Single molecule RNA sequencing uncovers trans-splicing and improves annotations in Anopheles stephensi.

Single molecule real-time (SMRT) sequencing has recently been used to obtain full-length cDNA sequences that improve genome annotation and reveal RNA isoforms. Here, we used one such method called isoform sequencing from Pacific Biosciences (PacBio) to sequence a cDNA library from the Asian malaria mosquito Anopheles stephensi. More than 600 000 full-length cDNAs, referred to as reads of insert, were identified. Owing to the inherently high error rate of PacBio sequencing, we tested different approaches for error correction. We found that error correction using Illumina RNA sequencing (RNA-seq) generated more data than using the default SMRT pipeline. The full-length error-corrected PacBio reads greatly improved the gene annotation of Anopheles stephensi: 4867 gene models were updated and 1785 alternatively spliced isoforms were added to the annotation. In addition, six trans-splicing events, where exons from different primary transcripts were joined together, were identified in An. stephensi. All six trans-splicing events appear to be conserved in Culicidae, as they are also found in Anopheles gambiae and Aedes aegypti. The proteins encoded by trans-splicing events are also highly conserved and the orthologues of these proteins are cis-spliced in outgroup species, indicating that trans-splicing may arise as a mechanism to rescue genes that broke up during evolution.

Bevacizumab as an adjunct to vitreoretinal surgery for diabetic retinopathy in East Africa.

PURPOSE: The purpose of this study is to evaluate the efficacy of preoperative intravitreal bevacizumab (IVB) for improving outcomes in vitrectomy for diabetic retinopathy-related non-clearing vitreous haemorrhage and/or tractional retinal detachment. METHODS: Medical record from patients undergoing vitrectomy for proliferative diabetic retinopathy (PDR) were retrospectively analysed (2003-2011). From 2007, IVB (1.25 mg 2-4 days before operating) was used on all eyes. Eyes receiving IVB were compared with those that did not receive IVB. Intraoperative complications, reoperation rates, and final visual acuity were the core outcome measures. RESULTS: Data were analysed for 88 patients (101 eyes). In all, 41 (41%) patients had received IVB, whereas 60 (59%) patients had not. Significant intraoperative haemorrhage occurred in six eyes (10%) in the non-IVB group and in one (2.4%) IVB eyes (P=0.24). Silicon oil was used in 29 (48%) non-IVB eyes and in 11 (27%) IVB eyes (P=0.03). The non-IVB eyes underwent significantly more vitreoretinal reoperations (P=0.01) and were significantly more likely to lose two or more lines of vision at the final follow-up (P=0.03). The numbers needed to treat (NNT) blindness (<3/60) was four for non-IVB eyes and two for the IVB group. CONCLUSIONS: IVB reduces surgical complications, the use of silicon oil, and the need for further retinal surgery. The NNT to restore useful vision (≥3/60) to a blind eye were significantly lower in the IVB group. Vitreoretinal surgery for the complications of PDR is effective in an East African context, and IVB should be considered a valuable adjunct.

Outcomes and reasons for dacryocystorhinostomy (DCR) at KCMC, a Tanzanian referral hospital, 2001-2006.

BACKGROUND: External dacryocystorhinostomy (DCR) is a surgical intervention aimed to treat blocked nasolacrimal ducts of almost all causes. To date there is only limited data available from the Sub Saharan African setting. OBJECTIVES: This study aimed to provide further information of the outcomes of DCR in Africa. METHODS: Records of all patients undergoing external DCR operations from January 2001 to April 2006 were systematically searched. 55 patients were identified and notes were available for 45 patients. RESULTS: Discharge and epiphora were resolved in 90.9% (30/33) and 84.4% (27/32) of patients respectively. Over half the cases (51.1%) were children. The commonest reason for operation was chronic dacryocystitis (51.1%). Outcomes for DCR were not significantly different for either children or adults and a clear improvement of symptoms was found in the vast majority of cases. CONCLUSION: This study provides information on the outcomes of DCR in the African population. An 84.4% cure rate of epiphora and 90.9% cure rate of discharge is comparable with findings in other developing countries. This study supports the continued use of this intervention in skilled hands for treatment of blocked nasolacrimal duct.

Use of cataract services in eastern Africa--a study from Tanzania.

PURPOSE: To establish the proportion of patients who are blind or have low vision prior to undergoing cataract surgery at tertiary referral centers in Tanzania. To assess which patient groups presenting for cataract surgery are more likely to be blind or visually impaired. METHODS: Using pre-existing computerized audit systems we gathered data on pre-operative visual status, age, gender and presentation mode (walk-in or outreach) for 3765 patients undergoing 4258 cataract operations at 2 hospitals in Tanzania. Visual status was defined based on vision in the better eye. RESULTS: 32% of operations were performed on blind patients, 37% on patients with low vision and 31% on normally sighted patients. Predictors of blindness at presentation were: female sex (OR 1.15; 95% CI 1.00-1.32); referral from a rural outreach program (OR 1.75; 95% CI 1.51-2.02) and older age (OR 1.02; 95% CI 1.01-1.02). CONCLUSIONS: It is not only the blind who present to cataract services in Tanzania. The demand for surgery amongst patients who or are normally sighted represents a positive move towards prevention, and not only cure of cataract blindness in Tanzania. However, it also highlights the need to target those left blind from cataract in order to deliver services to those most in need. Cataract programs targeting patients in rural areas and older patients are likely to increase the number of blind patients benefiting from cataract services.

Extent and economic effect of heat loads on dairy cattle production in Australia.

OBJECTIVE: To investigate the extent of heat load problems, caused by the combination of excessive temperature and humidity, in Holstein-Friesian cows in Australia. Also, to outline how milk production losses and consequent costs from this can be estimated and minimised. PROCEDURES: Long-term meteorological data for Australia were analysed to determine the distribution of hot conditions over space and time. Fifteen dairy production regions were identified for higher-resolution data analysis. Both the raw meteorological data and their integration into a temperature-humidity thermal index were compiled onto a computer program. This mapping software displays the distribution of climatic patterns, both Australia-wide and within the selected dairying regions. Graphical displays of the variation in historical records for 200 locations in the 15 dairying regions are also available. As a separate study, production data from research stations, on-farm trials and milk factory records were statistically analysed and correlated with the climatic indices, to estimate production losses due to hot conditions. RESULTS: Both milk yields and milk constituents declined with increases in the temperature-humidity index. The onset and rate of this decline are dependent on a number of factors, including location, level of production, adaptation, and management regime. These results have been integrated into a farm-level economic analysis for managers of dairy properties. CONCLUSION: By considering the historical patterns of hot conditions over time and space, along with expected production losses, managers of dairy farms can now conduct an economic evaluation of investment strategies to alleviate heat loads. These strategies include the provision of sprinklers, shade structures, or combinations of these.

LOCS III versus the Oxford Clinical Cataract Classification and Grading System for the assessment of nuclear, cortical and posterior subcapsular cataract.

PURPOSE: To compare two methods of cataract grading for nuclear cataract, cortical cataract and posterior subcapsular cataract. METHODS: The Melton Eye Study is an English community-based epidemiological study of the natural history of eye disease in people aged 55 to 74 years. The lenses of both eyes of 560 subjects were graded at the slit lamp using two cataract grading systems. The Oxford Clinical Cataract Classification and Grading System (OCCCGS) employs standard diagrams and Munsell colour samples for the grading of cortical, posterior subcapsular and nuclear cataract. The Lens Opacities Classification System III (LOCS III) uses photographic transparencies of the lens as standards. In both systems a decimalised score is assigned. We also graded the LOCS III standard images using the OCCCGS. Interobserver variation was calculated for both grading systems. Linear calibration lines are plotted for each type of lens opacity. RESULTS: The relationship between LOCS III and OCCCGS for nuclear cataract and posterior subcapsular cataract is linear. The relationship for cortical cataract is linear once the LOCS III scores are squared. The intervals between the LOCS III images when ranked by the human eye using the OCCCGS are linear. Interobserver variation for both systems is good. CONCLUSIONS: The linear calibration lines may be used to convert from one system to another and will be useful in comparing studies or performing meta-analysis. These results show that data from cataract studies using different clinical grading schemes can be compared.

Prevalence of lenticular abnormalities in a population-based study: Oxford Clinical Cataract Grading in the Melton Eye Study.

PURPOSE: To describe the distribution of the 11 features assessed by the Oxford Clinical Cataract Classification and Grading System (OCCCGS) in a population-based study. METHODS: The Melton Eye Study is an English community-based epidemiological study of the natural history of eye disease in people aged 55 to 74 years. Both lenses of 560 subjects were graded at the slit lamp using a decimalised version of the OCCCGS. Subject prevalences were estimated by logistic regression and the extent, when present, against normal errors regression. RESULTS: White nuclear scatter (WNS), brunescence, cortical spoke (CS), anterior subcapsular opacity (ASC), fibre folds (FF), waterclefts (WC) and perinuclear retro-dots all increased with age (p < 0.05). Posterior subcapsular opacity (PSC), vacuoles, focal dots (FD) and coronary flakes (CF) did not significantly increase with age. Subject prevalences of the features and the mean Oxford scores when present were: WNS (1.33), brunescence (0.88), CS 36% (0.34), PSC 11% (0.52), ASC 2% (0.53), FF 18% (0.53), WC 17% (0.29), retro-dots 11% (1.15), vacuoles 59% (0.43), FD 98% (1.79) and CF 39% (1.24). Significantly more common in women were both coronary flakes (p < 0.001) and waterclefts (p < 0.05). CONCLUSIONS: These are the first data on the distribution of these minor lesions in a population-based study. Coronary flakes and waterclefts are more common in women.

Associations between lens features assessed in the Oxford Clinical Cataract Classification and Grading System.

PURPOSE: To study the associations between eleven lens features graded according to the Oxford Clinical Cataract Classification and Grading System (OCCCGS). METHOD: 560 subjects taking part in the Melton Eye Study had their lenses graded according to the OCCCGS by one of two examiners. Associations between features were examined using log-linear models for categorised grades. Adjustment was made for age, sex and grader. RESULTS: Within subjects, the two nuclear features, white nuclear scatter and brunescence, are closely related with one another, as are coronary flakes and focal-dots, but these two groupings are negatively associated. Cortical spoke, fibrefolds and waterclefts are all associated with one another and this group is positively associated with coronary flakes and focal-dots. Posterior subcapsular and anterior subcapsular opacity are associated with one another and with cortical spokes. A within-eye analysis gives similar results. CONCLUSION: These associations may be important in defining cataract subtypes and in identifying minor features that indicate early cataract development.

Innovative clinical teaching.

Participating faculty are entering their third year using this type of student rotation and have found that it continues to be a positive teaching-learning experience for faculty, staff, and students. It has enabled a greater number of students to enjoy a positive clinical experience in areas that previously were limited to a smaller number of students. Perhaps this strategy, or a part of it, may be of benefit to other nurse educators as a method of improving undergraduate clinical experiences.

Stress modification of allelopathy ofHelianthus annuus L. debris on seedling biomass production ofAmaranthus retroflexus L.

When added to soil,Helianthus annuus debris from plants grown under various nutrient stresses in the greenhouse and field had a significant negative effect onAmaranthus retroflexus seedling dry weight production. The regression models for dry weight versus total phenolic compounds and the variation in N, P, and K contents of theHelianthus debris were significant. It was possible to partially simulate debris inhibition ofA. retroflexus seedling dry weight production when chlorogenic acid alone was added to the soil instead ofHelianthus debris. The inhibition ofAmaranthus growth byHelianthus debris and chlorogenic acid was not evident when nutrient solution was applied to the soil.

A thermodynamic view of polymer degradation, and its application to extensive sonication of DNA.

A simple thermodynamic theory is developed, which predicts (in agreement with a wide variety of other theoretical approaches and experimental results) that for simple polymers the most probable Schulz distribution of fragments will be obtained in a polymer degradation process that is allowed to proceed to a dynamic equilibrium. When the same method is applied to a double-stranded polymer like DNA, however, it predicts that some narrowing of this distribution may occur in conjunction with a limited amount of base unpairing at the fragment termini. The compatibility of this prediction with the experimental results of long-time sonication of DNA is considered.