Identification of Irpex and Rhodotorula on surveillance bronchoscopy in a pediatric lung transplant recipient: A case report and review of literature of these atypical fungal organisms.

BACKGROUND: Invasive fungal disease (IFD) is a frequent complication in pediatric lung transplant recipients, occurring in up to 12% of patients in the first year. Risk factors for infection include impaired lung defenses and intense immunosuppressive regimens. While most IFD occurs from Aspergillus, other fungal conidia are continuously inhaled, and infections with fungi on a spectrum of human pathogenicity can occur. CASE REPORT: We report a case of a 17-year-old lung transplant recipient in whom Irpex lacteus and Rhodotorula species were identified during surveillance bronchoscopy. She was asymptomatic and deemed to be colonized by Irpex lacteus and Rhodotorula species following transplant. 2 years after transplantation, she developed a fever, respiratory symptoms, abnormal lung imaging, and histological evidence of acute and chronic bronchitis on transbronchial biopsy. After developing symptoms concerning for a pulmonary infection and graft dysfunction, she was treated for a presumed IFD. Unfortunately, further diagnostic testing could not be performed at this time given her tenuous clinical status. Despite the initiation of antifungal therapy, her graft function continued to decline resulting in a second lung transplantation. CONCLUSIONS: This case raises the concern for IFD in lung transplant recipients from Irpex species. Further investigation is needed to understand the pathogenicity of this organism, reduce the incidence and mortality of IFD in lung transplant recipients, and refine the approach to diagnosis and manage the colonization and isolation of rare, atypical fungal pathogens in immunocompromised hosts.

Sudden Death in a Child With Ocular Lesions.

This case report describes a patient treated for ocular lesions who died suddenly at age 8 years and was diagnosed postmortem with Carney complex.

Sexual and reproductive health in military settings: A qualitative study.

Mission readiness is critical to the operational success of the United States (US) military and includes having a healthy and fit fighting force. Service members and their dependents have access to a wide range of sexual and reproductive health services with no out-of-pocket costs. Despite this access, negative outcomes such as sexually transmitted infections (STIs) and unintended pregnancy persist. Semi-structured, in-depth interviews were conducted with service members and stakeholders (e.g. medical providers). Interviews explored the individual, interpersonal, organizational, and institutional factors that inform sexual norms, behaviors, and healthcare experiences in the US military. Interview transcripts were coded manually; data were summarized for themes related to unique aspects of military culture and healthcare affecting sexual and reproductive health. Twenty-five (25) service members and 15 stakeholders completed interviews. Four themes emerged: 1) despite free access, both general and military-specific barriers to sexual and reproductive healthcare persist; 2) general and military-specific cultural norms apply to sexual behavior and care seeking; 3) sexual and reproductive health-related norms can be perceived as confusing and contradictory within the military; and 4) resources addressing sexual assault are ubiquitous in military settings, but resources addressing prevention of STIs and unintended pregnancy are limited. Both general and military-specific norms, behavior, and healthcare experiences need to be considered in clinical care, public health campaigns, and other efforts to promote sexual and reproductive health in military settings.

Large Malignant Testicular/Paratesticular Tumors in Adolescence: Assessment of Gross Tumor Size in a Vulnerable Age Group.

OBJECTIVE: We hypothesized that reticence to address a groin mass may result in late presentation of testicular/paratesticular malignancy in early puberty through adolescence. METHODS: Malignant testicular and paratesticular tumors (malignant germ cell tumors and rhabdomyosarcomas) diagnosed at our institution from 1994-2023 for patients aged 11-20 were included. Clinicopathologic features were recorded, and statistically analyzed. RESULTS: Eighty-five cases were identified. Patient ages ranged from 11 to 20 years (mean 17 years, median 16 years). The greatest tumor dimension ranged from 0.8 to 18.0 cm (mean 4.4 cm, median 3.5 cm). Ten tumors (11.8% of cases) were ≥10.0 cm. In the 11-13-year-old age group, 100% of tumors (3/3) were ≥10 cm. The proportion of tumors ≥10 cm was significantly higher in the 11-13-year-old age group than in either the 14-16-year-old (P<0.001) or 17-20-year-old (P<0.001) age groups. CONCLUSION: This adolescent cohort with malignant testicular and paratesticular tumors showed a high proportion (11.8%) of very large (≥10 cm) tumors. Although the reasons are unknown and likely multifactorial, this study suggests that adolescents, particularly the 11-13 year age group, are a vulnerable population.

Facilitators and Barriers to Breastfeeding Among Veterans Using Veterans Affairs Maternity Care Benefits.

INTRODUCTION: U.S. veterans of childbearing age represent one of the fastest growing populations using Veterans Affairs (VA) health care. The VA does not provide obstetric care directly but pays for VA-enrolled veterans to obtain outside obstetric care. The VA also provides maternity care coordination (MCC) services, including lactation support. Breastfeeding benefits mothers and babies; however, previous research shows that veteran mothers quit exclusive breastfeeding earlier than the American Academy of Pediatrics and World Health Organization recommendation of 6 months. This study aimed to understand facilitators and barriers to breastfeeding among a cohort of veterans who used VA maternity care benefits. METHODS: Qualitative data from an open-ended question from a national sample of postpartum veterans using VA pregnancy benefits were coded using deductive and inductive content analysis within a matrix framework. Quantitative data were used to contextualize the responses. RESULTS: Four themes emerged from the data: (1) impacts on health of baby/mother; (2) the ability to breastfeed; (3) early postnatal experiences breastfeeding; and (4) cost/convenience. Among those who responded to the open-ended breastfeeding question (329/669), most participants (n = 316; 96%) attempted breastfeeding their current baby. Respondents who did not initiate breastfeeding or who discontinued breastfeeding earlier than planned cited diverse reasons. These included low milk supply, poor latch, nipple pain, mental health factors, and low confidence in their ability to continue breastfeeding. Participants cited the MCC program as a facilitator to breastfeeding, and non-VA hospital experiences were mentioned as barriers. CONCLUSION: Veterans in this cohort of 329 veterans who responded to an open-ended breastfeeding question wanted and attempted to breastfeed; however, barriers such as lactation challenges and unsupportive health care providers made it difficult to continue the practice. As the MCC program grows to include more lactation professionals, MCCs may address barriers such as lactation challenges and unsupportive non-VA health care providers. Further program development should focus on addressing these challenges prenatally.

De novo mutations mediate phenotypic switching in an opportunistic human lung pathogen.

Bacteria evolving within human hosts encounter selective tradeoffs that render mutations adaptive in one context and deleterious in another. Here, we report that the cystic fibrosis-associated pathogen Burkholderia dolosa overcomes in-human selective tradeoffs by acquiring successive point mutations that alternate phenotypes. We sequenced the whole genomes of 931 respiratory isolates from two recently infected patients and an epidemiologically-linked, chronically-infected patient. These isolates are contextualized using 112 historical genomes from the same outbreak strain. Within both newly infected patients, diverse parallel mutations that disrupt O-antigen expression quickly arose, comprising 29% and 63% of their B. dolosa communities by 3 years. The selection for loss of O-antigen starkly contrasts with our previous observation of parallel O-antigen-restoring mutations after many years of chronic infection in the historical outbreak. Experimental characterization revealed that O-antigen loss increases uptake in immune cells while decreasing competitiveness in the mouse lung. We propose that the balance of these pressures, and thus whether O-antigen expression is advantageous, depends on tissue localization and infection duration. These results suggest that mutation-driven alternation during infection may be more frequent than appreciated and is underestimated without dense temporal sampling.

Increasing Incidence of Gonorrhea at an Urban STI Clinic in the United States.

Medical record data was extracted from a sexually transmitted infection (STI) clinic in Providence, Rhode Island to characterize trends in Neisseria gonorrhoeae (GC) infection and explore risk factors. Of 16,601 clinical encounters, 6% (n=991) tested GC positive: 5.28 GC case rate (per 100 encounters) in the first two years of data collection (2015-2016) and 7.04 in the last two years (2020-2021). Analysis suggested a single linear trend line over time (p<.05). Overall, in more recent years, patients were older and more like to identify as male, Black, and Hispanic/Latino, as well as to have reported a previous STI, current symptoms, and specific risk behaviors. GC-positive patients in 2020-2021 were older and more like to identify as female and Black compared to 2015-2016. Lower rates of condom use were especially salient among female patients. These findings may reflect GC trends in the community.

Pericytes contribute to pulmonary vascular remodeling via HIF2α signaling.

Vascular remodeling is the process of structural alteration and cell rearrangement of blood vessels in response to injury and is the cause of many of the world's most afflicted cardiovascular conditions, including pulmonary arterial hypertension (PAH). Many studies have focused on the effects of vascular endothelial cells and smooth muscle cells (SMCs) during vascular remodeling, but pericytes, an indispensable cell population residing largely in capillaries, are ignored in this maladaptive process. Here, we report that hypoxia-inducible factor 2α (HIF2α) expression is increased in the lung tissues of PAH patients, and HIF2α overexpressed pericytes result in greater contractility and an impaired endothelial-pericyte interaction. Using single-cell RNAseq and hypoxia-induced pulmonary hypertension (PH) models, we show that HIF2α is a major molecular regulator for the transformation of pericytes into SMC-like cells. Pericyte-selective HIF2α overexpression in mice exacerbates PH and right ventricular hypertrophy. Temporal cellular lineage tracing shows that HIF2α overexpressing reporter NG2+ cells (pericyte-selective) relocate from capillaries to arterioles and co-express SMA. This novel insight into the crucial role of NG2+ pericytes in pulmonary vascular remodeling via HIF2α signaling suggests a potential drug target for PH.

Allele-specific control of rodent and human lncRNA KMT2E-AS1 promotes hypoxic endothelial pathology in pulmonary hypertension.

Hypoxic reprogramming of vasculature relies on genetic, epigenetic, and metabolic circuitry, but the control points are unknown. In pulmonary arterial hypertension (PAH), a disease driven by hypoxia inducible factor (HIF)-dependent vascular dysfunction, HIF-2α promoted expression of neighboring genes, long noncoding RNA (lncRNA) histone lysine N-methyltransferase 2E-antisense 1 (KMT2E-AS1) and histone lysine N-methyltransferase 2E (KMT2E). KMT2E-AS1 stabilized KMT2E protein to increase epigenetic histone 3 lysine 4 trimethylation (H3K4me3), driving HIF-2α-dependent metabolic and pathogenic endothelial activity. This lncRNA axis also increased HIF-2α expression across epigenetic, transcriptional, and posttranscriptional contexts, thus promoting a positive feedback loop to further augment HIF-2α activity. We identified a genetic association between rs73184087, a single-nucleotide variant (SNV) within a KMT2E intron, and disease risk in PAH discovery and replication patient cohorts and in a global meta-analysis. This SNV displayed allele (G)-specific association with HIF-2α, engaged in long-range chromatin interactions, and induced the lncRNA-KMT2E tandem in hypoxic (G/G) cells. In vivo, KMT2E-AS1 deficiency protected against PAH in mice, as did pharmacologic inhibition of histone methylation in rats. Conversely, forced lncRNA expression promoted more severe PH. Thus, the KMT2E-AS1/KMT2E pair orchestrates across convergent multi-ome landscapes to mediate HIF-2α pathobiology and represents a key clinical target in pulmonary hypertension.

Epstein-Barr virus-positive mucocutaneous ulcer in a patient with ulcerative colitis: a recently described entity to take into account.

Epstein-Barr virus-positive mucocutaneous ulcer (EBVMCU) is a new entity recently included in the classification of B-cell lymphoproliferative disorders associated with Epstein-Barr virus (EBV). It is related to immunosuppression and it usually appears in the oropharynx or the skin, being the colon an uncommon location. We present the case of a 31-year-old man with ulcerative proctitis being treated with azathioprine (AZA) and adalimumab (ADA), who was admitted to the hospital due to suspicion of a moderate-severe flare of ulcerative proctitis. Microbiological stool analyses were negative, with a positive fecal calprotectin test (2700 mg/kg). Rectoscopy showed severe endoscopic activity, taking multiple biopsies. Intravenous steroids were started at a dose of 60 mg/day. He presented a favorable clinical and analytical evolution, being discharged from the hospital. The histological results were received at gastroenterology consultation, being compatible with EBVMCU. AZA and ADA were withdrawn, whereas descending steroid regimen and oral and topical mesalazine were continued, being the clinical response adequate.

Multiomic Analysis of Neuroinflammation and Occult Infection in Sudden Infant Death Syndrome.

Importance: Antemortem infection is a risk factor for sudden infant death syndrome (SIDS)-the leading postneonatal cause of infant mortality in the developed world. Manifestations of infection and inflammation are not always apparent in clinical settings or by standard autopsy; thus, enhanced resolution approaches are needed. Objective: To ascertain whether a subset of SIDS cases is associated with neuroinflammation and occult infection. Design, Setting, and Participants: In this case-control study, postmortem fluids from SIDS cases and controls collected between July 2011 and November 2018 were screened for elevated inflammatory markers, specifically cerebrospinal fluid (CSF) neopterin and CSF and serum cytokines. CSF, liver, and brain tissue from SIDS cases with elevated CSF neopterin were subjected to metagenomic next-generation sequencing (mNGS) to probe for infectious pathogens. Brainstem tissue from a subset of these cases was analyzed by single-nucleus RNA sequencing (snRNAseq) to measure cell type-specific gene expression associated with neuroinflammation and infection. All tissue and fluid analyses were performed from April 2019 to January 2023 in a pathology research laboratory. Included was autopsy material from infants dying of SIDS and age-matched controls dying of known causes. Exposures: There were no interventions or exposures. Main Outcomes and Measures: CSF neopterin levels were measured by high-performance liquid chromatography. Cytokines were measured by multiplex fluorometric assay. mNGS was performed on liver, CSF, brain, and brainstem tissue. snRNAseq was performed on brainstem tissue. Results: A cohort of 71 SIDS cases (mean [SD] age, 55.2 [11.4] postconceptional weeks; 42 male [59.2%]) and 20 controls (mean [SD] age, 63.2 [16.9] postconceptional weeks; 11 male [55.0%]) had CSF and/or serum available. CSF neopterin was screened in 64 SIDS cases and 15 controls, with no exclusions. Tissues from 6 SIDS cases were further analyzed. For CSF neopterin measures, SIDS samples were from infants with mean (SD) age of 54.5 (11.3) postconceptional weeks (38 male [59.4%]) and control samples were from infants with mean (SD) age of 61.5 (17.4) postconceptional weeks (7 male [46.7%]). A total of 6 SIDS cases (9.3%) with high CSF neopterin were identified, suggestive of neuroinflammation. mNGS detected human parechovirus 3 (HPeV3) in tissue and CSF from 1 of these 6 cases. snRNAseq of HPeV3-positive brainstem tissue (medulla) revealed dramatic enrichment of transcripts for genes with predominately inflammatory functions compared with 3 age-matched SIDS cases with normal CSF neopterin levels. Conclusions and Relevance: Next-generation molecular tools in autopsy tissue provide novel insight into pathogens that go unrecognized by normal autopsy methodology, including in infants dying suddenly and unexpectedly.

Neonatal and Infant Lung Disorders: Glossary, Practical Approach, and Diagnoses.

A multitude of lung disorders ranging from congenital and genetic anomalies to iatrogenic complications can affect the neonate or the infant within the first year of life. Neonatal and infant chest imaging, predominantly by plain radiography and computed tomography, is frequently employed to aid in diagnosis and management; however, these disorders can be challenging to differentiate due to their broad-ranging, and frequently overlapping radiographic features. A systematic and practical approach to imaging interpretation which includes recognition of radiologic patterns, utilization of commonly accepted nomenclature and classification, as well as interpretation of imaging findings in conjunction with clinical history can not only assist radiologists to suggest the diagnosis, but also aid clinicians in management planning. The contents of this article were endorsed by the leadership of both the World Federation of Pediatric Imaging (WFPI), and the International Society of Pediatric Thoracic Imaging (ISPTI).

Hepatosplenic schistosomiasis: an uncommon cause of liver disease in developed countries.

Schistosomiasis is a parasitic infection caused by trematode species of the genus Schistosoma. It is prevalent in tropical regions of Africa, Asia and South America, being rare in Europe, where it is usually diagnosed in immigrants and tourists from endemic areas. It has different clinical forms of presentation. Hepatosplenic schistosomiasis produces periportal fibrosis, which can progress to presinusoidal portal hypertension, with all its associated complications. We present the case of a 43-year-old female patient from the Philippines who was referred to gastroenterology consultation due to liver enzyme alteration with a predominantly cholestatic pattern. An aetiological study was performed, with negative results. An abdominal ultrasound revealed signs of chronic liver disease, with transient elastography of 9.5 kPa. A percutaneous liver biopsy was performed, with histological findings consistent with infestation by schistosome eggs, receiving treatment with praziquantel and subsequently verifying its eradication with a stool test.

CENTRE: a gradient boosting algorithm for Cell-type-specific ENhancer-Target pREdiction.

MOTIVATION: Identifying target promoters of active enhancers is a crucial step for realizing gene regulation and deciphering phenotypes and diseases. Up to now, several computational methods were developed to predict enhancer gene interactions, but they require either many epigenomic and transcriptomic experimental assays to generate cell-type (CT)-specific predictions or a single experiment applied to a large cohort of CTs to extract correlations between activities of regulatory elements. Thus, inferring CT-specific enhancer gene interactions in unstudied or poorly annotated CTs becomes a laborious and costly task. RESULTS: Here, we aim to infer CT-specific enhancer target interactions, using minimal experimental input. We introduce Cell-specific ENhancer Target pREdiction (CENTRE), a machine learning framework that predicts enhancer target interactions in a CT-specific manner, using only gene expression and ChIP-seq data for three histone modifications for the CT of interest. CENTRE exploits the wealth of available datasets and extracts cell-type agnostic statistics to complement the CT-specific information. CENTRE is thoroughly tested across many datasets and CTs and achieves equivalent or superior performance than existing algorithms that require massive experimental data. AVAILABILITY AND IMPLEMENTATION: CENTRE's open-source code is available at GitHub via https://github.com/slrvv/CENTRE.

Going to the Source: Discussions With Early and Mid-Career Faculty From Groups Underrepresented in Biomedical Research to Develop and Enhance CFAR Services.

BACKGROUND: To include, sustain, and retain HIV-focused early career faculty from groups historically excluded from biomedical research, the Providence/Boston Center for AIDS Research (CFAR) conducted focus groups and individual interviews with early and mid-career faculty to discern their needs. METHODS: We conducted focus groups and interviews with 15 faculty at institutions affiliated with Providence/Boston CFAR from groups underrepresented in biomedical research. The discussion was guided using the domains of an Asset Bundle Model encompassing scientific human capital, social capital, and financial capital. RESULT: Participants' identities, including their race, ethnicity, gender, sexual orientation, and being a parent affected their vision of themselves as scientists. Participants reported confusion or limited training on or access to resources for professional development, hiring staff, meeting NIH reporting requirements, international research, support for working parents, sabbaticals, and addressing workplace conflict or unsupportive work environments. Some described feeling like they were a burden on their mentors who seemed overextended. They identified attributes of effective mentors, such as believing in and investing in the mentee; having the requisite content area expertise and self-confidence; being able to identify mentees needs and meet them where they are; and being consistent, communicative, respectful, and kind. They described a need for additional education and support preresearch and postresearch grant award management. CONCLUSIONS: To learn how to equitably serve all interested in HIV research, CFARs should engage and include perspectives from scientists who have historically been excluded from biomedical research. Our future work will test, implement, and disseminate the ideas generated by these focus group discussions.

Lack of apoptosis leads to cellular senescence and tumorigenesis in Drosophila epithelial cells.

Programmed cell death (apoptosis) is a homeostasis program of animal tissues designed to remove cells that are unwanted or are damaged by physiological insults. To assess the functional role of apoptosis, we have studied the consequences of subjecting Drosophila epithelial cells defective in apoptosis to stress or genetic perturbations that normally cause massive cell death. We find that many of those cells acquire persistent activity of the JNK pathway, which drives them into senescent status, characterized by arrest of cell division, cell hypertrophy, Senescent Associated ß-gal activity (SA-ß-gal), reactive oxygen species (ROS) production, Senescent Associated Secretory Phenotype (SASP) and migratory behaviour. We have identified two classes of senescent cells in the wing disc: 1) those that localize to the appendage part of the disc, express the upd, wg and dpp signalling genes and generate tumour overgrowths, and 2) those located in the thoracic region do not express wg and dpp nor they induce tumour overgrowths. Whether to become tumorigenic or non-tumorigenic depends on the original identity of the cell prior to the transformation. We also find that the p53 gene contributes to senescence by enhancing the activity of JNK.

First Ecuadorian statement consensus for the evaluation and treatment of osteoporosis.

Osteoporosis management has become more relevant as the life expectancy increases. In Ecuador, approximately 19% of adults over 65 years of age have been diagnosed with osteoporosis. There is no national consensus for the management and prevention of the disease being this proposal the first Ecuadorian consensus. INTRODUCTION: In Ecuador, it is estimated that around 19% of adults over 65 years of age have osteoporosis. Due to the increase in life expectancy in the world population, the evaluation and management of osteoporosis has become more relevant. Currently, there is no national consensus for the management and prevention of the disease. The Ecuadorian Society of Rheumatology presented the project for the elaboration of the first Ecuadorian consensus for the management and prevention of osteoporosis. METHODS: A panel of experts in multiple areas and extensive experience was invited to participate. The consensus was carried out using the Delphi method. Six working dimensions were created: definition and epidemiology of osteoporosis, fracture risk prediction tools, non-pharmacological treatment, pharmacological treatment, calcium and vitamin D, and glucocorticoid-induced osteoporosis. RESULTS: The first round was held in December 2021, followed by the second round in February 2022 and the third round in March 2022. The data was shared with the specialists at the end of each round. After three rounds of work, a consensus was reached for the management and prevention of osteoporosis. CONCLUSION: This is the first Ecuadorian consensus for the management and treatment of postmenopausal osteoporosis.

An intronic variant in TBX4 in a single family with variable and severe pulmonary manifestations.

A male infant presented at term with neonatal respiratory failure and pulmonary hypertension. His respiratory symptoms improved initially, but he exhibited a biphasic clinical course, re-presenting at 15 months of age with tachypnea, interstitial lung disease, and progressive pulmonary hypertension. We identified an intronic TBX4 gene variant in close proximity to the canonical donor splice site of exon 3 (hg 19; chr17:59543302; c.401 + 3 A > T), also carried by his father who had a typical TBX4-associated skeletal phenotype and mild pulmonary hypertension, and by his deceased sister who died shortly after birth of acinar dysplasia. Analysis of patient-derived cells demonstrated a significant reduction in TBX4 expression resulting from this intronic variant. Our study illustrates the variable expressivity in cardiopulmonary phenotype conferred by TBX4 mutation and the utility of genetic diagnostics in enabling accurate identification and classification of more subtly affected family members.

Copy Number Variation and Structural Genomic Findings in 116 Cases of Sudden Unexplained Death between 1 and 28 Months of Age.

In sudden unexplained death in pediatrics (SUDP) the cause of death is unknown despite an autopsy and investigation. The role of copy number variations (CNVs) in SUDP has not been well-studied. Chromosomal microarray (CMA) data are generated for 116 SUDP cases with age at death between 1 and 28 months. CNVs are classified using the American College of Medical Genetics and Genomics guidelines and CNVs in our cohort are compared to an autism spectrum disorder (ASD) cohort, and to a control cohort. Pathogenic CNVs are identified in 5 of 116 cases (4.3%). Variants of uncertain significance (VUS) favoring pathogenic CNVs are identified in 9 cases (7.8%). Several CNVs are associated with neurodevelopmental phenotypes including seizures, ASD, developmental delay, and schizophrenia. The structural variant 47,XXY is identified in two cases (2/69 boys, 2.9%) not previously diagnosed with Klinefelter syndrome. Pathogenicity scores for deletions are significantly elevated in the SUDP cohort versus controls (p = 0.007) and are not significantly different from the ASD cohort. The finding of pathogenic or VUS favoring pathogenic CNVs, or structural variants, in 12.1% of cases, combined with the observation of higher pathogenicity scores for deletions in SUDP versus controls, suggests that CMA should be included in the genetic evaluation of SUDP.

Large cell calcifying Sertoli cell tumour: molecular and immunohistochemical assessment of a series comprising non-metastasising and metastasising neoplasms.

Large cell calcifying Sertoli cell tumour (LCCSCT) is a type of testicular sex cord-stromal tumour that may occur sporadically or in the context of Carney complex and other genetic syndromes. A subset is clinically malignant, and the molecular mechanisms that drive such aggressive behaviour remain unknown. METHODS AND RESULTS: We analysed 21 samples from 20 patients with LCCSCT (12 non-metastasising and eight metastasising) using PRKAR1A immunohistochemistry (IHC) and next-generation sequencing. All tumours except two (cases 17 and 20, both metastasising) demonstrated loss of PRKAR1A expression. Among 11 cases with interpretable sequencing results, all harboured pathogenic single nucleotide variants of PRKAR1A. Evidence of loss of heterozygosity (LOH) of PRKAR1A was present in all tumours with interpretable zygosity data, but the mechanisms of LOH were different for non-metastasising and metastasising tumours. Non-metastasising tumours demonstrated only copy-neutral LOH, while metastasising tumours demonstrated a spectrum of mechanisms of LOH, including copy-loss LOH, two concurrent mutations or copy-neutral LOH. Relevant molecular findings in non-metastasising LCCSCT were limited to PRKAR1A variants. In contrast, all metastasising LCCSCTs with interpretable data harboured additional pathogenic variants, including (but not restricted to) BRCA2 mutations with evidence of LOH and bi-allelic CDKN2A/B deletions. Three patients harboured PRKAR1A variants of inferred germline origin, including one with Carney complex and two without known syndromic features. CONCLUSIONS: This study further confirms that PRKAR1A IHC is a useful diagnostic tool for both non-metastasising and metastasising tumours and suggests that molecular analyses can be helpful to identify non-metastasising tumours with malignant potential in selected patients. Importantly, these results highlight that germline assessment could be beneficial for all patients presenting with LCCSCT.