Development of polymorphic microsatellite DNA markers from the Korean field mouse, Apodemus peninsulae

We isolated and characterized eight polymorphic microsatellite DNA markers from the Korean field mouse, Apodemus peninsulae. The primers developed in this study yielded an average polymorphic information content of 0.78 (range 0.44–0.90), with an average of 10.9 alleles per locus (range 5–16). Observed and expected heterozygosities ranged from 0.46 to 1.00 and from 0.49 to 0.93, respectively. These polymorphic loci may provide useful tools for understanding the species’ genetic structure and ecology.     

Risk of Fatal Bleeding in Episodes of Major Bleeding with New Oral Anticoagulants and Vitamin K Antagonists: A Systematic Review and Meta-Analysis

Background

The reversibility of new/novel oral anticoagulants (NOAC) is not well understood, whereas the reversal strategies for bleeding associated with vitamin k antagonists (VKA), such as warfarin, is well established. It is unknown whether outcomes are different between bleeds occurring with NOAC compared to VKA use.

Objectives

This systematic review and meta-analysis of randomized controlled trials determines the relative odds of fatal bleeding given that a patient suffered a major bleed while on NOAC versus VKA therapy.

Search Methods

Data on major and fatal bleeding events was sought from randomized controlled trials of NOAC agents compared to VKAs.

Main Results

20 trials were included in the meta-analysis. From which, 4056 first-time, major bleeding events were reported and included in the primary analysis. The summary odds ratio for the conditional odds of fatal bleeding given that a major bleeding event occurred was 0.65 [0.52, 0.81] favoring the NOAC agents (p = 0.0001). The reduced odds of fatal bleeding with NOACs was not demonstrated after controlling for bleeding location. Given that an intracranial bleeding event occurred, the summary odds ratio for the conditional odds of fatal bleeding was 0.96 [0.70, 1.32]. For extracranial bleeding events, the summary odds ratio was also statistically insignificant at 0.945 [0.66, 1.35].

Author’s Conclusions

The odds ratio calculated in this meta-analysis showed a reduced odds of death in major bleeding associated with NOAC use. This risk reduction was due to a disproportionate amount of intracranial bleeding in the VKA arms. For any given bleeding site, there was no evidence of a significant difference in fatal outcomes from bleeds associated with NOAC versus VKA use.

Protocol Registration

Protocol registered on PROSPERO under CRD42014013294.     

Assessment of variation of nest survival for grassland birds due to method of nest discovery

Capsule Interpretation of nest survival estimates may be improved by incorporating the search method used to locate nests as a covariate.

Aims To compare annual survival estimates for Dickcissel Spiza americana nests and determine if incorporating search method (structured, opportunistic, or behavioural searches) improved model fit.

Methods Dickcissel nests were located using structured, opportunistic, or behavioural searches over three years (2011–2013) in Mississippi, USA. Models were used to estimate daily survival rates (DSRs) and to analyse factors influencing nest survival.

Results DSRs for Dickcissels were best explained by quadratic date, nest age, age found, and year, but incorporating search method improved model fit. Daily survival was 1.51 times greater for nests located using opportunistic search methods relative to structured searches, but was not significantly different between structured and behavioural searches.

Conclusions Survival estimates varied by search method, specifically between structured searches and opportunistically located nests. This might have arisen because heterogeneity in nest placement or parental behaviour may influence the sample of nests located with a given search method. Researchers may be able to account for this potential source of bias by including search method as a model covariate when using standard survey designs or modelling approaches.     

Cross-species amplification of microsatellites in crocodilians: assessment and applications for the future

Microsatellite DNA loci have emerged as the dominant genetic tool for addressing questions associated with genetic diversity in many wildlife species, including crocodilians. Despite their usefulness, their isolation and development can be costly, as well as labour intensive, limiting their wider use in many crocodilian species. In this study, we investigate the cross-species amplification success of 82 existing microsatellites previously isolated for the saltwater crocodile (Crocodylus porosus) in 18 non-target crocodilian species; Alligator sinensis, Caiman crocodylus, Caiman latirostris, Caiman yacare, Melanosuchus niger, Paleosuchus palpebrosus, Crocodylus acutus, Mecistops cataphractus, Crocodylus intermedius, Crocodylus johnstoni, Crocodylus mindorensis, Crocodylus moreletii, Crocodylus niloticus, Crocodylus novaeguineae, Crocodylus palustis, Crocodylus rhombifer, Crocodylus siamensis, and Osteolaemus tetraspis. Our results show a high level of microsatellites cross-amplification making available polymorphic markers for a range of crocodilian species previously lacking informative genetic markers.     

Proposed tests for measuring the running velocity at the oxygen consumption and heart rate thresholds for treadmill exercise

The purposes of this study were to (a) determine if the mathematical model used to estimate the physical working capacity at the oxygen consumption threshold (PWC(VO(2))) and physical working capacity at the heart rate threshold (PWC(HRT)) for cycle ergometry could be applied to treadmill running; (b) propose new fatigue thresholds called the running velocity at the oxygen uptake threshold (RV(VO(2))) and running velocity at the heart rate threshold (RV(HRT)) for treadmill exercise; and (c) statistically compare the velocities at the RV(VO(2)), RV(HRT), and ventilatory threshold (VT). Seven aerobically trained adult volunteers (mean +/- SD: age 24.0 +/- 3.9 years, Vo(2) max 56.7 +/- 7.1 ml.kg(-1).min(-1)) performed a maximal treadmill test to determine Vo(2) peak and VT as well as four 8-minute submaximal workbouts for the determination of RV(VO(2)) and RV(HRT). One-way repeated-measures analysis of variance indicated that there were no significant (p > 0.05) mean differences among the running velocities for the RV(VO(2)), RV(HRT), and VT. The results of this study indicated that the mathematical model used to estimate PWC(VO(2)) and PWC(HRT) for cycle ergometry could be applied to treadmill running. Furthermore, the RV(VO(2)) and RV(HRT) test may provide submaximal techniques for estimating the VT.     

Regulation of oxygen delivery during induced polycythemia in exercising dogs

Previous studies have concluded that polycythemia decreases oxygen delivery primarily because of a large fall in cardiac output associated with a rise in systemic vascular resistance that has been attributed to increased blood viscosity. However, because other studies have shown that polycythemia may not reduce oxygen delivery, an alternative hypothesis is that cardiac output falls in response to a rising oxygen content, thereby maintaining oxygen delivery constant. To determine whether oxygen content participates in the regulation of cardiac output during polycythemia, we studied eight chronically instrumented dogs trained to exercise on a treadmill. The dogs underwent exchange transfusion with packed red blood cells containing methemoglobin, which caused an increase in hematocrit from 35 +/- 1 to 50 +/- 1% and in viscosity, with little change in oxygen content. The expected fall in exercise cardiac output failed to occur after exchange transfusion with red blood cells containing methemoglobin (7.5 +/- 4 vs. 6.8 +/- 0.5 l/min; P = not significant), and there was no rise in systemic vascular resistance. Methylene blue was then administered intravenously to facilitate conversion of methemoglobin to oxyhemoglobin, which increased oxygen content (12.8 +/- 0.9 vs. 18.4 +/- 0.9 vol%; P < 0.01) with no change in hematocrit or viscosity. Resting cardiac output did not change significantly, but there was a significant decrease in exercise output (6.8 +/- 0.5 vs. 5.8 +/- 0.4 l/min; P < 0.05). Thus we conclude that the fall in cardiac output seen in acute polycythemia results in part from the regulation of oxygen delivery and is not due solely to increased blood viscosity.     

Inhibition of coxsackievirus B3 in cell cultures and in mice by peptide-conjugated morpholino oligomers targeting the internal ribosome entry site

Coxsackievirus B3 (CVB3) is a primary cause of viral myocarditis, yet no effective therapeutic against CVB3 is available. Nucleic acid-based interventional strategies against various viruses, including CVB3, have shown promise experimentally, but limited stability and inefficient delivery in vivo remain as obstacles to their potential as therapeutics. We employed phosphorodiamidate morpholino oligomers (PMO) conjugated to a cell-penetrating arginine-rich peptide, P007 (to form PPMO), to address these issues. Eight CVB3-specific PPMO were evaluated with HeLa cells and HL-1 cardiomyocytes in culture and in a murine infection model. One of the PPMO (PPMO-6), designed to target a sequence in the 3' portion of the CVB3 internal ribosomal entry site, was found to be especially potent against CVB3. Treatment of cells with PPMO-6 prior to CVB3 infection produced an approximately 3-log(10) decrease in viral titer and largely protected cells from a virus-induced cytopathic effect. A similar antiviral effect was observed when PPMO-6 treatment began shortly after the virus infection period. A/J mice receiving intravenous administration of PPMO-6 once prior to and once after CVB3 infection showed an approximately 2-log(10)-decreased viral titer in the myocardium at 7 days postinfection and a significantly decreased level of cardiac tissue damage, compared to the controls. Thus, PPMO-6 provided potent inhibition of CVB3 amplification both in cell cultures and in vivo and appears worthy of further evaluation as a candidate for clinical development.     

Floxed allele for conditional inactivation of the voltage-gated sodium channel beta1 subunit Scn1b

The voltage-gated sodium channel gene Scn1b encodes the auxiliary subunit beta1, which is widely distributed in neurons and glia of the central and peripheral nervous systems, cardiac myocytes, skeletal muscle myocytes, and neuroendocrine cells. We showed previously that the Scn1b null mutation results in a complex and severe phenotype that includes retarded growth, seizures, ataxia, and death by postnatal day 21. We generated a floxed allele of Scn1b by inserting loxP sites surrounding the second coding exon. Ubiquitous deletion of the floxed exon by Cre recombinase using CMV-Cre-transgenic mice produced the Scn1b(del) allele. The null phenotype of Scn1b(del) homozygotes is indistinguishable from that of Scn1b nulls and confirms the invivo inactivation of Scn1b. Conditional inactivation ofthe floxed allele will make it possible to circumvent the lethality that results from complete loss of this gene, such that the physiological role of Scn1b in specific cell types and/or specific developmental time points can be investigated.     

Parturition prediction and timing of canine pregnancy

An accurate method of predicting the date of parturition in the bitch is clinically useful to minimize or prevent reproductive losses by timely intervention. Similarly, an accurate method of timing canine ovulation and gestation is critical for development of assisted reproductive technologies, e.g. estrous synchronization and embryo transfer. This review discusses present methods for accurately timing canine gestational age and outlines their use in clinical management of high-risk pregnancies and embryo transfer research.     

CheX in the three-phosphatase system of bacterial chemotaxis

Bacterial chemotaxis involves the regulation of motility by a modified two-component signal transduction system. In Escherichia coli, CheZ is the phosphatase of the response regulator CheY but many other bacteria, including Bacillus subtilis, use members of the CheC-FliY-CheX family for this purpose. While Bacillus subtilis has only CheC and FliY, many systems also have CheX. The effect of this three-phosphatase system on chemotaxis has not been studied previously. CheX was shown to be a stronger CheY-P phosphatase than either CheC or FliY. In Bacillus subtilis, a cheC mutant strain was nearly complemented by heterologous cheX expression. CheX was shown to overcome the DeltacheC adaptational defect but also generally lowered the counterclockwise flagellar rotational bias. The effect on rotational bias suggests that CheX reduced the overall levels of CheY-P in the cell and did not truly replicate the adaptational effects of CheC. Thus, CheX is not functionally redundant to CheC and, as outlined in the discussion, may be more analogous to CheZ.