Sample Size and Geometric Morphometrics Methodology Impact the Evaluation of Morphological Variation.

Geometric morphometrics has had a profound impact on our understanding of morphological evolution. However, factors such as sample size and the views and elements selected for two-dimensional geometric morphometric (2DGM) analyses, which are often dictated by specimen availability and time rather than study design, may affect the outcomes of those analyses. Leveraging large intraspecific sample sizes (n > 70) for two bat species, Lasiurus borealis and Nycticeius humeralis, we evaluate the impact of sample size on calculations of mean shape, shape variance, and centroid size. Additionally, we assessed the concordance of multiple skull 2D views with one another and characterized morphological variation in skull shape in L. borealis and N. humeralis, as well as a closely related species, Lasiurus seminolus. Given that L. seminolus is a morphologically cryptic species with L. borealis, we assessed whether differences in skull shape and in 2DGM approach would allow species discrimination. We found that reducing sample size impacted mean shape and increased shape variance, that shape differences were not consistent across views or skull elements, and that trends shown by the views and elements were not all strongly associated with one another. Further, we found that L. borealis and L. seminolus were statistically different in shape using 2DGM in all views and elements. These results underscore the importance of selecting appropriate sample sizes, 2D views, and elements based on the hypothesis being tested. While there is likely not a generalizable sample size or 2D view that can be employed given the wide variety of research questions and systems evaluated using 2DGM, a generalizable solution to issues with 2DGM presented here is to run preliminary analyses using multiple views, elements, and sample sizes, thus ensuring robust conclusions.

A morfometria geométrica teve um impacto profundo na compreensão da evolução morfológica. No entanto, fatores como o tamanho amostral, vista anatômica e os elementos selecionados para as análises de morfometria geométricas bidimensionais (MG2D), que geralmente são determinados pela disponibilidade de espécimes e de tempo ao invés do design de estudo, podem afetar os resultados dessas análises. Utilizando grandes tamanhos de amostra intraespecífica (n > 70) em duas espécies de morcegos, Lasiurus borealis e Nycticeius humeralis, avaliamos o impacto do tamanho amostral nos cálculos da média e da variância da forma, e do tamanho do centroide. Adicionalmente, avaliamos a concordância entre vários planos 2D do crânio e caracterizamos a variação morfológica da forma cranial em L. borealis e N. humeralis, bem como em uma outra espécie proximamente relacionada, Lasiurus seminolus. Dado que L. seminolus é uma espécie morfologicamente críptica em relação a L. borealis, avaliamos se as diferenças no formato do crânio e na abordagem de MG2D utilizada permitiriam discriminar as espécies. Descobrimos que a redução do tamanho da amostra impactou o formato médio e aumentou a variância da forma, que as diferenças na forma não foram consistentes entre as vistas ou elementos cranianos, e que as tendências apresentadas pelos planos anatômicos e pelos elementos não foram fortemente associadas umas às outras. Adicionalmente, constatamos que as formas de L. borealis e L. seminolus, quantificadas por MG2D, foram estatisticamente diferentes em todas as vistas e elementos. Esses resultados enfatizam a importância em selecionar tamanhos amostrais, vistas 2D e elementos anatômicos adequados, com base na hipótese testada. Devido à grande diversidade de perguntas de pesquisa e de sistemas avaliados com MG2D, provavelmente não exista um tamanho amostral ou vista 2D que possam ser aplicados de forma generalizada. No entanto, uma solução geral para os problemas envolvendo MG2D aqui apresentados é conduzir análises preliminares utilizando várias vistas anatômicas, elementos e tamanhos amostrais, garantindo assim conclusões mais robustas.

A New Era of Morphological Investigations: Reviewing Methods for Comparative Anatomical Studies.

The increased use of imaging technology in biological research has drastically altered morphological studies in recent decades and allowed for the preservation of important collection specimens alongside detailed visualization of bony and soft-tissue structures. Despite the benefits associated with these newer imaging techniques, there remains a need for more "traditional" methods of morphological examination in many comparative studies. In this paper, we describe the costs and benefits of the various methods of visualizing, examining, and comparing morphological structures. There are significant differences not only in the costs associated with these different methods (monetary, time, equipment, and software), but also in the degree to which specimens are destroyed. We argue not for any one particular method over another in morphological studies, but instead suggest a combination of methods is useful not only for breadth of visualization, but also for the financial and time constraints often imposed on early-career research scientists.

Divergent Genital Morphologies and Female-Male Covariation in Watersnakes.

Genital evolution can be driven by diverse selective pressures. Across taxa we see evidence of covariation between males and females, as well as divergent genital morphologies between closely related species. Quantitative analyses of morphological changes in coevolving male and female genitalia have not yet been shown in vertebrates. This study uses 2D and 3D geometric morphometrics to quantitatively compare the complex shapes of vaginal pouches and hemipenes across three species of watersnakes (the sister taxa Nerodia fasciata, N. sipedon, and a close relative N. rhombifer) to address the relationship between genital morphology and divergence time in a system where sexual conflict may have driven sexually antagonistic coevolution of genital traits. Our pairwise comparisons of shape differences across species show that the sister species have male and female genitalia that are significantly different from each other, but more similar to each other than to N. rhombifer. We also determine that the main axes of shape variation are the same for males and females, with changes that relate to deeper bilobation of the vaginal pouch and hemipenes. In males, the protrusion of the region of spines at the base of the hemipene trades off with the degree of bilobation, suggesting amelioration of sexual conflict, perhaps driven by changes in the relative size of the entrance of the vaginal pouch that could have made spines less effective.

Percutaneous heel cord release for clubfoot: a retrospective, multicentre cost analysis.

PURPOSE: The Ponseti method of treatment is the standard of care for idiopathic clubfoot. Following serial casting, percutaneous tendo-Achilles tenotomy (TAT) is performed to correct residual equinus. This procedure can be performed in either the outpatient clinic or the operating room. The purpose of this study was to evaluate the expense of this procedure by examining hospital charges in both settings. METHODS: We retrospectively reviewed charts of 382 idiopathic clubfoot patients with a mean age of 2.4 months (0.6 to 26.6) treated with the Ponseti method at three institutions. Patients were divided into three groups depending on the setting for the TAT procedure: 140 patients in the outpatient clinic (CL), 219 in the operating room with discharge following the procedure (OR) and 23 in the operating room with admission to hospital for observation (OR+). Medical records were reviewed to analyze age, deformity, perioperative complications and specific time spent in each setting. Hospital charges for all three groups were standardized to one institution's charge structure. RESULTS: Charges among the three groups undergoing TAT (CL, OR, OR+) were found to be significantly different ($3840.60 versus $7962.30 versus $9110.00, respectively; p ≤ 0.001), and remained significant when separating unilateral and bilateral deformities (p < 0.001). There were nine total perioperative complications (six returns to the ER and three unexpected admissions to the hospital): five (2.3%) in the OR group, four (17.4%) in the OR+ group and none in the CL group. The OR+ group statistically had a higher rate of complications compared with the other two groups (p = 0.006). The total event time of the CL group was significantly shorter compared with the OR and OR+ groups (129.1, 171.7 and 1571.6 minutes respectively; p < 0.001). CONCLUSION: Hospital charges and total event time were significantly less when percutaneous TAT was performed in the outpatient clinic compared with the operating room. In addition, performing the procedure in clinic was associated with the lowest rate of complications. LEVEL OF EVIDENCE: Therapeutic, Level III.

An Injured Psittacosaurus (Dinosauria: Ceratopsia) From the Yixian Formation (Liaoning, China): Implications for Psittacosaurus Biology.

We describe a Psittacosaurus specimen from the Lujiatun beds of the Yixian Formation in Liaoning, China with an abnormality on its left fibula. Although a large number of Psittacosaurus specimens are known, only a single example of a pathologic Psittacosaurus has been previously noted. The specific pathology in the current specimen is believed to be a healed fibular fracture as assessed through a combination of gross morphology, microcomputed tomography (microCT), and histology data. The fracture can be identified using microCT, but the degree of remodeling and the stage of fracture repair are best determined histologically. The fracture callus is made up of radially oriented spokes of woven bone in a cartilage matrix and the original cortical bone prior to the fracture has been largely eroded. A transverse histologic section taken at the level of the fracture shows the displacement of the proximal and distal parts of the fibula. The Psittacosaurus appears to have survived the break considering the deposition of circumferential non-pathologic bone at the periosteal surface outside of the callus. The combination of gross morphological description, microCT data, and histologic data allowed for a full diagnosis of the abnormality. While some previous authors have preferred gross morphological description above other methods for assessing paleopathologies, it is evident based on this specimen that an amalgam of techniques provides greater clarity to paleopathology diagnoses. Although this Psittacosaurus lived in an environment with many predators, it was able to survive with a fracture on its hindlimb, which undoubtedly would have impacted its locomotion. Anat Rec, 299:897-906, 2016. © 2016 Wiley Periodicals, Inc.

Safe and drug-free school coordinators' perceived needs to improve violence and drug prevention programs.

A random sample of 380 Ohio safe and drug-free school coordinators was mailed surveys regarding their perceived needs to improve safe and drug-free school programs. Of 298 who responded (78%), most were White (88%), female (71%), older than age 40 (72%), possessed a master's degree (56%), and worked at either a rural (43%) or suburban (40%) school. Training topics perceived as most beneficial in improving safe and drug-free school programs included training on effective techniques that enhance youth success, building school-parent-community partnerships, and providing conflict management/dispute resolution skills. Lack of time, poor parental attitudes, and lack of school personnel support were identified most often as barriers to program improvement. Findings can be used by state health education resource centers and training providers to more effectively meet the needs of safe and drug-free school coordinators.

Descriptive epidemiology of physical activity in university graduates with locomotor disabilities.

The descriptive epidemiology of physical activity in a sample of 577 University of Illinois graduates (1952-1991) with locomotor disabilities was assessed by mail survey. The survey requested basic demographic information, age, gender, marital status, household income. Respondents were asked to rate their current activity levels and activity levels during their college years compared to others their age on a 5 point scale: (1) much less active to (5) much more active. Completed surveys were received from 229 alumni (40%); 59 semi-ambulatory, 115 paraplegic, 55 quadriplegic. Results indicated current physical activity was associated with mobility limitation. With more severe mobility limitations the percentage reporting being less/much less active increased (42.4% semi-ambulatory, 56.5% paraplegic, 66.7% quadriplegic, P < 0.001) and the percentage reporting being more active decreased (20.3% semi-ambulatory, 16.5% paraplegic, 13.0% quadriplegic, P < 0.001). Current physical activity was significantly lower (P < 0.05) with increasing age, lower self-rated health, higher disability severity and among those who were sedentary during college. Physical activity did not differ by gender, marital status or household income. Multiple regression analysis indicated that health status was a significant predictor of current physical activity in all mobility categories (P < 0.001) after controlling for age, gender, income, disability severity and college activity. Among both paraplegics and quadriplegics physical activity during college was significantly associated (P < 0.001 paraplegic; P < 0.01 quadriplegic) with current physical activity. These results document a low level of physical activity in a well-educated sample of individuals with locomotor disabilities and suggest that exposure to physical activity in an educational setting may be an effective technique for increasing physical activity in individuals with locomotor disabilities.

Two forms of striatal tyrosine hydroxylase from DEAE-cellulose chromatography.

Tyrosine hydroxylase (TH) in freshly prepared 45,000 g supernatant from rat striatum was fractionated by DEAE-cellulose chromatography. The elution was made with 2 vols. of buffer (50 mM Tris, pH 7.4; 2 mM dithiothreitol) followed by 4 vols. of a linear NaCl gradient (0 0.3 M) in the same buffer. TH activity was eluted in two distinct peaks: one at about 0.1 M salt (I), and the other at 0.2 M salt (II). The relationship between the two enzymes peaks was examined as follows. (1) Incubation of the supernatant in the presence of cAMP-dependent protein kinase, 1 mM ATP, 10 mM Mg2+, and 0.1 mM cAMP resulted in the elimination of peak I, with a concomitant increase of peak II. This shift of TH peaks was prevented when the protein kinase was blocked by the addition of its inhibitory modulator. (2) Incubation of the supernatant with alkaline phosphatase, an enzyme known to dephosphorylate a variety of phosphoproteins, resulted in the elimination of peak II, with a concomitant increase of peak I. (3) Only freshly prepared supernatants showed two distinct TH peaks from DEAE-cellulose. From supernatants held at 0 degrees C for 24 h. peak II was markedly reduced and peak I concomitantly increased. Since peak II appears to be readily convertible to peak I, no further fractionation was attempted. From the data obtained here, we believe that peaks I and II are respectively the nonphosphorylated and phosphorylated forms of TH. Furthermore, the endogenous distribution of the two TH forms in striatum was altered by the administration of haloperidol (2 mg/kg. i.p.), a neuroleptic drug known to activate the enzyme via a cAMP-dependent mechanism. At 90 min after the treatment, there was a marked increase of peak II, with a concomitant decrease of peak I. Thus, this procedure provides a simple means for estimating the degree of phosphorylation of TH in vivo in catecholaminergic neurons under various physiological and pharmacological conditions.

Effects of dexamethasone and other glucocorticoid steroids on tyrosine hydroxylase activity in the superior cervical ganglion.

Dexamethasone is known to elicit an increase of tyrosine hydroxylase activity in the superior cervical ganglion. The details of such a glucocorticoid effect were investigated in the present study. Of 4 glucocorticoids (dexamethasone, corticosterone, hydrocortisone and triamcinolone) examined in rats, only the synthetic steroid dexamethasone was found to be effective in increasing ganglionic tyrosine hydroxylase activity (by 50% at 48 h after drug administration). Corticosterone even at doses as high as 50 mg/kg failed to show an effect. Since recent reports indicate that a cytoplasmic glucocorticoid receptor is not present in the sympathetic ganglion, it is unlikely that the dexamethasone effect involves a receptor-mediated mechanism. Moreover, the dexamethasone effect was totally blocked by chlorisondamine, a nicotinic cholinergic receptor antagonist. The possibility of an enhanced impulse flow from the CNS, however, was excluded by the finding that decentralization immediately prior to dexamethasone administration did not prevent the increase of ganglionic tyrosine hydroxylase activity, although earlier decentralization (24 h or longer) abolished the steroid effect. Significantly, in the freshly decentralized ganglia, the increase of tyrosine hydroxylase activity by dexamethasone was still blocked by chlorisondamine. Since synaptic activity in terminals is known to continue for a brief period following nerve transection, our data support the contention that the primary site of the dexamethasone effect may be the preganglionic terminals.