Bacterial dormancy: A subpopulation of viable but non-culturable cells demonstrates better fitness for revival.

The viable but non culturable (VBNC) state is a condition in which bacterial cells are viable and metabolically active, but resistant to cultivation using a routine growth medium. We investigated the ability of V. parahaemolyticus to form VBNC cells, and to subsequently become resuscitated. The ability to control VBNC cell formation in the laboratory allowed us to selectively isolate VBNC cells using fluorescence activated cell sorting, and to differentiate subpopulations based on their metabolic activity, cell shape and the ability to cause disease in Galleria mellonella. Our results showed that two subpopulations (P1 and P2) of V. parahaemolyticus VBNC cells exist and can remain dormant in the VBNC state for long periods. VBNC subpopulation P2, had a better fitness for survival under stressful conditions and showed 100% revival under favourable conditions. Proteomic analysis of these subpopulations (at two different time points: 12 days (T12) and 50 days (T50) post VBNC) revealed that the proteome of P2 was more similar to that of the starting microcosm culture (T0) than the proteome of P1. Proteins that were significantly up or down-regulated between the different VBNC populations were identified and differentially regulated proteins were assigned into 23 functional groups, the majority being assigned to metabolism functional categories. A lactate dehydrogenase (lldD) protein, responsible for converting lactate to pyruvate, was significantly upregulated in all subpopulations of VBNC cells. Deletion of the lactate dehydrogenase (RIMD2210633:ΔlldD) gene caused cells to enter the VBNC state significantly more quickly compared to the wild-type, and adding lactate to VBNC cells aided their resuscitation and extended the resuscitation window. Addition of pyruvate to the RIMD2210633:ΔlldD strain restored the wild-type VBNC formation profile. This study suggests that lactate dehydrogenase may play a role in regulating the VBNC state.

Linguistic Variation in Iranian University Student Graffiti: Examining the Role of Gender.

Research on the discursive features of graffiti in institutional settings is in its infancy and few studies have investigated the phenomenon and its implications in educational contexts. In this paper, we report on a study in which we employed systemic functional linguistics (Halliday in Learning how to mean, Edward Arnold, 1975). to probe communicative functions and gender differences in Iranian university student graffiti that appeared in all-male and all-female locations. The data comprised authentic instances of graffiti generated by students, analysis of which suggests that male and female university students each have their own distinctive motives for using graffiti, as realised in significant differences observed in the context-specific functions they perform. Graffiti pieces represented a distinctive and meaningful way of communicating, and its most salient features were creativity, simplicity and variation. Indications are that university students' graffiti reflects psychological and social challenges, and the thoughts, attitudes and feelings expressed through it serve students' personal and interactional purposes.

SMOKING STATUS AND TREATMENT OUTCOMES OF VASCULAR ENDOTHELIAL GROWTH FACTOR INHIBITORS FOR NEOVASCULAR AGE-RELATED MACULAR DEGENERATION.

PURPOSE: To assess whether smoking status affects 1-year visual outcomes in eyes treated with vascular endothelial growth factor inhibitors for neovascular age-related macular degeneration. METHODS: Retrospective analysis of data from a prospectively designed, multicenter, observational database. Nine hundred and eighty seven treatment-naive eyes of patients with neovascular age-related macular degeneration were tracked by the Fight Retinal Blindness! outcome registry in Australia, New Zealand, Singapore, and Switzerland who had documented smoking status at baseline and commenced vascular endothelial growth factor inhibitor therapy from January 2006 to December 2016. Generalized additive models were used to display visual acuity results. RESULTS: There was a significant difference in mean improvement in visual acuity at 12 months between nonsmokers, ex-smokers, and current smokers (7.7 vs. 6.1 vs. 3.5 letters of change; P = 0.046) among patients who completed 12 months of treatment when adjusted for age, baseline visual acuity, and choroidal neovascular membrane lesion type and nested for practice. There was no significant difference in the median number of injections over 12 months of treatment by smoking status. Current smokers were a mean of 6.2 years younger than nonsmokers when they started treatment (P < 0.001). CONCLUSION: This study found inferior 12-month visual outcomes in patients who continued to smoke while receiving vascular endothelial growth factor inhibitor therapy for neovascular age-related macular degeneration.

New Zealand ophthalmologists' opinions and behaviours on climate, carbon and sustainability.

IMPORTANCE: Ophthalmology faces imperatives to improve sustainability, but there is uncertainty about how to respond. BACKGROUND: We sought New Zealand ophthalmologists' opinions on climate change, sustainability and the role of ophthalmologists in responding to these issues, as well as information on the extent that ophthalmology practices are acting on sustainability. DESIGN: Anonymous online survey of New Zealand fellows and trainees (178) of the Royal Australian and New Zealand College of Ophthalmologists (RANZCO) was conducted. PARTICIPANTS: Forty-seven respondents (response rate 26%) were included in the study. METHODS: Respondents were asked their level of agreement with statements on climate, health and sustainability and invited to comment. Current sustainability activities were collected from clinical leaders and directors of hospital departments and private practices. MAIN OUTCOME MEASURE: Distribution of agreement scores was the main outcome measure. RESULTS: Agreement with mainstream positions on climate change was as expected. A minority of up to 19% expressed the opinion that climate change was not due to human activity, and did not require mitigation. Younger ophthalmologists tended to have greater agreement with the need for broad-based political action on climate mitigation than those aged over 50 years. Most practices had room to improve on reducing waste, travel and carbon footprints. CONCLUSIONS AND RELEVANCE: The majority of New Zealand ophthalmologists are concerned about anthropogenic climate change. Currently, sustainability is not a performance indicator for New Zealand district health boards, so there is limited incentive to drive improvements. These data form a reference point to compare future opinions and ophthalmology carbon footprinting.

Short-term outcomes of small incision cataract surgery provided by a regional population in the Pacific.

BACKGROUND: This study aims to describe patient demographics, visual and surgical outcomes of a cohort undergoing small incision cataract surgery at a Pacific regional ophthalmic training facility. DESIGN: This is a prospective, longitudinal study conducted at the Pacific Eye Institute, Fiji. PARTICIPANTS: One hundred fifty-six patients underwent small incision cataract surgery. METHODS: Preoperatively, a complete ophthalmic examination, including pachymetry and macular optical coherence tomography performed by two independent investigators. Temporal small incision cataract surgery with intraocular lens insertion was performed by ophthalmologists, and trainee ophthalmologists. Follow-up occurred at day one, four weeks and 3 months. MAIN OUTCOME MEASURES: Patient characteristics, visual outcomes including surgically induced astigmatism and complications. RESULTS: Ninety-one per cent (142/156) attended 3-month follow-up with median age 63 years (range 19-82), 52% female (81/156) and 58% (90/156) Fijian ethnicity. In 37% (57/156), the contralateral eye was pseudophakic. Mean preoperative best spectacle-corrected visual acuity was logMAR 1.44 (6/165). At 3 months, 74% (105/142) achieved ≥6/18 unaided vision (94% [133/142] best spectacle-corrected visual acuity). Mean postoperative spherical equivalent was -0.78 (SD 0.95) D and mean surgically induced astigmatism 1.04 (0.57) D. Complications included posterior capsular rupture with vitreous loss (4% [6/156]), optical coherence tomography confirmed macular oedema (8% [12/152] at 4 weeks), significant posterior capsular opacity (23% [31/136]) and >5% increase in preoperative pachymetry (1% [1/142]) at 3 months. Reduction in preoperative best spectacle-corrected visual acuity occurred in 2% (3/142). No cases of endophthalmitis. CONCLUSION AND RELEVANCE: Small incision cataract surgery provided to a regional population is effective with 94% achieving the World Health Organisation's definition of 'Good Vision' (≥6/18). Refinements in biometric and surgical techniques may allow a greater proportion of patients to achieve good unaided vision.

Scope for genetic rescue of an endangered subspecies though re-establishing natural gene flow with another subspecies.

Genetic diversity is positively linked to the viability and evolutionary potential of species but is often compromised in threatened taxa. Genetic rescue by gene flow from a more diverse or differentiated source population of the same species can be an effective strategy for alleviating inbreeding depression and boosting evolutionary potential. The helmeted honeyeater Lichenostomus melanops cassidix is a critically endangered subspecies of the common yellow-tufted honeyeater. Cassidix has declined to a single wild population of ~130 birds, despite being subject to intensive population management over recent decades. We assessed changes in microsatellite diversity in cassidix over the last four decades and used population viability analysis to explore whether genetic rescue through hybridization with the neighbouring Lichenostomus melanops gippslandicus subspecies constitutes a viable conservation strategy. The contemporary cassidix population is characterized by low genetic diversity and effective population size (N(e) < 50), suggesting it is vulnerable to inbreeding depression and will have limited capacity to evolve to changing environments. We find that gene flow from gippslandicus to cassidix has declined substantially relative to pre-1990 levels and argue that natural levels of gene flow between the two subspecies should be restored. Allowing gene flow (~4 migrants per generation) from gippslandicus into cassidix (i.e. genetic rescue), in combination with continued annual release of captive-bred cassidix (i.e. demographic rescue), should lead to positive demographic and genetic outcomes. Although we consider the risk of outbreeding depression to be low, we recommend that genetic rescue be managed within the context of the captive breeding programme, with monitoring of outcomes.

Koalas (Phascolarctos cinereus) From Queensland Are Genetically Distinct From 2 Populations in Victoria.

The koala (Phascolarctos cinereus) suffered population declines and local extirpation due to hunting in the early 20th century, especially in southern Australia. Koalas were subsequently reintroduced to the Brisbane Ranges (BR) and Stony Rises (SR) by translocating individuals from a population on French Island descended from a small number of founders. To examine genetic diversity and north-south differentiation, we genotyped 13 microsatellite markers in 46 wild koalas from the BR and SR, and 27 Queensland koalas kept at the US zoos. The Queensland koalas displayed much higher heterozygosity (H O = 0.73) than the 2 southern Australian koala populations examined: H O = 0.49 in the BR, whereas H O = 0.41 in the SR. This is consistent with the historical accounts of bottlenecks and founder events affecting the southern populations and contrasts with reports of high genetic diversity in some southern populations. The 2 southern Australian koala populations were genetically similar (F ST = 0.018, P = 0.052). By contrast, northern and southern Australian koalas were highly differentiated (F ST = 0.27, P < 0.001), thereby suggesting that geographic structuring should be considered in the conservation management of koalas. Sequencing of 648bp of the mtDNA control region in Queensland koalas found 8 distinct haplotypes, one of which had not been previously detected among koalas. Queensland koalas displayed high mitochondrial haplotype diversity (H = 0.753) and nucleotide diversity (π = 0.0072), indicating along with the microsatellite data that North American zoos have maintained high levels of genetic diversity among their Queensland koalas.

Evaluation of the World Health Organization outcome standards at the early and late post-operative visits following cataract surgery.

INTRODUCTION: This study was conducted to determine whether the World Health Organization (WHO) visual acuity standards are correlated between the early and late early post-operative periods following phacoemulsification (phaco) and small incision extracapsular cataract surgery (SICS). Secondary aims were to compare visual outcomes and complications following SICS and phaco. METHODS: Retrospective cohort study following phaco and SICS performed by one surgeon. Primary outcome measures included uncorrected visual acuity (UCVA) and best corrected visual acuity (BCVA) at the early (≤72 hours) and late (≥21 days) post-operative visits. Secondary outcome measures included complications and astigmatism. RESULTS: 705 eyes were studied (509 phaco, 196 SICS). The correlation for UCVA between early and late follow-up examinations was higher for phaco (rs=0.58) compared to SICS (rs=0.45, p=0.04) while correlation for BCVA was similar (phaco,rs=0.52; SICS, rs=0.47; p=0.44). At the early post-operative visit, a higher proportion in the phaco group achieved ≥6/18 UCVA (81.5% phaco vs 64.8% SICS, p<0.0001) and BCVA (87.8% phaco vs 73.5% SICS, p<0.0001). At the late post-operative visit, a higher proportion following phaco also achieved ≥6/18 UCVA (93.9% phaco vs 85.2% SICS, p=0.0004) and BCVA (96.9% phaco vs 91.3% SICS, p=0.004). After exclusion of eyes with pre-existing ocular comorbidities, a similar proportion had ≥6/18 late UCVA (98.9% phaco vs 96.9% SICS, p=0.22) and BCVA (100% phaco vs 99.2% SICS, p=0.27). CONCLUSIONS: Early and late post-operative WHO visual acuity levels are correlated, but not equivalent, following both phaco and SICS. In eyes without comorbidities, similar final visual outcomes can be achieved after phaco and SICS.

Ophthalmic Simulated Surgical Competency Assessment Rubric for manual small-incision cataract surgery.

PURPOSE: To develop and test the validity of a surgical competency assessment tool for simulated small-incision cataract surgery (SICS). SETTING: Participating ophthalmologists contributed from 8 countries. DESIGN: Qualitative and quantitative development and evaluation of face and content validity of an assessment rubric, and evaluation of construct validity and reliability. METHODS: The SICS Ophthalmic Simulated Surgical Competency Assessment Rubric (Sim-OSSCAR) was developed and assessed for face and content validity by an international group of experienced ophthalmologists. Groups of novice and competent surgeons from 4 countries were recorded performing surgery, and masked assessments were performed by 4 expert surgeons, to determine construct validity and reliability. RESULTS: The Sim-OSSCAR for SICS was assessed by a panel of 12 international experts from 8 countries. In response to the question, "Do you think the OSSCAR represents the surgical techniques and skills upon which trainees should be assessed?," all respondents either agreed or strongly agreed. Face validity was rated as 4.60 (out of 5.0). The content was iteratively agreed to by the panel of experts; final content validity was rated as 4.5. Interobserver reliability was assessed, and 17 of 20 items in the assessment matrix had a Krippendorff α correlation of more than 0.6. A Wilcoxon rank-sum test showed that competent surgeons perform better than novices (P = .02). CONCLUSIONS: This newly developed and validated assessment tool for simulation SICS, based on the International Council of Ophthalmology's Ophthalmology Surgical Competency Assessment Rubric, has good face and content validity. It can play a role in ophthalmic surgical education.

Improved Refractive Outcomes of Small-Incision Extracapsular Cataract Surgery after Implementation of a Biometry Training Course.

PURPOSE: To determine whether a biometry training course could improve refractive outcomes of patients undergoing manual small-incision extracapsular cataract surgery (SICS). MATERIALS AND METHODS: This was a prospective, interventional, cohort study at the Pacific Eye Institute, Fiji. SICS refractive outcomes were evaluated before and after a structured biometry teaching course. Eyes that underwent evaluation and subsequent SICS with placement of a posterior chamber intraocular lens (IOL) were included. Axial length measurements were obtained using A-scan applanation ultrasound and keratometry with a handheld keratometer. Main outcome measures included mean absolute prediction error of IOL calculations, percentage of eyes within ±0.5 D and ±1.0 D of intended spherical equivalent, and proportion of eyes with ≥6/18 uncorrected visual acuity. RESULTS: A total of 240 eyes were analyzed: 120 eyes before and 120 eyes after the structured biometry training. The mean absolute prediction error was 50% lower following the training (1.13 ± 0.84 D pre vs. 0.56 ± 0.44 D post; P < 0.001). A higher percentage of the eyes had a postoperative spherical equivalent within ±0.5 D (26.7% pre vs. 52.5% post; P < 0.001) and ±1.0 D (55.0% pre vs. 90.0% post; P < 0.001) of the intended target. A higher proportion of the eyes achieved ≥6/18 uncorrected visual acuity (77.5% pre vs. 91.7% post, P = 0.004), while the proportion with ≥6/18 corrected visual acuity was similar (94.4% pre vs. 98.3% post; P = 0.28). CONCLUSIONS: A structured biometry training course may improve the accuracy of preoperative IOL calculations to achieve the postoperative refractive target. Ophthalmology training programs should include structured biometry teaching in their curricula.

Lifetime Fitness Costs of Inbreeding and Being Inbred in a Critically Endangered Bird.

Reduced fitness as a result of inbreeding is a major threat facing many species of conservation concern [1-4]. However, few case studies for assessing the magnitude of inbreeding depression in the wild means that its relative importance as a risk factor for population persistence remains under-appreciated [5]. The increasing availability and affordability of genomic technologies provide new opportunities to address knowledge gaps around the magnitude and manifestation of inbreeding depression in wild populations [6-12]. Here, we combine over three decades of individual lifetime reproductive data and genomic data to estimate the relative lifetime and short-term fitness costs of both being inbred and engaging in inbreeding in the last wild population (<250 individuals remaining) of an iconic and critically endangered bird: the helmeted honeyeater Lichenostomus melanops cassidix. The magnitude of inbreeding depression was substantial: the mean predicted lifetime reproductive success of the most inbred (homozygosity = 0.82) individuals was on average 87%-90% lower than that of the least inbred (homozygosity = 0.75). For individual reproductive events and lifetime measures, we provide rare empirical evidence that pairing with a genetically dissimilar individual can reduce fitness costs associated with being an inbred individual. By comparing lifetime and short-term fitness measures, we demonstrate how short-term measures of reproductive success that are associated with only weak signatures of inbreeding depression can still underlie stronger lifetime effects. Our study represents a valuable case study, highlighting the critical importance of inbreeding depression as a factor influencing the immediate viability of populations in threatened species management.

Randomized, Controlled, Double-Masked, Multicenter, Pilot Study Evaluating Safety and Efficacy of Intranasal Neurostimulation for Dry Eye Disease.

Purpose: We assess the safety and effectiveness of intranasal neurostimulation to promote tear production via the nasolacrimal pathway in subjects with dry eye disease. Methods: A multicenter, randomized, controlled, double-masked pilot study was conducted in adults with dry eye diagnosis and at least one eye with corneal fluorescein staining ≥2 in at least one region or a sum of all regions ≥5 (National Eye Institute grading), basal Schirmer test score ≤10 mm, a cotton-swab stimulated Schirmer score ≥7 mm higher, and an Ocular Surface Disease Index score ≥23. Subjects were randomized to receive active intranasal neurostimulation or sham control intranasal stimulation 4 to 8 times per day. Assessments were scheduled before (unstimulated) and during (stimulated) device application at days 0, 7, 14, 30, and 90. The primary effectiveness endpoint was stimulation-induced change in Schirmer test (with anesthesia) score. Primary safety measure was incidence of device-related adverse events (AEs). Results: Fifty-eight subjects were randomized at nine sites in Australia and New Zealand; 56 completed the 90-day study. Stimulation-induced change in Schirmer score was significantly greater with active intranasal (mean ± SEM, 9.0 ± 2.0) than sham control intranasal stimulation (0.4 ± 0.6; P < 0.001) at day 90. Similar results were observed at days 0, 7, 14, and 30 (P < 0.001). No serious device-related AEs were observed. Mild nosebleed, the most common device-related AE, was reported in five (16.7%) subjects. Conclusions: Intranasal neurostimulation was effective in inducing acute tear production after 90 days of use and generally was well tolerated in subjects with dry eye disease.

Thrombocytopenia in the neonate.

Thrombocytopenia is one of the commonest haematological problems in neonates, affecting at least 25% of all admissions to neonatal intensive care units (NICUs) [Murray NA, Howarth LJ, McCloy MP et al. Platelet transfusion in the management of severe thrombocytopenia in neonatal intensive care unit patients. Transfus Med 2002;12:35-41; Garcia MG, Duenas E, Sola MC et al. Epidemiologic and outcome studies of patients who received platelet transfusions in the neonatal intensive care unit. J Perinatol 2001;21:415-20; Del Vecchio A, Sola MC, Theriaque DW et al. Platelet transfusions in the neonatal intensive care unit: factors predicting which patients will require multiple transfusions. Transfusion 2001;41:803-8]. Although a long list of disorders associated with neonatal thrombocytopenia can be found in many textbooks, newer classifications based on the timing of onset of thrombocytopenia (early vs. late) are more useful for planning diagnostic investigations and day-to-day management. The mainstay of treatment of neonatal thrombocytopenia remains platelet transfusion although it is important to note that no studies have yet shown clinical benefit of platelet transfusion in this setting. Indeed some reports even suggest that there may be significant adverse effects of platelet transfusion in neonates, including increased mortality, and that the effects of transfusion may differ in different groups of neonates with similar degrees of thrombocytopenia [Bonifacio L, Petrova A, Nanjundaswamy S, Mehta R. Thrombocytopenia related neonatal outcome in preterms. Indian J Pediatr 2007;74:269-74; Kenton AB, Hegemier S, Smith EO et al. Platelet transfusions in infants with necrotizing enterocolitis do not lower mortality but may increase morbidity. J Perinatol 2005;25:173-7]. There is also considerable variation in transfusion practice between different countries and between different neonatal units. Here we review recent progress in understanding the prevalence, causes and pathogenesis of thrombocytopenia in the newborn, the clinical consequences of thrombocytopenia and developments in neonatal platelet transfusion.

Neonatal thrombocytopenia.

Thrombocytopenia (platelets <150 x 10(9)/L) is one of the most common haematological problems in neonates, particularly those who are preterm and sick. In those preterm neonates with early thrombocytopenia who present within 72 h of birth, the most common cause is reduced platelet production secondary to intrauterine growth restriction and/or maternal hypertension. By contrast, the most common causes of thrombocytopenia arising after the first 72 h of life, both in preterm and term infants, are sepsis and necrotizing enterocolitis. The most important cause of severe thrombocytopenia (platelets <50 x 10(9)/L) is neonatal alloimmune thrombocytopenia (NAIT), as diagnosis can be delayed and death or long-term disability due to intracranial haemorrhage may occur. Platelet transfusion is the mainstay of treatment for severe thrombocytopenia. However, the correlation between thrombocytopenia and bleeding is unclear and no studies have yet shown clinical benefit for platelet transfusion in neonates. Studies to identify optimal platelet transfusion practice for neonatal thrombocytopenia are urgently required.

Nucleus extraction in sutureless surgery: getting hooked!

Simple modifications to a readily available disposable 25 gauge cannula facilitate nucleus extraction in sutureless extracapsular cataract surgery. A case series using the modified cannula, and a temporal incision approach, are presented. Outcomes and complications attributable to the use of the cannula are discussed.