Viral PB1-F2 and host IFN-γ guide ILC2 and T cell activity during influenza virus infection.

Functional plasticity of innate lymphoid cells (ILCs) and T cells is regulated by host environmental cues, but the influence of pathogen-derived virulence factors has not been described. We now report the interplay between host interferon (IFN)-γ and viral PB1-F2 virulence protein in regulating the functions of ILC2s and T cells that lead to recovery from influenza virus infection of mice. In the absence of IFN-γ, lung ILC2s from mice challenged with the A/California/04/2009 (CA04) H1N1 virus, containing nonfunctional viral PB1-F2, initiated a robust IL-5 response, which also led to improved tissue integrity and increased survival. Conversely, challenge with Puerto Rico/8/1934 (PR8) H1N1 virus expressing fully functional PB1-F2, suppressed IL-5+ ILC2 responses, and induced a dominant IL-13+ CD8 T cell response, regardless of host IFN-γ expression. IFN-γ-deficient mice had increased survival and improved tissue integrity following challenge with lethal doses of CA04, but not PR8 virus, and increased resistance was dependent on the presence of IFN-γR+ ILC2s. Reverse-engineered influenza viruses differing in functional PB1-F2 activity induced ILC2 and T cell phenotypes similar to the PB1-F2 donor strains, demonstrating the potent role of viral PB1-F2 in host resistance. These results show the ability of a pathogen virulence factor together with host IFN-γ to regulate protective pulmonary immunity during influenza infection.

Sequential targeting of interferon pathways for increased host resistance to bacterial superinfection during influenza.

Bacterial co-infections represent a major clinical complication of influenza. Host-derived interferon (IFN) increases susceptibility to bacterial infections following influenza, but the relative roles of type-I versus type-II IFN remain poorly understood. We have used novel mouse models of co-infection in which colonizing pneumococci were inoculated into the upper respiratory tract; subsequent sublethal influenza virus infection caused the bacteria to enter the lungs and mediate lethal disease. Compared to wild-type mice or mice deficient in only one pathway, mice lacking both IFN pathways demonstrated the least amount of lung tissue damage and mortality following pneumococcal-influenza virus superinfection. Therapeutic neutralization of both type-I and type-II IFN pathways similarly provided optimal protection to co-infected wild-type mice. The most effective treatment regimen was staggered neutralization of the type-I IFN pathway early during co-infection combined with later neutralization of type-II IFN, which was consistent with the expression and reported activities of these IFNs during superinfection. These results are the first to directly compare the activities of type-I and type-II IFN during superinfection and provide new insights into potential host-directed targets for treatment of secondary bacterial infections during influenza.

Training for outbreak response through the Global Outbreak Alert and Response Network.

For 20 years, the Global Outbreak Response and Alert Network (GOARN) has been a leader in the coordination of international outbreak response. On the premise that no single institution can provide all capacities required to successfully respond to a complex public health emergency or fulfil all outbreak response training needs, GOARN embarked on a capacity building journey that draws on the unique strengths of more than 250 partner institutions. Through extensive engagement and collaboration, GOARN Partners have created a bespoke, multifaceted, 3-tiered training programme which has evolved over the last 15 years and enhanced the competencies of thousands of multidisciplinary outbreak responders around the world.

Topical application of nebulized human IgG, IgA and IgAM in the lungs of rats and non-human primates.

BACKGROUND: Recurrent and persistent infections are known to affect airways of patients with Primary Immunodeficiency despite appropriate replacement immunoglobulin serum levels. Interestingly, patients with Chronic Obstructive Pulmonary Disease or with non-CF bronchiectasis also show similar susceptibility to such infections. This may be due to the limited availability of immunoglobulins from the systemic circulation in the conductive airways, resulting in local immunodeficiency. Topical application of nebulized plasma-derived immunoglobulins may represent a means to address this deficiency. In this study, we assessed the feasibility of nebulizing plasma-derived immunoglobulins and delivering them into the airways of rats and non-human primates. METHODS: Distinct human plasma-derived immunoglobulin isotype preparations were nebulized with an investigational eFlow® nebulizer and analyzed in vitro or deposited into animals. Biochemical and immunohistological analysis of nebulized immunoglobulins were then performed. Lastly, efficacy of topically applied human plasma-derived immunoglobulins was assessed in an acute Streptococcus pneumoniae respiratory infection in mice. RESULTS: Characteristics of the resulting aerosols were comparable between preparations, even when using solutions with elevated viscosity. Neither the structural integrity nor the biological function of nebulized immunoglobulins were compromised by the nebulization process. In animal studies, immunoglobulins levels were assessed in plasma, broncho-alveolar lavages (BAL) and on lung sections of rats and non-human primates in samples collected up to 72 h following application. Nebulized immunoglobulins were detectable over 48 h in the BAL samples and up to 72 h on lung sections. Immunoglobulins recovered from BAL fluid up to 24 h after inhalation remained structurally and functionally intact. Importantly, topical application of human plasma-derived immunoglobulin G into the airways of mice offered significant protection against acute pneumococcal pneumonia. CONCLUSION: Taken together our data demonstrate the feasibility of topically applying plasma-derived immunoglobulins into the lungs using a nebulized liquid formulation. Moreover, topically administered human plasma-derived immunoglobulins prevented acute respiratory infection.

Prevention of Influenza Virus-Induced Immunopathology by TGF-ß Produced during Allergic Asthma.

Asthma is believed to be a risk factor for influenza infection, however little experimental evidence exists to directly demonstrate the impact of asthma on susceptibility to influenza infection. Using a mouse model, we now report that asthmatic mice are actually significantly more resistant to a lethal influenza virus challenge. Notably, the observed increased resistance was not attributable to enhanced viral clearance, but instead, was due to reduced lung inflammation. Asthmatic mice exhibited a significantly reduced cytokine storm, as well as reduced total protein levels and cytotoxicity in the airways, indicators of decreased tissue injury. Further, asthmatic mice had significantly increased levels of TGF-ß1 and the heightened resistance of asthmatic mice was abrogated in the absence of TGF-ß receptor II. We conclude that a transient increase in TGF-ß expression following acute asthma can induce protection against influenza-induced immunopathology.

Expression of suppressor of cytokine signaling 1 (SOCS1) impairs viral clearance and exacerbates lung injury during influenza infection.

Suppressor of cytokine signaling (SOCS) proteins are inducible feedback inhibitors of cytokine signaling. SOCS1-/- mice die within three weeks postnatally due to IFN-γ-induced hyperinflammation. Since it is well established that IFN-γ is dispensable for protection against influenza infection, we generated SOCS1-/-IFN-γ-/- mice to determine whether SOCS1 regulates antiviral immunity in vivo. Here we show that SOCS1-/-IFN-γ-/- mice exhibited significantly enhanced resistance to influenza infection, as evidenced by improved viral clearance, attenuated acute lung damage, and consequently increased survival rates compared to either IFN-γ-/- or WT animals. Enhanced viral clearance in SOCS1-/-IFN-γ-/- mice coincided with a rapid onset of adaptive immune responses during acute infection, while their reduced lung injury was associated with decreased inflammatory cell infiltration at the resolution phase of infection. We further determined the contribution of SOCS1-deficient T cells to antiviral immunity. Anti-CD4 antibody treatment of SOCS1-/-IFN-γ-/- mice had no significant effect on their enhanced resistance to influenza infection, while CD8+ splenocytes from SOCS1-/-IFN-γ-/- mice were sufficient to rescue RAG1-/- animals from an otherwise lethal infection. Surprisingly, despite their markedly reduced viral burdens, RAG1-/- mice reconstituted with SOCS1-/-IFN-γ-/- adaptive immune cells failed to ameliorate influenza-induced lung injury. In conclusion, in the absence of IFN-γ, the cytoplasmic protein SOCS1 not only inhibits adaptive antiviral immune responses but also exacerbates inflammatory lung damage. Importantly, these detrimental effects of SOCS1 are conveyed through discrete cell populations. Specifically, while SOCS1 expression in adaptive immune cells is sufficient to inhibit antiviral immunity, SOCS1 in innate/stromal cells is responsible for aggravated lung injury.

Limited Efficacy of Antibacterial Vaccination Against Secondary Serotype 3 Pneumococcal Pneumonia Following Influenza Infection.

BACKGROUND: Secondary bacterial infections following influenza represent a major cause of mortality in the human population, which, in turn, has led to a call for stockpiling of bacterial vaccines for pandemic preparedness. METHODS: To investigate the efficacy of bacterial vaccination for protection against secondary pneumococcal infection, mice were immunized with pneumococcal capsular polysaccharide conjugate vaccine, and then sequentially coinfected 5 weeks later with PR8 influenza virus and A66.1 Streptococcus pneumoniae. RESULTS: In the absence of influenza virus exposure, vaccination with polysaccharide conjugate vaccine was highly effective, as indicated by 100% survival from lethal pneumococcal pneumonia and 10 000-fold greater efficiency in clearance of bacteria from the lung compared to unvaccinated mice. Enhanced clearance after vaccination was dependent upon Fc receptor (FcR) expression. However, following influenza, <40% of vaccinated mice survived bacterial coinfection and FcR-dependent clearance of antibody-opsonized bacteria reduced bacterial levels in the lungs only 5-10 fold. No differences in lung myeloid cell numbers or in FcR cell surface expression were observed following influenza. CONCLUSIONS: The results show that induction of antibacterial humoral immunity is only partially effective in protection against secondary bacterial infections that occur following influenza, and suggest that additional therapeutic strategies to overcome defective antibacterial immunity should be explored.

Allergic Lung Inflammation Reduces Tissue Invasion and Enhances Survival from Pulmonary Pneumococcal Infection in Mice, Which Correlates with Increased Expression of Transforming Growth Factor ß1 and SiglecF(low) Alveolar Macrophages.

Asthma is generally thought to confer an increased risk for invasive pneumococcal disease (IPD) in humans. However, recent reports suggest that mortality rates from IPD are unaffected in patients with asthma and that chronic obstructive pulmonary disease (COPD), a condition similar to asthma, protects against the development of complicated pneumonia. To clarify the effects of asthma on the subsequent susceptibility to pneumococcal infection, ovalbumin (OVA)-induced allergic lung inflammation (ALI) was induced in mice followed by intranasal infection with A66.1 serotype 3 Streptococcus pneumoniae. Surprisingly, mice with ALI were significantly more resistant to lethal infection than non-ALI mice. The heightened resistance observed following ALI correlated with enhanced early clearance of pneumococci from the lung, decreased bacterial invasion from the airway into the lung tissue, a blunted inflammatory cytokine and neutrophil response to infection, and enhanced expression of transforming growth factor ß1 (TGF-ß1). Neutrophil depletion prior to infection had no effect on enhanced early bacterial clearance or resistance to IPD in mice with ALI. Although eosinophils recruited into the lung during ALI appeared to be capable of phagocytizing bacteria, neutralization of interleukin-5 (IL-5) to inhibit eosinophil recruitment likewise had no effect on early clearance or survival following infection. However, enhanced resistance was associated with an increase in levels of clodronate-sensitive, phagocytic SiglecF(low) alveolar macrophages within the airways following ALI. These findings suggest that, while the risk of developing IPD may actually be decreased in patients with acute asthma, additional clinical data are needed to better understand the risk of IPD in patients with different asthma phenotypes.

Global Outbreak Alert and Response Network deployments during the COVID-19 pandemic, WHO Western Pacific Region.

Problem: The Global Outbreak Alert and Response Network (GOARN) has responded to more than 100 outbreaks during the past 23 years. The coronavirus disease (COVID-19) pandemic presented unprecedented operational constraints that challenged GOARN's core mission to rapidly deploy technical experts from its partners to support national in-country responses to public health emergencies. This paper describes the type and duration of GOARN deployments to and within the World Health Organization's (WHO's) Western Pacific Region during the COVID-19 pandemic. Context: Despite strict border closures and ever-changing vaccination and quarantine requirements, GOARN continued to deploy international technical assistance to strengthen COVID-19 response operations within the Region, as requested. Action: Data were analysed from the GOARN Knowledge Platform about deployments to and within the Region for responses to the COVID-19 pandemic between 1 January 2020 and 5 May 2023. Data were available about deployment duration, technical role requested, country or area, partner organization and deployed expert's demographics. Feedback from postdeployment briefings with the experts was collected and thematically analysed to determine ongoing needs and gaps to help improve deployment operations. Outcome: There were 72 experts deployed on 89 missions through GOARN to 12 countries and areas in the Region, for a total of 4558 field days, to support the response to the COVID-19 pandemic. Discussion: The volume of requests for assistance from countries and areas in the Region to respond to the COVID-19 pandemic uncovered a deficit in human resources available for domestic response to outbreaks and the reliance on international assistance. Strengthening the in-country capacity of ready-to-respond public health emergency staff is critical to meet the needs for outbreak response. The ongoing demand for technical experts to support national responses means that these lessons may have immediate implications.

One response: Global Outbreak Alert and Response Network supporting the COVID-19 response, Kiribati.

Problem: In January 2022, Kiribati experienced widespread community transmission of COVID-19, leading to high rates of infection among health-care workers (HCWs), which reduced essential HCWs during a period of increased hospital admissions. Context: Kiribati, a Pacific island country made up of a remote group of 33 low-lying atolls in the Pacific Ocean, experienced its first surge of COVID-19 cases beginning on 24 January 2022. Action: Reports of increasing numbers of COVID-19 cases in South Tarawa prompted the Kiribati Ministry of Health and Medical Services to request assistance from the international community, including the World Health Organization's Global Outbreak Alert and Response Network (GOARN), to support national COVID-19 response operations. Specialists in infection prevention and control (IPC) were deployed to Kiribati in February 2022 to assist the Ministry's National COVID-19 Taskforce in collaboration with national partners. These specialists helped review and strengthen IPC capacities to accommodate a potential patient surge and consequent demands for medical consumables in health-care facilities in South Tarawa. Outcome: Strengthened knowledge about and processes for IPC among HCWs prevented health care-associated infections and reduced community disease transmission during the first surge of COVID-19 cases in Kiribati. Discussion: GOARN has the capacity and ability to rapidly deploy experts to support requests for assistance. Outbreak response activities can be enhanced and sustained by using GOARN's resources and collaborating with all partners, as necessary.

Differing effects of interleukin-10 on cutaneous and pulmonary Francisella tularensis live vaccine strain infection.

We investigated the role of interleukin-10 (IL-10) in cutaneous and pulmonary infection with Francisella tularensis. We found that after intradermal challenge of mice with the live vaccine strain (LVS) of F. tularensis, splenic IL-10 levels increased rapidly and reached a peak 5 days after infection. However, IL-10 expression after infection was detrimental, since IL-10(-/-) mice showed increased bacterial clearance and were resistant to an infectious dose (>10(6) CFU/mouse) that was uniformly lethal for IL-10(+/+) mice. Furthermore, IL-10(+/+) mice treated with neutralizing anti-IL-10R monoclonal antibody were able to survive lethal cutaneous LVS challenge. The presence of IL-10 appeared to restrain the expression of IL-17, since high levels of splenic IL-17 were observed after intradermal LVS infection only in IL-10(-/-) mice. Furthermore, treatment with neutralizing anti-IL-17R antibody ablated the enhanced survival observed in IL-10(-/-) mice. However, neutralization of IL-10 activity in IL-17R(-/-) mice failed to provide protection. Thus, IL-10 suppresses a protective IL-17 response that is necessary for resistance to cutaneous LVS infection. Surprisingly, however, IL-10(-/-) mice were significantly more susceptible to pulmonary infection with LVS. Finally, although IL-10 is a critical and novel regulator of immunity to F. tularensis LVS infection, its effects were masked during infection with the highly virulent SchuS4 strain. Taken together, these findings suggest that differentially regulating expression of the IL-10 pathway in various tissues could ultimately have prophylactic and therapeutic benefits for protection against tularemia.

Progression from new methicillin-resistant Staphylococcus aureus colonisation to infection: an observational study in a hospital cohort.

BACKGROUND: Patients newly colonised with methicillin-resistant Staphylococcus aureus (MRSA) are at higher risk of clinical MRSA infection. At present, there are limited data on the duration or magnitude of this risk in a hospital population with a known time of MRSA acquisition. METHODS: A retrospective cohort study of 909 adult patients known to have newly identified MRSA colonisation during admission to National University Hospital, Singapore between 1 July 2007 and 30 June 2011 was undertaken. Patients were excluded if they had history of previous MRSA colonisation or infection, or if they had been a hospital inpatient in the preceding 12 months. Data were collected on the development of MRSA infection requiring hospitalisation up to 30 June 2012. RESULTS: Of 840 patients newly colonised with MRSA as identified on active surveillance and not clinical specimens, 546 were men (65.0%) and the median age was 65 years (range 18-103 years). Median follow up was 24 months (range 0 -64 months, 85.1% followed >6 months). Clinical infection occurred in 121 patients (14.4%) with median time to infection of 22 days (95% CI 14-31). Overall 71.9% (87/121) of infected patients developed infection within 60 days of the date MRSA colonisation was detected. However, 17/121 patients (14.0%) developed clinical infection more than six months after documented MRSA acquisition. The most common sites of clinical infection were skin and soft tissue (49/121, 40.5%, 95% CI 31.7-49.8), respiratory tract (37/121, 30.6%, 95% CI 22.5-39.6) and bone and joint infections (14/121, 11.6%, 95% CI 6.5-18.7). Thirteen patients (13/121, 10.7%, 95% CI 5.8-17.7) had bacteraemias, of which six (5.0% 95% CI 1.8-10.5) were primary and seven (5.7%, 95% CI 2.3-11.6) were secondary to infection at other sites. Crude mortality at 30 days and six months was higher in patients with MRSA infection than colonisation alone (aOR 5.49, 95% CI 2.75-10.95, p<0.001 and aOR 2.94, 95% CI 1.78-4.85, p<0.001 respectively). CONCLUSION: Risk of clinical infection is highest soon after MRSA acquisition. Prevention of MRSA acquisition in hospital will have significant impact on morbidity and mortality for patients.

Developing a partnership to improve health care delivery to children <18 years with cancer and blood disorders in the English-speaking Caribbean: lessons from the SickKids-Caribbean Initiative (SCI).

In 2013, the SickKids-Caribbean Initiative (SCI) was formalised among The Hospital for Sick Children in Toronto, Canada, the University of the West Indies, and Ministries of Health in six Caribbean countries (Barbados, The Bahamas, Jamaica, St. Lucia, St. Vincent and the Grenadines, and Trinidad and Tobago). The aim was to improve the outcomes and quality of life of children (<18 years) with cancer and blood disorders in the partner countries. Core activities included filling a human resource gap by training paediatric haematologists/oncologists and specialised registered nurses; improving capacity to diagnose and treat diverse haematology/oncology cases; developing and maintaining paediatric oncology databases; creating ongoing advocacy activities with international agencies, decision makers, and civil society; and establishing an integrated administration, management, and funding structure. We describe core program components, successes, and challenges to inform others seeking to improve health service delivery in a multidisciplinary and complex partnership.

Lack of immunogenicity of xenogeneic DNA from porcine biomaterials.

Background: Clinically useful biomaterials are derived from xenogeneic extracellular matrices, but extensive processes often used to remove all residual DNA are detrimental to their proper biological function. We hypothesized that deliberate and repeated injection of DNA extracted from clinically implantable, xenogeneic extracellular matrices might elicit an immune response in a well-established murine model that could ultimately lead to altered extracellular matrix remodeling. Methods: DNA was purified from unprocessed porcine extracellular matrices and processed extracellular matrices before sterilization (aseptic) and after sterilization. Groups of 10 mice were injected with these 3 purified DNAs and 3 controls: (1) DNA from E. coli; (2) DNA from unprocessed porcine extracellular matrices combined with interleukin-12 and methylated bovine serum albumin and emulsified in incomplete Freund's adjuvant; and (3) buffered saline. Immunizations occurred every 2 weeks for a total of 3 injections. Local cytokines and systemic anti-DNA antibodies were quantified 3 and 7 days after final injection. Results: The DNA extracted from unprocessed, aseptic, or sterilized porcine extracellular matrices failed to elicit a rejection response, and only with significant, proinflammatory adjuvant activation could such a response be seen. Without the adjuvants, biomaterial-derived DNA resulted in a mild accommodation cytokine response locally and no systemic anti-DNA antibody expression even at doses approximately 100-fold larger than would be clinically likely via extracellular matrix implantation. Conclusion: The immunological safety of porcine extracellular matrix biomaterials appears not to be related to DNA residues present. Such biomaterials need not be extensively processed, likely leading to detrimental changes in their bioactivity, solely in an effort to remove the mammalian DNA.

Lethal synergy between SARS-CoV-2 and Streptococcus pneumoniae in hACE2 mice and protective efficacy of vaccination.

Secondary infections are frequent complications of viral respiratory infections, but the potential consequence of SARS-CoV-2 coinfection with common pulmonary pathogens is poorly understood. We report that coinfection of human ACE2-transgenic mice with sublethal doses of SARS-CoV-2 and Streptococcus pneumoniae results in synergistic lung inflammation and lethality. Mortality was observed regardless of whether SARS-CoV-2 challenge occurred before or after establishment of sublethal pneumococcal infection. Increased bacterial levels following coinfection were associated with alveolar macrophage depletion, and treatment with murine GM-CSF reduced numbers of lung bacteria and pathology and partially protected from death. However, therapeutic targeting of IFNs, an approach that is effective against influenza coinfections, failed to increase survival. Combined vaccination against both SARS-CoV-2 and pneumococci resulted in 100% protection against subsequent coinfection. The results indicate that when seasonal respiratory infections return to prepandemic levels, they could lead to an increased incidence of lethal COVID-19 superinfections, especially among the unvaccinated population.

Ready to respond: adapting rapid response team training in Papua New Guinea during the COVID-19 pandemic.

Problem: Rapid response teams (RRTs) are critical for effective responses to acute public health events. While validated training packages and guidance on rolling out training for RRTs are available, they lack country-specific adaptations. Documentation is limited on RRT programming experiences in various contexts. Context: In Papua New Guinea, there remain gaps in implementing standardized, rapid mobilization of multidisciplinary RRTs at the national, provincial and district levels to investigate public health alerts. Action: The human resources needed to respond to the coronavirus disease (COVID-19) pandemic forced a review of the RRT training programme and its delivery. The training model was contextualized and adapted for implementation using a staged approach, with the initiation training phase designed to ensure RRT readiness to deploy immediately in response to COVID-19 and other public health events. Lessons learned: Selecting appropriate trainees and using a phased training approach, incorporating after-training reviews, and between-phase support from the national programme team were found to be important for programme design in Papua New Guinea. Using participatory training methods based on principles of adult learning, in which trainees draw on their own experiences, was integral to building confidence among team members in conducting outbreak investigations. Discussion: The RRT training experience in Papua New Guinea has highlighted the importance of codeveloping and delivering a context-specific training programme to meet a country's unique needs. A staged training approach that builds on knowledge and skills over time, used together with ongoing follow-up and support in the provinces, has been critical in operationalizing ready-to-respond RRTs.

Ward-level factors associated with methicillin-resistant Staphylococcus aureus acquisition-an electronic medical records study in Singapore.

BACKGROUND: Methicillin-Resistant Staphylococcus aureus (MRSA) is endemic in hospitals worldwide. Intrahospital transfers may impact MRSA acquisition risk experienced by patients. In this study, we investigated ward characteristics and connectivity that are associated with MRSA acquisition. METHODS: We analysed electronic medical records on patient transfers and MRSA screening of in-patients at an acute-care tertiary hospital in Singapore to investigate whether ward characteristics and connectivity within a network of in-patient wards were associated with MRSA acquisition rates over a period of four years. RESULTS: Most patient transfers concentrated in a stable core network of wards. Factors associated with increased rate of MRSA acquisition were MRSA prevalence among patients transferred from other wards (rate ratio (RR): 7.74 [95% confidence interval (CI): 3.88, 15.44], additional 5 percentage point), critical care ward (RR: 1.72 [95% CI: 1.09, 2.70]) and presence of MRSA cohorting beds (RR: 1.39 [95% CI: 1.03, 1.90]. Oncology ward (RR: 0.66 [95% CI: 0.46, 0.94]) (compared to medical ward), and median length of stay (RR: 0.70 [95% CI: 0.55, 0.90], additional 1.5 days) were associated with lower acquisition rates. In addition, we found evidence of interaction between MRSA prevalence among patients transferred from other wards and weighted in-degree although the latter was not associated with MRSA acquisition after controlling for confounders. CONCLUSION: Wards with higher MRSA prevalence among patients transferred from other wards were more likely to have higher MRSA acquisition rate. Its effect further increased in wards receiving greater number of patients. In addition, critical care ward, presence of MRSA cohorting beds, ward specialty, and median length of stay were associated with MRSA acquisition.

Determinants of access to childhood cancer medicines: a comparative, mixed-methods analysis of four Caribbean countries.

BACKGROUND: Equitable access to essential medicines is a key facet of childhood cancer care, recognised by WHO as vital to improved childhood cancer outcomes globally. In the Caribbean, childhood cancer outcomes are poorer than those in most high-income countries. We aimed to generate in-depth comparative evidence of the current challenges and opportunities related to access to childhood cancer medicines in the Caribbean to identify context-sensitive health systems strategies to improve drug access and inform evidence-based paediatric cancer policies in the region. METHODS: In this convergent, parallel, mixed-methods study, we mapped and analysed the determinants of access to childhood cancer medicines in four Caribbean countries (The Bahamas, Barbados, Jamaica, and Trinidad and Tobago). We analysed contextual determinants of access to medicines within and across study site jurisdictions, alignment of childhood cancer medicine inclusion between each country's national essential medicines list (NEML) and WHO's 2017 Essential Medicines List for Children, and availability and cost of chemotherapeutic agents at five tertiary care hospitals. We used a mixed-effects logistic regression model to analyse the association of medicine price, procurement efficiency (via median price ratio [MPR]), and site with drug availability. The fixed effect evaluated the effect of site and MPR on the probability of stockout in a given month. We assessed determinants of medicine access via thematic analysis of semi-structured qualitative interviews, literature, and policy documents. FINDINGS: We collected and analysed data for 28 childhood cancer medicines from Barbados, 32 from The Bahamas, 30 from Trinidad and Tobago, and 31 from Jamaica. Despite stepwise inclusion of childhood cancer medicines in NEMLs, all four countries had frequent and recurrent stockouts for many cytotoxic medicines, showing no consistent relationship between NEML inclusion and availability. A mean MPR of greater than 3·0 in Trinidad and Tobago, The Bahamas, and Barbados suggests uniformly high procurement inefficiency, resulting in significant effects on drug stockout days. For each one unit increase in MPR the adjusted odds ratio (OR) of stockout increased by 10% (adjusted OR 1·10, 95% CI 1·04-1·16; p<0·01). These challenges in access to childhood cancer medicines stem from health system and policy dynamics at institutional, national, and supranational levels that cause price volatility and erratic medicine availability. Key challenges include disparate policy commitments (eg, among sites), inefficient procurement and supply chain management practices, and local effects of international market pressures. INTERPRETATION: The Caribbean region exemplifies deficiencies in access to childhood cancer medicines that might be overcome by improved regional harmonisation of drug registration, pharmacovigilance, and procurement alongside national forecasting to strengthen global pharmaceutical planning and prioritisation. Focused political attention to address these challenges is required to ensure efficient, reliable, and sustained availability of cancer mediciness. FUNDING: The SickKids-Caribbean Initiative.

Direct role for the Rpd3 complex in transcriptional induction of the anaerobic DAN/TIR genes in yeast.

Saccharomyces cerevisiae adapts to hypoxia by expressing a large group of "anaerobic" genes. Among these, the eight DAN/TIR genes are regulated by the repressors Rox1 and Mot3 and the activator Upc2/Mox4. In attempting to identify factors recruited by the DNA binding repressor Mot3 to enhance repression of the DAN/TIR genes, we found that the histone deacetylase and global repressor complex, Rpd3-Sin3-Sap30, was not required for repression. Strikingly, the complex was instead required for activation. In addition, the histone H3 and H4 amino termini, which are targets of Rpd3, were also required for DAN1 expression. Epistasis tests demonstrated that the Rpd3 complex is not required in the absence of the repressor Mot3. Furthermore, the Rpd3 complex was required for normal function and stable binding of the activator Upc2 at the DAN1 promoter. Moreover, the Swi/Snf chromatin remodeling complex was strongly required for activation of DAN1, and chromatin immunoprecipitation analysis showed an Rpd3-dependent reduction in DAN1 promoter-associated nucleosomes upon induction. Taken together, these data provide evidence that during anaerobiosis, the Rpd3 complex acts at the DAN1 promoter to antagonize the chromatin-mediated repression caused by Mot3 and Rox1 and that chromatin remodeling by Swi/Snf is necessary for normal expression.

SON DNA-binding protein mediates macrophage autophagy and responses to intracellular infection.

Intracellular pathogens affect diverse host cellular defence and metabolic pathways. Here, we used infection with Francisella tularensis to identify SON DNA-binding protein as a central determinant of macrophage activities. RNAi knockdown of SON increases survival of human macrophages following F. tularensis infection or inflammasome stimulation. SON is required for macrophage autophagy, interferon response factor 3 expression, type I interferon response and inflammasome-associated readouts. SON knockdown has gene- and stimulus-specific effects on inflammatory gene expression. SON is required for accurate splicing and expression of GBF1, a key mediator of cis-Golgi structure and function. Chemical GBF1 inhibition has similar effects to SON knockdown, suggesting that SON controls macrophage functions at least in part by controlling Golgi-associated processes.