Frequent detection of monkeypox virus DNA in saliva, semen, and other clinical samples from 12 patients, Barcelona, Spain, May to June 2022.

A monkeypox (MPX) outbreak has expanded worldwide since May 2022. We tested 147 clinical samples collected at different time points from 12 patients by real-time PCR. MPX DNA was detected in saliva from all cases, sometimes with high viral loads. Other samples were frequently positive: rectal swab (11/12 cases), nasopharyngeal swab (10/12 cases), semen (7/9 cases), urine (9/12 cases) and faeces (8/12 cases). These results improve knowledge on virus shedding and the possible role of bodily fluids in disease transmission.

Minimally Invasive Tissue Sampling: A Tool to Guide Efforts to Reduce AIDS-Related Mortality in Resource-Limited Settings.

BACKGROUND: Available information on the causes of death among people living with human immunodeficiency virus (PLHIV) in low- and middle-income countries (LMICs) remains scarce. We aimed to provide data on causes of death in PLHIV from two LMICs, Brazil and Mozambique, to assess the impact of clinical misdiagnosis on mortality rates and to evaluate the accuracy of minimally invasive tissue sampling (MITS) in determining the cause of death in PLHIV. METHODS: We performed coupled MITS and complete autopsy on 164 deceased PLHIV (18 children, 36 maternal deaths, and 110 adults). HIV antibody levels and HIV RNA viral loads were determined from postmortem serum samples. RESULTS: Tuberculosis (22.7%), toxoplasmosis (13.9%), bacterial infections (13.9%), and cryptococcosis (10.9%) were the leading causes of death in adults. In maternal deaths, tuberculosis (13.9%), bacterial infections (13.9%), cryptococcosis (11.1%), and cerebral malaria (8.3%) were the most frequent infections, whereas viral infections, particularly cytomegalovirus (38.9%), bacterial infections (27.8%), pneumocystosis (11.1%), and HIV-associated malignant neoplasms (11.1%) were the leading cause among children. Agreement between the MITS and the complete autopsy was 100% in children, 91% in adults, and 78% in maternal deaths. The MITS correctly identified the microorganism causing death in 89% of cases. CONCLUSIONS: Postmortem studies provide highly granular data on the causes of death in PLHIV. The inaccuracy of clinical diagnosis may play a significant role in the high mortality rates observed among PLHIV in LMICs. MITS might be helpful in monitoring the causes of death in PLHIV and in highlighting the gaps in the management of the infections.

Minimally Invasive Tissue Sampling Findings in 12 Patients With Coronavirus Disease 2019.

BACKGROUND: Minimally invasive tissue sampling (MITS), a postmortem procedure that uses core needle biopsy samples and does not require opening the body, may be a valid alternative to complete autopsy (CA) in highly infectious diseases such as coronavirus disease-19 (COVID-19). This study aimed to (1) compare the performance of MITS and CA in a series of COVID-19 deaths and (2) evaluate the safety of the procedure. METHODS: From October 2020 to February 2021, MITS was conducted in 12 adults who tested positive before death for COVID-19, in a standard, well-ventilated autopsy room, where personnel used reinforced personal protective equipment. In 9 cases, a CA was performed after MITS. A thorough histological evaluation was conducted, and the presence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was evaluated by real-time reverse-transcription polymerase chain reaction (RT-PCR) and immunohistochemistry. RESULTS: The diagnoses provided by MITS and CA matched almost perfectly. In 9 patients, COVID-19 was in the chain of events leading to death, being responsible for diffuse alveolar damage and mononuclear T-cell inflammatory response in the lungs. No specific COVID-19 features were identified. Three deaths were not related to COVID-19. All personnel involved in MITS repeatedly tested negative for COVID-19. SARS-CoV-2 was identified by RT-PCR and immunohistochemistry in the MITS samples, particularly in the lungs. CONCLUSIONS: MITS is useful for evaluating COVID-19-related deaths in settings where a CA is not feasible. The results of this simplified and safer technique are comparable to those of CA.

Unmasking the hidden tuberculosis mortality burden in a large post mortem study in Maputo Central Hospital, Mozambique.

Sensitive tools are needed to accurately establish the diagnosis of tuberculosis (TB) at death, especially in low-income countries. The objective of this study was to evaluate the burden of TB in a series of patients who died in a tertiary referral hospital in sub-Saharan Africa using an in-house real time PCR (TB-PCR) and the Xpert MTB/RIF Ultra (Xpert Ultra) assay.Complete diagnostic autopsies were performed in a series of 223 deaths (56.5% being HIV-positive), including 54 children, 57 maternal deaths and 112 other adults occurring at the Maputo Central Hospital, Mozambique. TB-PCR was performed in all lung, cerebrospinal fluid and central nervous system samples in HIV-positive patients. All samples positive for TB-PCR or showing histological findings suggestive of TB were analysed with the Xpert Ultra assay.TB was identified as the cause of death in 31 patients: three out of 54 (6%) children, five out of 57 (9%)maternal deaths and 23 out of 112 (21%) other adults. The sensitivity of the main clinical diagnosis to detect TB as the cause of death was 19.4% (95% CI 7.5-37.5) and the specificity was 97.4% (94.0-99.1) compared to autopsy findings. Concomitant TB (TB disease in a patient dying of other causes) was found in 31 additional cases. Xpert Ultra helped to identify 15 cases of concomitant TB. In 18 patients, Mycobacterium tuberculosis DNA was identified by TB-PCR and Xpert Ultra in the absence of histological TB lesions. Overall, 62 (27.8%) cases had TB disease at death and 80 (35.9%) had TB findings.The use of highly sensitive, easy to perform molecular tests in complete diagnostic autopsies may contribute to identifying TB cases at death that would have otherwise been missed. Routine use of these tools in certain diagnostic algorithms for hospitalised patients needs to be considered. Clinical diagnosis showed poor sensitivity for the diagnosis of TB at death.

Validity of a minimally invasive autopsy for cause of death determination in maternal deaths in Mozambique: An observational study.

BACKGROUND: Despite global health efforts to reduce maternal mortality, rates continue to be unacceptably high in large parts of the world. Feasible, acceptable, and accurate postmortem sampling methods could provide the necessary evidence to improve the understanding of the real causes of maternal mortality, guiding the design of interventions to reduce this burden. METHODS AND FINDINGS: The validity of a minimally invasive autopsy (MIA) method in determining the cause of death was assessed in an observational study in 57 maternal deaths by comparing the results of the MIA with those of the gold standard (complete diagnostic autopsy [CDA], which includes any available clinical information). Concordance between the MIA and the gold standard diagnostic categories was assessed by the kappa statistic, and the sensitivity, specificity, positive and negative predictive values and their 95% confidence intervals (95% CI) to identify the categories of diagnoses were estimated. The main limitation of the study is that both the MIA and the CDA include some degree of subjective interpretation in the attribution of cause of death. A cause of death was identified in the CDA in 98% (56/57) of cases, with indirect obstetric conditions accounting for 32 (56%) deaths and direct obstetric complications for 24 (42%) deaths. Nonobstetric infectious diseases (22/32, 69%) and obstetric hemorrhage (13/24, 54%) were the most common causes of death among indirect and direct obstetric conditions, respectively. Thirty-six (63%) women were HIV positive, and HIV-related conditions accounted for 16 (28%) of all deaths. Cerebral malaria caused 4 (7%) deaths. The MIA identified a cause of death in 86% of women. The overall concordance of the MIA with the CDA was moderate (kappa = 0.48, 95% CI: 0.31-0.66). Both methods agreed in 68% of the diagnostic categories and the agreement was higher for indirect (91%) than for direct obstetric causes (38%). All HIV infections and cerebral malaria cases were identified in the MIA. The main limitation of the technique is its relatively low performance for identifying obstetric causes of death in the absence of clinical information. CONCLUSIONS: The MIA procedure could be a valuable tool to determine the causes of maternal death, especially for indirect obstetric conditions, most of which are infectious diseases. The information provided by the MIA could help to prioritize interventions to reduce maternal mortality and to monitor progress towards achieving global health targets.

Validity of a minimally invasive autopsy for cause of death determination in stillborn babies and neonates in Mozambique: An observational study.

BACKGROUND: Over 5 million stillbirths and neonatal deaths occur annually. Limited and imprecise information on the cause of these deaths hampers progress in achieving global health targets. Complete diagnostic autopsies (CDAs)-the gold standard for cause of death determination-are difficult to perform in most high-burden settings. Therefore, validation of simpler and more feasible methods is needed. METHODS AND FINDINGS: In this observational study, the validity of a minimally invasive autopsy (MIA) method in determining the cause of death was assessed in 18 stillbirths and 41 neonatal deaths by comparing the results of the MIA with those of the CDA. Concordance between the categories of diseases obtained by the 2 methods was assessed by the Kappa statistic, and the sensitivity, specificity, positive, and negative predictive values of the MIA diagnoses were calculated. A cause of death was identified in 16/18 (89%) and 15/18 (83%) stillborn babies in the CDA and the MIA, respectively. Fetal growth restriction accounted for 39%, infectious diseases for 22%, intrapartum hypoxia for 17%, and intrauterine hypoxia for 11% of stillborn babies. Overall, the MIA showed in this group a substantial concordance with the CDA (Kappa = 0.78, 95% CI [0.56-0.99]). A cause of death was identified in all (100%) and 35/41 (85%) neonatal deaths in the CDA and the MIA, respectively. In this group, the majority of deaths were due to infectious diseases (66%). The overall concordance of the MIA with the CDA in neonates was moderate (Kappa = 0.40, 95% CI [0.18-0.63]). A high percentage of accuracy was observed for the MIA in all the diagnostic categories in both stillbirths and neonates (>75%). The main limitation of this study is that some degree of subjective interpretation is inherent to cause-of-death attribution in both the MIA and the CDA; this is especially so in stillbirths and in relation to fetal growth restriction. CONCLUSIONS: The MIA could be a useful tool for cause-of-death determination in stillbirths and neonatal deaths. These findings may help to accelerate progress towards meeting global health targets by obtaining more accurate information on the causes of death in these age groups, which is essential in guiding the design of new interventions and increasing the effectiveness of those already implemented.

Validity of a minimally invasive autopsy tool for cause of death determination in pediatric deaths in Mozambique: An observational study.

BACKGROUND: In recent decades, the world has witnessed unprecedented progress in child survival. However, our knowledge of what is killing nearly 6 million children annually in low- and middle-income countries remains poor, partly because of the inadequacy and reduced precision of the methods currently utilized in these settings to investigate causes of death (CoDs). The study objective was to validate the use of a minimally invasive autopsy (MIA) approach as an adequate and more acceptable substitute for the complete diagnostic autopsy (CDA) for pediatric CoD investigation in a poor setting. METHODS AND FINDINGS: In this observational study, the validity of the MIA approach in determining the CoD was assessed in 54 post-neonatal pediatric deaths (age range: ≥1 mo to 15 y) in a referral hospital of Mozambique by comparing the results of the MIA with those of the CDA. Concordance in the category of disease obtained by the two methods was evaluated by the Kappa statistic, and the sensitivity, specificity, and positive and negative predictive values of the MIA diagnoses were calculated. A CoD was identified in all cases in the CDA and in 52/54 (96%) of the cases in the MIA, with infections and malignant tumors accounting for the majority of diagnoses. The MIA categorization of disease showed a substantial concordance with the CDA categorization (Kappa = 0.70, 95% CI 0.49-0.92), and sensitivity, specificity, and overall accuracy were high. The ICD-10 diagnoses were coincident in up to 75% (36/48) of the cases. The MIA allowed the identification of the specific pathogen deemed responsible for the death in two-thirds (21/32; 66%) of all deaths of infectious origin. Discrepancies between the MIA and the CDA in individual diagnoses could be minimized with the addition of some basic clinical information such as those ascertainable through a verbal autopsy or clinical record. The main limitation of the analysis is that both the MIA and the CDA include some degree of expert subjective interpretation. CONCLUSIONS: The MIA showed substantial concordance with CDA for CoD identification in this series of pediatric deaths in Mozambique. This minimally invasive approach, simpler and more readily acceptable than the more invasive CDA, could provide robust data for CoD surveillance, especially in resource-limited settings, which could be helpful for guiding child survival strategies in the future.

Controlled human malaria infection by intramuscular and direct venous inoculation of cryopreserved Plasmodium falciparum sporozoites in malaria-naïve volunteers: effect of injection volume and dose on infectivity rates.

BACKGROUND: Controlled human malaria infection (CHMI) by mosquito bite is a powerful tool for evaluation of vaccines and drugs against Plasmodium falciparum malaria. However, only a small number of research centres have the facilities required to perform such studies. CHMI by needle and syringe could help to accelerate the development of anti-malaria interventions by enabling centres worldwide to employ CHMI. METHODS: An open-label CHMI study was performed with aseptic, purified, cryopreserved P. falciparum sporozoites (PfSPZ Challenge) in 36 malaria naïve volunteers. In part A, the effect of the inoculation volume was assessed: 18 participants were injected intramuscularly (IM) with a dose of 2,500 PfSPZ divided into two injections of 10 µL (n = 6), 50 µL (n = 6) or 250 µL (n = 6), respectively. In part B, the injection volume that resulted in highest infectivity rates in part A (10 µL) was used to formulate IM doses of 25,000 PfSPZ (n = 6) and 75,000 PfSPZ (n = 6) divided into two 10-µL injections. Results from a parallel trial led to the decision to add a positive control group (n = 6), each volunteer receiving 3,200 PfSPZ in a single 500-µL injection by direct venous inoculation (DVI). RESULTS: Four/six participants in the 10-µL group, 1/6 in the 50-µL group and 2/6 in the 250-µL group developed parasitaemia. Geometric mean (GM) pre-patent periods were 13.9, 14.0 and 15.0 days, respectively. Six/six (100%) participants developed parasitaemia in the 25,000 and 75,000 PfSPZ IM and 3,200 PfSPZ DVI groups. GM pre-patent periods were 12.2, 11.4 and 11.4 days, respectively. Injection of PfSPZ Challenge was well tolerated and safe in all groups. CONCLUSIONS: IM injection of 75,000 PfSPZ and DVI injection of 3,200 PfSPZ resulted in infection rates and pre-patent periods comparable to the bite of five PfSPZ-infected mosquitoes. Remarkably, it required 23.4-fold more PfSPZ administered IM than DVI to achieve the same parasite kinetics. These results allow for translation of CHMI from research to routine use, and inoculation of PfSPZ by IM and DVI regimens. TRIAL REGISTRATION: ClinicalTrials.gov NCT01771848.

Validation and cross-cultural adaptation of the postoperative quality of recovery 15 (QoR-15) questionnaire for Spanish-speaking patients: A prospective cohort study.

BACKGROUND: The postoperative QoR-15 questionnaire may improve the detection of postoperative complications on an early basis and contribute to optimize treatment and recovery. No validated Spanish QoR-15 questionnaire has been available to date. METHODS: The Spanish QoR-15 questionnaire (QoR-15E), carried out by official bilingual translators, was administered to 242 adult patients undergoing elective surgery before and 24 h after surgery. Patients were asked about their perceived quality of general recovery using a visual analogue scale (VAS). A random subgroup of 36 patients completed a third questionnaire 30-60 min after having completed the first one, and under the same conditions. RESULTS: The Pearson correlation coefficient between QoR-15E and the VAS score was 0.759. Cronbach's alpha was 0.856 in the postoperative period. Reliability by the split-half method was 0.781. Test-retest correlation coefficient was 0.998. Cohen's d was 0.94. The mean time to complete the preoperative questionnaire was 2.9 ± 0.5 min. CONCLUSIONS: The QoR-15E is valid and reliable for assessing postoperative quality of recovery in Spanish-speaking patients, with psychometric and interpretative features similar to those of the original instrument.

Potentially toxic element bioaccumulation in consumed indoor shrimp farming associated with diet, water and sediment levels.

Shrimp production is an important industry for many countries and shrimp consumption is increasing worldwide. Shrimps are a highly nutritional food, but can pose a risk for human health if subject to high levels of environmental contaminants. This work studies the presence of As, Cd, Co, Cr, Cu, Hg, Ni, Pb and Zn in shrimps from Ecuador and compares them to such contents noted in other shrimp-production areas in the world to evaluate the possible risks associated with these elements for consumer health, and to relate them to potentially toxic element (PTE) contents in water, sediments and diets, and also to animal biometric parameters. The PTE levels (mg kg-1 DM) obtained are as follows: in the head-As (3.52-6.11), Cd (0.02-0.10), Co (0.14-0.49) Cr (0.23-4.89), Cu (99.9--233.0), Ni (0.52-1.86), Pb (0.24-1.09), Zn (51.8-100.5) and Hg (µg kg-1 DM) (10.00-66.81); in the tail-(0.91-3.21), Cd (0.01-0.02), Co (0.01-0.43) Cr (0.01-6.52), Cu (20.0-72.44), Ni (0.15-2.03), Pb (0.01-0.69), Zn (31.2-66.1) and Hg (µg kg-1 DM) (10.00-67.18). The concentration of all the PTEs is generally lower than the limits set for seafood by European regulations, except for As in the cephalothorax (4.63 mg kg-1). Different behaviours for PTE accumulation in shrimps were found, which preferentially tend to accumulate in the cephalothorax, except for Hg (40.13 µg kg-1 DM), which accumulates in muscle (body) and is associated with contents of proteins, lipids and total shrimp weight. Nonetheless, the target hazard quotient (THQ) values for PTEs indicate that the consumption of shrimp muscles from Ecuador does not pose a human health risk because the values of these indices are below 1 in all cases.

Diagnostic Performance of Six Rapid Antigen Tests for SARS-CoV-2.

Microbiological diagnosis of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been a challenge. Although real-time reverse transcription PCR (RT-PCR) represents the gold standard method, strategies that allow rapid and simple diagnosis are necessary for the early identification of cases. In this study, we evaluated the diagnostic performance of six different commercial rapid antigen tests (Coronavirus antigen [Ag] rapid test cassette [Healgen Scientific, Houston, TX, USA], COVID-19 Ag FIA [Vircell, SD Biosensor Inc., Gyeonggi-do, Republic of Korea], Clinitest rapid COVID-19 antigen test [Siemens, Healthineers, Erlangen, Germany], SARS-CoV-2 rapid antigen test [SD Biosensor; Roche Diagnostics, Basel, Switzerland], Panbio COVID-19 Ag rapid test device [Abbott, Chicago, IL, USA], and SARS-CoV-2 test [MonLab, Barcelona, Spain]) in 130 nasopharyngeal swab samples tested previously by RT-PCR. The overall sensitivity of the rapid tests ranged from 65% to 79%, and the specificity was 100% for all of them. The sensitivity was higher for those samples with RT-PCR cycle threshold (CT) values below 25 and those from patients presenting within the first week of symptoms. The Siemens test showed the highest sensitivity for patients with high viral loads while the Vircell test performed better than the rest for CT values of ≥25. IMPORTANCE The rapid detection of people infected with SARS-CoV-2 is essential for a correct and effective control of the disease it causes. This process must be sensitive, fast, and simple, and it must be possible to carry out in any type of health center. Rapid antigen tests are the answer to this need. Knowing its ability to detect the virus in different stages of the disease is essential for a correct diagnosis, which is why this study has been carried out to evaluate the sensitivity and specificity of 6 different antigens tests in nasopharyngeal smear samples.

High within-host diversity found from direct genotyping on post-mortem tuberculosis specimens in a high-burden setting.

OBJECTIVES: To characterize the clonal complexity in Mycobacterium tuberculosis (MTB) infections considering factors that help maximize the detection of coexisting strains/variants. METHODS: Genotypic analysis by Mycobacterial Interspersed Repetitive-Unit-Variable-Number Tandem-Repeats (MIRU-VNTR) was performed directly on 70 biopsy specimens from two or more different tissues involving 28 tuberculosis cases diagnosed post-mortem in Mozambique, a country with a high tuberculosis burden. RESULTS: Genotypic data from isolates collected from two or more tissues were obtained for 23 of the 28 cases (82.1%), allowing the analysis of within-patient diversity. MIRU-VNTR analysis revealed clonal diversity in ten cases (35.7%). Five cases showed allelic differences in three or more loci, suggesting mixed infection with two different strains. In half of the cases showing within-host diversity, one of the specimens associated with clonal heterogeneity was brain tissue. CONCLUSIONS: Direct MTB genotyping from post-mortem tissue samples revealed a frequent within-host Mycobacterium tuberculosis diversity, including mixed and polyclonal infections. Most of this diversity would have been overlooked if only standard analysis of respiratory specimens had been performed.

Accuracy of verbal autopsy, clinical data and minimally invasive autopsy in the evaluation of malaria-specific mortality: an observational study.

BACKGROUND: Global malaria mortality estimates are hindered by the low reliability of the verbal autopsy (VA) and the clinical records, the most common sources of information used to estimate malaria-specific mortality. We aimed to determine the accuracy of these tools, as well as of the minimally invasive autopsy (MIA), a needle-based postmortem sampling method, to identify malaria-specific mortality in a large series of deceased patients from Mozambique, using complete autopsy as the gold standard. METHODS: Observational study that included 264 deaths, occurring at a tertiary level hospital in Mozambique, from 1 November 2013 to 31 March 2015 (17 months-long period). Clinical data were abstracted, a computer coded VA was completed using the clinical data as source of information, and an MIA followed by a complete autopsy were performed. Screening for malaria infection was conducted postmortem to all participants using molecular and histological techniques (PCR and immunohistochemistry). FINDINGS: Malaria infection was considered the cause of death in 6/264 (2.3%) cases: 2/54 children (3.7%, both less than 5 years old) and 4/57 (7.0%) maternal deaths. The sensitivity and specificity of the VA, the clinical data and the MIA to identify malaria-specific deaths were 33.3% and 96.1%, 66.7% and 96.1%, and 100% and 100%, respectively. In addition, malaria was identified as a possible contributor in 14 additional patients who died of other diseases. These cases were also accurately identified by the MIA (sensitivity 82.4%, specificity 100%). INTERPRETATION: The high sensitivity and specificity of the MIA in identifying malaria may help to improve current estimates of malaria-specific mortality in endemic areas.

High prevalence and mortality due to Histoplasma capsulatum in the Brazilian Amazon: An autopsy study.

BACKGROUND: Histoplasmosis is acquired by inhalation of spores of the dimorphic fungus Histoplasma spp. Although this pathogen is distributed worldwide, it is more prevalent in the Americas. However, the real burden of histoplasmosis remains undefined in many endemic regions. METHODOLOGY: We conducted a series of 61 autopsies to individuals who died in a hospital in the Brazilian Amazon focused on infectious diseases. We performed a detailed histological and microbiological evaluation with genetic characterization of Histoplasma strains with the aim to evaluate the contribution of histoplasmosis to morbidity and mortality. Additionally, we assessed the clinicopathological correlation. PRINCIPAL FINDINGS: Evidence of Histoplasma infection was detected in 21 patients (34%). Eight cases were disseminated infections, all of them occurred in HIV-positive patients. Six cases were localized histoplasmosis, limited to the lungs. In seven patients Histoplasma DNA was detected by PCR in patients with no histological lesions. Histoplasma infection was detected in 38% of HIV-positive patients and was a major contributor to death in 22% of them. Lungs, liver and spleen were affected in all cases of disseminated histoplasmosis. Phylogenetic analysis of the strains suggested a high diversity of Histoplasma species circulating in the Brazilian Amazon. Histoplasmosis was clinically missed in 75% of the disseminated infections. CONCLUSIONS: The high incidence of histoplasmosis, the low index of clinical suspicion, and the severity of the disseminated disease highlight the need of proactively implementing sensitive routine screening methods for this pathogen in endemic areas. Antifungal prophylaxis against Histoplasma should be encouraged in the severely immunocompromised HIV patients in these areas. In conclusion, substantial mortality is associated with disseminated histoplasmosis among HIV-positive patients in the Brazilian Amazon.

Case Report: Partial Uniparental Disomy Unmasks a Novel Recessive Mutation in the LYST Gene in a Patient With a Severe Phenotype of Chédiak-Higashi Syndrome.

Chédiak-Higashi syndrome (CHS) is a rare autosomal recessive (AR) immune disorder that has usually been associated to missense, nonsense or indels mutations in the LYST gene. In this study, we describe for the first time the case of a CHS patient carrying a homozygous mutation in the LYST gene inherited as a result of a partial uniparental isodisomy (UPiD) of maternal origin. Sanger sequencing of the LYST cDNA and single nucleotide polymorphism (SNP)-arrays were performed to identify the causative mutation and to explain the molecular mechanism of inheritance, respectively. Partial-UPiD leads to a copy neutral loss of heterozygosity (CN-LOH) of the telomeric region of chromosome 1 (1q41q44), unmasking the potential effect of the mutation detected. The mutation (c.8380dupT) is an insertion located in exon 32 of the LYST gene resulting in a premature stop codon and leading to the loss of all the conserved domains at the C-terminal of the LYST protein. This would account for the severe phenotype observed. We also reviewed the only two previously reported cases of CHS as a result of a uniparental disomy. In this study, we show that the combination of different strategies, including the use of SNP-arrays, is pivotal to fine-tune the diagnosis of rare AR disorders, such as CHS. Moreover, this case highlights the relevance of uniparental disomy as a potential mechanism of CHS expression in non-consanguineous families.

Molecular Characterization of Imported and Autochthonous Dengue in Northeastern Spain.

Dengue is the most significant arbovirus worldwide and a public health threat to non-endemic areas in which Aedes vectors are present. Autochthonous dengue transmission has been reported in several European countries in the last decade. Infected travelers from endemic regions arriving to areas colonized by Aedes albopictus in Europe need to be monitored in surveillance and control programs. We aimed to perform molecular characterization of RT-PCR-positive dengue cases detected in Catalonia, northeastern Spain, from 2013 to 2018. The basic demographic information and the geographical regions of importation were also analyzed. One-hundred four dengue cases were studied (103 imported infections and the first autochthonous case in our region). The dengue virus strains detected were serotyped and genotyped using molecular methods, and phylogenetic analyses were conducted. All four dengue serotypes were detected in travelers, including up to 10 different genotypes, reflecting the global circulation of dengue in endemic areas. The primary travel-related case of the 2018 autochthonous transmission was not identified, but the molecular analysis revealed dengue serotype 1, genotype I of Asian origin. Our results highlight the diversity of imported dengue virus strains and the role of molecular epidemiology in supporting arbovirus surveillance programs.

Minimally Invasive Autopsy Practice in COVID-19 Cases: Biosafety and Findings.

Postmortem studies are crucial for providing insight into emergent diseases. However, a complete autopsy is frequently not feasible in highly transmissible diseases due to biohazard challenges. Minimally invasive autopsy (MIA) is a needle-based approach aimed at collecting samples of key organs without opening the body, which may be a valid alternative in these cases. We aimed to: (a) provide biosafety guidelines for conducting MIAs in COVID-19 cases, (b) compare the performance of MIA versus complete autopsy, and (c) evaluate the safety of the procedure. Between October and December 2020, MIAs were conducted in six deceased patients with PCR-confirmed COVID-19, in a basic autopsy room, with reinforced personal protective equipment. Samples from the lungs and key organs were successfully obtained in all cases. A complete autopsy was performed on the same body immediately after the MIA. The diagnoses of the MIA matched those of the complete autopsy. In four patients, COVID-19 was the main cause of death, being responsible for the different stages of diffuse alveolar damage. No COVID-19 infection was detected in the personnel performing the MIAs or complete autopsies. In conclusion, MIA might be a feasible, adequate and safe alternative for cause of death investigation in COVID-19 cases.

A Single-Run Next-Generation Sequencing (NGS) Assay for the Simultaneous Detection of Both Gene Mutations and Large Chromosomal Abnormalities in Patients with Myelodysplastic Syndromes (MDS) and Related Myeloid Neoplasms.

Myelodysplastic syndromes (MDS) and myelodysplastic/myeloproliferative neoplasms are clonal disorders that share most of their cytogenetic and molecular alterations. Despite the increased knowledge of the prognostic importance of genetics in these malignancies, next-generation sequencing (NGS) has not been incorporated into clinical practice in a validated manner, and the conventional karyotype remains mandatory in the evaluation of suspected cases. However, non-informative cytogenetics might lead to an inadequate estimation of the prognostic risk. Here, we present a novel targeted NGS-based assay for the simultaneous detection of all the clinically relevant genetic alterations associated with these disorders. We validated this platform in a large cohort of patients by performing a one-to-one comparison with the lesions from karyotype and single-nucleotide polymorphism (SNP) arrays. Our strategy demonstrated an approximately 97% concordance with standard clinical assays, showing sensitivity at least equivalent to that of SNP arrays and higher than that of conventional cytogenetics. In addition, this NGS assay was able to identify both copy-neutral loss of heterozygosity events distributed genome-wide and copy number alterations, as well as somatic mutations within significant driver genes. In summary, we show a novel NGS platform that represents a significant improvement to current strategies in defining diagnosis and risk stratification of patients with MDS and myeloid-related disorders.

Analysis of SNP Array Abnormalities in Patients with DE NOVO Acute Myeloid Leukemia with Normal Karyotype.

Nearly 50% of patients with de novo acute myeloid leukemia (AML) harbor an apparently normal karyotype (NK) by conventional cytogenetic techniques showing a very heterogeneous prognosis. This could be related to the presence of cryptic cytogenetic abnormalities (CCA) not detectable by conventional methods. The study of copy number alterations (CNA) and loss of heterozygozity (LOH) in hematological malignancies is possible using a high resolution SNP-array. Recently, in clinical practice the karyotype study has been complemented with the identification of point mutations in an increasing number of genes. We analyzed 252 de novo NK-AML patients from Hospital La Fe (n = 44) and from previously reported cohorts (n = 208) to identify CCA by SNP-array, and to integrate the analysis of CCA with molecular alterations detected by Next-Generation-sequencing. CCA were detected in 58% of patients. In addition, 49% of them harbored CNA or LOH and point mutations, simultaneously. Patients were grouped in 3 sets by their abnormalities: patients carrying several CCA simultaneously, patients with mutations in FLT3, NPM1 and/or DNMT3A and patients with an amalgam of mutations. We found a negative correlation between the number of CCA and the outcome of the patients. This study outlines that CCA are present in up to 50% of NK-AML patients and have a negative impact on the outcome. CCA may contribute to the heterogeneous prognosis.