Several recent studies have shown that amphibian populations may exhibit high genetic subdivision in areas with recent fragmentation and urban development. Less is known about the potential for genetic differentiation in continuous habitats. We studied genetic differentiation of red-backed salamanders (Plethodon cinereus) across a 2-km transect through continuous forest in Virginia, USA. Mark-recapture studies suggest very little dispersal for this species, whereas homing experiments and post-Pleistocene range expansion both suggest greater dispersal abilities. We used six microsatellite loci to examine genetic population structure and differentiation between eight subpopulations of red-backed salamanders at distances from 200 m to 2 km. We also used several methods to extrapolate dispersal frequencies and test for sex-biased dispersal. We found small, but detectable differentiation among populations, even at distances as small as 200 m. Differentiation was closely correlated with distance and both Mantel tests and assignment tests were consistent with an isolation-by-distance model for the population. Extrapolations of intergenerational variance in spatial position (sigma(2)<15 m(2)) and pair-wise dispersal frequencies (4 Nm < 25 for plots separated by 300 m) both suggest limited gene flow. Additionally, tests for sex-biased dispersal imply that dispersal frequency is similarly low for both sexes. We suggest that these low levels of gene flow and the infrequent dispersal observed in mark-recapture studies may be reconciled with homing ability and range expansion if dispersing animals rarely succeed in breeding in saturated habitats, if dispersal is flexible depending on the availability of habitat, or if dispersal frequency varies across the geographic range of red-backed salamanders.

Roads can fragment animal populations by reducing gene flow, which can lead to drift and the loss of genetic diversity. One of the principle signatures of reduced gene flow is increased genetic differentiation in isolated populations, and evidence that roads contribute to such differentiation has been reported for several species. We used microsatellites to examine whether six roads led to increased genetic differentiation in red-backed salamanders (Plethodon cinereus). These six roads included one divided interstate highway, one undivided four-lane highway, and four secondary roads. We found that the genetic distance between plots that were bisected by the interstate highway was significantly greater than the genetic distance between equidistant plots on the same side of the highway. However, for the five smaller roads, plots across the road were no more genetically distinct than were plots on the same side of the road. Bayesian clustering methods also supported both of these findings. The optimal clustering of plots for the interstate highway consisted of two clusters that corresponded to the two sides of highway. For the other five sites, the optimal grouping consisted of a single cluster containing all of the plots. Our findings suggest that gene flow across very large roads is rare and that bisected red-backed salamander populations are likely to diverge from one another. For smaller roads, our results imply that the indirect effects of roads on genetic population structure are probably less of a pressing concern for terrestrial salamanders than are the direct effects of mortality and habitat alteration.

While many studies have examined the barrier effects of large rivers on animal dispersal and gene flow, few studies have considered the barrier effects of small streams. We used displacement experiments and analyses of genetic population structure to examine the effects of first-order and second-order streams on the dispersal of terrestrial red-backed salamanders, Plethodon cinereus (Green, 1818). We marked red-backed salamanders from near the edges of one first-order stream and one second-order stream, and experimentally displaced them either across the stream or an equal distance farther into the forest. A comparison of return rates indicated that both streams were partial barriers to salamander movement, reducing return rates by approximately 50%. Analysis of six microsatellite loci from paired plots on the same side and on opposite sides of the second-order stream suggested that the stream did contribute to genetic differentiation of salamander populations. Collectively, our results imply that low-order streams do influence patterns of movement and gene flow in red-backed salamanders. We suggest that given the high density of first-order and second-order streams in most landscapes, these features may have important effects on species that, like red-backed salamanders, have limited dispersal and large geographic ranges.

Confirmation bias is the tendency of observers to see what they expect to see while conducting scientific research. Although confirmation bias has been well‐studied by psychologists in the context of qualitative judgments, it has been much less studied with respect to the kinds of quantitative observations made by behavioral biologists. We carried out two experiments that used multiple observers of the aggression and foraging behaviors of red‐backed salamanders (Plethodon cinereus) to determine whether behavioral observations were influenced by the a priori expectations of observers. In both experiments, one group of observers was given a specific set of expectations with respect to sex differences in salamander behavior, while a second group was given the opposite set of expectations. In one experiment, observers collected data on variable sets of live salamanders, while in the other experiment, observers collected data from identical videotaped trials. Across experiments and observed behaviors, the expectations of observers did appear to bias observations, but only to a small or moderate degree. Confirmation bias never accounted for more than 13% of the observed variation in behavior, and was generally equivalent to <20% of the mean value of each variable. The estimated magnitude of confirmation bias was quite similar for men and women, suggesting no relationship between observer gender and susceptibility to confirmation bias. We believe that these results are largely optimistic with respect to confirmation bias in behavioral ecology, in that they suggest the bias may often be small relative to individual variation in behavior, even for relatively inexperienced observers.

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