Robust and stable transcriptional repression in Giardia using CRISPRi.

Giardia lamblia is a binucleate protistan parasite causing significant diarrheal disease worldwide. An inability to target Cas9 to both nuclei, combined with the lack of nonhomologous end joining and markers for positive selection, has stalled the adaptation of CRISPR/Cas9-mediated genetic tools for this widespread parasite. CRISPR interference (CRISPRi) is a modification of the CRISPR/Cas9 system that directs catalytically inactive Cas9 (dCas9) to target loci for stable transcriptional repression. Using a Giardia nuclear localization signal to target dCas9 to both nuclei, we developed efficient and stable CRISPRi-mediated transcriptional repression of exogenous and endogenous genes in Giardia. Specifically, CRISPRi knockdown of kinesin-2a and kinesin-13 causes severe flagellar length defects that mirror defects with morpholino knockdown. Knockdown of the ventral disk MBP protein also causes severe structural defects that are highly prevalent and persist in the population more than 5 d longer than defects associated with transient morpholino-based knockdown. By expressing two guide RNAs in tandem to simultaneously knock down kinesin-13 and MBP, we created a stable dual knockdown strain with both flagellar length and disk defects. The efficiency and simplicity of CRISPRi in polyploid Giardia allows rapid evaluation of knockdown phenotypes and highlights the utility of CRISPRi for emerging model systems

Using drones and sirens to elicit avoidance behaviour in white rhinoceros as an anti-poaching tactic.

Poaching fuelled by international trade in horn caused the deaths of over 1000 African rhinoceros (Ceratotherium simum and Diceros bicornis) per year between 2013 and 2017. Deterrents, which act to establish avoidance behaviours in animals, have the potential to aid anti-poaching efforts by moving at-risk rhinos away from areas of danger (e.g. near perimeter fences). To evaluate the efficacy of deterrents, we exposed a population of southern white rhinos (C. simum simum) to acoustic- (honeybee, siren, turtle dove), olfactory- (chilli, sunflower), and drone-based stimuli on a game reserve in South Africa. We exposed rhinos to each stimulus up to four times. Stimuli were considered effective deterrents if they repeatedly elicited avoidance behaviour (locomotion away from the deterrent). Rhinos travelled significantly further in response to the siren than to the honeybee or turtle dove stimulus, and to low-altitude drone flights than to higher altitude flights. We found the drone to be superior at manipulating rhino movement than the siren owing to its longer transmission range and capability of pursuit. By contrast, the scent stimuli were ineffective at inciting avoidance behaviour. Our findings indicate that deterrents are a prospective low-cost and in situ method to manage rhino movement in game reserves

Identification of flavin-containing monooxygenase 5 (FMO5) as a regulator of glucose homeostasis and a potential sensor of gut bacteria

We have previously identified flavin-containing monooxygenase 5 (FMO5) as a regulator of metabolic aging. The aim of the present study was to investigate the role of FMO5 in glucose homeostasis and the impact of diet and gut flora on the phenotype of mice in which the Fmo5 gene has been disrupted (Fmo5−/− mice). In comparison with wild-type (WT) counterparts, Fmo5−/− mice are resistant to age-related changes in glucose homeostasis and maintain the higher glucose tolerance and insulin sensitivity characteristic of young animals. When fed a high-fat diet, they are protected against weight gain and reduction of insulin sensitivity. The phenotype of Fmo5−/− mice is independent of diet and the gut microbiome and is determined solely by the host genotype. Fmo5−/− mice have metabolic characteristics similar to those of germ-free mice, indicating that FMO5 plays a role in sensing or responding to gut bacteria. In WT mice, FMO5 is present in the mucosal epithelium of the gastrointestinal tract where it is induced in response to a high-fat diet. In comparison with WT mice, Fmo5−/− mice have fewer colonic goblet cells, and they differ in the production of the colonic hormone resistin-like molecule β. Fmo5−/− mice have lower concentrations of tumor necrosis factor α in plasma and of complement component 3 in epididymal white adipose tissue, indicative of improved inflammatory tone. Our results implicate FMO5 as a regulator of body weight and of glucose disposal and insulin sensitivity and, thus, identify FMO5 as a potential novel therapeutic target for obesity and insulin resistance

Association between glycaemic control and quality of life in diabetes mellitus

Background: Relationship between quality of life (QOL) and haemoglobin A1c (HbA1c) amongst diabetics in the community setting is unclear. Aims: Assess the association between QOL and change in HbA1c in diabetic patients over one year. Settings and Design: Cohort study of patients from four community clinics in California, USA. Methods: Diabetic patients identified from databases using International Classification of Disease (ICD-9) codes were asked to complete Short Form 36 (SF-36), which measures health-related QOL, and invited to attend monthly diabetes workshops. From December 2000 to December 2001, data were collected on multiple parameters, including HbA1c. SF-36 surveys were re-collected at project termination. Statistical Analysis: Regression analysis was used to correlate change in HbA1c with change in QOL physical component summary (PCS) and mental component summary (MCS) scores, while considering potential confounders. Results: Of 1679 eligible patients, 380 completed SF-36 at project initiation. 243 of those completed SF-36 at project termination. Pre and post HbA1c data were available for 170 of the 243 who completed SF-36 at both times. Average MCS increased by 8.46% and PCS decreased by 2.24%. After adjustment, a 5% decrease in HbA1c values was associated with a 1% increase in MCS. No association between changes in HbA1c and PCS was observed. Conclusions: Association between better HbA1c and improved mental, but not physical, QOL may reflect physical inconvenience of increased regimen complexity and mental empowerment from proactive disease management. Larger cohort studies with longer follow-up are needed to further elucidate the relationship between glycemic control and QOL

Negligible hormonal response following dehorning in free-ranging white rhinoceros (Ceratotherium simum)

The white rhinoceros (Ceratotherium simum) is experiencing unsustainable poaching losses fuelled by a demand for horn. Increasingly, private and state reserves are dehorning their rhinoceros populations in an attempt to reduce poaching pressure. Rhinoceroses use their horns in social interactions as well as during resource access and so its partial removal as part of reserve management practices may adversely influence these behaviours. Physiological stress can correlate with animal welfare, reproductive state and health and thus acts as a useful indicator of these parameters. To establish whether dehorning causes a physiological stress response, glucocorticoid and gonadal steroid profiles of free-ranging white rhinoceroses were determined through the collection and analysis of faecal steroid metabolites before and after dehorning. Faecal corticoid profiles were not influenced by the number of occasions a rhinoceros had been dehorned or by the number of days that had elapsed since dehorning. Furthermore, there was no apparent suppression in the concentrations of testosterone or progesterone metabolites in males and females, respectively, after exposure to multiple dehorning procedures. These findings should increase wildlife managers' confidence that dehorning does not negatively impact white rhinoceros physiology as measured hormonally

Hospital Episode Statistics and trends in ophthalmic surgery 1998 – 2004

BACKGROUND: Hospital episode statistics (HES) is a UK national database for the National Health Service (NHS), now available online. The purpose of this study was to observe trends in ophthalmic operations performed during the period from 1998 to 2004, using this data. METHODS: From the 'Main Operation' codes within the 'Free data' section of the HES website we analysed data in regard to 28 specific ophthalmic operations. These represented each sub speciality within ophthalmology. RESULTS: The figures show a change in the total number and proportions of operations performed for many of the procedures. For example, there was an increase in numbers of orbital decompressions, but a decrease in numbers of glaucoma filtering operations. Changing trends could be seen in different surgical areas such as the change in operations used for corneal grafting and in retinal surgery. CONCLUSION: The HES database represents an important, potentially useful source of information. There are imitations in interpretation of and validity of such data related to coding inconsistencies. We suggest the benefit of the data comes from observing trends rather than exact numbers. As other studies using this data have suggested, it is important that clinicians are involved in improving the quality of this data