Cough in Children and Adults: Diagnosis, Assessment and Management (CICADA). Summary of an updated position statement on chronic cough in Australia.

INTRODUCTION: Cough is the most common symptom leading to medical consultation. Chronic cough results in significant health care costs, impairs quality of life, and may indicate the presence of a serious underlying condition. Here, we present a summary of an updated position statement on cough management in the clinical consultation. MAIN RECOMMENDATIONS: Assessment of children and adults requires a focused history of chronic cough to identify any red flag cough pointers that may indicate an underlying disease. Further assessment with examination should include a chest x-ray and spirometry (when age > 6 years). Separate paediatric and adult diagnostic management algorithms should be followed. Management of the underlying condition(s) should follow specific disease guidelines, as well as address adverse environmental exposures and patient/carer concerns. First Nations adults and children should be considered a high risk group. The full statement from the Thoracic Society of Australia and New Zealand and Lung Foundation Australia for managing chronic cough is available at https://lungfoundation.com.au/resources/cicada-full-position-statement. CHANGES IN MANAGEMENT AS A RESULT OF THIS STATEMENT: Algorithms for assessment and diagnosis of adult and paediatric chronic cough are recommended. High quality evidence supports the use of child-specific chronic cough management algorithms to improve clinical outcomes, but none exist in adults. Red flags that indicate serious underlying conditions requiring investigation or referral should be identified. Early and effective treatment of chronic wet/productive cough in children is critical. Culturally specific strategies for facilitating the management of chronic cough in First Nations populations should be adopted. If the chronic cough does not resolve or is unexplained, the patient should be referred to a respiratory specialist or cough clinic.

Transitioning to home and beyond following stroke: a prospective cohort study of outcomes and needs.

INTRODUCTION: Understanding of the needs of people with stroke at hospital discharge and in the first six-months is limited. This study aim was to profile and document the needs of people with stroke at hospital discharge to home and thereafter. METHODS: A prospective cohort study recruiting individuals with stroke, from three hospitals, who transitioned home, either directly, through rehabilitation, or with early supported discharge teams. Their outcomes (global-health, cognition, function, quality of life, needs) were described using validated questionnaires and a needs survey, at 7-10 days, and at 3-, and 6-months, post-discharge. RESULTS: 72 patients were available at hospital discharge; mean age 70 (SD 13); 61% female; median NIHSS score of 4 (IQR 0-20). 62 (86%), 54 (75%), and 45 (63%) individuals were available respectively at each data collection time-point. Perceived disability was considerable at hospital discharge (51% with mRS ≥ 3), and while it improved at 3-months, it increased thereafter (35% with mRS ≥ 3 at 6-months). Mean physical health and social functioning were "fair" at hospital discharge and ongoing; while HR-QOL, although improved over time, remained impaired at 6-months (0.69+/-0.28). At 6-months cognitive impairment was present in 40%. Unmet needs included involvement in transition planning and care decisions, with ongoing rehabilitation, information, and support needs. The median number of unmet needs at discharge to home was four (range:1-9), and three (range:1-7) at 6-months. CONCLUSION: Stroke community reintegration is challenging for people with stroke and their families, with high levels of unmet need. Profiling outcomes and unmet needs for people with stroke at hospital-to-home transition and onwards are crucial for shaping the development of effective support interventions to be delivered at this juncture. ISRCTN REGISTRATION: 02/08/2022; ISRCTN44633579.

Global declines in coral reef calcium carbonate production under ocean acidification and warming.

Ocean warming and acidification threaten the future growth of coral reefs. This is because the calcifying coral reef taxa that construct the calcium carbonate frameworks and cement the reef together are highly sensitive to ocean warming and acidification. However, the global-scale effects of ocean warming and acidification on rates of coral reef net carbonate production remain poorly constrained despite a wealth of studies assessing their effects on the calcification of individual organisms. Here, we present global estimates of projected future changes in coral reef net carbonate production under ocean warming and acidification. We apply a meta-analysis of responses of coral reef taxa calcification and bioerosion rates to predicted changes in coral cover driven by climate change to estimate the net carbonate production rates of 183 reefs worldwide by 2050 and 2100. We forecast mean global reef net carbonate production under representative concentration pathways (RCP) 2.6, 4.5, and 8.5 will decline by 76, 149, and 156%, respectively, by 2100. While 63% of reefs are projected to continue to accrete by 2100 under RCP2.6, 94% will be eroding by 2050 under RCP8.5, and no reefs will continue to accrete at rates matching projected sea level rise under RCP4.5 or 8.5 by 2100. Projected reduced coral cover due to bleaching events predominately drives these declines rather than the direct physiological impacts of ocean warming and acidification on calcification or bioerosion. Presently degraded reefs were also more sensitive in our analysis. These findings highlight the low likelihood that the world's coral reefs will maintain their functional roles without near-term stabilization of atmospheric CO2 emissions.

Investigating the effect of ribavirin treatment on genetic mutations in Crimean-Congo haemorrhagic fever virus (CCHFV) through next-generation sequencing.

Crimean-Congo haemorrhagic fever (CCHF) is the most widespread tick-borne viral haemorrhagic fever affecting humans, and yet a licensed drug against the virus (CCHFV) is still not available. While several studies have suggested the efficacy of ribavirin against CCHFV, current literature remains inconclusive. In this study, we have utilised next-generation sequencing to investigate the mutagenic effect of ribavirin on the CCHFV genome during clinical disease. Samples collected from CCHF patients receiving ribavirin treatment or supportive care only at Sivas Cumhuriyet University Hospital, Turkey, were analysed. By comparing the frequency of mutations in each group, we found little evidence of an overall mutagenic effect. This suggests that ribavirin, administered at the acute stages of CCHFV infection (at the World Health Organization-recommended dose) is unable to induce lethal mutagenesis that would cause an extinction event in the CCHFV population and reduce viremia.

TLR9 expression in chronic lymphocytic leukemia identifies a promigratory subpopulation and novel therapeutic target.

Chronic lymphocytic leukemia (CLL) remains incurable despite B-cell receptor-targeted inhibitors revolutionizing treatment. This suggests that other signaling molecules are involved in disease escape mechanisms and resistance. Toll-like receptor 9 (TLR9) is a promising candidate that is activated by unmethylated cytosine guanine dinucleotide-DNA. Here, we show that plasma from patients with CLL contains significantly more unmethylated DNA than plasma from healthy control subjects (P < .0001) and that cell-free DNA levels correlate with the prognostic markers CD38, ß2-microglobulin, and lymphocyte doubling time. Furthermore, elevated cell-free DNA was associated with shorter time to first treatment (hazard ratio, 4.0; P = .003). We also show that TLR9 expression was associated with in vitro CLL cell migration (P < .001), and intracellular endosomal TLR9 strongly correlated with aberrant surface expression (sTLR9; r = 0.9). In addition, lymph node-derived CLL cells exhibited increased sTLR9 (P = .016), and RNA-sequencing of paired sTLR9hi and sTLR9lo CLL cells revealed differential transcription of genes involved in TLR signaling, adhesion, motility, and inflammation in sTLR9hi cells. Mechanistically, a TLR9 agonist, ODN2006, promoted CLL cell migration (P < .001) that was mediated by p65 NF-κB and STAT3 transcription factor activation. Importantly, autologous plasma induced the same effects, which were reversed by a TLR9 antagonist. Furthermore, high TLR9 expression promoted engraftment and rapid disease progression in a NOD/Shi-scid/IL-2Rγnull mouse xenograft model. Finally, we showed that dual targeting of TLR9 and Bruton's tyrosine kinase (BTK) was strongly synergistic (median combination index, 0.2 at half maximal effective dose), which highlights the distinct role for TLR9 signaling in CLL and the potential for combined targeting of TLR9 and BTK as a more effective treatment strategy in this incurable disease.

Global warming and recurrent mass bleaching of corals.

During 2015-2016, record temperatures triggered a pan-tropical episode of coral bleaching, the third global-scale event since mass bleaching was first documented in the 1980s. Here we examine how and why the severity of recurrent major bleaching events has varied at multiple scales, using aerial and underwater surveys of Australian reefs combined with satellite-derived sea surface temperatures. The distinctive geographic footprints of recurrent bleaching on the Great Barrier Reef in 1998, 2002 and 2016 were determined by the spatial pattern of sea temperatures in each year. Water quality and fishing pressure had minimal effect on the unprecedented bleaching in 2016, suggesting that local protection of reefs affords little or no resistance to extreme heat. Similarly, past exposure to bleaching in 1998 and 2002 did not lessen the severity of bleaching in 2016. Consequently, immediate global action to curb future warming is essential to secure a future for coral reefs.

Thoracic Society of Australia and New Zealand (TSANZ) position statement on chronic suppurative lung disease and bronchiectasis in children, adolescents and adults in Australia and New Zealand.

This position statement, updated from the 2015 guidelines for managing Australian and New Zealand children/adolescents and adults with chronic suppurative lung disease (CSLD) and bronchiectasis, resulted from systematic literature searches by a multi-disciplinary team that included consumers. The main statements are: Diagnose CSLD and bronchiectasis early; this requires awareness of bronchiectasis symptoms and its co-existence with other respiratory diseases (e.g., asthma, chronic obstructive pulmonary disease). Confirm bronchiectasis with a chest computed-tomography scan, using age-appropriate protocols and criteria in children. Undertake a baseline panel of investigations. Assess baseline severity, and health impact, and develop individualized management plans that include a multi-disciplinary approach and coordinated care between healthcare providers. Employ intensive treatment to improve symptom control, reduce exacerbation frequency, preserve lung function, optimize quality-of-life and enhance survival. In children, treatment also aims to optimize lung growth and, when possible, reverse bronchiectasis. Individualize airway clearance techniques (ACTs) taught by respiratory physiotherapists, encourage regular exercise, optimize nutrition, avoid air pollutants and administer vaccines following national schedules. Treat exacerbations with 14-day antibiotic courses based upon lower airway culture results, local antibiotic susceptibility patterns, clinical severity and patient tolerance. Patients with severe exacerbations and/or not responding to outpatient therapy are hospitalized for further treatments, including intravenous antibiotics and intensive ACTs. Eradicate Pseudomonas aeruginosa when newly detected in lower airway cultures. Individualize therapy for long-term antibiotics, inhaled corticosteroids, bronchodilators and mucoactive agents. Ensure ongoing care with 6-monthly monitoring for complications and co-morbidities. Undertake optimal care of under-served peoples, and despite its challenges, delivering best-practice treatment remains the overriding aim.

A Phase 3b Study for Management of Ocular Side Effects in Patients with Epidermal Growth Factor Receptor-Amplified Glioblastoma Receiving Depatuxizumab Mafodotin.

INTRODUCTION: The Understanding New Interventions with GBM ThErapy (UNITE) study was designed to assess the effect of prophylaxis for ocular side effects (OSEs) in patients with glioblastoma receiving the antibody-drug conjugate (ADC) depatuxizumab mafodotin. UNITE (NCT03419403) was a phase 3b, open-label, randomized, exploratory study performed at 18 research sites in 5 countries. METHODS: The study enrolled adult patients with epidermal growth factor receptor-amplified, histologically confirmed, newly diagnosed supratentorial glioblastoma or grade IV gliosarcoma, and a Karnofsky Performance Status ≥70, receiving depatuxizumab mafodotin. All patients were administered depatuxizumab mafodotin during concurrent radiotherapy and temozolomide and with adjuvant temozolomide. Ninety patients were to be randomized (1:1:1) to OSE prophylactic treatments with each depatuxizumab mafodotin infusion: (a) standard steroid eye drops, (b) standard steroid eye drops plus vasoconstrictor eye drops and cold compress, or (c) enhanced steroids plus vasoconstrictor eye drops and cold compress. A Corneal Epitheliopathy Adverse Event (CEAE) scale was devised to capture symptoms, grade OSEs (scale of 0-5), and inform ADC dose modifications. The primary endpoint was the frequency of a required change in OSE management due to inadequate control of OSEs, defined as decline from baseline in visual acuity (using logarithm of the minimum angle of resolution [LogMAR] scale) or a Grade ≥3 CEAE event, in the worst eye in the first 8 weeks of treatment; unless otherwise specified, the treatment period refers to both the chemoradiation and adjuvant phases. RESULTS: The UNITE study was stopped early after interim analysis of separate phase III trial showed no difference in survival from depatuxizumab mafodotin. Forty patients were randomized (38 received depatuxizumab mafodotin). Overall, 23 patients experienced inadequate control of OSEs that required change in OSE management within 8 weeks of treatment, with 21 (70.0%) experiencing ≥+0.3 change on LogMAR scale in baseline-adjusted visual acuity and 12 reporting a grade ≥3 CEAE. There were no definitive differences among prophylactic treatments. CONCLUSIONS: The premature cessation of the study precludes definitive conclusions regarding the OSE prophylaxis strategies. No new clinically significant safety findings were noted. Despite these limitations, this study highlights the need for novel assessment tools to better understand and mitigate OSEs associated with ADCs.

Plant-made dengue virus-like particles produced by co-expression of structural and non-structural proteins induce a humoral immune response in mice.

Dengue virus (DENV) is an emerging threat causing an estimated 390 million infections per year. Dengvaxia, the only licensed vaccine, may not be adequately safe in young and seronegative patients; hence, development of a safer, more effective vaccine is of great public health interest. Virus-like particles (VLPs) are a safe and very efficient vaccine strategy, and DENV VLPs have been produced in various expression systems. Here, we describe the production of DENV VLPs in Nicotiana benthamiana using transient expression. The co-expression of DENV structural proteins (SP) and a truncated version of the non-structural proteins (NSPs), lacking NS5 that contains the RNA-dependent RNA polymerase, led to the assembly of DENV VLPs in plants. These VLPs were comparable in appearance and size to VLPs produced in mammalian cells. Contrary to data from other expression systems, expression of the protein complex prM-E was not successful, and strategies used in other expression systems to improve the VLP yield did not result in increased yields in plants but, rather, increased purification difficulties. Immunogenicity assays in BALB/c mice revealed that plant-made DENV1-SP + NSP VLPs led to a higher antibody response in mice compared with DENV-E domain III displayed inside bluetongue virus core-like particles and a DENV-E domain III subunit. These results are consistent with the idea that VLPs could be the optimal approach to creating candidate vaccines against enveloped viruses.

Forecasting intensifying disturbance effects on coral reefs.

Anticipating future changes of an ecosystem's dynamics requires knowledge of how its key communities respond to current environmental regimes. The Great Barrier Reef (GBR) is under threat, with rapid changes of its reef-building hard coral (HC) community structure already evident across broad spatial scales. While several underlying relationships between HC and multiple disturbances have been documented, responses of other benthic communities to disturbances are not well understood. Here we used statistical modelling to explore the effects of broad-scale climate-related disturbances on benthic communities to predict their structure under scenarios of increasing disturbance frequency. We parameterized a multivariate model using the composition of benthic communities estimated by 145,000 observations from the northern GBR between 2012 and 2017. During this time, surveyed reefs were variously impacted by two tropical cyclones and two heat stress events that resulted in extensive HC mortality. This unprecedented sequence of disturbances was used to estimate the effects of discrete versus interacting disturbances on the compositional structure of HC, soft corals (SC) and algae. Discrete disturbances increased the prevalence of algae relative to HC while the interaction between cyclones and heat stress was the main driver of the increase in SC relative to algae and HC. Predictions from disturbance scenarios included relative increases in algae versus SC that varied by the frequency and types of disturbance interactions. However, high uncertainty of compositional changes in the presence of several disturbances shows that responses of algae and SC to the decline in HC needs further research. Better understanding of the effects of multiple disturbances on benthic communities as a whole is essential for predicting the future status of coral reefs and managing them in the light of new environmental regimes. The approach we develop here opens new opportunities for reaching this goal.

Feasibility of increasing the dosage of inpatient occupational therapy and physiotherapy rehabilitation via independent tasks and exercises: 'My Therapy'.

INTRODUCTION: The dosage of occupational therapy and physiotherapy positively correlates with rehabilitation patient and health service outcomes. Nevertheless, increasing the dosage during inpatient rehabilitation without additional resources can be challenging. This study aimed to determine feasibility of increasing the dosage of inpatient occupational therapy and physiotherapy rehabilitation with independent tasks and exercises outside of supervised sessions, the 'My Therapy' programme. METHODS: A two-group, quasi-experimental, pre-post-design examined feasibility of delivering My Therapy in addition to usual care, compared to usual care alone, for hospitalised musculoskeletal and frail older rehabilitation patients. My Therapy was prescribed by the occupational therapist and physiotherapist. A booklet was provided with an individually tailored set of tasks and exercises that were a sub-set of routine therapy, to be practised safely, effectively and independently outside of supervised sessions. The primary outcome was feasibility of My Therapy implementation to achieve at least 70% adherence. Secondary outcomes were self-reported daily My Therapy participation (minutes), total daily rehabilitation participation (minutes), adverse events, length of stay, 10-metre walk speed, FIM scores and discharge destination. RESULTS: Participation in My Therapy was achieved by 72% (83/116) of the My Therapy group, who averaged 14 min (SD 14) of daily practice outside of supervised sessions. Total daily rehabilitation participation was 177 min (SD 47) for My Therapy participants (n = 116) and 148 min (SD 88) for usual care participants (n = 89); mean difference 30 min (p = .00). A minimal clinically important difference in FIM was achieved for a significantly higher portion of the My Therapy group (22%, n = 26) compared to usual care (10%, n = 9; p = .02). There were no adverse events, safety concerns or group differences for other secondary outcomes. CONCLUSION: My Therapy was a feasible and safe way to increase the dosage of inpatient occupational therapy and physiotherapy rehabilitation via independent practice. Clinical Trial Registry: ACTRN12616000691448.

Symbiodinium biogeography tracks environmental patterns rather than host genetics in a key Caribbean reef-builder, Orbicella annularis.

The physiological performance of a reef-building coral is a combined outcome of both the coral host and its algal endosymbionts, Symbiodinium While Orbicella annularis-a dominant reef-building coral in the Wider Caribbean-is known to be a flexible host in terms of the diversity of Symbiodinium types it can associate with, it is uncertain how this diversity varies across the Caribbean, and whether spatial variability in the symbiont community is related to either O. annularis genotype or environment. Here, we target the Symbiodinium-ITS2 gene to characterize and map dominant Symbiodinium hosted by O. annularis at an unprecedented spatial scale. We reveal northwest-southeast partitioning across the Caribbean, both in terms of the dominant symbiont taxa hosted and in assemblage diversity. Multivariate regression analyses incorporating a suite of environmental and genetic factors reveal that observed spatial patterns are predominantly explained by chronic thermal stress (summer temperatures) and are unrelated to host genotype. Furthermore, we were able to associate the presence of specific Symbiodinium types with local environmental drivers (for example, Symbiodinium C7 with areas experiencing cooler summers, B1j with nutrient loading and B17 with turbidity), associations that have not previously been described.

Characterization of the invasiveness of monophasic and aphasic Salmonella Typhimurium strains in 1-day-old and point-of-lay chickens.

Egg-related outbreaks of salmonellosis are a significant health concern. Although Salmonella Enteritidis (SE) is the major egg-associated serotype, Salmonella Typhimurium (ST) can also infect the hen's reproductive tract and contaminate eggs. Recently, monophasic and aphasic variants of ST have been reported with increased frequency in Europe, and the isolation of these variants from laying flocks triggers the same legislative restrictions associated with biphasic ST strains. However, little is known about the colonization, invasiveness and persistence of monophasic and aphasic ST strains in laying hens. In this study, seven groups of 1-day-old and point-of-lay commercial Hy-line chicken layers were separately challenged with four different strains of monophasic ST, one aphasic ST, one biphasic ST and one egg-invasive SE strain. Tissue samples and cloacal swabs (point-of-lay chickens only) were collected at regular intervals post challenge in order to recover the Salmonella challenge strains. In 1-day-old chicks, only the aphasic ST strain and the SE strain were recovered after direct plating, suggesting that the number of salmonellas colonizing the tissues of the chicks infected with the other strains was likely to be low. Interestingly, all of the strains colonized well in the point-of-lay chickens, and there was no statistical difference in the overall number of positive samples or Salmonella counts between the seven strains. Salmonella was recovered from the point-of-lay birds to the end of the study (20 days after challenge). Monophasic and aphasic ST strains colonized point-of-lay birds as efficiently as biphasic ST and SE strains. Further studies are necessary to estimate the invasiveness of these strains in naturally-infected vaccinated laying hens, and to assess the impact of natural infection on egg contamination.

Communicating medical information with Aboriginal patients: lessons learned from GPs and GP registrars in Aboriginal primary health care.

BACKGROUND: Aboriginal culture stands as the oldest continuous culture in the world. It gives paramount importance to a harmonious balance between personal connections to the body, spirit, and mind, as well as collective relationships with family, land, and community, integral to the wellbeing of Aboriginal people. However, obstacles can emerge for patients due to language barriers, cultural differences, or a historical lack of trust in the healthcare system. The establishment of Aboriginal Community Controlled Health Organisations (ACCHOs) has undoubtedly improved the healthcare experience for Aboriginal patients, yet there is limited research on the specific approaches utilised by general practitioners (GPs) working in these clinics. METHODS: Twelve semi-structured interviews were conducted with two groups of GPs working in Aboriginal health. Each GP was presented with three scenarios and asked questions related to each scenario. Braun and Clarke's method of thematic analysis was applied to transcribed interviews. RESULTS: Patient-doctor relationship, health literacy, and engagement with the health system emerged as key factors influencing communication with Aboriginal patients. Experienced GPs, despite differing clinical backgrounds, shared concise yet similar ideas to their less experienced counterparts. Notably, experienced GPs prioritised non-medical conversations and mindful body language, emphasising the importance of building strong patient relationships over other consultation aspects. CONCLUSIONS: This research provides initial insights for GPs in Aboriginal health, comparing experienced GPs with more than 10years experience to novices. However, further research involving Aboriginal patients is needed to validate GP strategies and understand their significance from the patients' perspective.

Difficult-to-express antigen generation through a co-expression and disassociation methodology.

Extracellular domain (ECD) antigens are crucial components for antibody discovery, in vitro assays, and epitope mapping during therapeutical antibody development. Oftentimes, those antigens are difficult to produce while retaining the biologic function/activity upon extracellular secretion in commonly used expression systems. We have developed an effective method to cope with the challenge of generating quality antigen ECDs. In this method, a monoclonal antibody (Mab) or antibody fragment antigen-binding (Fab) region acts as a "chaperone" to stabilize the antigen ECD through forming an antibody:antigen complex. This methodology includes transient co-expression of the complex in Chinese hamster ovary cells and then dissociation of the purified complex into individual components by low pH treatment in the presence of arginine. The antigen is then separated from the chaperone on a preparative size exclusion chromatography (pSEC) followed by an optional affinity chromatography process to remove residual Mab or Fab. We demonstrate this co-expression/disassociation methodology on two difficult-to-express antigen ECDs from cluster-of-differentiation/cytokine family and were successful in producing stable, biologically active antigens when the common methods using Histidine-tagged and/or Fc-fused protein failed. This can be applied as a general approach for antigen production if a Mab or binding partner is available.

Regulation of inositol 5-phosphatase activity by the C2 domain of SHIP1 and SHIP2.

SHIP1, an inositol 5-phosphatase, plays a central role in cellular signaling. As such, it has been implicated in many conditions. Exploiting SHIP1 as a drug target will require structural knowledge and the design of selective small molecules. We have determined apo, and magnesium and phosphate-bound structures of the phosphatase and C2 domains of SHIP1. The C2 domains of SHIP1 and the related SHIP2 modulate the activity of the phosphatase domain. To understand the mechanism, we performed activity assays, hydrogen-deuterium exchange mass spectrometry, and molecular dynamics on SHIP1 and SHIP2. Our findings demonstrate that the influence of the C2 domain is more pronounced for SHIP2 than SHIP1. We determined 91 structures of SHIP1 with fragments bound, with some near the interface between the two domains. We performed a mass spectrometry screen and determined four structures with covalent fragments. These structures could act as starting points for the development of potent, selective probes.

Convalescent human plasma candidate reference materials protect against Crimean-Congo haemorrhagic fever virus (CCHFV) challenge in an A129 mouse model.

Crimean-Congo Haemorrhagic Fever Virus (CCHFV) is spread by infected ticks or direct contact with blood, tissues and fluids from infected patients or livestock. Infection with CCHFV causes severe haemorrhagic fever in humans which is fatal in up to 83 % of cases. CCHFV is listed as a priority pathogen by the World Health Organization (WHO) and there are currently no widely-approved vaccines. Defining a serological correlate of protection against CCHFV infection would support the development of vaccines by providing a 'target threshold' for pre-clinical and clinical immunogenicity studies to achieve in subjects and potentially obviate the need for in vivo protection studies. We therefore sought to establish titratable protection against CCHFV using pooled human convalescent plasma, in a mouse model. Convalescent plasma collected from seven individuals with a known previous CCHFV virus infection were characterised using binding antibody and neutralisation assays. All plasma recognised nucleoprotein and the Gc glycoprotein, but some had a lower Gn glycoprotein response by ELISA. Pooled plasma and two individual donations from convalescent donors were administered intraperitoneally to A129 mice 24 h prior to intradermal challenge with CCHFV (strain IbAr10200). A partial protective effect was observed with all three convalescent plasmas characterised by longer survival post-challenge and reduced clinical score. These protective responses were titratable. Further characterisation of the serological reactivities within these samples will establish their value as reference materials to support assay harmonisation and accelerate vaccine development for CCHFV.

Development of humanised antibodies for Crimean-Congo Haemorrhagic fever virus: Comparison of hybridoma-based versus phage library techniques.

Humanised antibodies targeting Crimean-Congo Haemorrhagic virus (CCHFV) are needed for the development and standardisation of serological assays. These assays are needed to address a shortfall in available tests that meet regulatory diagnostic standards and to aid surveillance activities to extend knowledge on the distribution of CCHFV. To generate a humanised monoclonal antibody against CCHFV, we have compared two methods: the traditional mouse hybridoma approach with subsequent sequencing and humanisation of antibodies versus a non-animal alternative using a human combinatorial antibody library (HuCAL). Our results demonstrated that the mouse hybridoma followed by humanisation protocol gave higher affinity antibodies. Whilst not yet able to demonstrate the generation of equivalent humanised antibodies without the use of animals, sequencing data enables the subsequent production of recombinant antibodies, thus providing a reduction in future animal usage for this application. Ultimately, our report provides information on development of a humanised standardised control, which can form an important positive control component of serological assays against CCHFV.