27,570. Furthermore, there is a second open reading frame immediately after cds, implying that cds may be part of an operon. We have also constructed a runaway replication cds-plasmid that directs approximately 50-fold overproduction of CDP-diglyceride synthetase. This overproduction has been utilized in the purification of the enzyme to homogeneity, as described in the accompanying paper (Sparrow, C.P., and Raetz, C.R.H., J. Biol. Chem. 260, 12084-12091). Finally, the molecular cloning work reported herein allows the exact placement of the cds gene on the E. coli genetic map.

Amino Acid Sequence↗

Purification and properties of the membrane-bound CDP-diglyceride synthetase from Escherichia coli.

The enzyme CDP-diglyceride synthetase (CTP: phosphatidate cytidylyltransferase; EC 2.7.7.41) has been purified to 90% homogeneity from Escherichia coli cells that overproduce the enzyme 50-fold through the use of recombinant DNA technology. The purification required the use of different detergents at each step, illustrating the refractory hydrophobic nature of this protein. Apparent physical effects of EDTA on the enzyme were also utilized in the purification. The enzyme has an apparent minimum subunit mass of 27,000 daltons, as estimated by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. The amino acid composition of the protein was determined, and it correlates well with the theoretical protein product of the cds gene, the sequence of which is reported in the accompanying paper (Icho, T., Sparrow, C. P., and Raetz, C. R. H. (1985) J. Biol. Chem. 260, 12078-12083). The pure enzyme displays surface dilution kinetics when assayed in the presence of Triton X-100. As previously suggested on the basis of studies using partially purified preparations, the enzyme mechanism is sequential, and computer-calculated kinetic constants are reported herein. The substrate specificity of the enzyme is also investigated. This is the first time this enzyme has been purified to homogeneity from any source, despite the fact that it is essential for phospholipid biosynthesis in all organisms.

Amino Acids↗

Seasonal changes in the size of the avian song control nucleus HVC defined by multiple histological markers.

Bird song is controlled by a discrete network of brain nuclei. The size of several song control nuclei changes seasonally in many seasonally breeding songbird species. Reports of seasonal changes in the size of song nuclei have relied primarily on Nissl stains to define the borders of these regions. Recent studies found that the size of the song nucleus higher vocal center (HVC) in male canaries did not change seasonally when its borders were defined by histological markers other than Nissl staining. We used three labels to define the borders of the HVC in male Gambel's white-crowned sparrows (Zonotrichia leucophrys gambelii): Nissl staining, the distribution of acetylcholinesterase-positive neuropil, and the distribution of neurons projecting to another song nucleus, area X. The HVC was larger in males exposed to a breeding photoperiod and testosterone concentrations than in males exposed to a nonbreeding photoperiod and testosterone concentrations, regardless of which of these three methods was used to define the borders of the HVC. This result suggests that seasonal changes in the Nissl-defined borders of the HVC reflect changes in the distribution of physiologically relevant markers of the nucleus and are not merely artifacts of the Nissl-staining method.

Acetylcholinesterase↗

Organogenesis of the Harderian gland: a comparative survey.

Although research interest in the Harderian gland (HG) has increased during the last few years, only a small amount of information exists about its organogenesis. In mouse the HG appears in the posterior part of eye region, in the form of nonluminated tubules between the sixteenth and eighteenth days of gestation. At birth it is still not differentiated histologically. In birds the HG originates from the conjunctival epithelium at a late embryonic stage. In the English sparrow, Passer domesticus (incubation period of about 13 days), it appears between the seventh and the eighth days of incubation. In the chick embryo (incubation period of about 21 days) it originates between the eleventh and the twelfth days. Among reptiles the lizard Podarcis s. sicula has proved to be a useful model to clarify the embryological origin of the orbital glands since it possesses the anterior lacrimal gland contiguous to the HG in the medial corner of the orbit. The anlage of the orbital glands appears on about the twenty-second day of development (incubation period of about 43 days) in the form of a short tubule projecting from the conjunctival epithelium, at the time of development of the nictitating membrane. At this stage the mesenchymal cells surrounding the glandular blastema form a well-defined sac, later occupied by the orbital glands. From this stage until hatching the growth of the glandular blastema continues with the formation of acini which move posteriorly into the preformed mesenchymal sac. At the thirty-sixth day of development the more lateral acini differentiate into the HG. Only at the forty-first day do the more medial acini differentiate into the anterior lacrimal gland. At hatching the HG is fully differentiated. In anuran amphibia the primordium of the HG appears during the metamorphosis at the time of development of the nictitating membrane.

Amphibians↗