Homozygous MFN2 variants causing severe antenatal encephalopathy with clumped mitochondria.

Pathogenic variants in the MFN2 gene are commonly associated with autosomal dominant (CMT2A2A) or recessive (CMT2A2B) Charcot-Marie-Tooth disease, with possible involvement of the CNS. Here, we present a case of severe antenatal encephalopathy with lissencephaly, polymicrogyria and cerebellar atrophy. Whole genome analysis revealed a homozygous deletion c.1717-274_1734 del (NM_014874.4) in the MFN2 gene, leading to exon 16 skipping and in-frame loss of 50 amino acids (p.Gln574_Val624del), removing the proline-rich domain and the transmembrane domain 1 (TM1). MFN2 is a transmembrane GTPase located on the mitochondrial outer membrane that contributes to mitochondrial fusion, shaping large mitochondrial networks within cells. In silico modelling showed that the loss of the TM1 domain resulted in a drastically altered topological insertion of the protein in the mitochondrial outer membrane. Fetus fibroblasts, investigated by fluorescent cell imaging, electron microscopy and time-lapse recording, showed a sharp alteration of the mitochondrial network, with clumped mitochondria and clusters of tethered mitochondria unable to fuse. Multiple deficiencies of respiratory chain complexes with severe impairment of complex I were also evidenced in patient fibroblasts, without involvement of mitochondrial DNA instability. This is the first reported case of a severe developmental defect due to MFN2 deficiency with clumped mitochondria.

Correction of a knock-in mouse model of acrodysostosis with gene therapy using a rAAV9-CAG-human PRKAR1A vector.

Acrodysostosis is a rare skeletal dysplasia caused by loss-of-function mutations in the regulatory subunit of protein kinase A (PRKAR1A). In a knock-in mouse model (PRKAR1Awt/mut) expressing one copy of the recurrent R368X mutation, we tested the effects of a rAAV9-CAG-human PRKR1A (hPRKAR1A) vector intravenously administered at 4 weeks of age. Caudal vertebrae and tibial diaphyses contained 0.52 ± 0.7 and 0.13 ± 0.3 vector genome per cell (VGC), respectively, at 10 weeks of age and 0.22 ± 0.04 and 0.020 ± 0.04 at 16 weeks while renal cortex contained 0.57 ± 0.14 and 0.26 ± 0.05 VGC. Vector-mediated hPRKAR1A expression was found in growth plate chondrocytes, osteoclasts, osteoblasts, and kidney tubular cells. Chondrocyte architecture was restored in the growth plates. Body length, tail length, and body weight were improved in vector treated PRKAR1Awt/mut mice, not the bone length of their limbs. These results provide one of the few proofs for gene therapy efficacy in a mouse model of chondrodysplasia. In addition, the increased urinary cAMP of PRKAR1Awt/mut mice was corrected almost to normal. In conclusion, gene therapy with hPRKAR1A improved skeletal growth and kidney dysfunction, the hallmarks of acrodysostosis in R368X mutated mice and humans.

Blastomycosis in Africa and the Middle East: A Comprehensive Review of Reported Cases and Reanalysis of Historical Isolates Based on Molecular Data.

BACKGROUND: Blastomycosis has been reported from countries in Africa and the Middle East, but a decades-long debate has persisted regarding whether this is the same disease known in North America and caused by Blastomyces dermatitidis and Blastomyces gilchristii. METHODS: We reviewed published cases of human and veterinary blastomycosis from Africa and the Middle East. We abstracted epidemiological and clinical features of cases, including sites of disease, diagnosis, management, outcomes, and, where available, genetic and antigenic typing of case isolates. In addition, we sequenced nucleic acids from 9 clinical isolates from Africa deposited in global collections as B. dermatitidis; for 5, we sequenced the internal transcribed spacer regions, and for the other 4 we sequenced the whole genomes. RESULTS: We identified 172 unique human patients with blastomycosis, including 159 patients from 25 African countries and 12 patients from 5 Middle Eastern countries, and also identified 7 reports of veterinary blastomycosis. In humans, cutaneous disease predominated (n = 100/137, 73%), followed by pulmonary (n = 73/129, 57%) and osteoarticular involvement (n = 61/128, 48%). Unusual direct microscopy/histopathological presentations included short hyphal fragments in tissues (n = 23/129, 18%). There were 34 genotyped case isolates that comprised 4 species: Blastomyces percursus (n = 22, 65%), from 8 countries throughout all regions; Blastomyces emzantsi (n = 9, 26%), from South Africa; B. dermatitidis (n = 1, 3%), from the Democratic Republic of Congo; and B. gilchristii (n = 2, 6%), from South Africa and Zimbabwe. CONCLUSIONS: Blastomycosis occurs throughout Africa and the Middle East and is caused predominantly by B. percursus and, at least in South Africa, B. emzantsi, resulting in distinct clinical and pathological patterns of disease.

Identification of Molds with Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry: Performance of the Newly Developed MSI-2 Application in Comparison with the Bruker Filamentous Fungi Database and MSI-1.

Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) represents a promising tool for the rapid and efficient identification of molds, but improvements are still necessary to achieve satisfactory results when identifying cryptic species. Here, we aimed to validate a new web application, MSI-2, which replaces MSI-1, an application that was built and deployed online in 2017. For the evaluation, we gathered 633 challenging isolates obtained from daily hospital practice that were first identified with DNA-based methods, and we submitted their corresponding mass spectra to three identification programs (Bruker, MSI-1, and MSI-2). The MSI-2 application had a better identification performance at the species level than MSI-1 and Bruker, reaching 83.25% correct identifications, compared with 63.19% (MSI-1), 38.07% (Bruker with a 1.7 threshold), and 21.8% (Bruker with a 2.0 threshold). The MSI-2 application performed especially well for Aspergillus and Fusarium species, including for many cryptic species, reaching 90% correct identifications for Aspergillus species and 78% for Fusarium species compared to 69% and 43% with MSI-1. Such an improvement may have a positive impact on patient management by facilitating the identification of cryptic species potentially associated with a specific antifungal resistance profile.

Cerebrospinal fluid exposure to bictegravir/emtricitabine/tenofovir in HIV-1-infected patients with CNS impairment.

OBJECTIVES: The penetration of antiretroviral drugs into deep compartments, such as the CNS, is a crucial component of strategies towards an HIV cure. This study aimed to determine CSF concentrations of bictegravir, emtricitabine and tenofovir in patients with HIV-related CNS impairment (HCI) enrolled in a real-life observational study. METHODS: Patients with HCI treated by optimized ART, including bictegravir/emtricitabine/tenofovir alafenamide (BIC/FTC/TAF) for at least 1 month were enrolled. Plasma and CSF concentrations were measured by quality control-validated assays (LC-MS/MS). The inhibitory quotient (IQARV) was calculated as the ratio of unbound (bictegravir) or total (emtricitabine and tenofovir) concentration to half (or 90%) maximal inhibitory concentration for bictegravir (or emtricitabine and tenofovir). All numerical variables are expressed as median (range). RESULTS: Twenty-four patients (nine women) were enrolled. The age was 45 (26-68) years. Unbound bictegravir and total emtricitabine and tenofovir CSF concentrations were 4.4 (1.6-9.6), 84.4 (28.6-337.4) and 1.6 (0.7-4.3) ng/mL, respectively. The unbound bictegravir CSF fraction was 34% (15%-82%) versus 0.33% (0.11%-0.92%) in plasma. Three patients had an IQARV above unity for the three antiretrovirals. Factors positively associated with the CSF concentration (unbound for bictegravir) were age and total plasma concentration for the three antiretrovirals. Patients aged over 51 years had higher CSF concentrations (unbound for bictegravir). CONCLUSIONS: We observed low CSF exposure to bictegravir, emtricitabine and tenofovir. These results suggest that BIC/FTC/TAF should be used with caution as first-line treatment for people living with HIV with HCI under 51 years of age.

Adenosine kinase deficiency: Three new cases and diagnostic value of hypermethioninemia.

Adenosine kinase (ADK) deficiency is characterized by liver disease, dysmorphic features, epilepsy and developmental delay. This defect disrupts the adenosine/AMP futile cycle and interferes with the upstream methionine cycle. We report the clinical, histological and biochemical courses of three ADK children carrying two new mutations and presenting with neonatal cholestasis and neurological disorders. One of them died of liver failure whereas the other two recovered from their liver damage. As the phenotype was consistent with a mitochondrial disorder, we studied liver mitochondrial respiratory chain activities in two patients and revealed a combined defect of several complexes. In addition, we retrospectively analyzed methionine plasma concentration, a hallmark of ADK deficiency, in a cohort of children and showed that methionine level in patients with ADK deficiency was strongly increased compared with patients with other liver diseases. ADK deficiency is a cause of neonatal or early infantile liver disease that may mimic primary mitochondrial disorders. In this context, an elevation of methionine plasma levels over twice the upper limit should not be considered as a nonspecific finding. ADK deficiency induced-liver dysfunction is most often transient, but could be life-threatening.

Characteristics of Dolutegravir and Bictegravir Plasma Protein Binding: a First Approach for the Study of Pharmacologic Sanctuaries.

This study aimed to characterize in vitro dolutegravir (DTG) and bictegravir (BIC) binding. They had a preferential binding to human serum albumin (HSA) with two classes of albumin sites. Human alpha-1-acid glycoprotein (HAAG) binding of DTG and BIC showed an atypical nonlinear binding. The low-affinity site on HSA, the main plasma binding protein, suggests that the high protein binding rate should not impair passive diffusion.

Mitochondrial dysfunction caused by novel ATAD3A mutations.

Mitochondrial respiratory chain integrity depends on a number of proteins encoded by nuclear and mitochondrial genomes. Mutations of such factors can result in isolated or combined respiratory chain deficits, some of which can induce abnormal morphology of the mitochondrial network or accumulation of intermediary metabolites. Consequently, affected patients are clinically heterogeneous, presenting with central nervous system, muscular, or neurodegenerative disorders. ATAD3A is a nuclear-encoded ATPase protein of the AAA+ family and has been localized to the inner mitochondrial membrane. Recently reported mutations or large deletions in the ATDA3A gene in patients have been shown to induce altered mitochondrial structure and function and abnormal cholesterol metabolism in a recessive or dominant manner. Here, we report two siblings presenting axonal sensory-motor neuropathy associated with neonatal cataract. Genetic analyses identified two novel mutations in ATAD3A; a point mutation and an intronic 15 bp deletion affecting splicing and leading to exon skipping. Biochemical analysis in patient cells and tissues showed abnormal function of the mitochondrial respiratory chain in muscle and abnormal mitochondrial cristae structure. These new cases underline the large spectrum of biochemical and clinical presentations of ATAD3A deficiency and the different modes of inheritance, making it an atypical mitochondrial disorder.

Superficial mycoses in Belgium: Burden, costs and antifungal drugs consumption.

BACKGROUND: Monitoring of superficial mycoses requires more attention due to their important incidence, health costs and antifungal drugs consumption. OBJECTIVES: The objectives were to estimate the burden of superficial mycoses in Belgium and to assess trends in associated antifungal consumption. METHODS: The burden of dermatophytoses (including onychomycosis), as well as skin and genital candidiasis, was estimated using disability-adjusted life years (DALY). Moreover, trends in systemic and topical antifungal consumption in ambulatory care were examined for the period 2010-2017, together with their associated costs. RESULTS: Due to their high incidence and long treatment duration, dermatophytoses represented the bulk of the burden, accounting for 92.2% of the total DALYs of superficial mycoses. Terbinafine was the most prescribed antifungal in terms of doses (35.4% of the total doses) while fluconazole was the most delivered drug in terms of packages (29.1% of the total packages). More than 70% of the prescriptions were made by general practitioners while consumption varied according to age and gender of the patients. A global 12% decrease in antifungal prescriptions was observed between 2011 and 2017. However, this reduction would result mainly from packaging changes and increased self-medication. A significant decrease in itraconazole treatments was notably compensated by an increased prescription of fluconazole packages. CONCLUSION: This study emphasises that dermatological presentations of superficial mycoses are the most important in terms of both burden and antifungal consumption in Belgium. Further reduction in antifungals use can be achieved by applying the adequate treatment after identification of the causative agent.

Resistance to Insulin in Patients with Gitelman Syndrome and a Subtle Intermediate Phenotype in Heterozygous Carriers: A Cross-Sectional Study.

BACKGROUND: Gitelman syndrome is a salt-losing tubulopathy caused by mutations in the SLC12A3 gene, which encodes the thiazide-sensitive sodium-chloride cotransporter. Previous studies suggested an intermediate phenotype for heterozygous carriers. METHODS: To evaluate the phenotype of heterozygous carriers of pathogenic SLC12A3 mutations, we performed a cross-sectional study of patients with Gitelman syndrome, heterozygous carriers, and healthy noncarriers. Participants measured their BP at home for three consecutive days before hospital admission for blood and urine sampling and an oral glucose tolerance test. RESULTS: We enrolled 242 participants, aged 18-75 years, including 81 heterozygous carriers, 82 healthy noncarriers, and 79 patients with Gitelman syndrome. The three groups had similar age, sex ratio, and body mass index. Compared with healthy noncarriers, heterozygous carriers showed significantly higher serum calcium concentration (P=0.01) and a trend for higher plasma aldosterone (P=0.06), but measures of home BP, plasma and urine electrolytes, renin, parathyroid hormone, vitamin D, and response to oral glucose tolerance testing were similar. Patients with Gitelman syndrome had lower systolic BP and higher heart rate than noncarriers and heterozygote carriers; they also had significantly higher fasting serum glucose concentration, higher levels of markers of insulin resistance, and a three-fold higher sensitivity to overweight. According to oral glucose tolerance testing, approximately 14% of patients with Gitelman syndrome were prediabetic, compared with 5% of heterozygous carriers and 4% of healthy noncarriers. CONCLUSIONS: Heterozygous carriers had a weak intermediate phenotype, between that of healthy noncarriers and patients with Gitelman syndrome. Moreover, the latter are at risk for development of type 2 diabetes, indicating the heightened importance of body weight control in these patients.

Black aspergilli: A remaining challenge in fungal taxonomy?

Aspergillus section Nigri is a taxonomically difficult but medically and economically important group. In this study, an update of the taxonomy of A. section Nigri strains within the BCCM/IHEM collection has been conducted. The identification accuracy of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) was tested and the antifungal susceptibilities of clinical isolates were evaluated. A total of 175 strains were molecularly analyzed. Three regions were amplified (ITS, benA, and caM) and a multi-locus phylogeny of the combined loci was created by using maximum likelihood analysis. The in-house MALDI-TOF MS reference database was extended and an identification data set of 135 strains was run against a reference data set. Antifungal susceptibility was tested for voriconazole, itraconazole, and amphotericin B, using the EUCAST method. Phylogenetic analysis revealed 18 species in our data set. MALDI-TOF MS was able to distinguish between A. brasiliensis, A. brunneoviolaceus, A. neoniger, A. niger, A. tubingensis, and A. welwitschiae of A. sect. Nigri. In the routine clinical lab, isolates of A. sect. Nigri are often identified as A. niger. However, in the clinical isolates of our data set, A. tubingensis (n = 35) and A. welwitschiae (n = 34) are more common than A. niger (n = 9). Decreased antifungal susceptibility to azoles was observed in clinical isolates of the /tubingensis clade. This emphasizes the importance of identification up to species level or at least up to clade level in the clinical lab. Our results indicate that MALDI-TOF MS can be a powerful tool to replace classical morphology.

Systemic antifungal drug use in Belgium-One of the biggest antifungal consumers in Europe.

BACKGROUND: Reports on the consumption of systemic antifungal drugs on a national level are scarce although of high interest to compare trends and the associated epidemiology in other countries and to assess the need for antifungal stewardship programmes. OBJECTIVES: To estimate patterns of Belgian inpatient and outpatient antifungal use and provide reference data for other countries. METHODS: Consumption records of antifungals were collected in Belgian hospitals between 2003 and 2016. Primary healthcare data were available for the azoles for the period 2010-2016. RESULTS: The majority of the antifungal consumption resulted from prescriptions of fluconazole and itraconazole in the ambulatory care while hospitals were responsible for only 6.4% of the total national consumption and echinocandin use was limited. The annual average antifungal consumption in hospitals decreased significantly by nearly 25% between 2003 and 2016, due to a decrease solely in non-university hospitals. With the exception of specialised burn centres, antifungals are mostly consumed at ICUs and internal medicine wards. A significant decline was also observed in the consumption of azoles in primary health care, attributed to itraconazole. The major part of azoles was prescribed by generalists followed by dermatologists. CONCLUSIONS: In spite of the downward trend in annual use of systemic antifungal drugs, Belgium remains one of the biggest consumers in Europe.

Clonal Spread of Candida glabrata Bloodstream Isolates and Fluconazole Resistance Affected by Prolonged Exposure: a 12-Year Single-Center Study in Belgium.

Candida glabrata is a major cause of candidemia in immunocompromised patients and is characterized by a high-level of fluconazole resistance. In the present study, the acquisition of antifungal resistance and potential clonal spread of C. glabrata were explored at a single center over a 12-year period by analyzing 187 independent clinical C. glabrata bloodstream isolates. One strain was found to be micafungin resistant due to a mutation in the FKS2 gene. Fluconazole resistance remained stable throughout the period and was observed in 20 (10.7%) of the isolates. An analysis of the antifungal consumption data revealed that recent prior exposure to fluconazole increased the risk to be infected by a resistant strain. In particular, the duration of the treatment was significantly longer for patients infected by a resistant isolate, while the total and mean daily doses received did not impact the acquisition of resistance in C. glabrata No link between genotype and resistance was found. However, multilocus variable-number tandem-repeat analyses indicated a potential intrahospital spread of some isolates between patients. These isolates shared the same genetic profiles, and infected patients were hospitalized in the same unit during an overlapping period. Finally, quantitative real-time PCR analyses showed that, unlike that for other ABC efflux pumps, the expression of CgCDR1 was significantly greater in resistant strains, suggesting that it would be more involved in fluconazole (FLC) resistance. Our study provides additional evidence that the proper administration of fluconazole is required to limit resistance and that strict hand hygiene is necessary to avoid the possible spreading of C. glabrata isolates between patients.

Correlating Schiff Moments in the Light Actinides with Octupole Moments.

We show that the measured intrinsic octupole moments of ^{220}Rn, ^{224}Ra, and ^{226}Ra constrain the intrinsic Schiff moments of ^{225}Ra, ^{221}Rn, ^{223}Rn, ^{223}Fr, ^{225}Ra, and ^{229}Pa. The result is a dramatically reduced uncertainty in intrinsic Schiff moments. Direct measurements of octupole moments in odd nuclei will reduce the uncertainty even more. The only significant source of nuclear-physics error in the laboratory Schiff moments will then be the intrinsic matrix elements of the time-reversal noninvariant interaction produced by CP-violating fundamental physics. Those matrix elements are also correlated with octupole moments, but with a larger systematic uncertainty.

Increasing candidaemia incidence from 2004 to 2015 with a shift in epidemiology in patients preexposed to antifungals.

Candidaemia is an important health problem in immunocompromised patients with an epidemiology varying with region, period and patient population involved. The occurrence of candidaemia and the associated species distribution over a 12-year period at a large tertiary care centre in Belgium were analysed. The trend in incidence in the intensive care units (ICUs) and non-ICUs was investigated as well as the influence of antifungal exposure on the species distribution. From 2004 until 2015, 865 candidaemia episodes occurred in 826 patients at the University Hospitals Leuven. Candida albicans (59%) remained the most important cause of candidaemia, followed by C. glabrata (22.4%) and C. parapsilosis (8%). The mean incidence in the whole hospital was 1.48 per 10 000 patient days (PD). The incidence in ICUs increased reaching up to 10.7 per 10 000 PD whereas in the non-ICUs, the incidence decreased. Prior exposure to fluconazole and echinocandins was associated with candidaemia caused by less susceptible species. Candidaemia incidence increased in the whole hospital, driven by ICUs. Surveillance of candidaemia epidemiology on a local scale is of high value to guide empirical treatment strategies.

Decision criteria for MALDI-TOF MS-based identification of filamentous fungi using commercial and in-house reference databases.

BACKGROUND: Several Matrix-Assisted Laser Desorption/Ionization Time-of-Flight mass spectrometry protocols, which differ in identification criteria, have been developed for mold and dermatophyte identification. Currently, the most widely used approach is Bruker technology, although no consensus concerning the log(score) threshold has been established. Furthermore, it remains unknown how far increasing the number of spots to compare results might improve identification performance. In this study, we used in-house and Bruker reference databases as well as a panel of 422 isolates belonging to 126 species to test various thresholds. Ten distinct identification algorithms requiring one to four spots were tested. RESULTS: Our findings indicate that optimal results were obtained by applying a decisional algorithm in which only the highest score of four spots was taken into account with a 1.7 log(score) threshold. Testing the entire panel enabled identification of 87.41% (in-house database) and 35.15% (Bruker database) of isolates, with a positive predictive value (PPV) of 1 at the genus level for both databases as well as 0.89 PPV (in-house database) and 0.72 PPV (Bruker database) at the species level. Applying the same rules to the isolates for which the species were represented by at least three strains in the database enabled identification of 92.1% (in-house database) and 46.6% (Bruker database) of isolates, with 1 PPV at the genus level for both databases as well as 0.95 PPV (in-house database) and 0.93 PPV (Bruker database) at the species level. CONCLUSIONS: Depositing four spots per extract and lowering the threshold to 1.7, a threshold which is notably lower than that recommended for bacterial identification, decreased the number of unidentified specimens without altering the reliability of the accepted results. Nevertheless, regardless of the criteria used for mold and dermatophyte identification, commercial databases require optimization.

Droplet digital PCR combined with minisequencing, a new approach to analyze fetal DNA from maternal blood: application to the non-invasive prenatal diagnosis of achondroplasia.

BACKGROUND: Achondroplasia is generally detected by abnormal prenatal ultrasound findings in the third trimester of pregnancy and then confirmed by molecular genetic testing of fetal genomic DNA obtained by aspiration of amniotic fluid. This invasive procedure presents a small but significant risk for both the fetus and mother. Therefore, non-invasive procedures using cell-free fetal DNA in maternal plasma have been developed for the detection of the fetal achondroplasia mutations. METHODS: To determine whether the fetus carries the de novo mis-sense genetic mutation at nucleotide 1138 in FGFR3 gene involved in >99% of achondroplasia cases, we developed two independent methods: digital-droplet PCR combined with minisequencing, which are very sensitive methods allowing detection of rare alleles. RESULTS: We collected 26 plasmatic samples from women carrying fetus at risk of achondroplasia and diagnosed to date a total of five affected fetuses in maternal blood. The sensitivity and specificity of our test are respectively 100% [95% confidence interval, 56.6-100%] and 100% [95% confidence interval, 84.5-100%]. CONCLUSIONS: This novel, original strategy for non-invasive prenatal diagnosis of achondroplasia is suitable for implementation in routine clinical testing and allows considering extending the applications of these technologies in non-invasive prenatal diagnosis of many other monogenic diseases. © 2016 John Wiley & Sons, Ltd.

Identification of filamentous fungi isolates by MALDI-TOF mass spectrometry: clinical evaluation of an extended reference spectra library.

The identification of filamentous fungi by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) relies mainly on a robust and extensive database of reference spectra. To this end, a large in-house library containing 760 strains and representing 472 species was built and evaluated on 390 clinical isolates by comparing MALDI-TOF MS with the classical identification method based on morphological observations. The use of MALDI-TOF MS resulted in the correct identification of 95.4% of the isolates at species level, without considering LogScore values. Taking into account the Brukers' cutoff value for reliability (LogScore >1.70), 85.6% of the isolates were correctly identified. For a number of isolates, microscopic identification was limited to the genus, resulting in only 61.5% of the isolates correctly identified at species level while the correctness reached 94.6% at genus level. Using this extended in-house database, MALDI-TOF MS thus appears superior to morphology in order to obtain a robust and accurate identification of filamentous fungi. A continuous extension of the library is however necessary to further improve its reliability. Indeed, 15 isolates were still not represented while an additional three isolates were not recognized, probably because of a lack of intraspecific variability of the corresponding species in the database.

Targeting mitochondrial function in macrophages: A novel treatment strategy for atherosclerotic cardiovascular disease?

Atherosclerotic cardiovascular disease is a major cause of morbidity and mortality due to chronic arterial injury caused by hyperlipidemia, hypertension, inflammation and oxidative stress. Recent studies have shown that the progression of this disease is associated with mitochondrial dysfunction and with the accumulation of mitochondrial alterations within macrophages of atherosclerotic plaques. These alterations contribute to processes of inflammation and oxidative stress. Among the many players involved, macrophages play a pivotal role in atherogenesis as they can exert both beneficial and deleterious effects due to their anti- and pro-inflammatory properties. Their atheroprotective functions, such as cholesterol efflux and efferocytosis, as well as the maintenance of their polarization towards an anti-inflammatory state, are particularly dependent on mitochondrial metabolism. Moreover, in vitro studies have demonstrated deleterious effects of oxidized LDL on macrophage mitochondrial function, resulting in a switch to a pro-inflammatory state and to a potential loss of atheroprotective capacity. Therefore, preservation of mitochondrial function is now considered a legitimate therapeutic strategy. This review focuses on the potential therapeutic strategies that could improve the mitochondrial function of macrophages, enabling them to maintain their atheroprotective capacity. These emerging therapies could play a valuable role in counteracting the progression of atherosclerotic lesions and possibly inducing their regression.

Pulmonary Aspergillosis in Humboldt Penguins-Susceptibility Patterns and Molecular Epidemiology of Clinical and Environmental Aspergillus fumigatus Isolates from a Belgian Zoo, 2017-2022.

Aspergillus fumigatus is the main causative agent of avian aspergillosis and results in significant health problems in birds, especially those living in captivity. The fungal contamination by A. fumigatus in the environment of Humboldt penguins (Spheniscus humboldti), located in a Belgian zoo, was assessed through the analysis of air, water, sand and nest samples during four non-consecutive days in 2021-2022. From these samples, potential azole-resistant A. fumigatus (ARAF) isolates were detected using a selective culture medium. A total of 28 veterinary isolates obtained after necropsy of Humboldt penguins and other avian species from the zoo were also included. All veterinary and suspected ARAF isolates from the environment were characterized for their azole-resistance profile by broth microdilution. Isolates displaying phenotypic resistance against at least one medical azole were systematically screened for mutations in the cyp51A gene. A total of 14 (13.6%) ARAF isolates were identified from the environment (n = 8) and from Humboldt penguins (n = 6). The TR34/L98H mutation was observed in all resistant environmental strains, and in two resistant veterinary strains. To the best of our knowledge, this is the first description of this mutation in A. fumigatus isolates from Humboldt penguins. During the period 2017-2022, pulmonary aspergillosis was confirmed in 51 necropsied penguins, which reflects a death rate due to aspergillosis of 68.0%, mostly affecting adults. Microsatellite polymorphism analysis revealed a high level of diversity among environmental and veterinary A. fumigatus isolates. However, a cluster was observed between one veterinary isolate and six environmental strains, all resistant to medical azoles. In conclusion, the environment of the Humboldt penguins is a potential contamination source of ARAF, making their management even more complex.