High-resolution protein structure determination by serial femtosecond crystallography

Structure determination of proteins and other macromolecules has historically required the growth of high-quality crystals sufficiently large to diffract x-rays efficiently while withstanding radiation damage. We applied serial femtosecond crystallography (SFX) using an x-ray free-electron laser (XFEL) to obtain high-resolution structural information from microcrystals (less than 1 micrometer by 1 micrometer by 3 micrometers) of the well-characterized model protein lysozyme. The agreement with synchrotron data demonstrates the immediate relevance of SFX for analyzing the structure of the large group of difficult-to-crystallize molecules.     

Continental-scale effects of nutrient pollution on stream ecosystem functioning

Excessive nutrient loading is a major threat to aquatic ecosystems worldwide that leads to profound changes in aquatic biodiversity and biogeochemical processes. Systematic quantitative assessment of functional ecosystem measures for river networks is, however, lacking, especially at continental scales. Here, we narrow this gap by means of a pan-European field experiment on a fundamental ecosystem process--leaf-litter breakdown--in 100 streams across a greater than 1000-fold nutrient gradient. Dramatically slowed breakdown at both extremes of the gradient indicated strong nutrient limitation in unaffected systems, potential for strong stimulation in moderately altered systems, and inhibition in highly polluted streams. This large-scale response pattern emphasizes the need to complement established structural approaches (such as water chemistry, hydrogeomorphology, and biological diversity metrics) with functional measures (such as litter-breakdown rate, whole-system metabolism, and nutrient spiraling) for assessing ecosystem health.     

The role of driving energy and delocalized States for charge separation in organic semiconductors

The electron-hole pair created via photon absorption in organic photoconversion systems must overcome the Coulomb attraction to achieve long-range charge separation. We show that this process is facilitated through the formation of excited, delocalized band states. In our experiments on organic photovoltaic cells, these states were accessed for a short time (<1 picosecond) via infrared (IR) optical excitation of electron-hole pairs bound at the heterojunction. Atomistic modeling showed that the IR photons promote bound charge pairs to delocalized band states, similar to those formed just after singlet exciton dissociation, which indicates that such states act as the gateway for charge separation. Our results suggest that charge separation in efficient organic photoconversion systems occurs through hot-state charge delocalization rather than energy-gradient-driven intermolecular hopping.     

Occurrence and molecular analysis of Campylobacter in wildlife on livestock farms

Wildlife harbor a variety of Campylobacter spp. and may play a significant role in the transmission of Campylobacter to livestock. Although studies have been conducted on wildlife-associated Campylobacter isolates from farms in other countries, there are little data available for livestock farms in the United States. In addition, the critical questions of whether wildlife harbor Campylobacter that is pathogenic to ruminants and/or antibiotic-resistant Campylobacter have yet to be addressed. We captured wild small mammals (n=142) and small birds (n=188) at livestock farms in central Iowa and sampled them for thermophilic Campylobacter during autumn 2009, spring 2010, and autumn 2010. Overall prevalence was 4.79%, with isolates found only in wild birds. Molecular typing revealed four multilocus sequence types (STs), three of which are novel. The remaining ST (ST-806) was found in two house sparrows and is an ST previously associated with ruminant abortion cases. Further analysis of ST-806 wild bird and ruminant abortion isolates by pulsed-field gel electrophoresis, resistance gene location, and antibiotic susceptibility tests indicated that the isolates are nearly identical. This is the first account of isolation of Campylobacter types from wild birds that are known to be pathogenic to ruminants. Furthermore, these same two wild bird isolates are resistant to the antibiotic fluoroquinolone. Our results indicate there is an overall low prevalence of Campylobacter in selected wildlife in Iowa, but suggest that wildlife may play a role in the epidemiology of pathogenic Campylobacter for domestic livestock, and may also serve as a reservoir for antibiotic-resistant Campylobacter.     

Effect of Restricted Grazing on Hindgut pH and Fluid Balance

Six mature idle geldings were used in a crossover design to determine the effects of restricted grazing on hindgut pH and fluid balance. Initially, horses were randomly assigned to a control group (CTRL: n = 3) having access to warm-season grass pasture continuously, or a restricted grazing (RG: n = 3) group having access to pasture for 12 consecutive hours (1900-700 hours) per 24-hour period for 7 days; they were then reassigned to the opposite treatment for an additional 7 days (i.e., CTRL: n = 6; RG: n = 6). Fecal samples were collected from each horse at 700 hours on day 7 of each period and analyzed for pH and dry matter (DM) as indicators of hindgut pH and fluid balance, respectively. Jugular blood samples were also collected at 700, 1300, and 1900 hours on day 7 and were analyzed for plasma protein as an indicator of systematic fluid balance. Fecal pH and DM data were analyzed using a paired t test. Plasma protein data were analyzed as a repeated-measures design. The mean (± SE) difference between CTRL and RG for fecal pH (.01 ± .16) and fecal DM (.68 ± .6%) was not significant (P = .93 and .52, respectively). Mean plasma protein concentrations were not affected by treatment or by treatment × sample time interaction, but tended to increase (P = .07) during the sampling period regardless of treatment. In conclusion, 12 hours of grazing restriction followed by 12 hours of grazing did not negatively impact hindgut pH or fluid balance.     

Acceptability and compliance of atenolol tablet, compounded paste and compounded suspension prescribed to healthy cats*

This study was designed to evaluate the cats' acceptance and compliance of the owners and cats towards an extemporaneously prepared palatable compounded atenolol (paste and suspension) formulation in comparison to the commercially obtained tablet, in a randomised, cross-over study design.The three formulations were prescribed twice daily for 6 days to 13 healthy privately-owned cats of 13 different owners, with varying levels of experience in medicating cats. Daily compliance was evaluated via an owner-completed diary, completed after each dose administered. Owner's experience and preference of the formulation was evaluated via questionnaires given prior to, at the end of each treatment protocol, and upon completion of the study. Although compounded suspension was association with fewest missed doses, the majority of cat owners expressed a preference for the divided tablet. Atenolol tablets, compounded paste and suspension acceptance and compliance were comparable. Further work is now required to assess the amount and stability of the active ingredient and the robustness of the paste and suspension formulations prior to any bioavailability comparisons between the formulations.     

Development of a novel paradigm for the measurement of olfactory discrimination in dogs (Canis familiaris): A pilot study

Olfactory dysfunction in older human beings has been shown to be associated with an increased risk of cognitive decline, yet age-related changes in olfactory behavior have received little attention in the dog model of human aging. We developed an odor habituation and fine odor discrimination paradigm to test the hypothesis that dogs would show a novelty response toward unfamiliar urine from entire male conspecifics. We tested 26 odor detection dogs (14 females, 12 males) from the New South Wales police dog unit, ranging in age from 1 year 2 months to 11 years 10 months. First, dogs were familiarized with a master odor over 2 presentations. Second, we measured difference in investigation time of a master odor as compared with 5 odor mixtures using the following ratios of novel-to-master odor: 100:0, 80:20, 60:40, 40:60, and 20:80. Dogs habituated to the master odor after the first presentation (t(25) = 6.048, P < 0.001). After 2 dogs that failed to habituate were excluded, there was a nonsignificant trend (t(21) = ?1.968, P = 0.062) for aged dogs (>8 years, N = 6) to show reduced habituation as compared with middle-aged dogs (5-8 years, N = 9) and with all dogs aged <8 years (N = 18, t(21) = ?1.883, P = 0.072). Approximately half of the dogs tested (N = 11) failed to show a novelty response toward the 100:0, novel:master odor. The remaining dogs (N = 15) showed a significant novelty response toward this odor (mean difference = 1.89 seconds, confidence interval = 0.86-2.84). Investigation of the remaining odor mixtures was not significantly different from investigation of the master odor in all dogs. Further development of this paradigm is needed using naive pet dogs before it can be used as a reliable measure of fine odor discrimination. The current, weak trend for an age effect in habituation warrants further investigation in a larger cohort to determine if this effect becomes significant or if it is simply a manifestation of small sample size and low statistical power. It is recommended that future studies use dogs that have not been trained against or actively discouraged from investigating urine because previous learning may have had a significant effect on the outcomes of this study.     

Looks can deceive: Molecular identity of an intraerythrocytic apicomplexan parasite in Australian gliders

Two yellow-bellied gliders (Petaurus australis) had an intraerythrocytic parasite closely related to the cyst-forming coccidia (Apicomplexa: Sarcocystidae). The parasitaemia persisted for 3 months or more but was observed to clear within 3 years in captivity. The parasite appears not to significantly debilitate its infected host. Traditionally, using morphological identification, the intraerythrocytic parasite would have been classified within the Hepatozoon species typically found in red blood cells. However, molecular diagnostic techniques targeting the parasite's SSU rDNA and LSU rDNA demonstrated the unusual identity of this blood parasite and disputed its identity as a haemogregarine parasite of the genus Hepatozoon. The sequence was compared with available sequences from diverse mammalian and non-mammalian blood parasites (malaria, piroplasms, hemosporidia and sarcosporidia). The intraerythrocytic blood parasite was found to be most closely related to the cyst-forming coccidia including Besnoitia spp., Cystoisospora spp., Hammondia spp., Hyaloklossia lieberkuehni, Neospora caninum, Sarcocystis spp. and Toxoplasma gondii. The life cycle of this intraerythrocytic parasite remains unknown. The presented DNA identification demonstrates its suitability for an improved identification of blood parasites.