Platypus and echidna genomes reveal mammalian biology and evolution.

Egg-laying mammals (monotremes) are the only extant mammalian outgroup to therians (marsupial and eutherian animals) and provide key insights into mammalian evolution1,2. Here we generate and analyse reference genomes of the platypus (Ornithorhynchus anatinus) and echidna (Tachyglossus aculeatus), which represent the only two extant monotreme lineages. The nearly complete platypus genome assembly has anchored almost the entire genome onto chromosomes, markedly improving the genome continuity and gene annotation. Together with our echidna sequence, the genomes of the two species allow us to detect the ancestral and lineage-specific genomic changes that shape both monotreme and mammalian evolution. We provide evidence that the monotreme sex chromosome complex originated from an ancestral chromosome ring configuration. The formation of such a unique chromosome complex may have been facilitated by the unusually extensive interactions between the multi-X and multi-Y chromosomes that are shared by the autosomal homologues in humans. Further comparative genomic analyses unravel marked differences between monotremes and therians in haptoglobin genes, lactation genes and chemosensory receptor genes for smell and taste that underlie the ecological adaptation of monotremes.

Aspergillus fumigatus sensu stricto genetic diversity from cystic fibrosis patients.

We aimed to access the genetic diversity of Apergillus fumigatus strains obtained from cystic fibrosis (CF) patients from southern Brazil. A. fumigatus sensu stricto isolates from respiratory clinical specimens were genotyped by microsatellite markers and azole resistance was evaluated by azole-agar screening. Twenty-seven isolates from twenty-seven patients showed a high genetic diversity, with the differentiation of 25 different genotypes (25 unique and one common to two isolates). All isolates were susceptible to the azoles tested. We believe that prospectively monitoring A. fumigatus genetic diversity is essential to identify interpatient transmission and outbreaks, as is the identification of resistant strains.

Humane Endpoint: Example from a Murine Model of Disseminated Sporotrichosis.

Vertebrate animal models are essential in research; however, efforts need to be made to decrease animal suffering as much as possible. It could be useful to determine humane endpoints that could serve as surrogates for a fatal outcome. We address this issue with respect to infectious diseases. We propose a humane endpoint for studies of Sporothrix brasiliensis infection. BALB/c mice were inoculated subcutaneously in the footpad. To define a humane endpoint, we considered two groups: animals who died during the experiment, and those euthanized at the end of the experiment. The groups were compared for colony-forming units (CFU) in internal organs, clinical condition, and body weight. Thirteen (22%) animals died before the end of the experiment due to the progression of local infection to severe and disseminated sporotrichosis. Analyzing data of the groups, we propose the following future criteria for euthanasia as a humane endpoint: neurological impairment OR necrosis of the footpad OR loss of ≥ 20% body weight AND moderate to severe dehydration. In view of the current global epidemiological impact of zoonotic sporotrichosis caused by S. brasiliensis, our data could facilitate the utility of models used to study the disease, particularly therapy.

Aspergillosis in Critically Ill Patients with and Without COVID-19 in a Tertiary Hospital in Southern Brazil.

The impact of invasive pulmonary aspergillosis (IPA) on non-neutropenic critically ill patients in intensive care units (ICU) has been demonstrated in recent decades. Furthermore, after the start of the COVID-19 pandemic, COVID-19 associated with pulmonary aspergillosis (CAPA) has become a major concern in ICUs. However, epidemiological data from different regions are scarce. We evaluated the prevalence and clinical-epidemiological data of IPA in patients with COVID-19 requiring mechanical ventilation (MV) in the ICU ("severe COVID-19") and non-COVID ICU patients in MV of a tertiary hospital in the southern region of Brazil. Eighty-seven patients admitted between June 2020 and August 2022 were included; 31 with severe COVID-19. For the diagnosis of IPA or CAPA, algorithms including host factors and mycological criteria (positive culture for Aspergillus spp., immunoassay for galactomannan detection, and/or qPCR) were utilized. The overall incidence of IPA and CAPA in our ICU was 73 cases/1000 ICU hospitalizations. Aspergillosis occurred in 13% (4/31) of the COVID-19 patients, and in 16% (9/56) of the critically ill patients without COVID-19, with mortality rates of 75% (3/4) and 67% (6/9), respectively. Our results highlight the need for physicians enrolled in ICU care to be aware of aspergillosis and for more access of the patients to sensitive and robust diagnostic tests by biomarkers detection.

Certolizumab pegol, abatacept, tocilizumab or active conventional treatment in early rheumatoid arthritis: 48-week clinical and radiographic results of the investigator-initiated randomised controlled NORD-STAR trial.

BACKGROUND: The optimal first-line treatment in early rheumatoid arthritis (RA) is debated. We compared clinical and radiographic outcomes of active conventional therapy with each of three biological treatments with different modes of action. METHODS: Investigator-initiated, randomised, blinded-assessor study. Patients with treatment-naïve early RA with moderate-severe disease activity were randomised 1:1:1:1 to methotrexate combined with (1) active conventional therapy: oral prednisolone (tapered quickly, discontinued at week 36) or sulfasalazine, hydroxychloroquine and intra-articular glucocorticoid injections in swollen joints; (2) certolizumab pegol; (3) abatacept or (4) tocilizumab. Coprimary endpoints were week 48 Clinical Disease Activity Index (CDAI) remission (CDAI ≤2.8) and change in radiographic van der Heijde-modified Sharp Score, estimated using logistic regression and analysis of covariance, adjusted for sex, anticitrullinated protein antibody status and country. Bonferroni's and Dunnet's procedures adjusted for multiple testing (significance level: 0.025). RESULTS: Eight hundred and twelve patients were randomised. Adjusted CDAI remission rates at week 48 were: 59.3% (abatacept), 52.3% (certolizumab), 51.9% (tocilizumab) and 39.2% (active conventional therapy). Compared with active conventional therapy, CDAI remission rates were significantly higher for abatacept (adjusted difference +20.1%, p<0.001) and certolizumab (+13.1%, p=0.021), but not for tocilizumab (+12.7%, p=0.030). Key secondary clinical outcomes were consistently better in biological groups. Radiographic progression was low, without group differences.The proportions of patients with serious adverse events were abatacept, 8.3%; certolizumab, 12.4%; tocilizumab, 9.2%; and active conventional therapy, 10.7%. CONCLUSIONS: Compared with active conventional therapy, clinical remission rates were superior for abatacept and certolizumab pegol, but not for tocilizumab. Radiographic progression was low and similar between treatments. TRIAL REGISTRATION NUMBER: NCT01491815.

Associations of baseline obstructive sleep apnea and sleep macroarchitecture with cognitive function after 8 years in middle-aged and older men from a community-based cohort study.

Previous prospective studies examining associations of obstructive sleep apnea and sleep macroarchitecture with future cognitive function recruited older participants, many demonstrating baseline cognitive impairment. This study examined obstructive sleep apnea and sleep macroarchitecture predictors of visual attention, processing speed, and executive function after 8 years among younger community-dwelling men. Florey Adelaide Male Ageing Study participants (n = 477) underwent home-based polysomnography, with 157 completing Trail-Making Tests A and B and the Mini-Mental State Examination. Associations of obstructive sleep apnea (apnea-hypopnea index, oxygen desaturation index, and hypoxic burden index) and sleep macroarchitecture (sleep stage percentages and total sleep time) parameters with future cognitive function were examined using regression models adjusted for baseline demographic, biomedical, and behavioural factors, and cognitive task performance. The mean (standard deviation) age of the men at baseline was 58.9 (8.9) years, with severe obstructive sleep apnea (apnea-hypopnea index ≥30 events/h) in 9.6%. The median (interquartile range) follow-up was 8.3 (7.9-8.6) years. A minority of men (14.6%) were cognitively impaired at baseline (Mini-Mental State Examination score <28/30). A higher percentage of light sleep was associated with better Trail-Making Test A performance (B = -0.04, 95% confidence interval [CI] -0.06, -0.01; p = 0.003), whereas higher mean oxygen saturation was associated with worse performance (B = 0.11, 95% CI 0.02, 0.19; p = 0.012). While obstructive sleep apnea and sleep macroarchitecture might predict cognitive decline, future studies should consider arousal events and non-routine hypoxaemia measures, which may show associations with cognitive decline.

Effectiveness of diphenyl diselenide against experimental sporotrichosis caused by Sporothrix brasiliensis.

Diphenyl diselenide (PhSe)2 is a stable organoselenium compound with promising in vitro antifungal activity against several fungi, including Sporothrix brasiliensis. This species is associated with feline and zoonotic sporotrichosis, an emergent mycosis in Latin America. We evaluated the activity of (PhSe)2, alone and in association with itraconazole, in the treatment of sporotrichosis caused by S. brasiliensis, in a murine model. Sixty mice were subcutaneously infected with S. brasiliensis in the footpad and treated by gavage for 30 consecutive days. The six treatment groups received: no active treatment, itraconazole (50 mg/kg), (PhSe)2 at 1, 5, and 10 mg/kg dosages, or itraconazole (50 mg/kg) + (PhSe)2 1 mg/kg, once a day, starting seven days post-inoculation. A significant reduction in the fungal burden of internal organs was achieved in the groups treated with (PhSe)2 1 mg/kg or itraconazole alone in comparison with the untreated group. Higher dosages (5 and 10 mg/kg) of (PhSe)2 increased the clinical manifestation of sporotrichosis and mortality rate. Treatment with both itraconazole and (PhSe)2 1 mg/kg was better than their activities alone (P < .001). This is the first demonstration of the potential use of (PhSe)2, alone or with the present drug of choice, in the treatment of sporotrichosis.

We evaluated the activity of diphenyl diselenide (PhSe)2, alone and in association with itraconazole, in the treatment of sporotrichosis caused by S. brasiliensis, in a murine model. This is the first demonstration of the potential use of (PhSe)2, alone or in an association against sporotrichosis.

Aspergillosis in free-ranging Magellanic penguins.

We evaluated the mortality due to aspergillosis in free-ranging Magellanic penguins during their migration and the reproductive season. A total of 98 carcasses of penguins were collected along 370 km of coastline in Southern Brazil, between June 2017 and October 2019, and from reproductive colonies in Patagonian Argentina, in January 2019. All animals were necropsied, and only proven cases were computed. Aspergillosis was diagnosed in 2.5% of the penguins evaluated during their migration route. Our study, of the Southern coast of Brazil, is the first to demonstrate that aspergillosis is an important cause of mortality in free-ranging penguins. The implications of these findings in the One Health context are discussed.

We evaluated the mortality due to aspergillosis in free-ranging Magellanic penguins during their migration and the reproductive season. The mortality rate of penguins was 2.5% during their migration route. Our study is the first to demonstrate aspergillosis as an important cause of mortality in free-ranging penguins.

Continuous dosing of nikkomycin Z against systemic candidiasis, in vivo and in vitro correlates.

BACKGROUND: Candidiasis is the most common cause of fungal sepsis, and new agents are of interest to ameliorate current deficiencies in therapy. Nikkomycin Z (NIKZ) is an inhibitor of chitin synthase, interfering with fungal cell wall development. OBJECTIVES/METHODS: We studied NIKZ therapy of disseminated murine candidiasis, via continuous drug exposure, in drinking water, to compensate for rapid clearance of the drug. RESULTS: Drinking, and thus drug intake in the NIKZ groups, as well as body weight, was affected by the degree of illness. NIKZ effect on survival, despite reduced drinking initially after infection, was highly efficacious and dose-related, and comparable to fluconazole, though neither were curative with the regimens employed. The challenge was rapidly lethal to all untreated animals, whereas NIKZ groups achieved >50% survival. Assays of residual fungal infection were consistent with impressions of efficacy based on survival. Although NIKZ MIC for Candida albicans appeared unpromising, mycelial formation assays more closely correlated with in vivo observations. CONCLUSIONS: In vitro-in vivo disparity may be explained by NIKZ tissue concentration in the target tissue and/or by enhanced NIKZ action on mycelial formation, a morphological change in vivo wherein chitin synthesis is more critical, compared to NIKZ activity in inhibiting planktonic growth. A sustained release oral form of NIKZ in drug development for humans could hold promise, possibly also in future exploring previously demonstrated synergy in vitro with other antifungals.

Nikkomycin Z for the treatment of experimental sporotrichosis caused by Sporothrix brasiliensis.

Sporotrichosis caused by Sporothrix brasiliensis is a global emergent infectious disease. Due to the scarcity of therapeutic options for fungal diseases, new antifungals are urgently needed. Nikkomycin Z (NikZ) is a future option as an agent against dimorphic fungi. We evaluated NikZ monotherapy and in combination with itraconazole (ITZ; the conventional therapy) in the treatment of experimental sporotrichosis caused by S. brasiliensis in a murine model. Animals were subcutaneously infected, and treated orally for 30 days. The study groups were as follows control (untreated), ITZ group (50 mg/kg/day), and three groups treated with NikZ, two by monotherapy (200 or 400 mg/kg/day), and one combining NikZ (400 mg/kg/day) and ITZ. Efficacy of treatments was evaluated via body weight gain, mortality and fungal burden in tissues. Efficacy was noted in all treatment groups, and the group receiving the drug combination showed even better results than those with monotherapy. Our study shows for the first time the high potential of NikZ to be used in the treatment of sporotrichosis caused by S. brasiliensis.

Exploiting evolutionary trade-offs for posttreatment management of drug-resistant populations.

Antibiotic resistance frequently evolves through fitness trade-offs in which the genetic alterations that confer resistance to a drug can also cause growth defects in resistant cells. Here, through experimental evolution in a microfluidics-based turbidostat, we demonstrate that antibiotic-resistant cells can be efficiently inhibited by amplifying the fitness costs associated with drug-resistance evolution. Using tavaborole-resistant Escherichia coli as a model, we show that genetic mutations in leucyl-tRNA synthetase (that underlie tavaborole resistance) make resistant cells intolerant to norvaline, a chemical analog of leucine that is mistakenly used by tavaborole-resistant cells for protein synthesis. We then show that tavaborole-sensitive cells quickly outcompete tavaborole-resistant cells in the presence of norvaline due to the amplified cost of the molecular defect of tavaborole resistance. This finding illustrates that understanding molecular mechanisms of drug resistance allows us to effectively amplify even small evolutionary vulnerabilities of resistant cells to potentially enhance or enable adaptive therapies by accelerating posttreatment competition between resistant and susceptible cells.

Synergy In Vitro of Nikkomycin Z with Azole Against the Invasive Form of Candida albicans.

In a previous study, therapeutic activity of nikkomycin Z (NZ) in a model of invasive candidiasis did not appear to correlate with lesser activity in vitro (using classical MIC methods) with planktonic organisms. However, NZ potency was much greater assaying activity in vitro against germ tubes, the initiator of the invasive mycelial form of the fungus, as occurs in infected tissues. Synergy has been demonstrated for NZ and other drugs, notably fluconazole (the most commonly used drug against candidiasis), in planktonic testing, which correlated with results in vivo. This raised the question whether activity shown by NZ alone against germ tubes would be reflected in drug combinations, and even whether synergy testing against germ tubes might be a better correlate of synergy in future in vivo studies. We show in this study significant NZ synergy with fluconazole against germ tubes, for several C. albicans isolates, with testing in many drug ratios. This observation opens the way for further explorations of this method of susceptibility testing for synergy, and correlation with combination therapy against candidiasis.

Controversies in the Management of Central Nervous System Coccidioidomycosis.

Central nervous system infection with Coccidioides spp. is fatal if untreated and complications occur even when therapy is directed by experienced clinicians. We convened a panel of clinicians experienced in the management of coccidioidal meningitis to summarize current controversies and provide consensus for the management of this difficult infection.

FDA Public Workshop Summary-Coccidioidomycosis (Valley Fever): Considerations for Development of Antifungal Drugs.

Coccidioidomycosis is a fungal disease endemic to the southwestern United States, Mexico, and Central and South America. Prevalence rates are increasing steadily, and new endemic areas of Coccidioides are emerging. Standard treatment is often administered for months to decades, and intolerance to medications and treatment failures are common. No new treatments for coccidioidomycosis have been approved in the United States in nearly 40 years. On 5 August 2020, the US Food and Drug Administration convened experts in coccidioidomycosis from academia, industry, patient groups, and other government agencies to discuss the disease landscape and strategies to facilitate product development for treatment of coccidioidomycosis. This article summarizes the key topics concerning drug development for coccidioidomycosis presented by speakers and panelists during the workshop, such as unmet need, trial designs, endpoints, incentives, research and development support, and collaborations to facilitate antifungal drug development.

Disseminated Histoplasmosis in Persons with HIV/AIDS, Southern Brazil, 2010-2019.

We evaluated disseminated histoplasmosis (DH) in HIV patients over 10 years in southern Brazil. The incidence was 12 cases/1,000 hospitalizations (2010-2019); the mortality rate was 35%. Tuberculosis frequently obscured the diagnosis of DH. We emphasize the need in our region to suspect and investigate DH using more sensitive methods.

Association between hypertensive disorders of pregnancy and later risk of cardiovascular outcomes.

BACKGROUND: Hypertensive disorders of pregnancy are common pregnancy complications that are associated with greater cardiovascular disease risk for mothers. However, risk of cardiovascular disease subtypes associated with gestational hypertension or pre-eclampsia is unclear. The present study aims to compare the risk of cardiovascular disease outcomes for women with and without a history of gestational hypertension and pre-eclampsia using national hospital admissions data. METHODS: This was a retrospective cohort study of national medical records from all National Health Service hospitals in England. Women who had one or more singleton live births in England between 1997 and 2015 were included in the analysis. Risk of total cardiovascular disease and 19 pre-specified cardiovascular disease subtypes, including stroke, coronary heart disease, cardiomyopathy and peripheral arterial disease, was calculated separately for women with a history of gestational hypertension and pre-eclampsia compared to normotensive pregnancies. RESULTS: Amongst 2,359,386 first live births, there were 85,277 and 74,542 hospital admissions with a diagnosis of gestational hypertension and pre-eclampsia, respectively. During 18 years (16,309,386 person-years) of follow-up, the number and incidence of total CVD for normotensive women, women with prior gestational hypertension and women with prior pre-eclampsia were n = 8668, 57.1 (95% CI: 55.9-58.3) per 100,000 person-years; n = 521, 85.8 (78.6-93.5) per 100,000 person-years; and n = 518, 99.3 (90.9-108.2) per 100,000 person-years, respectively. Adjusted HRs (aHR) for total CVD were aHR (95% CI) = 1.45 (1.33-1.59) for women with prior gestational hypertension and aHR = 1.62 (1.48-1.78) for women with prior pre-eclampsia. Gestational hypertension was strongly associated with dilated cardiomyopathy, aHR = 2.85 (1.67-4.86), and unstable angina, aHR = 1.92 (1.33-2.77). Pre-eclampsia was strongly associated with hypertrophic cardiomyopathy, aHR = 3.27 (1.49-7.19), and acute myocardial infarction, aHR = 2.46 (1.72-3.53). Associations were broadly homogenous across cardiovascular disease subtypes and increased with a greater number of affected pregnancies. CONCLUSIONS: Women with either previous gestational hypertension or pre-eclampsia are at greater risk of a range of cardiovascular outcomes. These women may benefit from clinical risk assessment or early interventions to mitigate their greater risk of various cardiovascular outcomes.

Anti-Aspergillus fumigatus IgG in patients with bronchiectasis and its relationship with clinical outcome.

Aspergillosis is a mycosis, most commonly affecting the airways. This mycosis can worsen the clinical condition of patients with concurrent lung diseases. We assayed for the presence of serum anti-A. fumigatus IgG in bronchiectasis patients from a tertiary hospital in south Brazil and evaluated the relationship with clinical outcome. Thirty-one patients with bronchiectasis, without cystic fibrosis, were included. Clinical and epidemiological data were collected from all participants. Positive serological tests were detected in 13% (4/31) of the patients. The mortality rate for the year following the assay was, in the seropositive group, 75% (3/4), whereas in the seronegative group, 15% (4/27). An illustrative case is also shown and discussed. Our study highlights the diagnostic challenge and the possible impact of Aspergillus infection on these patients, indicating the necessity of more and larger investigations in the field.

Brain mitochondrial dysfunction and driving simulator performance in untreated obstructive sleep apnea.

It is challenging to determine which patients with obstructive sleep apnea (OSA) have impaired driving ability. Vulnerability to this neurobehavioral impairment may be explained by lower brain metabolites levels involved in mitochondrial metabolism. This study compared markers of brain energy metabolism in OSA patients identified as vulnerable vs resistant to driving impairment following extended wakefulness. 44 patients with moderate-severe OSA underwent 28hr extended wakefulness with three 90min driving simulation assessments. Using a two-step cluster analysis, objective driving data (steering deviation and crashes) from the 2nd driving assessment (22.5 h awake) was used to categorise patients into vulnerable (poor driving, n = 21) or resistant groups (good driving, n = 23). 1 H magnetic resonance spectra were acquired at baseline using two scan sequences (short echo PRESS and longer echo-time asymmetric PRESS), focusing on key metabolites, creatine, glutamate, N-acetylaspartate (NAA) in the hippocampus, anterior cingulate cortex and left orbito-frontal cortex. Based on cluster analysis, the vulnerable group had impaired driving performance compared with the resistant group and had lower levels of creatine (PRESS p = ns, APRESS p = 0.039), glutamate, (PRESS p < 0.01, APRESS p < 0.01), NAA (PRESS p = 0.038, APRESS p = 0.035) exclusively in the left orbito-frontal cortex. Adjusted analysis, higher glutamate was associated with a 21% (PRESS) and 36% (APRESS) reduced risk of vulnerable classification. Brain mitochondrial bioenergetics in the frontal brain regions are impaired in OSA patients who are vulnerable to driving impairment following sleep loss. These findings provide a potential way to identify at risk OSA phenotype when assessing fitness to drive, but this requires confirmation in larger future studies.

Update of the Epidemiology of the Sporotrichosis Epidemic in the state of Rio Grande do Sul, Brazil.

BACKGROUND: Sporotrichosis is a disease not requiring jurisdictional notification and consequently is underreported in Brazil. Therefore, the epidemiological picture even in hyperendemic states is unknown. Thus, we evaluated the occurrence of sporotrichosis throughout the territory of the southern state of Brazil, Rio Grande do Sul (RS). METHODS: We update the epidemiological situation of sporotrichosis in the southern region of this state and describe the emergence of this disease in the Metropolitan region. We engaged professionals from RS enrolled in animal health care in answering a questionnaire regarding sporotrichosis. RESULTS: The occurrence of local cases of feline sporotrichosis was reported by 83% of the participants from 40 cities, distributed through the seven health districts of RS. Human sporotrichosis cases, transmitted by cats, were also reported by professionals from four regions of the state. The frequency of the disease in both the South and Metropolitan regions showed a marked increase in recent years. CONCLUSION: Feline and cat-transmitted human sporotrichosis is an underreported mycosis in RS, widely distributed in the territory of this state and increasing. Aggressive public health policies are urgently necessary to control the geographical expansion of this spreading mycosis.

CATastrophe: Response to the challenges of zoonotic sporotrichosis in southern Brazil.

BACKGROUND: In Brazil, zoonotic sporotrichosis became a national public health problem, with thousands of cases in the last decade in several regions of the country. In this context, health education activities are critical, especially in promoting early diagnosis and access to proper health care in sporotrichosis hyperendemic areas. Therefore, we report the implementation of a public specialised reference service (SRS) for diagnosis and treatment of sporotrichosis in southern Brazil. We evaluated the impact of the SRS on diagnostic confirmation and speed of diagnosis. METHODS: The SRS was implemented in Rio Grande City. We implemented a public service to promote the correct diagnosis, treatment and follow-up of human sporotrichosis cases. To study the impact of implementing SRS, the annual number of cases and the period between the appearance of lesions until diagnosis were compared, using prior data and that post-implementation. RESULTS: The implementation of the SRS directly benefited almost 50 patients in only four years, with the collaboration of almost 50% of the local health groups, together with an increase of more than 200% in diagnosis confirmation and speed of diagnosis, showing the reach, impact and importance of the SRS. CONCLUSION: The impact on the individual and collective health of the local population highlights the value of this public health approach in facing the epidemiological threat of zoonotic sporotrichosis.