Discovery of a novel non-steroidal GR antagonist with in vivo efficacy in the olanzapine-induced weight gain model in the rat

We report the optimization of a series of non-steroidal GR antagonists that led to the identification of compound 7. This compound is efficacious when dosed orally in an olanzapine-induced weight gain model in rats.     

Population Genetics of the Eastern Hellbender (Cryptobranchus alleganiensis alleganiensis) across Multiple Spatial Scales

Conservation genetics is a powerful tool to assess the population structure of species and provides a framework for informing management of freshwater ecosystems. As lotic habitats become fragmented, the need to assess gene flow for species of conservation management becomes a priority. The eastern hellbender (Cryptobranchus alleganiensis alleganiensis) is a large, fully aquatic paedamorphic salamander. Many populations are experiencing declines throughout their geographic range, yet the genetic ramifications of these declines are currently unknown. To this end, we examined levels of genetic variation and genetic structure at both range-wide and drainage (hierarchical) scales. We collected 1,203 individuals from 77 rivers throughout nine states from June 2007 to August 2011. Levels of genetic diversity were relatively high among all sampling locations. We detected significant genetic structure across populations (F_st values ranged from 0.001 between rivers within a single watershed to 0.218 between states). We identified two genetically differentiated groups at the range-wide scale: 1) the Ohio River drainage and 2) the Tennessee River drainage. An analysis of molecular variance (AMOVA) based on landscape-scale sampling of basins within the Tennessee River drainage revealed the majority of genetic variation (∼94–98%) occurs within rivers. Eastern hellbenders show a strong pattern of isolation by stream distance (IBSD) at the drainage level. Understanding levels of genetic variation and differentiation at multiple spatial and biological scales will enable natural resource managers to make more informed decisions and plan effective conservation strategies for cryptic, lotic species.     

Oxygen transport during steady-state submaximal exercise in chronic hypoxia

Arterial O2 delivery during short-term submaximal exercise falls on arrival at high altitude but thereafter remains constant. As arterial O2 content increases with acclimatization, blood flow falls. We evaluated several factors that could influence O2 delivery during more prolonged submaximal exercise after acclimatization at 4,300 m. Seven men (23 +/- 2 yr) performed 45 min of steady-state submaximal exercise at sea level (barometric pressure 751 Torr), on acute ascent to 4,300 m (barometric pressure 463 Torr), and after 21 days of residence at altitude. The O2 uptake (VO2) was constant during exercise, 51 +/- 1% of maximal VO2 at sea level, and 65 +/- 2% VO2 at 4,300 m. After acclimatization, exercise cardiac output decreased 25 +/- 3% compared with arrival and leg blood flow decreased 18 +/- 3% (P less than 0.05), with no change in the percentage of cardiac output to the leg. Hemoglobin concentration and arterial O2 saturation increased, but total body and leg O2 delivery remained unchanged. After acclimatization, a reduction in plasma volume was offset by an increase in erythrocyte volume, and total blood volume did not change. Mean systemic arterial pressure, systemic vascular resistance, and leg vascular resistance were all greater after acclimatization (P less than 0.05). Mean plasma norepinephrine levels also increased during exercise in a parallel fashion with increased vascular resistance. Thus we conclude that both total body and leg O2 delivery decrease after arrival at 4,300 m and remain unchanged with acclimatization as a result of a parallel fall in both cardiac output and leg blood flow and an increase in arterial O2 content.(ABSTRACT TRUNCATED AT 250 WORDS)     

Whole genome sequencing and de novo assembly identifies Sydney-like variant noroviruses and recombinants during the winter 2012/2013 outbreak in England

Background

Influenza B viruses can cause morbidity and mortality in humans but due to the lack of an animal reservoir are not associated with pandemics. Because of this, there is relatively limited genetic sequences available for influenza B viruses, especially from developing countries. Complete genome analysis of one influenza B virus and several gene segments of other influenza B viruses isolated from Uganda from May 2009 through December 2010 was therefore undertaken in this study.

Methods

Samples were collected from patients showing influenza like illness and screened for influenza A and B by PCR. Influenza B viruses were isolated on Madin-Darby Canine Kidney cells and selected isolates were subsequently sequenced and analyzed phylogenetically.

Findings

Of the 2,089 samples collected during the period, 292 were positive by PCR for influenza A or B; 12.3% of the PCR positives were influenza B. Thirty influenza B viruses were recovered and of these 25 that grew well consistently on subculture were subjected to further analysis. All the isolates belonged to the B/Victoria-lineage as identified by hemagglutination inhibition assay and genetic analysis except one isolate that grouped with the B-Yamagata-lineage. The Ugandan B/Victoria-lineage isolates grouped in clade 1 which was defined by the N75K, N165K and S172P substitutions in hemagglutinin (HA) protein clustered together with the B/Brisbane/60/2008 vaccine strain. The Yamagata-like Ugandan strain, B/Uganda/MUWRP-053/2009, clustered with clade 3 Yamagata viruses such as B/Bangladesh/3333/2007 which is characterized by S150I and N166Y substitutions in HA.

Conclusion

In general there was limited variation among the Ugandan isolates but they were interestingly closer to viruses from West and North Africa than from neighboring Kenya. Our isolates closely matched the World Health Organization recommended vaccines for the seasons.     

A new genus of Nippostrongylinae (Nematoda: Heligmonellidae) from the water rat Scapteromys aquaticus (Sigmodontinae) in Argentina

A new genus of Nippostrongylinae, Malvinema n. gen., with 3 coparasitic species M. frederici n. sp., M. carolinae n. sp., and M. victoriae n. sp. from the intestine of the water rat, Scapteromys aquaticus Thomas (Rodentia: Muridae), from the northeast of Buenos Aires Province, Argentina, is proposed in this study. The new genus shows similarities to 2 Neotropical Nippostrongylinae: Carolinensis (Travassos, 1937) by some characters of the synlophe and Stilestrongylus Freitas, Lent and Almeida, 1937, by the pattern of the caudal bursa. It is characterized by a synlophe with triple or quadruple gradient of size of the ridges, lateromedian, decreasing from the largest left and right ridges. The gradient situated in the right ventral quadrant is always present. The caudal bursa shows a pattern of type 1-4. Malvinema frederici possesses a synlophe with 17 ridges and an axis of orientation inclined at 45 degrees from the sagittal axis; M. carolinae possesses a synlophe with 22-24 ridges and an axis of orientation almost merged with the sagittal axis. Both species have a caudal bursa with the right lobe enlarged transversally. Malvinema victoriae possesses a synlophe with 22-24 ridges, an axis of orientation inclined at 45 degrees from the sagittal axis, and a caudal bursa with the right lobe enlarged vertically.     

The pressure-flow relation for plasma in whole organ skeletal muscle and its experimental verification.

The whole-organ pressure-flow relation in resting rat skeletal muscle is examined for the flow of plasma. Due to the small size of the blood vessels in this organ, inertia and convective forces in the blood are negligible and viscous forces dominate. Direct measurements in the past have shown that skeletal muscle blood vessels are distensible. Theoretical formulations based on these measurements lead to a third order polynomial model for the pressure-flow relation. The purpose of the current study is to examine this relation experimentally in an isolated muscle organ. A high precision feedback controlled pump is used to perfuse artificial plasma into the vasodilated rat gracilis muscle. The results indicate that the pressure-flow curve in this tissue is nonlinear in the low flow region and almost linear at physiological flow rates, following closely the third order polynomial function. Vessel fixation with glutaraldehyde causes the curves to become linear at all pressures, indicating that vessel distention is the primary mechanism causing the nonlinearity. Furthermore, the resistance of the post-fixed tissue is determined by the pressure at which the fixative is perfused. At fixation pressures below 10 mmHg, the resistance is three times higher than in vessels fixed at normal physiological pressures. Dextran (229,000 Dalton) is used to obtain Newtonian perfusates at different viscosities. The pressure-flow relation is found to be linearly dependent on viscosity for all flow rates.(ABSTRACT TRUNCATED AT 250 WORDS)     

Identification of an oestrus-associated glycoprotein in oviducal fluid of the sheep

Samples of oviducal fluid were collected daily from sheep with indwelling catheters. Fluid samples taken from both oviducts of 2 sheep for 2 cycles during the middle of the breeding season (April/May) (8 sets of data) were compared with 9 sets of data generated from 2 cycles in 3 sheep later in the breeding season (June/July). Around the period of oestrus, the output of oviducal fluid increased to a peak volume of 1.56 +/- 0.35 ml per day (mean +/- s.d.) compared with a mid-cycle volume of 0.49 +/- 0.29 ml. Later in the breeding season, the flow rates were lower, but showed the same trend (0.91 +/- 0.24 ml at the peak and 0.25 +/- 0.18 ml 7 days later). The total amount of protein secreted by the oviduct each day increased 2-4-fold around the time of oestrus, with higher levels in mid-season ewes. When oviducal fluids were fractionated by SDS electrophoresis, a novel glycoprotein, subunit size of Mr 80-90 000 was identified in samples for 3-6 days of each cycle, coinciding with the period of high fluid flow rate. This protein first appeared in the oviducal fluid on the day of oestrus or the following day and it represented 1 of the 2 major glycoproteins in oviducal fluid as assessed by periodic acid-Schiff (PAS) staining. A PAS-positive protein (Mr 80-90 000) was also detected in fluid taken after oestrus on native highly cross-linked gradient gels after electrophoresis at pH 3.1 but not at pH 8.3. Both gradient gel systems showed an increase in high molecular weight material (Mr greater than 10(6] in fluid taken soon after oestrus.     

Residual active granzyme B in cathepsin C-null lymphocytes is sufficient for perforin-dependent target cell apoptosis

Cathepsin C activates serine proteases expressed in hematopoietic cells by cleaving an N-terminal dipeptide from the proenzyme upon granule packaging. The lymphocytes of cathepsin C-null mice are therefore proposed to totally lack granzyme B activity and perforin-dependent cytotoxicity. Surprisingly, we show, using live cell microscopy and other methodologies, that cells targeted by allogenic CD8(+) cytotoxic T lymphocyte (CTL) raised in cathepsin C-null mice die through perforin-dependent apoptosis indistinguishable from that induced by wild-type CTL. The cathepsin C-null CTL expressed reduced but still appreciable granzyme B activity, but minimal granzyme A activity. Also, in contrast to mice with inactivation of both their granzyme A/B genes, cathepsin C deficiency did not confer susceptibility to ectromelia virus infection in vivo. Overall, our results indicate that although cathepsin C clearly generates the majority of granzyme B activity, some is still generated in its absence, pointing to alternative mechanisms for granzyme B processing and activation. Cathepsin C deficiency also results in considerably milder immune deficiency than perforin or granzyme A/B deficiency.     

Considering increased mouse stomach mass when calculating prophylactic vaccine efficacy against Helicobacter pylori

BACKGROUND: Calculating the level of protection follow vaccination against Helicobacter pylori in mice is conventionally performed based on stomach mass. However, prophylactic vaccination is associated with a post-immunisation gastritis which results in considerable thickening of the gastric mucosa. RESULTS AND CONCLUSIONS: Here, an analysis of nine experiments demonstrates that this post-immunisation gastritis can actually induce a significant increase in stomach mass. To determine whether this mass increase impacts upon the perceived level of protection, calculations were performed comparing colony-forming units (CFU) either per gram of tissue or per whole stomach. A trend was apparent towards increased stomach mass influencing the degree of protection if CFU were calculated per gram. However, as this effect was not significant, it appears valid to express CFU either per gram or per stomach (if age and gender matched mice are used).     

Assembling millions of short DNA sequences using SSAKE

Novel DNA sequencing technologies with the potential for up to three orders magnitude more sequence throughput than conventional Sanger sequencing are emerging. The instrument now available from Solexa Ltd, produces millions of short DNA sequences of 25 nt each. Due to ubiquitous repeats in large genomes and the inability of short sequences to uniquely and unambiguously characterize them, the short read length limits applicability for de novo sequencing. However, given the sequencing depth and the throughput of this instrument, stringent assembly of highly identical sequences can be achieved. We describe SSAKE, a tool for aggressively assembling millions of short nucleotide sequences by progressively searching through a prefix tree for the longest possible overlap between any two sequences. SSAKE is designed to help leverage the information from short sequence reads by stringently assembling them into contiguous sequences that can be used to characterize novel sequencing targets. Availability: http://www.bcgsc.ca/bioinfo/software/ssake.