Degenerate codon mixing for PCR-based manipulation of highly repetitive sequences

Objective: Repeat expansion of polyglutamine tracks leads to a group of inherited human neurodegenerative disorders. Studying such repetitive sequences is required to gain insight into the pathophysiology of these diseases. PCR-based manipulation of repetitive sequences, however, is challenging due to the absence of unique primer binding sites or the generation of non-specific products. Results: We have utilised the degeneracy of the genetic code to generate a polyglutamine sequence with low repeat similarity. This strategy allowed us to use conventional PCR to generate multiple constructs with approximately defined numbers of glutamine repeats. We then used these constructs to measure the in vivo variation in autophagic degradation activity related to the different numbers of glutamine repeats, providing an example of their applicability to study repeat expansion diseases. Our simple and easily generalised method of generating low repetition DNA sequences coding for uniform stretches of amino acid residues provides a strategy for generating particular lengths of polyglutamine tracts using standard PCR and cloning protocols.Dhanushika Ratnayake, Morgan Newman and Michael Lardell

Post-Disaster Housing Reconstruction in Sri Lanka: What Methodology?

Research methodology is the procedural framework within which the research is conducted. This includes the overall approach to a problem that could be put into practice in a research process, from the theoretical underpinning to the collection and analysis of data. Choice of methodology depends on the primary drivers: topic to be researched and the specific research questions. Hence, methodological perspectives of managing stakeholder expectations of PDHR context are composed of research philosophies, research strategy, research design, and research techniques. This research belonged to social constructivism or interpretivism within a philosophical continuum. The nature of the study was more toward subjectivism where human behavior favored voluntary stance. Ontological, methodological, epistemological, and axiological positioning carried the characteristics of idealism, ideographic, anti-positivism, and value laden, respectively. Data collection comprises two phases, preliminary and secondary. Exploratory interviews with construction experts in the United Kingdom and Sri Lanka were carried out to refine the interview questions and identify the case studies. Case study interviews during the secondary phase took place in Sri Lanka. Data collected at the preliminary stage were used to assess the attributes of power, legitimacy/proximity, and urgency of stakeholders to the project using Stakeholder Circle™ software. Moreover, the data collected at secondary phase via case studies will be analyzed with NVivo 8. This article aims to discuss these methodological underpinnings in detail applied in a post-disaster housing reconstruction context in Sri Lanka

Birds Learn Socially to Recognize Heterospecific Alarm Calls by Acoustic Association

Animals in natural communities gain information from members of other species facing similar ecological challenges [1, 2, 3, 4, 5], including many vertebrates that recognize the alarm calls of heterospecifics vulnerable to the same predators [6]. Learning is critical in explaining this widespread recognition [7, 8, 9, 10, 11, 12, 13], but there has been no test of the role of social learning in alarm-call recognition, despite the fact that it is predicted to be important in this context [14, 15]. We show experimentally that wild superb fairy-wrens, Malurus cyaneus, learn socially to recognize new alarm calls and can do so through the previously undemonstrated mechanism of acoustic-acoustic association of unfamiliar with known alarm calls. Birds were trained in the absence of any predator by broadcasting unfamiliar sounds, to which they did not originally flee, in combination with a chorus of conspecific and heterospecific aerial alarm calls (typically given to hawks in flight). The fairy-wrens responded to the new sounds after training, usually by fleeing to cover, and responded equally as strongly in repeated tests over a week. Control playbacks showed that the response was not due simply to greater wariness. Fairy-wrens therefore learnt to associate new calls with known alarm calls, without having to see the callers or a predator. This acoustic-acoustic association mechanism of social learning could result in the rapid spread of alarm-call recognition in natural communities, even when callers or predators are difficult to observe. Moreover, this mechanism offers potential for use in conservation by enhancing training of captive-bred individuals before release into the wild

Duchenne muscular dystrophy from brain to muscle: The role of brain dystrophin isoforms in motor functions

Brain function and its effect on motor performance in Duchenne muscular dystrophy (DMD) is an emerging concept. The present study explored how cumulative dystrophin isoform loss, age, and a corticosteroid treatment affect DMD motor outcomes. A total of 133 genetically confirmed DMD patients from Sri Lanka were divided into two groups based on whether their shorter dystrophin isoforms (Dp140, Dp116, and Dp71) were affected: Group 1, containing patients with Dp140, Dp116, and Dp71 affected (n = 98), and Group 2, containing unaffected patients (n = 35). A subset of 52 patients (Group 1, n = 38; Group 2, n = 14) was followed for up to three follow-ups performed in an average of 28-month intervals. The effect of the cumulative loss of shorter dystrophin isoforms on the natural history of DMD was analyzed. A total of 74/133 (56%) patients encountered developmental delays, with 66/74 (89%) being in Group 1 and 8/74 (11%) being in Group 2 (p \u3c 0.001). Motor developmental delays were predominant. The hip and knee muscular strength, according to the Medical Research Council (MRC) scale and the North Star Ambulatory Assessment (NSAA) activities, “standing on one leg R”, “standing on one leg L”, and “walk”, declined rapidly in Group 1 (p \u3c 0.001 In the follow-up analysis, Group 1 patients became wheelchair-bound at a younger age than those of Group 2 (p = 0.004). DMD motor dysfunction is linked to DMD mutations that affect shorter dystrophin isoforms. When stratifying individuals for clinical trials, considering the DMD mutation site and its impact on a shorter dystrophin isoform is crucial

The dynamics and outcomes of AKI progression during the COVID-19 pandemic.

PURPOSE: Acute kidney injury (AKI) associated with COVID-19 is associated with poor prognosis. This study assessed the hitherto uninvestigated impact of COVID-19 on the progression and clinical outcomes of patients with AKI. METHODS: Data from 576 patients with AKI admitted between 13/3/20 and 13/5/20 were studied. Increasingly complex analyses, from logistic regressions to competing-risk and multi-state models, have revealed insights into AKI progression dynamics associated with PCR-confirmed COVID-19 acquisition and death. Meta-analyses of case fatality ratios among patients with AKI were also conducted. RESULTS: The overall case-fatality ratio was 0.33 [95% CI (0.20-0.36)]; higher in COVID-19 positive (COVID+) patients 0.52 [95% CI (0.46-0.58)] than in their negative (COVID-) counterparts 0.16 [95% CI (0.12-0.20)]. In AKI Stage-3 patients, that was 0.71 [95% CI (0.64-0.79)] among COVID+ patients with 45% dead within 14 days and 0.35 [95% CI (0.25-0.44)] in the COVID- group and 28% died within 14 days. Among patients diagnosed with AKI Stage-1 within 24 h, the probability of progression to AKI Stage-3 on day 7 post admission was 0.22 [95% CI (0.17-0.27)] among COVID+ patients, and 0.06 [95% CI (0.03, 0.09)] among those who tested negative. The probability of discharge by day 7 was 0.71 [95% CI (0.66, 0.75)] in COVID- patients, and 0.27 [95% CI (0.21, 0.32)] in COVID+ patients. By day 14, in AKI Stage-3 COVID+ patients, that was 0.35 [95% CI (0.25, 0.44)] with little change by day 10, that is, 0.38 [95% CI (0.29, 0.47)]. CONCLUSION: These results are consistent with either a rapid progression in severity, prolonged hospital care, or high case fatality ratio among AKI Stage-3 patients, significantly exacerbated by COVID-19 infection

Formulation of a fertilizer package for hybrid varieties of pumpkin (Cucurbita maxima)

Pumpkin (Cucurbita maxima) is a popular low country vegetable in Sri Lanka and it has become afundamental crop species in the farming systems of dry and intermediate zones. Introduced hybridpumpkin varieties are becoming popular in Sri Lanka because ofthe higher yield. However, there isno fertilizer recommendation for hybrid pumpkin. Local farmers apply high dosages ofNitrogen(N),Phosphorus(P) and Potassium(K) levels which causes increased cost of production and reducedprofit. Therefore, a field experiment was conducted to identify the effects of higher N, P and K levelson yield of hybrid pumpkin. The hybrid variety Arjuna was tested under eight different fertilizer levelswhere the Department of Agriculture (DOA) recommendation was taken as the control and one andhalf times of that was used as the higher levels of N, P and K. The experiment was arranged in aconfounding design with three replicates with two blocks per each. Reproductive parameters andyield parameters viz. yield, number of fruits per vine, fruit width and diameter were recorded. Thetested fertilizer levels did not show any significant effect on reproductive parameters and yieldparameters of hybrid pumpkin, thus, indicating that the application of higher dosages ofN, P and K isnot economical. Hence, the DOA fertilizer recommendation is adequate for hybrid pumpkins thoughthe yield is higher when compared to local pumpkin varieties