Study protocol for an individually randomized control trial for India's first roleplay-based mobile game for reproductive health for adolescent girls.

BACKGROUND: Go Nisha Go™ (GNG), is a mobile game combining behavioural science, human-centric design, game-based learning, and interactive storytelling. The model uses a direct-to-consumer (DTC) approach to deliver information, products, services, interactive learning, and agency-building experiences directly to girls. The game's five episodes focus on issues of menstrual health management, fertility awareness, consent, contraception, and negotiation for delay of marriage and career. The game's effectiveness on indicators linked to these issues will be measured using an encouragement design in a randomized controlled trial (RCT). METHODS: A two-arm RCT will be conducted in three cities in India: Patna, Jaipur, and Delhi-NCR. The first arm is the treatment (encouragement) arm (n = 975) where the participants will be encouraged to download and play the game, and the second arm (n = 975) where the participants will not receive any nudges/encouragement to play the game. They may or may not have access to the game. After the baseline recruitment, participants will be randomly assigned to these two arms across the three locations. Participants of the treatment/encouragement arm will receive continuous support as part of the encouragement design to adopt, install the game from the Google Play Store at no cost, and play all levels on their Android devices. The encouragement activity will continue for ten weeks, during which participants will receive creative messages via weekly phone calls and WhatsApp messages. We will conduct the follow-up survey with all the participants (n = 1950) from the baseline survey after ten weeks of exposure. We will conduct 60 in-depth qualitative interviews (20 at each location) with a sub-sample of the participants from the encouragement arm to augment the quantitative surveys. DISCUSSION: Following pre-testing of survey tools for feasibility of methodologies, we will recruit participants, randomize, collect baseline data, execute the encouragement design, and conduct the follow-up survey with eligible adolescents as written in the study protocol. Our study will add insights for the implementation of an encouragement design in RCTs with adolescent girls in the spectrum of game-based learning on sexual and reproductive health in India. Our study will provide evidence to support the outcome evaluation of the digital mobile game app, GNG. To our knowledge this is the first ever outcome evaluation study for a game-based application, and this study is expected to facilitate scalability of a direct-to-consumer approach to improve adolescent sexual and reproductive health outcomes in India. TRIAL REGISTRATION NUMBER: ctri.nic.in: CTRI/2023/03/050447.

Our paper describes implementation of a study protocol for an outcome evaluation of a mobile game app called Go Nisha Go™, produced by the Game of Choice, Not Chance™ project, funded by USAID. Consenting adolescent girls, aged 15­19, from three cities in India will be enrolled to participate in an encouragement design led RCT. Girls will be randomly assigned to either, a) a treatment (encouragement) arm where they will be nudged to play the game for ten weeks, or b) a control arm where participants will not be provided any encouragement to download or play the game. The study will be evaluated using surveys at baseline and follow-up. The findings from this study will support the measurement of effectiveness of the digital intervention and facilitate scalability of a direct-to-consumer approach, using a game-based application to improve adolescent sexual and reproductive health outcomes in India.

A Design-Led Theory of Change for a Mobile Game App (Go Nisha Go) for Adolescent Girls in India: Multimix Methodology Study.

BACKGROUND: India has one of the largest adolescent populations in the world. Yet adolescents, particularly adolescent girls, have limited access to correct sexual and reproductive health information and services. The context in which adolescent girls live is one of gender inequity where they contend with early marriage and early pregnancy and have few opportunities for quality education and labor force participation. The digital revolution has expanded the penetration of mobile phones across India, increasingly being used by adolescent girls. Health interventions are also moving onto digital platforms. Evidence has shown that applications of game elements and game-based learning can be powerful tools in behavior change and health interventions. This provides a unique opportunity, particularly for the private sector, to reach and empower adolescent girls directly with information, products, and services in a private and fun manner. OBJECTIVE: The objective of this paper is to describe how a design-led Theory of Change (ToC) was formulated for a mobile game app that is not only underpinned by theories of various behavior change models but also identifies variables and triggers for in-game behavioral intentions that can be tracked and measured within the game and validated through a rigorous post-gameplay outcome evaluation. METHODS: We describe the use of a multimix methodology to formulate a ToC informing behavioral frameworks and co-design approaches in our proof-of-concept product development journey. This process created a statement of hypothesis and "pathways to impact" with a continuous, cumulative, and iterative design process that included key stakeholders in the production of a smartphone app. With theoretical underpinnings of social behavior and modeling frameworks, systematic research, and other creative methods, we developed a design-led ToC pathway that can delineate complex and multidisciplinary outputs for measuring impact. RESULTS: The statement of hypothesis that emerged posits that "If girls virtually experience the outcomes of choices that they make for their avatar in the mobile game, then they can make informed decisions that direct the course of their own life." Four learning pathways (DISCOVER, PLAY, DECIDE, and ACT) are scaffolded on 3 pillars of evidence, engagement, and evaluation to support the ToC-led framework. It informs decision-making and life outcomes through game-based objectives and in-game triggers that offer direct access to information, products, and services. CONCLUSIONS: This approach of using a multimix methodology for identifying varied and multidisciplinary pathways to change is of particular interest to measuring the impact of innovations, especially digital products, that do not necessarily conform with traditional behavioral change models or standard co-design approaches. We also explain the benefits of using iterative and cumulative inputs to integrate ongoing user feedback, while identifying pathways to various impacts, and not limiting it to only the design and development phase.

'Real-world' priority setting for service improvement in English primary care: a decentred approach.

This article develops an analysis of population-level priority setting informed by Bevir's decentred theory of governance and drawing on a qualitative study of priority setting for service improvement conducted in the complex multi-layered governance context of English primary care. We show how powerful actors, operating at the meso-level, utilize pluralistic and contradictory elements of complex governance networks to discursively construct, legitimize and enact service improvement priorities. Our analysis highlights the role of situated agency in integrating top-down, bottom-up and horizontal influences on priority setting, which leads to variation in local priorities despite the continuous presence of strong hierarchical influences.

Implementation of corneal confocal microscopy for screening and early detection of diabetic neuropathy in primary care alongside retinopathy screening: Results from a feasibility study.

Objective: Screening for diabetic peripheral neuropathy (DPN) is essential for early detection and timely intervention. Quantitative assessment of small nerve fiber damage is key to the early diagnosis and assessment of its progression. Corneal confocal microscopy (CCM) is a non-invasive, in-vivo diagnostic technique that provides an accurate surrogate biomarker for small-fiber neuropathy. In this novel study for the first time, we introduced CCM to primary care as a screening tool for DPN alongside retinopathy screening to assess the level of neuropathy in this novel cohort. Research design and methods: 450 consecutive subjects with type 1 or type 2 diabetes attending for annual eye screening in primary care optometry settings underwent assessment with CCM to establish the prevalence of sub-clinical diabetic peripheral neuropathy. Subjects underwent assessment for neurological and ocular symptoms of diabetes and a history of diabetic foot disease, neuropathy and diabetic retinopathy (DR). Results: CCM examination was completed successfully in 427 (94.9%) subjects, 22% of whom had neuropathy according to Diabetic Neuropathy Symptom (DNS) score. The prevalence of sub-clinical neuropathy as defined by abnormal corneal nerve fiber length (CNFL) was 12.9%. In the subjects with a short duration of type 2 diabetes, 9.2% had abnormal CNFL. CCM showed significant abnormalities in corneal nerve parameters in this cohort of subjects with reduction of corneal nerve fiber density (CNFD, p<0.001), CNFL (p<0.001) and corneal nerve branch density (CNBD, p<0.001) compared to healthy subjects. In subjects who had no evidence of DR (67% of all subjects), 12.0% had abnormal CNFL. Conclusions: CCM may be a sensitive biomarker for early detection and screening of DPN in primary care alongside retinopathy screening.

Post-discharge care following acute kidney injury: quality improvement in primary care.

BACKGROUND: Over the past decade, targeting acute kidney injury (AKI) has become a priority to improve patient safety and health outcomes. Illness complicated by AKI is common and is associated with adverse outcomes including high rates of unplanned hospital readmission. Through national patient safety directives, NHS England has mandated the implementation of an AKI clinical decision support system in hospitals. In order to improve care following AKI, hospitals have also been incentivised to improve discharge summaries and general practices are recommended to establish registers of people who have had an episode of illness complicated by AKI. However, to date, there is limited evidence surrounding the development and impact of interventions following AKI. DESIGN: We conducted a quality improvement project in primary care aiming to improve the management of patients following an episode of hospital care complicated by AKI. All 31 general practices within a single NHS Clinical Commissioning Group were incentivised by a locally commissioned service to engage in audit and feedback, education training and to develop an action plan at each practice to improve management of AKI. RESULTS: AKI coding in general practice increased from 28% of cases in 2015/2016 to 50% in 2017/2018. Coding of AKI was associated with significant improvements in downstream patient management in terms of conducting a medication review within 1 month of hospital discharge, monitoring kidney function within 3 months and providing written information about AKI to patients. However, there was no effect on unplanned hospitalisation and mortality. CONCLUSION: The findings suggest that the quality improvement intervention successfully engaged a primary care workforce in AKI-related care, but that a higher intensity intervention is likely to be required to improve health outcomes. Development of a real-time audit tool is necessary to better understand and minimise the impact of the high mortality rate following AKI.

Implementing post-discharge care following acute kidney injury in England: a single-centre qualitative evaluation.

OBJECTIVES: We sought to understand the factors influencing the implementation of a primary care intervention to improve post-discharge care following acute kidney injury (AKI). DESIGN: Qualitative study using semi-structured interviews and thematic analysis. SETTING: General practices in one Clinical Commissioning Group area in England. PARTICIPANTS: A total of 18 healthcare staff took part in interviews. Participants were practice pharmacists, general practitioners, practice managers and administrators involved in implementing the intervention. RESULTS: We identified three main factors influencing implementation: differentiation of the new intervention from other practice work; development of skill mix and communication across organisations. Overall, post-AKI processes of care were deemed straightforward to embed into existing practice. However, it was also important to separate the intervention from other work in general practice. Dedicating staff time to proactively identify AKI on discharge summaries and to coordinate the provision of care enabled implementation of the intervention. The post-AKI intervention provided an opportunity for practice pharmacists to expand their primary care role. Working in a new setting also brought challenges; time to develop trusting relationships including an understanding of boundaries of clinical expertise influenced pharmacists' roles. Unclear and inconsistent information on discharge summaries contributed to concerns about additional work in primary care. CONCLUSIONS: The research highlights challenges around post-discharge management in the primary care context. Coordination and communication were key factors for improving follow-up care following AKI. Further consideration is required to understand patient experiences of the interface between secondary and primary care. The issues pertaining to discharge care following AKI are relevant to practitioners and commissioners as they work to improve transitions of care for vulnerable patient populations.

More that unites us than divides us? A qualitative study of integration of community health and social care services.

BACKGROUND: The integration of community health and social care services has been widely promoted nationally as a vital step to improve patient centred care, reduce costs, reduce admissions to hospital and facilitate timely and effective discharge from hospital. The complexities of integration raise questions about the practical challenges of integrating health and care given embedded professional and organisational boundaries in both sectors. We describe how an English city created a single, integrated care partnership, to integrate community health and social care services. This led to the development of 12 integrated neighbourhood teams, combining and co-locating professionals across three separate localities. The aim of this research is to identify the context and the factors enabling and hindering integration from a qualitative process evaluation. METHODS: Twenty-four semi-structured interviews were conducted with equal numbers of health and social care staff at strategic and operational level. The data was subjected to thematic analysis. RESULTS: We describe three key themes: 1) shared vision and leadership; 2) organisational factors; 3) professional workforce factors. We found a clarity of vision and purpose of integration throughout the partnership, but there were challenges related to the introduction of devolved leadership. There were widespread concerns that the specified outcome measures did not capture the complexities of integration. Organisational challenges included a lack of detail around clinical and service delivery planning, tensions around variable human resource practices and barriers to data sharing. A lack of understanding and trust meant professional workforce integration remained a key challenge, although integration was also seen as a potential solution to engender relationship building. CONCLUSIONS: Given the long-term national policy focus on integration this ambitious approach to integrate community health and social care has highlighted implications for leadership, organisational design and inter-professional working. Given the ethos of valuing the local assets of individuals and networks within the new partnership we found the integrated neighbourhood teams could all learn from each other. Many of the challenges of integration could benefit from embracing the inherent capabilities across the integrated neighbourhood teams and localities of this city.

Implementation of clinical decision support to manage acute kidney injury in secondary care: an ethnographic study.

BACKGROUND: Over the past decade, acute kidney injury (AKI) has become a global priority for improving patient safety and health outcomes. In the UK, a confidential inquiry into AKI led to the publication of clinical guidance and a range of policy initiatives. National patient safety directives have focused on the mandatory establishment of clinical decision support systems (CDSSs) within all acute National Health Service (NHS) trusts to improve the detection, alerting and response to AKI. We studied the organisational work of implementing AKI CDSSs within routine hospital care. METHODS: An ethnographic study comprising non-participant observation and interviews was conducted in two NHS hospitals, delivering AKI quality improvement programmes, located in one region of England. Three researchers conducted a total of 49 interviews and 150 hours of observation over an 18-month period. Analysis was conducted collaboratively and iteratively around emergent themes, relating to the organisational work of technology adoption. RESULTS: The two hospitals developed and implemented AKI CDSSs using very different approaches. Nevertheless, both resulted in adaptive work and trade-offs relating to the technology, the users, the organisation and the wider system of care. A common tension was associated with attempts to maximise benefit while minimise additional burden. In both hospitals, resource pressures exacerbated the tensions of translating AKI recommendations into routine practice. CONCLUSIONS: Our analysis highlights a conflicted relationship between external context (policy and resources), and organisational structure and culture (eg, digital capability, attitudes to quality improvement). Greater consideration is required to the long-term effectiveness of the approaches taken, particularly in light of the ongoing need for adaptation to incorporate new practices into routine work.

Outcomes Comparison of Enculturating Advance Directives Process at a Health System.

The Center for Medicare and Medicaid Services requires organizations to comply with the Patient Self-Determination Act by having processes that inform patients about their rights to execute an advance directive (AD) and engage in shared decision-making. The aim of this study was to compare AD data from a previous study (1999-2002) to a postenculturation (2011-2015) of a structured process for documented patient's preferences. Second, to conduct a descriptive, bivariate analysis of the enculturated structured ADs process during 2011 and 2015. This descriptive, comparative analysis included 500 random patients from four hospitals, and the enculturated descriptive analysis included 302 patients from six hospitals. Comparisons showed less no ADs and a greater institutional ADs post compared with pre (p < .05). Fifty-four percent of patients from 2011 to 2015 had an AD, and none of them had resuscitative measures when Do-Not-Resuscitate status was ordered. This enculturated process which includes education for health-care professionals and the community facilitates optimal patient, family-centered care.

Getting messier with TIDieR: embracing context and complexity in intervention reporting.

BACKGROUND: The Template for Intervention Description and Replication (TIDieR) checklist and guide was developed by an international team of experts to promote full and accurate description of trial interventions. It is now widely used in health research. The aim of this paper is to describe the experience of using TIDieR outside of trials, in a range of applied health research contexts, and make recommendations on its usefulness in such settings. MAIN BODY: We used the TIDieR template for intervention description in six applied health research projects. The six cases comprise a diverse sample in terms of clinical problems, population, settings, stage of intervention development and whether the intervention was led by researchers or the service deliverers. There was also variation in how the TIDieR description was produced in terms of contributors and time point in the project. Researchers involved in the six cases met in two workshops to identify issues and themes arising from their experience of using TIDieR. We identified four themes which capture the difficulties or complexities of using TIDieR in applied health research: (i) fidelity and adaptation: all aspects of an intervention can change over time; (ii) voice: the importance of clarity on whose voice the TIDieR description represents; (iii) communication beyond the immediate context: the usefulness of TIDieR for wider dissemination and sharing; (iv) the use of TIDieR as a research tool. CONCLUSION: We found TIDieR to be a useful tool for applied research outside the context of clinical trials and we suggest four revisions or additions to the original TIDieR which would enable it to better capture these complexities in applied health research: An additional item, 'voice' conveys who was involved in preparing the TIDieR template, such as researchers, service users or service deliverers. An additional item, 'stage of implementation' conveys what stage the intervention has reached, using a continuum of implementation research suggested by the World Health Organisation. A new column, 'modification' reminds authors to describe modifications to any item in the checklist. An extension of the 'how well' item encourages researchers to describe how contextual factors affected intervention delivery.

Understanding the implementation of 'sick day guidance' to prevent acute kidney injury across a primary care setting in England: a qualitative evaluation.

OBJECTIVES: The study sought to examine the implementation of sick day guidance cards designed to prevent acute kidney injury (AKI), in primary care settings. DESIGN: Qualitative semistructured interviews were conducted and comparative analysis informed by normalisation process theory was undertaken to understand sense-making, implementation and appraisal of the cards and associated guidance. SETTING: A single primary care health setting in the North of England. PARTICIPANTS: 29 participants took part in the qualitative evaluation: seven general practitioners, five practice nurses, five community pharmacists, four practice pharmacists, two administrators, one healthcare assistant and five patients. INTERVENTION: The sick day guidance intervention was rolled out (2015-2016) in general practices (n=48) and community pharmacies (n=60). The materials consisted of a 'medicine sick day guidance' card, provided to patients who were taking the listed drugs. The card provided advice about medicines management during episodes of acute illness. An information leaflet was provided to healthcare practitioners and administrators suggesting how to use and give the cards. RESULTS: Implementation of sick day guidance cards to prevent AKI entailed a new set of working practises across primary care. A tension existed between ensuring reach in administration of the cards to at risk populations while being confident to ensure patient understanding of their purpose and use. Communicating the concept of temporary cessation of medicines was a particular challenge and limited their administration to patient populations at higher risk of AKI, particularly those with less capacity to self-manage. CONCLUSIONS: Sick day guidance cards that focus solely on medicines management may be of limited patient benefit without adequate resourcing or if delivered as a standalone intervention. Development and evaluation of primary care interventions is urgently warranted to tackle the harm associated with AKI.

Development and Validation of a Self-Report Measure of Mentalizing: The Reflective Functioning Questionnaire.

Reflective functioning or mentalizing is the capacity to interpret both the self and others in terms of internal mental states such as feelings, wishes, goals, desires, and attitudes. This paper is part of a series of papers outlining the development and psychometric features of a new self-report measure, the Reflective Functioning Questionnaire (RFQ), designed to provide an easy to administer self-report measure of mentalizing. We describe the development and initial validation of the RFQ in three studies. Study 1 focuses on the development of the RFQ, its factor structure and construct validity in a sample of patients with Borderline Personality Disorder (BPD) and Eating Disorder (ED) (n = 108) and normal controls (n = 295). Study 2 aims to replicate these findings in a fresh sample of 129 patients with personality disorder and 281 normal controls. Study 3 addresses the relationship between the RFQ, parental reflective functioning and infant attachment status as assessed with the Strange Situation Procedure (SSP) in a sample of 136 community mothers and their infants. In both Study 1 and 2, confirmatory factor analyses yielded two factors assessing Certainty (RFQ_C) and Uncertainty (RFQ_U) about the mental states of self and others. These two factors were relatively distinct, invariant across clinical and non-clinical samples, had satisfactory internal consistency and test-retest stability, and were largely unrelated to demographic features. The scales discriminated between patients and controls, and were significantly and in theoretically predicted ways correlated with measures of empathy, mindfulness and perspective-taking, and with both self-reported and clinician-reported measures of borderline personality features and other indices of maladaptive personality functioning. Furthermore, the RFQ scales were associated with levels of parental reflective functioning, which in turn predicted infant attachment status in the SSP. Overall, this study lends preliminary support for the RFQ as a screening measure of reflective functioning. Further research is needed, however, to investigate in more detail the psychometric qualities of the RFQ.

Variable-Number Tandem-Repeat Analysis of Respiratory and Household Water Biofilm Isolates of "Mycobacterium avium subsp. hominissuis" with Establishment of a PCR Database.

"Mycobacterium aviumsubsp.hominissuis" is an important cause of pulmonary disease. It is acquired from environmental sources, but there is no methodology for large population studies. We evaluated the potential of variable-number tandem-repeat (VNTR) analysis. Clinical and household biofilmM. aviumisolates underwent molecular identification. Testing for IS901was done to separateM. aviumsubsp.aviumfromM. aviumsubsp.hominissuis VNTR types were defined using VNTR loci, and subtyping was performed using 3'hsp65and internal transcribed spacer (ITS) sequencing. Forty-nine VNTR types and eight subtypes ofM. aviumsubsp.hominissuis(IS901negative) were identified among 416 isolates ofM. aviumfrom 121 patients and 80 biofilm sites. Of those types, 67% were found only among patient isolates, 11% only among household water isolates, and 23% among both. Of 13 VNTR types that included ≥4 patients, the majority (61.5%) represented geographic clustering (same city). Most VNTR types with multiple patients belonged to the same 3'hsp65sequence code (sequevar). A total of 44 isolates belonging to fourM. aviumsubsp.hominissuisVNTR types (8%), including three with the rare Mav-F ITS sequence and 0/8 subspecies, produced amplicons with IS901PCR primers. By sequencing, all 44 amplicons were not IS901but ISMav6, which was recently observed in Japan but had not been previously described among U.S. isolates. VNTR analysis ofM. aviumsubsp.hominissuisisolates is easier and faster than pulsed-field gel electrophoresis. Seven VNTR loci separated 417 isolates into 49 types. No isolates ofM. aviumsubsp.aviumwere identified. The distributions of the VNTR copy numbers, the allelic diversity, and the low prevalence of ISMav6 differed from the findings for respiratory isolates reported from Japan.

Utility of sequencing the erm(41) gene in isolates of Mycobacterium abscessus subsp. abscessus with low and intermediate clarithromycin MICs.

The erm(41) gene confers inducible macrolide resistance in Mycobacterium abscessus subsp. abscessus, calling into question the usefulness of macrolides for treating M. abscessus subsp. abscessus infections. With an extended incubation (14 days), isolates with MICs of ≥8 µg/ml are considered macrolide resistant by current CLSI guidelines. Our goals were to determine the incidence of macrolide susceptibility in U.S. isolates, the validity of currently accepted MIC breakpoints, and the erm(41) sequences associated with susceptibility. Of 349 isolates (excluding those with 23S rRNA gene mutations), 85 (24%) had clarithromycin MICs of ≤8 µg/ml. Sequencing of the erm(41) genes from these isolates, as well as from isolates with MICs of ≥16 µg/ml, including ATCC 19977T, revealed 10 sequevars. The sequence in ATCC 19977T was designated sequevar (type) 1; most macrolide-resistant isolates were of this type. Seven sequevars contained isolates with MICs of >16 µg/ml. The T28C substitution in erm(41), previously associated with macrolide susceptibility, was identified in 62 isolates (18%) comprising three sequevars, with MICs of ≤2 (80%), 4 (10%), and 8 (10%) µg/ml. No other nucleotide substitution was associated with macrolide susceptibility. We recommend that clarithromycin susceptibility breakpoints for M. abscessus subsp. abscessus be changed from ≤2 to ≤4 µg/ml and that isolates with an MIC of 8 µg/ml have repeat MIC testing or erm sequencing performed. Our studies suggest that macrolides are useful for treating approximately 20% of U.S. isolates of M. abscessus subsp. abscessus. Sequencing of the erm gene of M. abscessus subsp. abscessus will predict inducible macrolide susceptibility.

EspR, a regulator of the ESX-1 secretion system in Mycobacterium tuberculosis, is directly regulated by the two-component systems MprAB and PhoPR.

The regulatory mechanisms that control the ESX-1 secretion system, a key player in the pathogenesis of Mycobacterium tuberculosis, have not been fully elucidated. However, factors that regulate the ESX-1 substrate EspA usually affect ESX-1 function. Previous studies showed that espA is directly regulated by the nucleoid-associated protein EspR and the two-component system (TCS) MprAB. The PhoPR TCS also activates espA, but the direct target of PhoP was unknown. In this report, we reveal that EspR is directly regulated by MprA and PhoP-Rv, but not by PhoP-Ra. PhoP-Rv and MprA binding sites in the espR promoter were determined by gel-shift and DNase I footprinting assays, which identified a PhoP-protected region centred approximately 205 bp before the espR start codon and that encompasses MprA Region-1, one of two MprA-protected regions. MprA Region-2 is located approximately 60 bp downstream of MprA Region-1 and overlaps a known EspR binding site. Nucleotides essential for the binding of PhoP and/or MprA were identified through site-directed DNA mutagenesis. Our studies also indicate that MprA Region-2, but not MprA Region-1/PhoP region, is required for the full expression of espR. Recombinant strains carrying mutations at MprA Region-2 exhibited lower transcription levels for espR, espA and espD, and had reduced EspR and EspA levels in cell lysates. These findings indicate that EspR may mediate the regulatory effect of PhoPR and MprAB, and provide more insight into the mechanisms underlying ESX-1 control.

Mechanisms of Antifungal Drug Resistance.

Antifungal therapy is a central component of patient management for acute and chronic mycoses. Yet, treatment choices are restricted because of the sparse number of antifungal drug classes. Clinical management of fungal diseases is further compromised by the emergence of antifungal drug resistance, which eliminates available drug classes as treatment options. Once considered a rare occurrence, antifungal drug resistance is on the rise in many high-risk medical centers. Most concerning is the evolution of multidrug- resistant organisms refractory to several different classes of antifungal agents, especially among common Candida species. The mechanisms responsible are mostly shared by both resistant strains displaying inherently reduced susceptibility and those acquiring resistance during therapy. The molecular mechanisms include altered drug affinity and target abundance, reduced intracellular drug levels caused by efflux pumps, and formation of biofilms. New insights into genetic factors regulating these mechanisms, as well as cellular factors important for stress adaptation, provide a foundation to better understand the emergence of antifungal drug resistance.

EUCAST testing of Isavuconazole susceptibility in Aspergillus: comparison of results for Inoculum standardization using Conidium counting versus optical density.

The EUCAST E.DEF9.1 standard recommends standardization of the inoculum concentration by conidium counting using a hemocytometer rather than a spectrophotometer. In this study, we investigated whether the choice of these methods influenced isavuconazole MICs. A blinded collection of 30 molecularly characterized azole-resistant isolates and 10 wild-type Aspergillus fumigatus isolates was shared with four different laboratories. Additionally, each laboratory selected approximately 100 A. fumigatus isolates and 50 isolates each of A. flavus, A. nidulans, A. niger, and A. terreus (1,237 isolates in total). Three laboratories (laboratories 1 to 3) used conidium counting. One laboratory standardized the inoculum using a spectrophotometer (that is, by use of the optical density [OD]) and is referred to as the OD laboratory. Correlation coefficients, intraclass correlation coefficients, and essential agreement were calculated, and 2-log-unit differences were assessed (paired t test). The MIC range for the blinded collection was 0.25 to 16 mg/liter, and a 1-dilution-step difference between the MIC50 and MIC90 across the four laboratories was detected and a 2-dilution-step difference between the modal MICs was detected. Compared to the results for laboratories 1 and 2, a significant correlation was found for the OD laboratory MIC data (correlation coefficients, 0.85 and 0.93, respectively; intraclass correlation coefficients, 0.88 and 0.96, respectively). The number of mutant isolates whose MICs overlapped those of the wild-type isolates was the lowest for the OD laboratory (14/30 [46.7%] mutant isolates), whereas the numbers were 18/30 (60%) isolates for laboratory 1, 17/30 (56.7%) isolates for laboratory 2, and 21/30 (70%) isolates for laboratory 3. For the A. flavus, A. fumigatus, A. nidulans, A. niger, and A. terreus isolates, comparative analysis again defined the MIC distributions from the OD laboratory to be in excellent agreement with those from laboratories 1 and 2 across all five Aspergillus spp. The findings suggest that EUCAST testing using OD determination is an appropriate alternative for standardization of Aspergillus inoculum concentrations.

Multi-resistant aspergillosis due to cryptic species.

Reports of cryptic species causing aspergillosis in humans are increasing in the literature. Cryptic species are defined as those which are morphologically indistinguishable, although their identifications can be confirmed using molecular or other techniques which continue to become more widely available in the clinical setting. Antifungal resistance has often been noted in these cases, and indeed there does appear to be a higher prevalence of reduced susceptibility in cryptic species. Many of these observations are published as individual case reports or as a small component of larger data sets, making it challenging to review and compare the data fully. This review article seeks to describe the susceptibility trends and key learning outcomes of specific cases of infections caused by cryptic species, including Aspergillus alliaceus, Aspergillus calidoustus, Aspergillus felis, Aspergillus lentulus, Aspergillus tubingensis, Aspergillus viridinutans and Neosartorya pseudofischeri. These reports highlight the clinical need for full accurate identification and susceptibility testing to guide patient care.

Efficacy of an abbreviated induction regimen of amphotericin B deoxycholate for cryptococcal meningoencephalitis: 3 days of therapy is equivalent to 14 days.

UNLABELLED: Cryptococcal meningoencephalitis is an urgent global health problem. Induction regimens using 14 days of amphotericin B deoxycholate (dAmB) are considered the standard of care but may not be suitable for resource-poor settings. We investigated the efficacy of conventional and abbreviated regimens of dAmB for cryptococcal meningoencephalitis in both murine and rabbit models of cryptococcal meningoencephalitis. We examined the extent to which immunological effectors contribute to the antifungal effect. We bridged the results to humans as a first critical step to define regimens suitable for further study in clinical trials. There were significant differences in the murine plasma-versus-cerebrum dAmB concentration-time profiles. dAmB was detectable in the cerebrum throughout the experimental period, even following the administration of only three doses of 3 mg/kg. dAmB induced a fungistatic effect in the cerebrum with a 2- to 3-log10 CFU/g reduction compared with controls. The effect of 3 days of therapy was the same as that of daily therapy for 14 days. There was no evidence of increased numbers of CD3(+) CD4(+) or CD3(+) CD8(+) cells in treated mice to account for the persistent antifungal effect of an abbreviated regimen. The administration of dAmB at 1 mg/kg/day for 3 days was the same as daily therapy in rabbits. The bridging studies suggested that a human regimen of 0.7 mg/kg/day for 3 days resulted in nearly maximal antifungal activity in both the cerebrum and cerebrospinal fluid. An abbreviated regimen (3 days of therapy) of dAmB appears to be just as effective as conventional induction therapy for cryptococcal meningoencephalitis. IMPORTANCE: Cryptococcal meningitis is a significant and neglected infection that is associated with excessive morbidity and mortality. In well-resourced health care settings, induction therapy with at least 2 weeks of amphotericin B deoxycholate (dAmB) is advocated. Multiple clinical studies suggest that dAmB is the drug of choice for cryptococcal meningitis. In many parts of the world where the burden of cryptococcal meningitis is highest, it is infeasible to administer dAmB for prolonged periods. This paper provides the experimental basis for the efficacy of abbreviated regimens of dAmB for cryptococcal meningitis. The concept was explored in two well-validated laboratory animal models of cryptococcal meningitis, and the results were bridged to humans by using a range of mathematical modeling techniques. A 3-day regimen is as effective as the standard 14-day course. An abbreviated regimen is significantly more feasible and may enable better antifungal therapy to be administered to many patients with this frequently lethal disease.