Reported Acquisition Practices of Australian Dog Owners

In Australia, the UK and the US dog ownership is prevalent with an estimated 40% ofAustralian households, 25% of UK households, and 50% of US households owning a dog. Onceacquired, a dog usually becomes a family companion so, unlike a faulty product, it can rarely bereturned or resold without some emotional impact on both the acquirer and the dog. Regarding thereality of dog relinquishment, there is a growing need for cross-disciplinary research that considershow dog owners are making their acquisition choices and, if prioritising different attributes, leads tomore optimal acquisition choices. This research collected data from 2840 dog owners via an onlinesurvey and examines how owners prioritised various attributes when acquiring their latest dog.The Pearson-Blotchky analysis of survey results show owners are split into two groups, with eachgroup prioritising different attributes or characteristics in their search for a new dog. The first groupare those dog owners who prioritised: the ability to rescue a dog, how compatible the dog wason the first meeting, and how compatible they believed the dog would be with their household.The second group are those owners who prioritised: a dog’s morphology, temperament predictability,and breeding practices. While each group prioritised different attributes, neither group madesubstantially more optimal acquisition choices in terms of overall satisfaction with the dog that theyultimately selected

Sarcoidosis of the hypothalamus and pituitary stalk

We report a rare case of sarcoidosis of the hypothalamic and suprasellar region, with clinical course and the magnetic resonance imaging follow-up

Knockout studies reveal an important role of <i>plasmodium</i> lipoic acid protein ligase a1 for asexual blood stage parasite survival

Lipoic acid (LA) is a dithiol-containing cofactor that is essential for the function of a-keto acid dehydrogenase complexes. LA acts as a reversible acyl group acceptor and 'swinging arm' during acyl-coenzyme A formation. The cofactor is post-translationally attached to the acyl-transferase subunits of the multienzyme complexes through the action of octanoyl (lipoyl): &lt;i&gt;N&lt;/i&gt;-octanoyl (lipoyl) transferase (LipB) or lipoic acid protein ligases (LplA). Remarkably, apicomplexan parasites possess LA biosynthesis as well as scavenging pathways and the two pathways are distributed between mitochondrion and a vestigial organelle, the apicoplast. The apicoplast-specific LipB is dispensable for parasite growth due to functional redundancy of the parasite's lipoic acid/octanoic acid ligases/transferases. In this study, we show that &lt;i&gt;LplA1&lt;/i&gt; plays a pivotal role during the development of the erythrocytic stages of the malaria parasite. Gene disruptions in the human malaria parasite &lt;i&gt;P.falciparum&lt;/i&gt; consistently were unsuccessful while in the rodent malaria model parasite &lt;i&gt;P. berghei&lt;/i&gt; the &lt;i&gt;LplA1&lt;/i&gt; gene locus was targeted by knock-in and knockout constructs. However, the &lt;i&gt;LplA1&lt;/i&gt; &lt;sup&gt;(-)&lt;/sup&gt; mutant could not be cloned suggesting a critical role of LplA1 for asexual parasite growth &lt;i&gt;in vitro&lt;/i&gt; and &lt;i&gt;in vivo&lt;/i&gt;. These experimental genetics data suggest that lipoylation during expansion in red blood cells largely occurs through salvage from the host erythrocytes and subsequent ligation of LA to the target proteins of the malaria parasite

Clinical biological and genetic heterogeneity of the inborn errors of pulmonary surfactant metabolism

Pulmonary surfactant is a multimolecular complex located at the air-water interface within the alveolus to which a range of physical (surface-active properties) and immune functions has been assigned. This complex consists of a surface-active lipid layer (consisting mainly of phospholipids), and of an aqueous subphase. From discrete surfactant sub-fractions one can isolate strongly hydrophobic surf acta nt proteins B (SP-B) and C (SP-C) as well as collectins SP-A and SP-D, which were shown to have specific structural, metabolic, or immune properties. Inborn or acquired abnormalities of the surfactant, qualitative or quantitative in nature, account for a number of human diseases. Beside hyaline membrane disease of the preterm neonate, a cluster of hereditary or acquired lung diseases has been characterized by periodic acid-Schiff-positive material filling the alveoli. From this heterogeneous nosologic group, at least two discrete entities presently emerge. The first is the SP-B deficiency, in which an essentially proteinaceous material is stored within the alveoli, and which represents an autosomal recessive Mendelian entity linked to the SFTPB gene (MIM 1786640). The disease usually generally entails neonatal respiratory distress with rapid fatal outcome, although partial or transient deficiencies have also been observed. The second is alveolar proteinosis, characterized by the storage of a mixed protein and lipid material, which constitutes a relatively heterogeneous clinical and biological syndrome, especially with regard to age at onset (from the neonate through to adulthood) as well as the severity of associated signs. Murine models, with a targeted mutation of the gene encoding granulocyte macrophage colony-stimulating factor (GM-CSF) (Csfgm) or the beta subunit of its receptor (II3rb1) support the hypothesis of an abnormality of surfactant turnover in which the alveolar macrophage is a key player. Apart from SP-B deficiency, in which a near-consensus diagnostic chart can be designed, the ascertainment of other abnormalities of surfactant metabolism is not straightforward. The disentanglement of this disease cluster is however essential to propose specific therapeutic procedures: repeated broncho-alveolar ravages, GM-CSF replacement, bone marrow grafting or lung transplantation

Negative DNA supercoiling induces genome-wide Cas9 off-target activity

CRISPR-Cas9 is a powerful gene-editing technology; however, off-target activity remains an important consideration for therapeutic applications. We have previously shown that force-stretching DNA induces off-target activity and hypothesized that distortions of the DNA topology in vivo, such as negative DNA supercoiling, could reduce Cas9 specificity. Using single-molecule optical-tweezers, we demonstrate that negative supercoiling λ-DNA induces sequence-specific Cas9 off-target binding at multiple sites, even at low forces. Using an adapted CIRCLE-seq approach, we detect over 10,000 negative-supercoiling-induced Cas9 off-target double-strand breaks genome-wide caused by increased mismatch tolerance. We further demonstrate in vivo that directed local DNA distortion increases off-target activity in cells and that induced off-target events can be detected during Cas9 genome editing. These data demonstrate that Cas9 off-target activity is regulated by DNA topology in vitro and in vivo, suggesting that cellular processes, such as transcription and replication, could induce off-target activity at previously overlooked sites

Stage and treatment variation with age in postmenopausal women with breast cancer: compliance with guidelines

Breast cancer-specific mortality is static in older women despite having fallen in younger age groups, possibly due to lack of screening and differences in treatment. This study compared stage and treatment between two cohorts of postmenopausal women (55–69 vs 470 years) in a single cancer network over 6 months. A total of 378 patients were studied (470: N ¼ 167, 55–69 years: N ¼ 210). Older women presented with more advanced disease (470: metastatic/locally advanced 12%, 55–69 years: 3%, Po0.01). Those with operable cancer had a worse prognosis (Nottingham Prognostic Index (NPI) 470: median NPI 4.4, 55–69 years: 4.25, Po0.03). These stage differences were partially explained by higher screening rates in the younger cohort. Primary endocrine therapy was used in 42% of older patients compared with 3% in the younger group (Po0.001). Older women with cancers suitable for breast conservation were more likely to choose mastectomy (470: 57.5% mastectomy rate vs 55–69 years: 20.6%, Po0.01). Nodal surgery was less frequent in older patients (470: 6.7% no nodal surgery, 55–69 years: 0.5%, Po0.01) and was more likely to be inadequate (470: 10.7% o4 nodes excised, 55–69 years: 3.4%, Po0.02). In summary, older women presented with more advanced breast cancer, than younger postmenopausal women and were treated less comprehensively

Characterization of growth and metabolism of the haloalkaliphile Natronomonas pharaonis

Natronomonas pharaonis is an archaeon adapted to two extreme conditions: high salt concentration and alkaline pH. It has become one of the model organisms for the study of extremophilic life. Here, we present a genome-scale, manually curated metabolic reconstruction for the microorganism. The reconstruction itself represents a knowledge base of the haloalkaliphile's metabolism and, as such, would greatly assist further investigations on archaeal pathways. In addition, we experimentally determined several parameters relevant to growth, including a characterization of the biomass composition and a quantification of carbon and oxygen consumption. Using the metabolic reconstruction and the experimental data, we formulated a constraints-based model which we used to analyze the behavior of the archaeon when grown on a single carbon source. Results of the analysis include the finding that Natronomonas pharaonis, when grown aerobically on acetate, uses a carbon to oxygen consumption ratio that is theoretically near-optimal with respect to growth and energy production. This supports the hypothesis that, under simple conditions, the microorganism optimizes its metabolism with respect to the two objectives. We also found that the archaeon has a very low carbon efficiency of only about 35%. This inefficiency is probably due to a very low P/O ratio as well as to the other difficulties posed by its extreme environment

Mutation detection by analysis of DNA heteroduplexes in TILLING populations of diploid species

In the beginning of mutation research, mutations could only be detected indirectly through the analysis of the phenotypic alterations that they caused. The detection of mutations at the DNA level became possible with the development of sequencing methods. Nowadays, there are many different methods and strategies that have been created for mutation detection, both in natural and mutagenised populations. The strategies differ in accuracy and sensitivity, as well as in the laboratory facilities, time, costs and efforts that are required. The majority of them involve the pooling of DNA samples and the amplification of a gene (fragment) of interest followed by heteroduplex formation. One of the popular strategies for mutation identification takes advantage of the specific endonuclease (e.g. CEL I) that recognises and cuts heteroduplexes precisely at the 3′ position of the mismatch site. The cleaved fragments are usually visualised through electrophoresis in a polyacrylamide gel using LI-COR sequencers, but agarose electrophoresis may also be used for this purpose, although with less sensitivity. A different mutation identification strategy, which is based on the high-resolution melting (HRM) technique, may be the method of choice when working with a short gene or a gene fragment whose length optimally does not exceed 400 bp

Development of a prototype lateral flow immunoassay (LFI) for the rapid diagnosis of melioidosis.

Burkholderia pseudomallei is a soil-dwelling bacterium and the causative agent of melioidosis. Isolation of B. pseudomallei from clinical samples is the "gold standard" for the diagnosis of melioidosis; results can take 3-7 days to produce. Alternatively, antibody-based tests have low specificity due to a high percentage of seropositive individuals in endemic areas. There is a clear need to develop a rapid point-of-care antigen detection assay for the diagnosis of melioidosis. Previously, we employed In vivo Microbial Antigen Discovery (InMAD) to identify potential B. pseudomallei diagnostic biomarkers. The B. pseudomallei capsular polysaccharide (CPS) and numerous protein antigens were identified as potential candidates. Here, we describe the development of a diagnostic immunoassay based on the detection of CPS. Following production of a CPS-specific monoclonal antibody (mAb), an antigen-capture immunoassay was developed to determine the concentration of CPS within a panel of melioidosis patient serum and urine samples. The same mAb was used to produce a prototype Active Melioidosis Detect Lateral Flow Immunoassay (AMD LFI); the limit of detection of the LFI for CPS is comparable to the antigen-capture immunoassay (∼0.2 ng/ml). The analytical reactivity (inclusivity) of the AMD LFI was 98.7% (76/77) when tested against a large panel of B. pseudomallei isolates. Analytical specificity (cross-reactivity) testing determined that 97.2% of B. pseudomallei near neighbor species (35/36) were not reactive. The non-reactive B. pseudomallei strain and the reactive near neighbor strain can be explained through genetic sequence analysis. Importantly, we show the AMD LFI is capable of detecting CPS in a variety of patient samples. The LFI is currently being evaluated in Thailand and Australia; the focus is to optimize and validate testing procedures on melioidosis patient samples prior to initiation of a large, multisite pre-clinical evaluation

Signal-Regulated Pre-mRNA Occupancy by the General Splicing Factor U2AF

Alternative splicing of transcripts in a signal-dependent manner has emerged as an important concept to ensure appropriate expression of splice variants under different conditions. Binding of the general splicing factor U2AF to splice sites preceding alternatively spliced exons has been suggested to be an important step for splice site recognition. For splicing to proceed, U2AF has to be replaced by other factors. We show here that U2AF interacts with the signal-dependent splice regulator Sam68 and that forced expression of Sam68 results in enhanced binding of the U2AF65 subunit to an alternatively spliced pre-mRNA sequence in vivo. Conversely, the rapid signal-induced and phosphorylation-dependent interference with Sam68 binding to RNA was accompanied by reduced pre-mRNA occupancy of U2AF in vivo. Our data suggest that Sam68 can affect splice site occupancy by U2AF in signal-dependent splicing. We propose that the induced release of U2AF from pre-mRNA provides a regulatory step to control alternative splicing