Chromosome-length genome assemblies and cytogenomic analyses of pangolins reveal remarkable chromosome counts and plasticity.

We report the first chromosome-length genome assemblies for three species in the mammalian order Pholidota: the white-bellied, Chinese, and Sunda pangolins. Surprisingly, we observe extraordinary karyotypic plasticity within this order and, in female white-bellied pangolins, the largest number of chromosomes reported in a Laurasiatherian mammal: 2n = 114. We perform the first karyotype analysis of an African pangolin and report a Y-autosome fusion in white-bellied pangolins, resulting in 2n = 113 for males. We employ a novel strategy to confirm the fusion and identify the autosome involved by finding the pseudoautosomal region (PAR) in the female genome assembly and analyzing the 3D contact frequency between PAR sequences and the rest of the genome in male and female white-bellied pangolins. Analyses of genetic variability show that white-bellied pangolins have intermediate levels of genome-wide heterozygosity relative to Chinese and Sunda pangolins, consistent with two moderate declines of historical effective population size. Our results reveal a remarkable feature of pangolin genome biology and highlight the need for further studies of these unique and endangered mammals.

Selective­logging and oil palm: multitaxon impacts, biodiversity indicators, and trade­offs for conservation planning.

Strong global demand for tropical timber and agricultural products has driven large-scale logging and subsequent conversion of tropical forests. Given that the majority of tropical landscapes have been or will likely be logged, the protection of biodiversity within tropical forests thus depends on whether species can persist in these economically exploited lands, and if species cannot persist, whether we can protect enough primary forest from logging and conversion. However, our knowledge of the impact of logging and conversion on biodiversity is limited to a few taxa, often sampled in different locations with complex land-use histories, hampering attempts to plan cost-effective conservation strategies and to draw conclusions across taxa. Spanning a land-use gradient of primary forest, once- and twice-logged forests, and oil palm plantations, we used traditional sampling and DNA metabarcoding to compile an extensive data set in Sabah, Malaysian Borneo for nine vertebrate and invertebrate taxa to quantify the biological impacts of logging and oil palm, develop cost-effective methods of protecting biodiversity, and examine whether there is congruence in response among taxa. Logged forests retained high species richness, including, on average, 70% of species found in primary forest. In contrast, conversion to oil palm dramatically reduces species richness, with significantly fewer primary-forest species than found on logged forest transects for seven taxa. Using a systematic conservation planning analysis, we show that efficient protection of primary-forest species is achieved with land portfolios that include a large proportion of logged-forest plots. Protecting logged forests is thus a cost-effective method of protecting an ecologically and taxonomically diverse range of species, particularly when conservation budgets are limited. Six indicator groups (birds, leaf-litter ants, beetles, aerial hymenopterans, flies, and true bugs) proved to be consistently good predictors of the response of the other taxa to logging and oil palm. Our results confidently establish the high conservation value of logged forests and the low value of oil palm. Cross-taxon congruence in responses to disturbance also suggests that the practice of focusing on key indicator taxa yields important information of general biodiversity in studies of logging and oil palm.

Gliding saves time but not energy in Malayan colugos.

Gliding is thought to be an economical form of locomotion. However, few data on the climbing and gliding of free-ranging gliding mammals are available. This study employed an animal-borne three-dimensional acceleration data-logging system to collect continuous data on the climbing and gliding of free-ranging Malayan colugos, Galeopterus variegatus. We combined these movement data with empirical estimates of the metabolic costs to move horizontally or vertically to test this long-standing hypothesis by determining whether the metabolic cost to climb to sufficient height to glide a given distance was less than the cost to move an equivalent distance horizontally through the canopy. On average, colugos climb a short distance to initiate glides. However, due to the high energetic cost of climbing, gliding is more energetically costly to move a given horizontal distance than would be predicted for an animal travelling the same distance through the canopy. Furthermore, because colugos spend a small fraction of their time engaged in locomotor activity, the high costs have little effect on their overall energy budget. As a result, the energetic economy hypothesis for the origins of gliding is not supported. It is likely that other ecologically relevant factors have played a greater role in the origins of gliding in colugos and other mammals.

Vertebrate Scavengers Control Abundance of Diarrhea-causing Bacteria in Tropical Plantations.

Scavenging is a common phenomenon, particularly amongst carnivorous vertebrates. By consuming carrion, vertebrate scavengers reduce resource availability for both pathogenic bacteria and their insect vectors. We investigated the ability of wild vertebrate scavengers to control agents of human diarrheal diseases (specifically Salmonella spp. and Shiga toxin-producing Escherichia coli [STEC]) in oil palm plantations in Sabah (East Malaysia), and the existence of spillover effect whereby additional vertebrate scavengers from adjacent forest patches result in greater disease control in plantation sections near these forest edges. Experimental carcasses were removed by common scavengers (Varanus salvator, Canis lupus familiaris, and Viverra tangalunga) at different time points, and this determined the length of time that the carcasses persisted in the environment. The amount of pathogenic bacteria on the surfaces of filth flies collected above the experimental carcasses was positively correlated to the duration of carcass persistence, and reduction in pathogenic bacterial abundances was largely due to carcass consumption by these vertebrate scavengers. Instead of a predicted positive spillover effect (greater scavenger activity near forest edges, hence reduced pathogen abundance), we detected a weak inverse spillover effect in which STEC counts were marginally higher in plantation sections near forest patches, and human hunting along the forest-plantation boundaries could explain this. We propose that making oil palm plantations scavenger-friendly could yield great human health benefits for the millions of workers employed in this rapidly-expanding industry, without drastically changing current management practices.

Take-off and landing kinetics of a free-ranging gliding mammal, the Malayan colugo (Galeopterus variegatus).

Arboreal animals negotiate a highly three-dimensional world that is discontinuous on many spatial scales. As the scale of substrate discontinuity increases, many arboreal animals rely on leaping or gliding locomotion between distant supports. In order to successfully move through their habitat, gliding animals must actively modulate both propulsive and aerodynamic forces. Here we examined the take-off and landing kinetics of a free-ranging gliding mammal, the Malayan colugo (Galeopterus variegatus) using a custom-designed three-dimensional accelerometry system. We found that colugos increase the propulsive impulse to affect longer glides. However, we also found that landing forces are negatively associated with glide distance. Landing forces decrease rapidly as glide distance increases from the shortest glides, then level off, suggesting that the ability to reorient the aerodynamic forces prior to landing is an important mechanism to reduce velocity and thus landing forces. This ability to substantially alter the aerodynamic forces acting on the patagial wing in order to reorient the body is a key to the transition between leaping and gliding and allows gliding mammals to travel long distances between trees with reduced risk of injury. Longer glides may increase the access to distributed resources and reduce the exposure to predators in the canopy or on the forest floor.

Genomic analysis reveals hidden biodiversity within colugos, the sister group to primates.

Colugos are among the most poorly studied mammals despite their centrality to resolving supraordinal primate relationships. Two described species of these gliding mammals are the sole living members of the order Dermoptera, distributed throughout Southeast Asia. We generated a draft genome sequence for a Sunda colugo and a Philippine colugo reference alignment, and used these to identify colugo-specific genetic changes that were enriched in sensory and musculoskeletal-related genes that likely underlie their nocturnal and gliding adaptations. Phylogenomic analysis and catalogs of rare genomic changes overwhelmingly support the contested hypothesis that colugos are the sister group to primates (Primatomorpha), to the exclusion of treeshrews. We captured ~140 kb of orthologous sequence data from colugo museum specimens sampled across their range and identified large genetic differences between many geographically isolated populations that may result in a >300% increase in the number of recognized colugo species. Our results identify conservation units to mitigate future losses of this enigmatic mammalian order.

Fracture toughness of human femoral neck: effect of microstructure, composition, and age.

The effects of porosity and pore size; osteonal area, size, and density; mineral content; water content; wet and dry apparent densities; and age on mode I (tensile) and mode II (shear) strain energy release rate were investigated for femoral neck cortical bone from human cadavers aged >/=50 years. The results suggest that porosity- and density-based parameters that are related to bone quantity are more consistently determinant for femoral neck fracture toughness than morphology-based parameters that are related to microstructural organization. Bone features examined here were more explanatory for shear than tension fracture toughness. Tension and shear fracture toughness did not change with age, unlike in previous reports investigating the femoral and tibial shaft. It was concluded that the femoral neck is different from the femoral and tibial shaft in terms of its microstructure and composition and in its relationship of fracture toughness to its constituents and age.

Microdamage of human cortical bone: incidence and morphology in long bones.

Increased bone fragility and osteoporotic fracture in human bone has been attributed to the accumulation of microdamage. According to Martin and Burr (J Biomech 15:137-139; 1982), microcracks from interstitial bone propagate to the cement line or concentric lamellae and debond or separate the Haversian canal from the surrounding bone which leads to repair of the damaged region. If this is true, we would expect to find a greater incidence of microdamage existing at the cement line than at other locations within the bone microstructure. The incidence of such an occurrence, however, is not established. The purpose of this investigation was to determine the incidence and morphology of microcracks in human cortical bone from the midshaft of the tibia and proximal femur. We investigated the hypothesis that osteons arrest and trap microcracks in bone. We were also interested in determining if relationships exist between microdamage and bone type (tibia or femur), cortex location (anterior, posterior, medial, and lateral), gender, and donor age. It was found that 62.4% of all microcracks run between the surrounding interstitial bone and the cement line supporting the hypothesis by Martin and Burr. It was also found that microdamage increased with age, was significantly greater in females than males, and was significantly greater in the proximal femur than the midshaft of the tibia.

Influence of bone composition and apparent density on fracture toughness of the human femur and tibia.

The influence of wet and dry apparent density, apparent and real percentage of mineral, organic and water contents on the tension and shear fracture toughness, i.e., the mode I and mode II strain energy release rate (GIc and GIIc), respectively, was investigated for the human femur and the tibia. The results suggest that the water content, apparent density, and age were the best explanatory parameters for GIc and GIIc. Both GIc and GIIc significantly increase with increasing wet or dry apparent density. They also decrease with increasing water content; the decrease is nonsignificant for GIIc in the femur only. Mineral and organic percentage did not change in the bone with age, while the apparent percentages did change. Compositional parameters altogether can explain 35%-59% of the variation in fracture toughness. We conclude that bone composition and density have an important influence on fracture resistance of the cortical bone.

The influence of bone morphology on fracture toughness of the human femur and tibia.

The influence of porosity, osteon density, osteonal area, osteonal lamellar area, osteon size, and haversian canal size on the tension and shear fracture toughness, that is, the mode I and mode II strain energy release rate (GIc and GIIc), respectively, were investigated for the human femur and the tibia. The results suggest that porosity and osteon density were the best explanatory morphological parameters for GIc and GIIc. Both GIc and GIIc significantly decrease with increasing porosity. They also increase with increasing osteon density, the increase being significant for the femur only. Morphological parameters, altogether, can explain 49%-68% of the variation in fracture toughness. We concluded that, although there must be other factors such as biochemical components and microdamage, osteon morphology has an important influence on fracture resistance of the cortical bone.

Influence of microdamage on fracture toughness of the human femur and tibia.

The relationship between microdamage accumulation and bone fragility is not well understood. Previous work has demonstrated a positive relationship between microdamage and age in human cortical bone. Prior investigations have also demonstrated that fracture toughness decreases with age in the same bone. Based on these findings, the objective of this study was to test the hypothesis that a decrease in fracture toughness can be attributed to an increase in microdamage density. Microdamage parameters (density, surface density, and average crack length) were measured from bone taken from the shaft of the human femur and tibia and correlated with results from fracture toughness tests of the same bone. Results indicated that there was a weak but significant inverse relationship between fracture toughness and microdamage parameters for tension loading of the femur. These findings suggest that in vivo microdamage observable at the transmitted light level (100x) plays a secondary role to other contributory factors to decreased fracture toughness with age. Results also indicate that this relationship depends on bone ductility that apparently differs between the femur and the tibia. This study, in addition to prior investigations, suggests that crack size (microscopic vs. submicroscopic) and crack origin or type (in vivo vs. stress induced de novo) may influence the relationship between microdamage and bone toughness.

Nerve growth factor gene and hypertension in spontaneously hypertensive rats.

OBJECTIVE: High blood pressure in spontaneously hypertensive rat (SHR) is associated with increased sympathetic innervation of key tissues, possibly as the result of increased nerve growth factor (NGF). The aim of this study was to test for genetic linkage of the NGF gene to high blood pressure. DESIGN: We studied NGF gene expression in young SHR and examined linkage of the NGF locus to mean arterial pressure in genetically segregating crosses of SHR and normotensive Donryu (DRY) rats. METHODS: NGF mRNA was measured by Northern blot, and a restriction fragment length polymorphism of the NGF gene revealed after digestion with the NsiI restriction enzyme was used to study inheritance. RESULTS: Levels of NGF mRNA were detected easily in the kidneys of 2-, 4- and 10-week-old SHR but not in age-matched DRY rats. In an F2 population, the blood pressure of rats homozygous for the DRY NGF allele was 6 mmHg less than in heterozygotes and 8 mmHg less than in rats homozygous for the SHR NGF allele (analysis of variance, P < 0.004). In backcross rats the blood pressure of NGF heterozygotes was not significantly different from that of SHR homozygotes. CONCLUSION: These results indicate differences in renal NGF mRNA in SHR during the development of hypertension and suggest that a genetic locus in or near the NGF gene contributes in a Mendelian dominant pattern to a significant increment in blood pressure in SHR.

Fracture toughness of human bone under tension.

The longitudinal fracture toughnesses of human cortical bone were compared to those of bovine cortical bone to test the hypothesis that although human osteonal bone is significantly weaker and more compliant than primary (plexiform) bone, it is not less tough than primary bone. The fracture toughness indices, critical strain energy release rate (Gc) and critical stress intensity factor (Kc), were determined for human Haversian bone and bovine bone under tension (Mode I) loading using the compact tension method. The effects of thickness, crack growth range and anisotropy on fracture indices for slow stable crack growth in cortical bone were determined. Plane strain assumptions required for application of linear elastic fracture mechanics (LEFM) to bone were investigated. Longitudinal oriented fracture toughness tests were used to assess the crack inhibiting effect of human bone microstructure on fracture resistance. Human bone Kc calculated from the stress concentration formula for 2 and 3 mm thick specimens equaled 4.32 and 4.05 MN m-3/2, respectively. Human bone Gc calculated from the compliance method equaled 827 N m-1 for 2 mm thick specimens and 595 N m-1 for 3 mm thick specimens. It was found that crack growth range, thickness and material assumptions affect fracture toughness. Kc calculated from Gc using an anisotropic relation provided the lowest estimate of Kc and equaled 3.31 MN m-3/2 for 2 mm thick specimens and 2.81 MN m-3/2 for 3 mm thick specimens. Both Kc and Gc were significantly reduced after being adjusted to ASTM standard thickness using ratios determined from bovine bone. The fracture toughness of bovine bone relative to human bone ranged from 1.08 to 1.66. This was compared to the longitudinal strength of bovine bone relative to the longitudinal strength of human bone which is approximately equal to 1.5. We found that even though human bone is significantly weaker than bovine bone, relative to its strength, the toughness of human and bovine bone are roughly similar, but the data were not sufficiently definitive to answer the question of which is tougher.

Effect of groove on bone fracture toughness.

When testing for the effects of bone orientation on mode I fracture toughness, compact tension specimens are grooved with a V-notch to provide a crack guide. The effect of grooving on the expressions for the critical stress intensity factor (Kc) and the critical strain energy release rate (Gc) for mode I fracture toughness was investigated. Experiments were performed using grooved and ungrooved bovine compact tension specimens. The results indicate that the standard expression used to determine Kc for a compact tension specimen requires modification. The thickness (B) must be modified to account for the thickness between the grooves (Bn). The thickness used in the standard expression is replaced by an effective thickness written as (BBn)0.5. It was also found that the thickness between the grooves should be used in the standard formula for Gc.

Resistance to crack growth in human cortical bone is greater in shear than in tension.

It has been proposed that longitudinal shear stresses create bone microdamage, which suggests that bone is weak in shear and may not be adapted to prevent crack growth under shear loading. However, based on the similarities between bone and other fiber-reinforced composites that are tough, i.e. resistant to crack growth, we hypothesized that resistance of human bone to crack growth under shear loading is greater than under tensile loading. Because bone from older individuals and women has demonstrated increased propensity to fracture, we also hypothesized that bone from these individuals has less resistance to crack growth under shear and tension loading. Using compact shear and compact tension specimens, the critical strain energy release rate (Gc) of human bone was measured for longitudinally oriented cracks under tension (mode I) and shear (mode II) loading for male and female cadavers ranging from 55 to 89 y. Average tensile fracture toughness (GIc) of male and female human bone was 339 Nm-1 (S.D. = +/- 132). Average shear fracture toughness (GIIc) of human bone over the same range was 4200 Nm-1 (S.D. = +/- 2516 Nm-1). Shear toughness was greater than tensile toughness (approximately 13 times), which is consistent with other fibrous composite materials. Fracture toughness decreased with age, but the fits were weak and significant for shear loading only. Tension and shear toughness did not depend on gender. We concluded that the resistance to crack propagation under shear loading is greater than under tensile loading, a finding which suggests that bone adapts to prevent crack growth in shear. We also found that bone toughness is equivalent in men and women and that bone toughness gradually decreases with age between 55 and 89 y.

Stem surface roughness alters creep induced subsidence and 'taper-lock' in a cemented femoral hip prosthesis.

The clinical success of polished tapered stems has been widely reported in numerous long term studies. The mechanical environment that exists for polished tapered stems, however, is not fully understood. In this investigation, a collarless, tapered femoral total hip stem with an unsupported distal tip was evaluated using a 'physiological' three-dimensional (3D) finite element analysis. It was hypothesized that stem-cement interface friction, which alters the magnitude and orientation of the cement mantle stress, would subsequently influence stem 'taper-lock' and viscoelastic relaxation of bone cement stresses. The hypothesis that creep-induced subsidence would result in increases to stem-cement normal (radial) interface stresses was also examined. Utilizing a viscoelastic material model for the bone cement in the analysis, three different stem-cement interface conditions were considered: debonded stem with zero friction coefficient (mu=0) (frictionless), debonded stem with stem-cement interface friction (mu=0.22) ('smooth' or polished) and a completely bonded stem ('rough'). Stem roughness had a profound influence on cement mantle stress, stem subsidence and cement mantle stress relaxation over the 24-h test period. The frictionless and smooth tapered stems generated compressive normal stress at the stem-cement interface creating a mechanical environment indicative of 'taper-lock'. The normal stress increased with decreasing stem-cement interface friction but decreased proximally with time and stem subsidence. Stem subsidence also increased with decreasing stem-cement interface friction. We conclude that polished stems have a greater potential to develop 'taper-lock' fixation than do rough stems. However, subsidence is not an important determinant of the maintenance of 'taper-lock'. Rather subsidence is a function of stem-cement interface friction and bone cement creep.

The efficacy of direct current stimulation for lumbar intertransverse process fusions in an animal model.

STUDY DESIGN: Posterolateral lumbar intertransverse process fusion using a rabbit model with autologous bone graft and direct current stimulation was compared with fusion achieved by using autologous bone graft alone. OBJECTIVES: To determine the efficacy of direct current electrical stimulation for the posterolateral lumbar intertransverse process fusion technique by using a 20-microA current and the more recently developed 60-microA current delivered by an implantable direct current stimulator. SUMMARY OF BACKGROUND DATA: Previous studies have demonstrated a positive effect of direct current electrical stimulation on posterior spinal fusion techniques. However, until recently, the environment of an intertransverse fusion was not well simulated. The current research examined the posterolateral lumbar intertransverse process fusion technique with direct current electrical stimulation using a rabbit model. This appears to parallel human fusion techniques more closely and allows for lower cost and technical ease. METHODS: In this study, 44 adult New Zealand white rabbits underwent an L5-L6 intertransverse process fusion. All the fusions used an autologous bone graft obtained from bilateral posterior iliac crests. A device was implanted in all the rabbits subcutaneously, and they were divided randomly into three groups: a sham or nonfunctioning group, a 20-microA low-current stimulator group, and a 60-microA higher-current stimulator group. Spinal fusion was evaluated radiographically, histologically, and manually as well as by biomechanical testing 5 weeks after surgery. RESULTS: Radiographic grades, manual palpation, biomechanical strength, and stiffness showed an increasing trend from sham or inactive stimulator groups to low-current and then to high-current stimulator groups. Histologic analysis revealed that the higher-current stimulator showed that, statistically, the healing response of the host tissue to the autograft had increased significantly, as compared with the sham. CONCLUSIONS: Direct current electrical stimulation is efficacious in improving both the healing rate and strength in this posterolateral lumbar fusion model. In addition, it appears that this effect is enhanced by increasing the stimulation current from 20 microA to 60 microA.

Mechanical behavior of thermo-responsive orthodontic archwires.

OBJECTIVES: This investigation was conducted to describe the mechanical behavior of thermo-responsive nitinol archwires in flexure at 5 degrees and 37 degrees C. METHODS: Four same-sized (but different force level) rectangular archwires were examined using a three-point bend test. Samples were tested at 5 degrees and 37 degrees C. Linear regressions were fit to different segments of the load-deflection plots. Regression parameters of the segments and other properties were statistically analyzed. RESULTS: Superelastic behavior was exhibited by all wires tested at 37 degrees but not at 5 degrees C. Permanent deformation was greater at 5 degrees than 37 degrees. The initial slope of the load-deflection data averaged 1230 g/mm at 37 degrees, which was significantly different from the average at 5 degrees (500 g/mm). Loads at the apparent yield point and the loads at 1, 2, and 3 mm of deflection were greater at 37 degrees C than at 5 degrees. While the slope and length of the superelastic region were not judged to be clinically significantly different, the average load of the superelastic region was significantly different: F300 (340 g) > F200 (250 g) > F100 (180 g) and Bioforce (180 g). When loaded at 5 degrees and then unloaded at 37 degrees, the mechanical hysteresis of the wires tested at 37 degrees and the permanent deformation of the wires tested at 5 degrees were reduced for all wires. SIGNIFICANCE: Nitinol wires are available with a variety of mechanical properties. The different mechanical properties of thermo-responsive wires at 5 degrees and 37 degrees C result in clinically useful shape-memory behavior. Utilizing the superelastic and shape-memory features of thermo-responsive wires has clinical advantages.

Aerobic exercise as a countermeasure for microgravity-induced bone loss and muscle atrophy in a rat hindlimb suspension model.

BACKGROUND: Loss of bone and skeletal muscle atrophy resulting from non-weight-bearing are major concerns associated with microgravity environment and spaceflight deconditioning. The objective of this research was to address the fundamental issue of whether bone loss and muscle atrophy could be attenuated using weight-bearing aerobic exercise on a treadmill as a countermeasure in rats subjected to simulated weightlessness by hindlimb suspension. METHOD: Bone and muscle from control and hindlimb-suspended groups with and without exercise were evaluated by bone mineral density (BMD), mechanical tests, bone histomorphometry and muscle mass. RESULTS: Femoral BMD of hindlimb-suspended (HS) rats subjected to treadmill exercise was significantly greater than femoral BMD of HS rats without exercise and also was equivalent to that of weight-bearing controls. Muscle mass from HS rats exercised on a treadmill was significantly greater than muscle mass from HS rats that did not exercise. Exercise did not result in muscle mass equal to that of controls, however. In addition, histomorphometric analysis of the metaphysis of the proximal tibia revealed that HS rats that exercised did not maintain bone formation equivalent to controls. No other bone parameters were found to vary significantly between groups. CONCLUSIONS: It was concluded that moderate aerobic exercise on a treadmill did attenuate bone loss and muscle atrophy due to simulated weightlessness by hindlimb suspension, however its effectiveness differed by tissue, anatomical site and parameter investigated.