In Vivo Quantification Reveals Extensive Natural Variation in Mitochondrial Form and Function in Caenorhabditis briggsae

In We have analyzed natural variation in mitochondrial form and function among a set of Caenorhabditis briggsae isolates known to harbor mitochondrial DNA structural variation in the form of a heteroplasmic nad5 gene deletion (nad5 Delta) that correlates negatively with organismal fitness. We performed in vivo quantification of 24 mitochondrial phenotypes including reactive oxygen species level, membrane potential, and aspects of organelle morphology, and observed significant amongisolate variation in 18 traits. Although several mitochondrial phenotypes were non-linearly associated with nad5D levels, most of the among-isolate phenotypic variation could be accounted for by phylogeographic clade membership. In particular, isolate-specific mitochondrial membrane potential was an excellent predictor of clade membership. We interpret this result in light of recent evidence for local adaptation to temperature in C. briggsae. Analysis of mitochondrial-nuclear hybrid strains provided support for both mtDNA and nuclear genetic variation as drivers of natural mitochondrial phenotype variation. This study demonstrates that multicellular eukaryotic species are capable of extensive natural variation in organellar phenotypes and highlights the potential of integrating evolutionary and cell biology perspectives

Boredom-like states in mink and their behavioural correlates: a replicate study

Scientists and laypeople have long expressed concern that animals in non-enriched, unchanging environments might experience boredom. However, this had attracted little empirical study: the state is difficult to assess without verbal self-reports, and spontaneous behavioural signs of boredom can vary in humans, making it hard to identify signs likely to be valid in other species. We operationally define boredom as a negative state that causes an increased, generalised interest in diverse stimuli. Previously, we demonstrated that this state existed in mink housed in non-enriched cages, compared to those in preferred, stress-reducing enriched enclosures; and that this heightened interest in stimuli positively correlated with time spent lying still but awake, while negatively correlating with locomotor stereotypic behaviour. However, these results needed replication. The current study tested for the same effects, in a new cohort of 20 male mink, by presenting 11 stimuli ranging from those predicted to typically be aversive (e.g. predator cues) to those predicted to be rewarding (e.g. food rewards; moving objects to chase). Where housing treatments differed, non-enriched mink were again more interested in the stimuli presented, spending longer oriented towards and in contact with them (e.g. for aversive stimuli: F1,9=6.27, p=0.034 and F1,9=8.24, p=0.019, respectively). Lying still but awake again correlated with interest in the stimuli (shorter latencies to contact rewarding stimuli: F1,17=3.70, p=0.036; in enriched mink only, more time oriented to and in contact with all stimuli: F1,8=9.49, p=0.015 and F1,8=15.9, p=0.004). In contrast, the previous correlations with stereotypic behaviour were not replicated. We therefore conclude that mink housed in non-enriched cages likely experience boredom-like states, and that time spent lying still while awake could potentially be used as a cage-side indicator of these states. We also suggest how future researchers might address further fundamental and practical questions about animal boredom, in mink and other species

In Vivo Quantification Reveals Extensive Natural Variation in Mitochondrial Form and Function in Caenorhabditis briggsae

We have analyzed natural variation in mitochondrial form and function among a set of Caenorhabditis briggsae isolates known to harbor mitochondrial DNA structural variation in the form of a heteroplasmic nad5 gene deletion (nad5Δ) that correlates negatively with organismal fitness. We performed in vivo quantification of 24 mitochondrial phenotypes including reactive oxygen species level, membrane potential, and aspects of organelle morphology, and observed significant among- isolate variation in 18 traits. Although several mitochondrial phenotypes were non-linearly associated with nad5Δ levels, most of the among-isolate phenotypic variation could be accounted for by phylogeographic clade membership. In particular, isolate-specific mitochondrial membrane potential was an excellent predictor of clade membership. We interpret this result in light of recent evidence for local adaptation to temperature in C. briggsae. Analysis of mitochondrial-nuclear hybrid strains provided support for both mtDNA and nuclear genetic variation as drivers of natural mitochondrial phenotype variation. This study demonstrates that multicellular eukaryotic species are capable of extensive natural variation in organellar phenotypes and highlights the potential of integrating evolutionary and cell biology perspectives

Mitonuclear hybrid strains more often resemble their mitochondrial parental isolate.

<p>Averages of maximum pharyngeal bulb fluorescence for mitochondrial (PB800 and HK105) and nuclear (AF16) parent isolates are on either side of the two hybrid strains (AFPB800 and AFHK105) (Fig. 1). Letters denote significantly different groups as determined by Tukey HSD analysis. Bars show one SEM for 15–20 independent samples.</p

Associations between mitochondrial function and morphology traits and isolate-specific <i>nad5Δ</i> level.

<p>Natural variation among <i>C. briggsae</i> isolates in (A) the total area of functional mitochondria, (B) the average area of individual non-functional mitochondria, (C) the total area of non-functional mitochondria, the (D) aspect ratio, (E) circularity, (F) circularity variance of non-functional mitochondria, in (G) relative ΔΨM, (I) the ratio of functional to non-functional organelles, and (H) relative ROS levels. Column colors corresponding to phylogenetic clade (orange = Kenya, white = Temperate, blue = Tropical), and isolates are ordered by deletion frequency along the x-axis. ED3101 and ED3092 do not experience the deletion and were assigned arbitrary x-values of −7 and −5, respectively, for this figure. Averages of maximum pharyngeal bulb fluorescence in <i>C. briggsae</i> natural isolates are plotted in relative fluorescence units (RFU). Bars represent one SEM for 15–20 independent samples.</p

Natural and experimental <i>C. briggsae</i> strains and description of the <i>nad5Δ</i> mtDNA deletion.

<p>A. Phylogenetic relationship and <i>nad5Δ</i> heteroplasmy level of <i>C. briggsae</i> isolates studied here. GL = global superclade; KE = Kenya clade; TE and TR = temperate and tropical subclades of GL; C(+) = isolates bearing compensatory Ψ<i>nad5Δ</i>-2 allele; C(-) = isolates bearing ancestral alleles. <i>nad5Δ</i> heteroplasmy categories were assigned to each <i>C. briggsae</i> natural isolate for statistical analysis following Estes et al. (2011): High = underlined font, medium = italicized, low = regular, and zero-<i>nad5Δ</i>="N/A”. Note that we assayed the natural HK104 isolate here instead of the mutation-accumulation line progenitor reported in Estes et al. (2011), which had evolved high <i>nad5Δ</i> levels in the lab (See <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0043837#s2" target="_blank">Materials and Methods</a>). B. Positions of the <i>nad5Δ</i> deletion (dashed line at top) and Ψ<i>nad5Δ</i>-2 elements in the mitochondrial genome. Primers are indicated by arrows (adapted from Howe and Denver, 2008). C. Mitochondrial and nuclear parent isolates for each mitochondrial-nuclear hybrid. <i>nad5Δ</i> heteroplasmy for each hybrid strain matches that of the maternal isolates as expected. Mitochondrial phenotypes are expected to match those of the maternal isolate if measured traits are predominantly determined by the mitochondrial genotype.</p

<em>In Vivo</em> Quantification Reveals Extensive Natural Variation in Mitochondrial Form and Function in <em>Caenorhabditis briggsae</em>

<div><p>We have analyzed natural variation in mitochondrial form and function among a set of <em>Caenorhabditis briggsae</em> isolates known to harbor mitochondrial DNA structural variation in the form of a heteroplasmic <em>nad5</em> gene deletion (<em>nad5Δ</em>) that correlates negatively with organismal fitness. We performed <em>in vivo</em> quantification of 24 mitochondrial phenotypes including reactive oxygen species level, membrane potential, and aspects of organelle morphology, and observed significant among-isolate variation in 18 traits. Although several mitochondrial phenotypes were non-linearly associated with <em>nad5Δ</em> levels, most of the among-isolate phenotypic variation could be accounted for by phylogeographic clade membership. In particular, isolate-specific mitochondrial membrane potential was an excellent predictor of clade membership. We interpret this result in light of recent evidence for local adaptation to temperature in <em>C. briggsae</em>. Analysis of mitochondrial-nuclear hybrid strains provided support for both mtDNA and nuclear genetic variation as drivers of natural mitochondrial phenotype variation. This study demonstrates that multicellular eukaryotic species are capable of extensive natural variation in organellar phenotypes and highlights the potential of integrating evolutionary and cell biology perspectives.</p> </div

Assigned labels and descriptions of all mitochondrial traits measured for <i>C. briggsae</i> natural isolates.

<p>The grand mean, F-ratio and degrees of freedom for one-way ANOVA testing for phenotypic differences among <i>C. briggsae</i> isolates. Bold font identifies the nine traits retained in the classification tree analysis when using categories based on isolate-specific <i>nad5Δ</i> % (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0043837#pone.0043837.s003" target="_blank">Table S3</a>). *, **, and *** denote p<0.05, 0.01, 0.001, respectively. Subscripts N, F, and T indicate whether the measure refers to Non-functional, Functional, or Total mitochondria. Subscript P and V denote that the measure refers to the entire mitochondrial population (not individual mitochondria), or the average individual variance in that trait, respectively.</p

Contemporary Practice as a Board-Certified Pediatric Clinical Specialist: A Practice Analysis

The purpose of the 2019 practice analysis was to identify the elements of contemporary practice as a board-certified pediatric clinical specialist. Consistent with the processes of the American Board of Physical Therapy Specialties (ABPTS), a subject matter expert panel used consensus-based processes to develop a survey to gather information concerning the knowledge areas, professional roles and responsibilities, practice expectations, and practice demographics of board-certified pediatric clinical specialists. The web-based survey was divided into 3 parts and administered to 3 separate groups of board-certified pediatric clinical specialists. Survey responses from 323 clinical specialists provided data to support confirmation and revision of the Description of Specialty Practice (DSP) for pediatrics. The revised DSP will provide contemporary practice information to inform the ABPTS specialist examination blueprint and the curricula of credentialed residency programs in pediatric physical therapy