Multicenter Prospective Study of Biomarkers for Diagnosis of Invasive Candidiasis in Children and Adolescents.

BACKGROUND: Diagnosis of invasive candidiasis (IC) relies on insensitive cultures; the relative utility of fungal biomarkers in children is unclear. METHODS: This multinational observational cohort study enrolled patients aged >120 days and <18 years with concern for IC from 1 January 2015 to 26 September 2019 at 25 centers. Blood collected at onset of symptoms was tested using T2Candida, Fungitell (1→3)-ß-D-glucan, Platelia Candida Antigen (Ag) Plus, and Platelia Candida Antibody (Ab) Plus assays. Operating characteristics were determined for each biomarker, and assays meeting a defined threshold considered in combination. Sterile site cultures were the reference standard. RESULTS: Five hundred participants were enrolled at 22 centers in 3 countries, and IC was diagnosed in 13 (2.6%). Thirteen additional blood specimens were collected and successfully spiked with Candida species, to achieve a 5.0% event rate. Valid T2Candida, Fungitell, Platelia Candida Ag Plus, and Platelia Candida Ab Plus assay results were available for 438, 467, 473, and 473 specimens, respectively. Operating characteristics for T2Candida were most optimal for detecting IC due to any Candida species, with results as follows: sensitivity, 80.0% (95% confidence interval, 59.3%-93.2%), specificity 97.1% (95.0%-98.5%), positive predictive value, 62.5% (43.7%-78.9%), and negative predictive value, 98.8% (97.2%-99.6%). Only T2Candida and Platelia Candida Ag Plus assays met the threshold for combination testing. Positive result for either yielded the following results: sensitivity, 86.4% (95% confidence interval, 65.1%- 97.1%); specificity, 94.7% (92.0%-96.7%); positive predictive value, 47.5% (31.5%-63.9%); and negative predictive value, 99.2% (97.7%-99.8%). CONCLUSIONS: T2Candida alone or in combination with Platelia Candida Ag Plus may be beneficial for rapid detection of Candida species in children with concern for IC. CLINICAL TRIALS REGISTRATION: NCT02220790.

Quality improvement project to improve vaccinations in the pediatric liver transplant population.

OBJECTIVES: A quality improvement approach was used to increase pediatric liver transplant recipient live and inactive vaccination rates by assessing titers and recommending vaccinations. METHODS: A new screening and immunization process for both live and inactive vaccines was discussed with families at their annual visit. Antibody titers for varicella, measles, mumps, rubella, Haemophilus influenzae type B, hepatitis A, and hepatitis B were obtained. Specific criteria were developed for live virus vaccination candidacy. Vaccines were recommended based on patient titers and vaccination candidacy criteria. Surveillance for adverse effects to live vaccines was performed. Repeat titers were obtained approximately 1-month post-vaccine administration. RESULTS: After PDSA cycle 1, 99% (71/72) of pediatric liver transplant patients had titers obtained. Live vaccines were recommended for 32 patients and 16 (50%) were vaccinated. Inactive vaccines were recommended to 64 patients, and 31 (48%) were vaccinated. Eight of 13 (62%) patients with follow-up titers achieved immunity for inactive vaccines. Zero patients had an adverse reaction to any live vaccine. Ten of 12 (83%) patients with follow-up titers achieved immunity from live vaccines. The most common barriers to receive live vaccines included not scheduling appointment with primary care provider (n = 3) and "non-vaccinators" (n = 3). CONCLUSIONS: Administering live and inactive vaccines to select pediatric liver transplant patients appears to be safe and effective in our studied population. For PDSA cycle 2, we will continue our current practice and consider offering vaccines in transplant clinic, since this was a barrier to vaccination identified during PDSA cycle 1.

Chemokine CCL28 Is a Potent Therapeutic Agent for Oropharyngeal Candidiasis.

Candida albicans is a commensal organism that causes life-threatening or life-altering opportunistic infections. Treatment of Candida infections is limited by the paucity of antifungal drug classes. Naturally occurring antimicrobial peptides are promising agents for drug development. CCL28 is a CC chemokine that is abundant in saliva and has in vitro antimicrobial activity. In this study, we examine the in vivo Candida killing capacity of CCL28 in oropharyngeal candidiasis as well as the spectrum and mechanism of anti-Candida activity. In the mouse model of oropharyngeal candidiasis, application of wild-type CCL28 reduces oral fungal burden in severely immunodeficient mice without causing excessive inflammation or altering tissue neutrophil recruitment. CCL28 is effective against multiple clinical strains of C. albicans Polyamine protein transporters are not required for CCL28 anti-Candida activity. Both structured and unstructured CCL28 proteins show rapid and sustained fungicidal activity that is superior to that of clinical antifungal agents. Application of wild-type CCL28 to C. albicans results in membrane disruption as measured by solute movement, enzyme leakage, and induction of negative Gaussian curvature on model membranes. Membrane disruption is reduced in CCL28 lacking the functional C-terminal tail. Our results strongly suggest that CCL28 can exert antifungal activity in part via membrane permeation and has potential for development as an anti-Candida therapeutic agent without inflammatory side effects.

Attenuation of Helper T Cell Capacity for TH1 and TH17 Differentiation in Children With Nontuberculous Mycobacterial Infection.

Nontuberculous mycobacteria (NTM) infect children with increasing frequency worldwide. Using blood and lymph node tissue from children with NTM lymphadenitis, and uninfected lymph node tissue from community controls, we evaluated helper T (TH) cells in functional assays of TH1/TH17 differentiation and measured the concentration of their associated cytokines at the site of infection. Circulating TH cells from infected children were attenuated in their TH1/TH17 differentiation capacity and expressed less interferon γ and interleukin 17 after polyclonal stimulation. Similar differences were observed at the site of infection, where most cytokine concentrations were unchanged relative to controls. Our data are consistent with a model wherein TH1/TH17 differentiation is attenuated in NTM-infected children.

Successful Treatment of Invasive Conidiobolus Infection During Therapy for Acute Lymphoblastic Leukemia.

Invasive fungal infections are a serious cause of morbidity and mortality in patients with hematologic malignancies. Conidiobolus species are molds within the order Entomophthorales and may disseminate to become rapidly fatal in immunocompromised individuals. This species of fungal infections are often multidrug resistant (MDR) and present unique therapeutic challenges. Reports of Conidiobolus infections are rare in pediatric oncology. We report the successful treatment of an adolescent male with B-cell lymphoblastic leukemia and MDR invasive sinopulmonary Conidiobolus infection with emphasis on early and aggressive neutrophil support with surgical debridement. The strategies described could be applied to other MDR fungal infections.

The Goldilocks model of immune symbiosis with Mycobacteria and Candida colonizers.

Mycobacteria and Candida species include significant human pathogens that can cause localized or disseminated infections. Although these organisms may appear to have little in common, several shared pathways of immune recognition and response are important for both control and infection-related pathology. In this article, we compare and contrast the innate and adaptive components of the immune system that pertain to these infections in humans and animal models. We also explore a relatively new concept in the mycobacterial field: biological commensalism. Similar to the well-established model of Candida infection, Mycobacteria species colonize their human hosts in equilibrium with the immune response. Perturbations in the immune response permit the progression to pathologic disease at the expense of the host. Understanding the immune factors required to maintain commensalism may aid with the development of diagnostic and treatment strategies for both categories of pathogens.

Role of neutrophils in IL-17-dependent immunity to mucosal candidiasis.

Oropharyngeal candidiasis (OPC), caused by the commensal fungus Candida albicans, is an opportunistic infection associated with infancy, AIDS, and IL-17-related primary immunodeficiencies. The Th17-associated cytokines IL-23 and IL-17 are crucial for immunity to OPC, but the mechanisms by which they mediate immunity are poorly defined. IL-17RA-deficient humans and mice are strongly susceptible to OPC, with reduced levels of CXC chemokines and concomitantly impaired neutrophil recruitment to the oral mucosa. Paradoxically, humans with isolated neutropenia are typically not susceptible to candidiasis. To determine whether immunity to OPC is mediated via neutrophil recruitment, mice lacking CXCR2 were subjected to OPC and were found to be highly susceptible, although there was no dissemination of fungi to peripheral organs. To assess whether the entire neutrophil response is IL-17 dependent, IL-17RA(-/-) and IL-23(-/-) mice were administered neutrophil-depleting Abs and subjected to OPC. These mice displayed increased oral fungal burdens compared with IL-17RA(-/-) or IL-23(-/-) mice alone, indicating that additional IL-17-independent signals contribute to the neutrophil response. WT mice treated with anti-Gr-1 Abs exhibited a robust infiltrate of CD11b(+)Ly-6G(low)F4/80(-) cells to the oral mucosa but were nonetheless highly susceptible to OPC, indicating that this monocytic influx is insufficient for host defense. Surprisingly, Ly-6G Ab treatment did not induce the same strong susceptibility to OPC in WT mice. Thus, CXCR2(+) and Gr-1(+) neutrophils play a vital role in host defense against OPC. Moreover, defects in the IL-23/17 axis cause a potent but incomplete deficiency in the neutrophil response to oral candidiasis.

A Candida albicans Strain Expressing Mammalian Interleukin-17A Results in Early Control of Fungal Growth during Disseminated Infection.

Candida albicans is normally a commensal fungus of the human mucosae and skin, but it causes life-threatening systemic infections in hospital settings in the face of predisposing conditions, such as indwelling catheters, abdominal surgery, or antibiotic use. Immunity to C. albicans involves various immune parameters, but the cytokine interleukin-17A (IL-17A) (also known as IL-17) has emerged as a centrally important mediator of immune defense against both mucosal and systemic candidiasis. Conversely, IL-17A has been suggested to enhance the virulence of C. albicans, indicating that it may exert detrimental effects on pathogenesis. In this study, we hypothesized that a C. albicans strain expressing IL-17A would exhibit reduced virulence in vivo. To that end, we created a Candida-optimized expression cassette encoding murine IL-17A, which was transformed into the DAY286 strain of C. albicans. Candida-derived IL-17A was indistinguishable from murine IL-17A in terms of biological activity and detection in standard enzyme-linked immunosorbent assays (ELISAs). Expression of IL-17A did not negatively impact the growth of these strains in vitro. Moreover, the IL-17A-expressing C. albicans strains showed significantly reduced pathogenicity in a systemic model of Candida infection, mainly evident during the early stages of disease. Collectively, these findings suggest that IL-17A mitigates the virulence of C. albicans.

The adaptor CARD9 is required for adaptive but not innate immunity to oral mucosal Candida albicans infections.

Oropharyngeal candidiasis (OPC [thrush]) is an opportunistic infection caused by the commensal fungus Candida albicans. OPC is common in individuals with HIV/AIDS, infants, patients on chemotherapy, and individuals with congenital immune defects. Immunity to OPC is strongly dependent on the interleukin-23 (IL-23)/IL-17R axis, as mice and humans with defects in IL-17R signaling (IL17F, ACT1, IL-17RA) or in genes that direct Th17 differentiation (STAT3, STAT1, CARD9) are prone to mucocutaneous candidiasis. Conventional Th17 cells are induced in response to C. albicans infection via signals from C-type lectin receptors, which signal through the adaptor CARD9, leading to production of Th17-inducing cytokines such as IL-6, IL-1ß, and IL-23. Recent data indicate that IL-17 can also be made by numerous innate cell subsets. These innate "type 17" cells resemble conventional Th17 cells, but they can be activated without need for prior antigen exposure. Because C. albicans is not a commensal organism in rodents and mice are thus naive to this fungus, we had the opportunity to assess the role of CARD9 in innate versus adaptive responses using an OPC infection model. As expected, CARD9(-/-) mice failed to mount an adaptive Th17 response following oral Candida infection. Surprisingly, however, CARD9(-/-) mice had preserved innate IL-17-dependent responses to Candida and were almost fully resistant to OPC. Thus, CARD9 is important primarily for adaptive immunity to C. albicans, whereas alternate recognition systems appear to be needed for effective innate responses.

Pipeline of Novel Antifungals for Invasive Fungal Disease in Transplant Recipients: A Pediatric Perspective.

Invasive fungal disease (IFD) remains a significant cause of morbidity and mortality in children undergoing transplantation. There is a growing armamentarium of novel antifungal agents recently approved for use or in late stages of clinical development. The overarching goal of this review is to discuss the mechanisms of action, spectrum of activity, stage of development, and pediatric-specific data for the following agents: encochleated amphotericin B deoxycholate, fosmanogepix, ibrexafungerp, isavuconazole, olorofim, opelconazole, oteseconazole, and rezafungin. Additionally, key drug attributes of these novel agents and their potential future therapeutic roles in pediatric transplant recipients are discussed.

Higher Weight-Based Doses Are Required to Achieve and Maintain Therapeutic Voriconazole Serum Trough Concentrations in Children.

OBJECTIVE: Children require weight-based voriconazole doses proportionately larger than adults to achieve therapeutic serum trough concentrations (1-6 mcg/mL). The objective of this quality improvement project was to determine the initial dose, proportion of patients achieving target concentrations with initial dosing, and subsequent therapeutic drug monitoring and dose modifications needed to achieve and maintain therapeutic voriconazole concentrations in children. METHODS: This retrospective study evaluated children aged <18 years treated with voriconazole during the study period. Dosing and therapeutic drug monitoring (TDM) values were collected and compared by age. Data are presented as median (IQR), unless otherwise stated. RESULTS: Fifty-nine patients, aged 10.4 (3.7-14.7) years and 49% female, met inclusion criteria; 42 had at least 1 steady-state voriconazole serum trough concentration measured. Twenty-one of 42 (50%) achieved the target concentration at the first steady-state measurement. An additional 13 of 42 (31%) achieved the target following 2 to 4 dose modifications. The dose required to first achieve a value in the target range was 22.3 (18.0-27.1) mg/kg/day in children aged <12 years and 12.0 (9.8-14.0) mg/kg/day in children aged ≥12 years. After reaching the target, 59% and 81% of repeated steady-state measurements were in the therapeutic range in patients aged <12 years and ≥12 years, respectively. CONCLUSIONS: Reaching therapeutic voriconazole serum trough concentrations required doses larger than currently recommended by the American Academy of Pediatrics. Multiple dose adjustments and TDM measurements were required to achieve and maintain therapeutic voriconazole serum concentrations.

Use of Clinical Practice Guideline to Improve Management of Osteoarticular Infections in Children.

OBJECTIVES: Osteoarticular infections (OAIs) in children pose significant risks if incorrectly managed. We introduced a clinical practice guideline (CPG) to decrease use of broad-spectrum and intravenous (IV) antibiotics for OAI treatment. The primary aims of our project were to decrease the percent of patients with empirical broad cephalosporin use to 10% and decrease IV antibiotic therapy on discharge to 20% while increasing narrow-spectrum oral antibiotic use to 80% within 24 months. METHODS: We used quality improvement methodology to study patients diagnosed with OAIs. Interventions included multidisciplinary workgroup planning, CPG implementation, education, information technology, and stakeholder feedback. Outcome measures were the percentage of patients prescribed empirical broad-spectrum cephalosporins, percent discharged on IV antibiotics, and percent discharged on narrow-spectrum oral antibiotics. Process measures included percent of patients hospitalized on medicine service and infectious diseases consultation. Balancing measures included rates of adverse drug reactions, disease complications, length of stay, and readmission within 90 days. The impact of the interventions was assessed with run and control charts. RESULTS: A total of 330 patients were included over 96 months. The percentage of patients with empirical broad cephalosporin coverage decreased from 47% to 10%, percent discharged on IV antibiotics decreased from 75% to 11%, and percent discharged on narrow-spectrum oral antibiotics increased from 24% to 84%. Adverse drug reactions decreased from 31% to 10%. Rates of complications, readmissions, and length of stay were unchanged. CONCLUSIONS: Through development and implementation of a CPG for OAI management, we demonstrated decreased use of empirical broad-spectrum antibiotics and improved definitive antibiotic management.

Safety and Immunogenicity of Live Viral Vaccines in a Multicenter Cohort of Pediatric Transplant Recipients.

Importance: Live vaccines (measles-mumps-rubella [MMR] and varicella-zoster virus [VZV]) have not been recommended after solid organ transplant due to concern for inciting vaccine strain infection in an immunocompromised host. However, the rates of measles, mumps, and varicella are rising nationally and internationally, leaving susceptible immunocompromised children at risk for life-threating conditions. Objective: To determine the safety and immunogenicity of live vaccines in pediatric liver and kidney transplant recipients. Design, Setting, and Participants: This cohort study included select pediatric liver and kidney transplant recipients who had not completed their primary MMR and VZV vaccine series and/or who displayed nonprotective serum antibody levels at enrollment between January 1, 2002, and February 28, 2023. Eligibility for live vaccine was determined by individual US pediatric solid organ transplant center protocols. Exposures: Exposure was defined as receipt of a posttransplant live vaccine. Transplant recipients received 1 to 3 doses of MMR vaccine and/or 1 to 3 doses of VZV vaccine. Main Outcome and Measure: Safety data were collected following each vaccination, and antibody levels were obtained at 0 to 3 months and 1 year following vaccination. Comparisons were performed using Mann-Whitney U test, and factors associated with development of postvaccination protective antibodies were explored using univariate analysis. Results: The cohort included 281 children (270 [96%] liver, 9 [3%] kidney, 2 [1%] liver-kidney recipients) from 18 centers. The median time from transplant to enrollment was 6.3 years (IQR, 3.4-11.1 years). The median age at first posttransplant vaccine was 8.9 years (IQR, 4.7-13.8 years). A total of 202 of 275 (73%) children were receiving low-level monotherapy immunosuppression at the time of vaccination. The majority of children developed protective antibodies following vaccination (107 of 149 [72%] varicella, 130 of 152 [86%] measles, 100 of 120 [83%] mumps, and 124 of 125 [99%] rubella). One year post vaccination, the majority of children who initially mounted protective antibodies maintained this protection (34 of 44 [77%] varicella, 45 of 49 [92%] measles, 35 of 42 [83%] mumps, 51 of 54 [94%] rubella). Five children developed clinical varicella, all of which resolved within 1 week. There were no cases of measles or rubella and no episodes of graft rejection within 1 month of vaccination. There was no association between antibody response and immunosuppression level at the time of vaccination. Conclusions and Relevance: The findings suggest that live vaccinations may be safe and immunogenic after solid organ transplant in select pediatric recipients and can offer protection against circulating measles, mumps, and varicella.

Adjunctive Diagnostic Studies Completed Following Detection of Candidemia in Children: Secondary Analysis of Observed Practice From a Multicenter Cohort Study Conducted by the Pediatric Fungal Network.

BACKGROUND: Adjunctive diagnostic studies (aDS) are recommended to identify occult dissemination in patients with candidemia. Patterns of evaluation with aDS across pediatric settings are unknown. METHODS: Candidemia episodes were included in a secondary analysis of a multicenter comparative effectiveness study that prospectively enrolled participants age 120 days to 17 years with invasive candidiasis (predominantly candidemia) from 2014 to 2017. Ophthalmologic examination (OE), abdominal imaging (AbdImg), echocardiogram, neuroimaging, and lumbar puncture (LP) were performed per clinician discretion. Adjunctive diagnostic studies performance and positive results were determined per episode, within 30 days from candidemia onset. Associations of aDS performance with episode characteristics were evaluated via mixed-effects logistic regression. RESULTS: In 662 pediatric candidemia episodes, 490 (74%) underwent AbdImg, 450 (68%) OE, 426 (64%) echocardiogram, 160 (24%) neuroimaging, and 76 (11%) LP; performance of each aDS per episode varied across sites up to 16-fold. Longer durations of candidemia were associated with undergoing OE, AbdImg, and echocardiogram. Immunocompromised status (58% of episodes) was associated with undergoing AbdImg (adjusted odds ratio [aOR] 2.38; 95% confidence intervals [95% CI] 1.51-3.74). Intensive care at candidemia onset (30% of episodes) was associated with undergoing echocardiogram (aOR 2.42; 95% CI 1.51-3.88). Among evaluated episodes, positive OE was reported in 15 (3%), AbdImg in 30 (6%), echocardiogram in 14 (3%), neuroimaging in 9 (6%), and LP in 3 (4%). CONCLUSIONS: Our findings show heterogeneity in practice, with some clinicians performing aDS selectively, potentially influenced by clinical factors. The low frequency of positive results suggests that targeted application of aDS is warranted.

COVID-19 Pandemic Exacerbates Childhood Immunization Disparities.

BACKGROUND: The objective of this study was to measure the recovery of routine pediatric immunization after a period of reduced vaccine administrations in the early weeks of the COVID-19 pandemic. METHODS: We recorded data on vaccines administered in Children's Wisconsin primary care or urgent care clinics from January 2019 through December 2020 and aggregated data by date and insurance type. RESULTS: During the gradual reopening period after week 21 in 2020, vaccine administration returned to prepandemic levels for children with commercial insurance but remained below baseline rates until the end of 2020 for children with Medicaid insurance. DISCUSSION: The decline in pediatric vaccination in 2020 disproportionately affected children with Medicaid insurance.

Metamorphic Protein Folding Encodes Multiple Anti-Candida Mechanisms in XCL1.

Candida species cause serious infections requiring prolonged and sometimes toxic therapy. Antimicrobial proteins, such as chemokines, hold great interest as potential additions to the small number of available antifungal drugs. Metamorphic proteins reversibly switch between multiple different folded structures. XCL1 is a metamorphic, antimicrobial chemokine that interconverts between the conserved chemokine fold (an α-ß monomer) and an alternate fold (an all-ß dimer). Previous work has shown that human XCL1 kills C. albicans but has not assessed whether one or both XCL1 folds perform this activity. Here, we use structurally locked engineered XCL1 variants and Candida killing assays, adenylate kinase release assays, and propidium iodide uptake assays to demonstrate that both XCL1 folds kill Candida, but they do so via different mechanisms. Our results suggest that the alternate fold kills via membrane disruption, consistent with previous work, and the chemokine fold does not. XCL1 fold-switching thus provides a mechanism to regulate the XCL1 mode of antifungal killing, which could protect surrounding tissue from damage associated with fungal membrane disruption and could allow XCL1 to overcome candidal resistance by switching folds. This work provides inspiration for the future design of switchable, multifunctional antifungal therapeutics.

Blastomycosis in 64 Wisconsin Children: Unanticipated Infection Risk and Severity in Urban Residents.

BACKGROUND: Blastomycosis, an endemic mycosis of immunocompetent individuals, is typically seen after exposure to waterways within rural wooded regions. It is not considered a disease of urban environments. Infection can be solely pneumonic or disseminate to skin, bone or central nervous system. Unknown factors influence disease acquisition and severity in children. METHODS: We analyzed acquisition risks and disease characteristics of blastomycosis in children seen at a tertiary care center from 1998 to 2018 to identify potential exposure sources, measure disease severity and assess the effect of race upon disease severity. RESULTS: Of 64 infected children, mean age was 12.9 years, with median time to diagnosis 38.5 days. About 72% were male, 38% resided in urban counties and 50% had typical environmental exposure. Isolated pulmonary infection occurred in 33 (52%). The remainder had evidence of dissemination to skin (N = 13), bone (N = 16; 7 clinically silent) and cranium (N = 7; 3 clinically silent). Infection was moderate/severe in 19 (30%). Two children (3%) died. About 79% of children with moderate/severe disease (P = 0.008) and 71% of urban children (P = 0.007) lacked typical environmental exposure. Comparing children from urban counties to other residences, 63% versus 5% were black (P < 0.001) and 71% versus 35% developed extrapulmonary dissemination (P = 0.006). Moderate/severe disease was seen in 7/17 (42%) black children but only 12/47 (26%) children of other races (P = 0.23). CONCLUSIONS: Blastomycosis, can be endemic in urban children in the absence of typical exposure history, have frequent, sometimes clinically silent, extrapulmonary dissemination and possibly produces more severe disease in black children.