Phylogeography of a widespread small carnivore, the western spotted skunk (Spilogale gracilis) reveals temporally variable signatures of isolation across western North America.

We analyzed phylogeographic patterns in the western spotted skunk, Spilogale gracilis Merriam, 1890 (Carnivora: Mephitidae) in relation to historical events associated with Pre-Pleistocene Divergence (PPD) and Quaternary climate change (QCC) using mitochondrial DNA from 97 individuals distributed across Western North America. Divergence times were generated using BEAST to estimate when isolation in putative refugia occurred. Patterns and timing of demographic expansion was performed using Bayesian skyline plot. Putative climatic refugia resulting from Quaternary climate change were identified using paleoecological niche modeling and divergence dates compared to major vicariant events associated with Pre-Pleistocene conditions. We recovered three major mitochondrial clades corresponding to western North America (California, Baja, and across the Great Basin), east-central North America (Texas, central Mexico, New Mexico), and southwestern Arizona/northwestern Mexico. Time to most recent common ancestor for S. gracilis occurred ~1.36 Ma. Divergence times for each major clade occurred between 0.25 and 0.12 Ma, with signature of population expansion occurring 0.15 and 0.10 Ma. Ecological niche models identified three potential climatic refugia during the Last Interglacial, (1) west coast of California and Oregon, (2) northwestern Mexico, and (3) southern Texas/northeastern Mexico as well as two refugia during the Last Glacial Maximum, (1) western USA and (2) southern Texas/northeastern Mexico. This study supports PPD in shaping species-level diversity compared to QCC-driven changes at the intraspecific level for Spilogale, similar to the patterns reported for other small mammals (e.g., rodents and bats). Phylogeographic patterns also appear to have been shaped by both habitat and river vicariance, especially across the desert southwest. Further, continuing climate change during the Holocene coupled with anthropogenic modifications during the Anthropocene appears to be removing both of these barriers to current dispersal of western spotted skunks.

A minimally invasive method for gender determination in the prehensile-tailed porcupine (Coendou prehensilis).

Prehensile-tailed porcupines (Coendou prehensilis), like other rodents, lack external sexual traits, making it difficult to non-invasively determine their gender. By exploiting genetic differences between the X and the Y chromosome, we developed a simple genetic test to determine the gender of Coendous from shed quills. We Sanger sequenced a short portion (195 bp) of the zinc finger protein gene of known male (XY) Coendous to identify positions that are polymorphic between the X and Y chromosomes at this locus. By directly sequencing this fragment, we were able to correctly determine (confirmed via anatomical sexing) the gender of male and female Coendous by the presences or absence of polymorphisms in the resulting chromatograms. This assay is simple, quick and is applicable to other porcupine species.

Role for a somatically diversified lectin in resistance of an invertebrate to parasite infection.

Invertebrates lack adaptive immune systems homologous to those of vertebrates, yet it is becoming increasingly clear that they can produce diversified antigen recognition molecules. We have previously noted that the snail Biomphalaria glabrata produces a secreted lectin, fibrinogen-related protein 3 (FREP3), unusual among invertebrate defense molecules because it is somatically diversified by gene conversion and point mutation. Here we implicate FREP3 in playing a central role in resistance to a major group of snail pathogens, digenetic trematodes. FREP3 is up-regulated in three models of resistance of B. glabrata to infection with Schistosoma mansoni or Echinostoma paraensei, and functions as an opsonin favoring phagocytosis by hemocytes. Knock-down of FREP3 in resistant snails using siRNA-mediated interference resulted in increased susceptibility to E. paraensei, providing a direct link between a gastropod immune molecule and resistance to trematodes. FREP3 up-regulation is also associated with heightened responsiveness following priming with attenuated digenetic trematodes (acquired resistance) in this model invertebrate immune system.

Confirmation of Choclo virus as the cause of hantavirus cardiopulmonary syndrome and high serum antibody prevalence in Panama.

Choclo virus (CHOV) was described in sigmodontine rodents, Oligoryzomys fulvescens, and humans during an outbreak of hantavirus cardiopulmonary syndrome (HCPS) in 1999-2000 in western Panama. Although HCPS is rare, hantavirus-specific serum antibody prevalence among the general population is high suggesting that CHOV may cause many mild or asymptomatic infections. The goals of this study were to confirm the role of CHOV in HCPS and in the frequently detected serum antibody and to establish the phylogenetic relationship with other New World hantaviruses. CHOV was cultured to facilitate the sequencing of the small (S) and medium (M) segments and to perform CHOV-specific serum neutralization antibody assays. Sequences of the S and M segments found a close relationship to other Oligoryzomys-borne hantaviruses in the Americas, highly conserved terminal nucleotides, and no evidence for recombination events. The maximum likelihood and maximum parsimony analyses of complete M segment nucleotide sequences indicate a close relationship to Maporal and Laguna Negra viruses, found at the base of the South American clade. In a focus neutralization assay acute and convalescent sera from six Panamanian HCPS patients neutralized CHOV in dilutions from 1:200 to 1:6,400. In a sample of antibody-positive adults without a history of HCPS, 9 of 10 sera neutralized CHOV in dilutions ranging from 1:100 to 1:6,400. Although cross-neutralization with other sympatric hantaviruses not yet associated with human disease is possible, CHOV appears to be the causal agent for most of the mild or asymptomatic hantavirus infections, as well as HCPS, in Panama.

Pattern and timing of diversification of the mammalian order Carnivora inferred from multiple nuclear gene sequences.

The mammalian order Carnivora has attracted the attention of scientists of various disciplines for decades, leading to intense interest in defining its supra-familial relationships. In the last few years, major changes to the topological structure of the carnivoran tree have been proposed and supported by various molecular data sets, radically changing the traditional view of family composition in this order. Although a sequence of molecular studies have established a growing consensus with respect to most inter-familial relationships, no analysis so far has included all carnivoran lineages (both feliform and caniform) in an integrated data set, so as to determine comparative patterns of diversification. Moreover, no study conducted thus far has estimated divergence dates among all carnivoran families, which is an important requirement in the attempt to understand the patterns and tempo of diversification in this group. In this study, we have investigated the phylogenetic relationships among carnivoran families, and performed molecular dating analyses of the inferred nodes. We assembled a molecular supermatrix containing 14 genes (7765 bp), most of which have not been previously used in supra-familial carnivoran phylogenetics, for 50 different genera representing all carnivoran families. Analysis of this data set led to consistent and robust resolution of all supra-familial nodes in the carnivoran tree, and allowed the construction of a molecular timescale for the evolution of this mammalian order.

Nutrition and behavior of striped skunks.

Skunks are an integral part of the environment and a fascinating component of the earth's biodiversity. Their behavioral idiosyncrasies, made possible by their unique method of defense, make them entertaining to watch in the wild, and their beneficial habits far outweigh any potential negative attributes. Striped skunks can be a benefit in urban and agricultural areas because they prey on harmful and damaging insects and rodents that plague gardens and crops. The general public should be encouraged to live in harmony with skunks, as opposed to viewing them as nuisances. Keeping captive or pet skunks should be considered only after feeding, care, disease, and legal information has been investigated.

Polymorphic microsatellite markers for the striped skunk, Mephitis mephitis, and other mephitids.

We report 10 polymorphic microsatellite loci primers developed for striped skunks (Mephitis mephitis), a widespread mesocarnivore in North America. Numbers of alleles in these loci ranged from seven to 14 and the observed heterozygosity ranged from 0.76 to 1.0. These primers will be useful for studying population dynamics of skunks where rabies is endemic and will be useful to estimate genetic relatedness among females sharing winter dens. Most of these primers amplify across species within the Mephitidae.

Phylogeography of the deer mouse (Peromyscus maniculatus) provides a predictive framework for research on hantaviruses.

Phylogeographical partitioning of Sin Nombre and Monongahela viruses (hantaviruses) may reflect that of their primary rodent host, the deer mouse (Peromyscus maniculatus). Lack of a comprehensive assessment of phylogeographical variation of the host has precluded the possibility of predicting spatial limits of existing strains of these viruses or geographical regions where novel viral strains might emerge. The complete cytochrome b gene was sequenced for 206 deer mice collected from sites throughout North America to provide a foundation for future studies of spatial structure and evolution of this ubiquitous host. Bayesian analyses of these sequences partitioned deer mice into six largely allopatric lineages, some of which may represent unrecognized species. The geographical distributions of these lineages were probably shaped by Quaternary climatic events. Populations of mice were apparently restricted to refugia during glacial advances, where they experienced genetic divergence. Expansion of these populations, following climatic amelioration, brought genetically distinctive forms into contact. Occurrence of parallel changes in virus strains can now be explored in appropriate regions. In New Mexico, for example, near the location where Sin Nombre virus was first discovered, there are three genetically distinctive lineages of deer mice whose geographical ranges need to be delineated precisely. The phylogeography of P. maniculatus provides a framework for interpreting geographical variability, not only in hosts, but also in associated viral variants and disease transmission, and an opportunity to predict the potential geographical distribution of newly emerging viral strains.

Rabies in hooded and striped skunks in Arizona.

Arizona is home to four species of skunks, and rabies is enzootic in the region in which their ranges overlap. Examination of state health data from 1985 to 2004 revealed an irregular 4-10 yr periodicity in the number of cases annually, which may be related to past precipitation patterns. The number of rabid skunks peaked during springtime. Locations of rabies epizootics changed over time, but there was no evidence of a large-scale geographic spread. Skunks live-trapped during 1996-2002 had a low prevalence of rabies-virus neutralizing antibodies. This study was the first to document rabies in hooded skunks (Mephitis macroura).

Identification of skunk species submitted for rabies testing in the desert southwest.

Skunks usually are identified by their common name (skunk) when submitted for rabies testing. In the desert southwest (Arizona, New Mexico, Texas, USA; and northern Mexico), there are five species of skunks; four of which can occur in sympatry. To better understand the ecology of skunk rabies in these areas, it is imperative that species be properly identified. We used the displacement loop (d-loop) of the mitochondrial genome to identify to species 24 skunk brain samples submitted for rabies testing in New Mexico from 2001 to 2002. Most were identified as striped skunks (Mephitis mephitis), but hooded (Mephitis macroura) and hog-nosed (Conepatus leuconotus) skunks were also found.

Volatile components in defensive spray of the hooded skunk, Mephitis macroura.

GC-MS analysis of the anal sac secretion from the hooded skunk, Mephitis macroura, showed the following seven major components comprised 99% of the volatiles in this secretion: (E)-2-butene-1-thiol, 3-methyl-1-butanethiol, S-(E)-2-butenyl thioacetate, S-3-methylbutenyl thioacetate, 2-phenylethanethiol, 2-methylquinoline, and 2-quinolinemethanethiol. Minor volatile components identified in this secretion are phenylmethanethiol, S-phenylmethyl thioacetate. S-2-phenylethyl thioacetate, bis[(E)-2-butenyl] disulfide, (E)-2-butenyl 3-methylbutyl disulfide, bis(3-methylbutyl) disulfide, and S-2-quinolinemethyl thioacetate. This secretion is similar to that of the striped skunk, Mephitis mephitis, differing only in that it contains four compounds not reported from the striped skunk: phenylmethanethiol, S-phenylmethyl thioacetate, 2-phenylethanethiol, and S-2-phenylethyl thioacetate.

Natural nidality in Bolivian hemorrhagic fever and the systematics of the reservoir species.

Zoonoses within wild reservoir host populations often occur focally obeying Pavlovskii's rules of "natural nidality". What appears to be a clear example is Bolivian hemorrhagic fever (BHF), a disease endemic to northeastern Bolivia. The etiological agent is Machupo virus (MACV, Arenaviridae). The vertebrate reservoir, identified 30 years ago, was Calomys callosus a wild rodent common to open biomes in the lowlands of southeastern South America. The lack of concordance between the occurrence of MACV and the range of its rodent host has puzzled cadres of researchers and could be used as an exemplar of natural nidality. Here, we show that the populations of rodents responsible for the maintenance and transmission of MACV are an independent monophyletic lineage, different from those in other areas of South America. Therefore a clearer understanding of the systematics of the host species explains the apparent natural nidality of BHF. Similar studies may prove to be informative in other zoonoses.