Interstitial lung disease as an indication for pediatric lung transplant.

Interstitial lung disease can be an indication for lung transplant at any age, but it is a particularly common indication for lung transplant in infants. Nevertheless, not all interstitial lung diseases will lead to lung transplant in childhood. Genetic testing has aided the identification of these diseases in children. In severely affected patients, however, definitive diagnosis is not always necessary to consider referral to a transplant center. At experienced transplant centers, a multidisciplinary team educates patient families and aids in the transplant evaluation of children with interstitial lung disease. Children who have undergone transplant require lifetime immunosuppression and close surveillance, but can enjoy good quality of life for years following surgery.

The Highly Conserved Stem-Loop II Motif Is Dispensable for SARS-CoV-2.

The stem-loop II motif (s2m) is an RNA structural element that is found in the 3' untranslated region (UTR) of many RNA viruses, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Though the motif was discovered over 25 years ago, its functional significance is unknown. In order to understand the importance of s2m, we created viruses with deletions or mutations of the s2m by reverse genetics and also evaluated a clinical isolate harboring a unique s2m deletion. Deletion or mutation of the s2m had no effect on growth in vitro or on growth and viral fitness in Syrian hamsters in vivo. We also compared the secondary structure of the 3' UTR of wild-type and s2m deletion viruses using selective 2'-hydroxyl acylation analyzed by primer extension and mutational profiling (SHAPE-MaP) and dimethyl sulfate mutational profiling and sequencing (DMS-MaPseq). These experiments demonstrate that the s2m forms an independent structure and that its deletion does not alter the overall remaining 3'-UTR RNA structure. Together, these findings suggest that s2m is dispensable for SARS-CoV-2. IMPORTANCE RNA viruses, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), contain functional structures to support virus replication, translation, and evasion of the host antiviral immune response. The 3' untranslated region of early isolates of SARS-CoV-2 contained a stem-loop II motif (s2m), which is an RNA structural element that is found in many RNA viruses. This motif was discovered over 25 years ago, but its functional significance is unknown. We created SARS-CoV-2 with deletions or mutations of the s2m and determined the effect of these changes on viral growth in tissue culture and in rodent models of infection. Deletion or mutation of the s2m element had no effect on growth in vitro or on growth and viral fitness in Syrian hamsters in vivo. We also observed no impact of the deletion on other known RNA structures in the same region of the genome. These experiments demonstrate that s2m is dispensable for SARS-CoV-2.

Position paper: Models of post-transplant care for individuals with cystic fibrosis.

There is no consensus on the best model of care for individuals with CF to manage the non-pulmonary complications that persist after lung transplant. The CF Foundation virtually convened a group of international experts in CF and lung-transplant care. The committee reviewed literature and shared the post-lung transplant model of care practiced by their programs. The committee then developed a survey that was distributed internationally to both the clinical and individual with CF/family audiences to determine the strengths, weaknesses, and preferences for various models of transplant care. Discussion generated two models to accomplish optimal CF care after transplant. The first model incorporates the CF team into care and proposes delineation of responsibilities for the CF and transplant teams. This model is reliant on outstanding communication between the teams, while leveraging the expertise of the CF team for management of the non-pulmonary manifestations of CF. The transplant team manages all aspects of the transplant, including pulmonary concerns and management of immunosuppression. The second model consolidates care in one center and may be more practical for transplant programs that have expertise managing CF and have access to CF multidisciplinary care team members (e.g., located in the same institution). The best model for each program is influenced by several factors and model selection needs to be decided between the transplant and the CF center and may vary from center to center. In either model, CF lung transplant recipients require a clear delineation of the roles and responsibilities of their providers and mechanisms for effective communication.

The highly conserved stem-loop II motif is dispensable for SARS-CoV-2.

The stem-loop II motif (s2m) is a RNA structural element that is found in the 3' untranslated region (UTR) of many RNA viruses including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Though the motif was discovered over twenty-five years ago, its functional significance is unknown. In order to understand the importance of s2m, we created viruses with deletions or mutations of the s2m by reverse genetics and also evaluated a clinical isolate harboring a unique s2m deletion. Deletion or mutation of the s2m had no effect on growth in vitro , or growth and viral fitness in Syrian hamsters in vivo . We also compared the secondary structure of the 3' UTR of wild type and s2m deletion viruses using SHAPE-MaP and DMS-MaPseq. These experiments demonstrate that the s2m forms an independent structure and that its deletion does not alter the overall remaining 3'UTR RNA structure. Together, these findings suggest that s2m is dispensable for SARS-CoV-2. IMPORTANCE: RNA viruses, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) contain functional structures to support virus replication, translation and evasion of the host antiviral immune response. The 3' untranslated region of early isolates of SARS-CoV-2 contained a stem-loop II motif (s2m), which is a RNA structural element that is found in many RNA viruses. This motif was discovered over twenty-five years ago, but its functional significance is unknown. We created SARS-CoV-2 with deletions or mutations of the s2m and determined the effect of these changes on viral growth in tissue culture and in rodent models of infection. Deletion or mutation of the s2m element had no effect on growth in vitro , or growth and viral fitness in Syrian hamsters in vivo . We also observed no impact of the deletion on other known RNA structures in the same region of the genome. These experiments demonstrate that the s2m is dispensable for SARS-CoV-2.

The highly conserved stem-loop II motif is important for the lifecycle of astroviruses but dispensable for SARS-CoV-2.

The stem-loop II motif (s2m) is an RNA element present in viruses from divergent viral families, including astroviruses and coronaviruses, but its functional significance is unknown. We created deletions or substitutions of the s2m in astrovirus VA1 (VA1), classic human astrovirus 1 (HAstV1) and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). For VA1, recombinant virus could not be rescued upon partial deletion of the s2m or substitutions of G-C base pairs. Compensatory substitutions that restored the G-C base-pair enabled recovery of VA1. For HAstV1, a partial deletion of the s2m resulted in decreased viral titers compared to wild-type virus, and reduced activity in a replicon system. In contrast, deletion or mutation of the SARS-CoV-2 s2m had no effect on the ability to rescue the virus, growth in vitro , or growth in Syrian hamsters. Our study demonstrates the importance of the s2m is virus-dependent.

Violence-related distress and lung function in two longitudinal studies of youth.

BACKGROUND: Exposure to violence has been associated with lower lung function in cross-sectional studies. METHODS: We examined whether increasing violence-related distress over time is associated with worse lung function and worse asthma control or quality of life in a secondary analysis of a 48-week randomised clinical trial in 98 youth with asthma (aged 9-16 years) treated with low-dose inhaled corticosteroids (Vitamin D Kids Asthma Study (VDKA)). We then replicated our findings for lung function in a prospective study of 232 Puerto Rican youth followed for an average of 5.4 years. Violence-related distress was assessed using the Checklist of Children's Distress Symptoms (CCDS) scale. Our outcomes of interest were percent predicted lung function measures and (in VDKA only) asthma control (assessed using the Asthma Control Test) and asthma-related quality of life (assessed using the Pediatric Asthma Quality of Life Questionnaire (PAQLQ)). RESULTS: In a multivariable analysis in VDKA, each 1-point increment in CCDS score was associated with decrements of 3.27% in forced expiratory volume in 1 s (FEV1) % pred (95% CI -6.44- -0.22%; p=0.04), 2.65% in forced vital capacity (FVC) % pred (95% CI -4.86- -0.45%; p=0.02) and 0.30 points in the overall PAQLQ score (95% CI -0.50- -0.10 points; p<0.01). Similar findings for FEV1 and FVC were obtained in the prospective study of Puerto Rican youth. CONCLUSIONS: Our findings suggest that violence-related distress may worsen lung function and quality of life in youth with asthma (even those treated with low-dose inhaled corticosteroids), and further support policies to reduce exposure to violence among children in the USA and Puerto Rico.

Effect of vitamin D supplementation on total and allergen-specific IgE in children with asthma and low vitamin D levels.

BACKGROUND: Observational studies have yielded inconsistent findings for the relation between vitamin D level and total IgE or allergic sensitization. OBJECTIVE: To determine whether vitamin D supplementation reduces levels of total IgE and IgE to each of 2 common indoor allergens in children with asthma and low vitamin D levels. METHODS: Total IgE, IgE to Dermatophagoides pteronyssinus, and IgE to Blattella germanica were measured at the randomization and exit visits for 174 participants in the Vitamin D Kids Asthma Study, a multicenter, double-blind, randomized placebo-controlled trial of vitamin D3 supplementation (4000 IU/d) to prevent severe exacerbations in children with persistent asthma and vitamin D levels less than 30 ng/mL. Multivariable linear regression was used for the analysis of the effect of vitamin D supplementation on change in each IgE measure. RESULTS: Participants were followed for an average of 316 days. At the exit visit, more subjects in the vitamin D arm achieved a vitamin D level equal to or more than 30 ng/mL compared with those in the placebo arm (87% vs 30%; P < .001). In a multivariable analysis, vitamin D3 supplementation had no significant effect on change in total IgE, IgE to Dermatophagoides pteronyssinus, or IgE to Blattella germanica between the exit and randomization visits (eg, for log10 total IgE, ß = 0.007; 95% CI, -0.061 to 0.074; P = .85). CONCLUSIONS: Vitamin D supplementation, compared with placebo, has no significant effect on serum levels of total IgE, IgE to dust mite, or IgE to cockroach in children with asthma and low vitamin D levels.

CTOTC-08: A multicenter randomized controlled trial of rituximab induction to reduce antibody development and improve outcomes in pediatric lung transplant recipients.

We conducted a randomized, placebo-controlled, double-blind study of pediatric lung transplant recipients, hypothesizing that rituximab plus rabbit anti-thymocyte globulin induction would reduce de novo donor-specific human leukocyte antigen antibodies (DSA) development and improve outcomes. We serially obtained clinical data, blood, and respiratory samples for at least one year posttransplant. We analyzed peripheral blood lymphocytes by flow cytometry, serum for antibody development, and respiratory samples for viral infections using multiplex PCR. Of 45 subjects enrolled, 34 were transplanted and 27 randomized to rituximab (n = 15) or placebo (n = 12). No rituximab-treated subjects versus five placebo-treated subjects developed de novo DSA with mean fluorescence intensity >2000. There was no difference between treatment groups in time to the primary composite outcome endpoint (death, bronchiolitis obliterans syndrome [BOS] grade 0-p, obliterative bronchiolitis or listing for retransplant). A post-hoc analysis substituting more stringent chronic lung allograft dysfunction criteria for BOS 0-p showed no difference in outcome (p = .118). The incidence of adverse events including infection and rejection episodes was no different between treatment groups. Although the study was underpowered, we conclude that rituximab induction may have prevented early DSA development in pediatric lung transplant recipients without adverse effects and may improve outcomes (Clinical Trials: NCT02266888).

Remote intervention engagement and outcomes in the Clinical Trials in Organ Transplantation in Children consortium multisite trial.

Remote interventions are increasingly used in transplant medicine but have rarely been rigorously evaluated. We investigated a remote intervention targeting immunosuppressant management in pediatric lung transplant recipients. Patients were recruited from a larger multisite trial if they had a Medication Level Variability Index (MLVI) ≥2.0, indicating worrisome tacrolimus level fluctuation. The manualized intervention included three weekly phone calls and regular follow-up calls. A comparison group included patients who met enrollment criteria after the subprotocol ended. Outcomes were defined before the intent-to-treat analysis. Feasibility was defined as ≥50% of participants completing the weekly calls. MLVI was compared pre- and 180 days postenrollment and between intervention and comparison groups. Of 18 eligible patients, 15 enrolled. Seven additional patients served as the comparison. Seventy-five percent of participants completed ≥3 weekly calls; average time on protocol was 257.7 days. Average intervention group MLVI was significantly lower (indicating improved blood level stability) at 180 days postenrollment (2.9 ± 1.29) compared with pre-enrollment (4.6 ± 2.10), p = .02. At 180 days, MLVI decreased by 1.6 points in the intervention group but increased by 0.6 in the comparison group (p = .054). Participants successfully engaged in a long-term remote intervention, and their medication blood levels stabilized. NCT02266888.

Effect of Vitamin D3 Supplementation on Severe Asthma Exacerbations in Children With Asthma and Low Vitamin D Levels: The VDKA Randomized Clinical Trial.

Importance: Severe asthma exacerbations cause significant morbidity and costs. Whether vitamin D3 supplementation reduces severe childhood asthma exacerbations is unclear. Objective: To determine whether vitamin D3 supplementation improves the time to a severe exacerbation in children with asthma and low vitamin D levels. Design, Setting, and Participants: The Vitamin D to Prevent Severe Asthma Exacerbations (VDKA) Study was a randomized, double-blind, placebo-controlled clinical trial of vitamin D3 supplementation to improve the time to severe exacerbations in high-risk children with asthma aged 6 to 16 years taking low-dose inhaled corticosteroids and with serum 25-hydroxyvitamin D levels less than 30 ng/mL. Participants were recruited from 7 US centers. Enrollment started in February 2016, with a goal of 400 participants; the trial was terminated early (March 2019) due to futility, and follow-up ended in September 2019. Interventions: Participants were randomized to vitamin D3, 4000 IU/d (n = 96), or placebo (n = 96) for 48 weeks and maintained with fluticasone propionate, 176 µg/d (6-11 years old), or 220 µg/d (12-16 years old). Main Outcomes and Measures: The primary outcome was the time to a severe asthma exacerbation. Secondary outcomes included the time to a viral-induced severe exacerbation, the proportion of participants in whom the dose of inhaled corticosteroid was reduced halfway through the trial, and the cumulative fluticasone dose during the trial. Results: Among 192 randomized participants (mean age, 9.8 years; 77 girls [40%]), 180 (93.8%) completed the trial. A total of 36 participants (37.5%) in the vitamin D3 group and 33 (34.4%) in the placebo group had 1 or more severe exacerbations. Compared with placebo, vitamin D3 supplementation did not significantly improve the time to a severe exacerbation: the mean time to exacerbation was 240 days in the vitamin D3 group vs 253 days in the placebo group (mean group difference, -13.1 days [95% CI, -42.6 to 16.4]; adjusted hazard ratio, 1.13 [95% CI, 0.69 to 1.85]; P = .63). Vitamin D3 supplementation, compared with placebo, likewise did not significantly improve the time to a viral-induced severe exacerbation, the proportion of participants whose dose of inhaled corticosteroid was reduced, or the cumulative fluticasone dose during the trial. Serious adverse events were similar in both groups (vitamin D3 group, n = 11; placebo group, n = 9). Conclusions and Relevance: Among children with persistent asthma and low vitamin D levels, vitamin D3 supplementation, compared with placebo, did not significantly improve the time to a severe asthma exacerbation. The findings do not support the use of vitamin D3 supplementation to prevent severe asthma exacerbations in this group of patients. Trial Registration: ClinicalTrials.gov Identifier: NCT02687815.

Anellovirus Dynamics Are Associated With Primary Graft Dysfunction in Lung Transplantation.

Primary graft dysfunction (PGD) is the leading cause of early death in lung transplant. Anelloviruses are small circular DNA viruses that have been noted to be present at elevated levels in immunosuppressed patients. They have been associated with both short- and long-term outcomes in lung transplant, and we hypothesized that anellovirus dynamics might be associated with the development of PGD. METHODS: We analyzed alphatorquevirus (ie, an anellovirus genus) levels in whole blood samples from 64 adult lung transplant recipients. RESULTS: Patients with a relatively rapid rise in alphatorquevirus levels in the week following transplant were less likely to develop higher-grade PGD over the first 3 days following transplant (P = 0.031). CONCLUSIONS: This study is the first to establish an association between the development of PGD and a component of the blood virome. While it is not known whether anelloviruses directly affect outcomes in lung transplant, they may serve as a biomarker of immune status in lung transplant recipients.

Vertical transmission in Caenorhabditis nematodes of RNA molecules encoding a viral RNA-dependent RNA polymerase.

Here, we report on the discovery in Caenorhabditis nematodes of multiple vertically transmitted RNAs coding for putative RNA-dependent RNA polymerases. Their sequences share similarity to distinct RNA viruses, including bunyaviruses, narnaviruses, and sobemoviruses. The sequences are present exclusively as RNA and are not found in DNA form. The RNAs persist in progeny after bleach treatment of adult animals, indicating vertical transmission of the RNAs. We tested one of the infected strains for transmission to an uninfected strain and found that mating of infected animals with uninfected animals resulted in infected progeny. By in situ hybridization, we detected several of these RNAs in the cytoplasm of the male and female germline of the nematode host. The Caenorhabditis hosts were found defective in degrading exogenous double-stranded RNAs, which may explain retention of viral-like RNAs. Strikingly, one strain, QG551, harbored three distinct virus-like RNA elements. Specific patterns of small RNAs complementary to the different viral-like RNAs were observed, suggesting that the different RNAs are differentially recognized by the RNA interference (RNAi) machinery. While vertical transmission of viruses in the family Narnaviridae, which are known as capsidless viruses, has been described in fungi, these observations provide evidence that multicellular animal cells harbor similar viruses.

Laboratory mice born to wild mice have natural microbiota and model human immune responses.

Laboratory mouse studies are paramount for understanding basic biological phenomena but also have limitations. These include conflicting results caused by divergent microbiota and limited translational research value. To address both shortcomings, we transferred C57BL/6 embryos into wild mice, creating "wildlings." These mice have a natural microbiota and pathogens at all body sites and the tractable genetics of C57BL/6 mice. The bacterial microbiome, mycobiome, and virome of wildlings affect the immune landscape of multiple organs. Their gut microbiota outcompete laboratory microbiota and demonstrate resilience to environmental challenges. Wildlings, but not conventional laboratory mice, phenocopied human immune responses in two preclinical studies. A combined natural microbiota- and pathogen-based model may enhance the reproducibility of biomedical studies and increase the bench-to-bedside safety and success of immunological studies.

American Thoracic Society 2019 Pediatric Core Curriculum.

The American Thoracic Society Pediatric Core Curriculum updates clinicians annually in pediatric pulmonary disease in a 3 to 4 year recurring cycle of topics. The 2019 course was presented in May during the Annual International Conference. An American Board of Pediatrics Maintenance of Certification module and a continuing medical education exercise covering the contents of the Core Curriculum can be accessed online at www.thoracic.org.

Anellovirus loads are associated with outcomes in pediatric lung transplantation.

Anelloviruses are DNA viruses ubiquitously present in human blood. Due to their elevated levels in immunosuppressed patients, anellovirus levels have been proposed as a marker of immune status. We hypothesized that low anellovirus levels, reflecting relative immunocompetence, would be associated with adverse outcomes in pediatric lung transplantation. We assayed blood samples from 57 patients in a multicenter study for alpha- and betatorquevirus, two anellovirus genera. The primary short-term outcome of interest was acute rejection, and longer-term outcomes were analyzed individually and as "composite" (death, chronic rejection, or retransplant within 2 years). Patients with low alphatorquevirus levels at 2 weeks post-transplantation were more likely to develop acute rejection within 3 months after transplant (P = .013). Low betatorquevirus levels at 6 weeks and 6 months after transplant were associated with death (P = .047) and the composite outcome (P = .017), respectively. There was an association between low anellovirus levels and adverse outcomes in pediatric lung transplantation. Alphatorquevirus levels were associated with short-term outcomes (ie, acute rejection), while betatorquevirus levels were associated with longer-term outcomes (ie, death, or composite outcome within 2 years). These observations suggest that anelloviruses may serve as useful biomarkers of immune status and predictors of adverse outcomes.

Folate Deficiency, Atopy, and Severe Asthma Exacerbations in Puerto Rican Children.

BACKGROUND: Little is known about folate and atopy or severe asthma exacerbations. We examined whether folate deficiency is associated with number of positive skin tests to allergens or severe asthma exacerbations in a high-risk population and further assessed whether such association is explained or modified by vitamin D status. METHODS: Cross-sectional study of 582 children aged 6 to 14 years with (n = 304) and without (n = 278) asthma in San Juan, Puerto Rico. Folate deficiency was defined as plasma folate less than or equal to 20 ng/ml. Our outcomes were the number of positive skin tests to allergens (range, 0-15) in all children and (in children with asthma) one or more severe exacerbations in the previous year. Logistic and negative binomial regression models were used for the multivariate analysis. All multivariate models were adjusted for age, sex, household income, residential proximity to a major road, and (for atopy) case/control status; those for severe exacerbations were also adjusted for use of inhaled corticosteroids and vitamin D insufficiency (a plasma 25[OH]D < 30 ng/ml). MEASUREMENTS AND MAIN RESULTS: In a multivariate analysis, folate deficiency was significantly associated with an increased degree of atopy and 2.2 times increased odds of at least one severe asthma exacerbation (95% confidence interval for odds ratio, 1.1-4.6). Compared with children who had normal levels of both folate and vitamin D, those with both folate deficiency and vitamin D insufficiency had nearly eightfold increased odds of one or more severe asthma exacerbation (95% confidence interval for adjusted odds ratio, 2.7-21.6). CONCLUSIONS: Folate deficiency is associated with increased degree of atopy and severe asthma exacerbations in school-aged Puerto Ricans. Vitamin D insufficiency may further increase detrimental effects of folate deficiency on severe asthma exacerbations.

Lung Transplantation in Cystic Fibrosis: Trends and Controversies.

This article is not an overview of all facets of lung transplantation in cystic fibrosis (CF), but rather it is intended as a review of current allocation controversies, as well as of trends in diagnostics and management in lung transplant recipients and in patients with end-stage lung disease. Despite changes in donor and recipient selection, long-term survival in pediatric lung transplant has continued to be limited by chronic lung allograft dysfunction (CLAD). Due to, in part, this short survival benefit, transplant continues to be an appropriate option for only a subset of pediatric patients with CF. The feasibility of transplant as a therapeutic option is also affected by the limited pediatric organ supply, which has moreover contributed to controversy over lung allocation. Debates over the allocation of this scarce resource, however, may also help to drive innovation in the field of lung transplant. Longer pretransplant survival-as aided by new lung bypass technologies, for example-could help to alleviate organ shortages, as well as facilitate the transport of organs to suitable pediatric recipients. Improved diagnosis and treatment for CLAD and for antibody-mediated rejection have the potential to extend survival in pediatric lung transplant. Regardless, the relative rarity of transplant could pose future challenges for pediatric lung transplant programs, which require adequate numbers of patients to maintain proper expertise.

Mouse allergen exposure and decreased risk of allergic rhinitis in school-aged children.

BACKGROUND: Little is known about exposure to mouse allergen (Mus m 1) and allergic rhinitis (AR). OBJECTIVE: To evaluate the association between mouse allergen exposure and AR in children. METHODS: We examined the relation between mouse allergen level in house dust and AR in 511 children aged 6 to 14 years in San Juan, Puerto Rico. Study participants were chosen from randomly selected households using a multistage probability sample design. The study protocol included questionnaires, allergy skin testing, and collection of blood and dust samples. AR was defined as current rhinitis symptoms and skin test reactivity to at least one allergen. RESULTS: In the multivariate analyses, mouse allergen level was associated with a 25% decreased odds of AR in participating children (95% confidence interval, 0.62-0.92). Although endotoxin and mouse allergen levels were significantly correlated (r = 0.184, P < .001), the observed inverse association between Mus m 1 and AR was not explained by levels of endotoxin or other markers of microbial or fungal exposure (peptidoglycan and glucan). CONCLUSION: Mouse allergen exposure is associated with decreased odds of AR in Puerto Rican school-aged children.