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Monkeypox DNA has been detected in skin lesions, saliva, oropharynx, urine, semen, and stool of patients infected during the 2022 clade IIb outbreak; however, the viral dynamics within these compartments remain unknown. We aimed to characterise the viral load kinetics over time in various parts of the body.This was an observational, prospective, multicentre study of outpatients diagnosed with monkeypox in two hospitals and two sexual health clinics in Spain between June 28, 2022, and Sept 22, 2022. Men and women aged over 18 years were eligible if they reported having symptom onset within the previous 10 days of presentation, and were ineligible if disease was severe enough to be admitted to hospital. Samples were collected from five body locations (skin lesions, oropharynx, rectum, semen or vagina, and a dried blood spot) at six time points up to 57 days after the screening visit. Samples were analysed by quantitative PCR and a subset by cell culture. The primary endpoint was time from symptom onset to viral DNA clearance.Overall, 1663 samples were collected from 77 study participants. 75 (97%) participants were men, the median age was 35·0 years (IQR 29·0-46·0), and 39 (51%) participants were living with HIV. The median time from symptom onset to viral clearance was 25 days (95% CI 23-28) in the skin lesions, 16 days (13-19) in the oropharynx, 16 days (13-23) in the rectum, 13 days in semen (9-18), and 1 day in blood (0-5). The time from symptom onset to viral clearance for 90% of cases was 41 days (95% CI 34-47) in skin lesions and 39 days (27-56) in semen. The median viral load in skin lesions was 7·3 logIn immunocompetent patients with mild monkeypox disease, PCR data alone would suggest a contact isolation period of 3 to 6 weeks but, based on detection of replication-competent virus, this time could be reduced. Based on findings from this cohort of patients, semen testing and prolonged use of condoms after recovery from monkeypox might not be necessary.University Hospital Germans Trias i Pujol and the YoMeCorono.For the Spanish translation of the abstract see Supplementary Materials section.

We evaluated the accuracy of patient-collected skin lesions, oropharyngeal, and rectal swabs among 50 individuals enrolled in a study of mpox viral dynamics. We found that the performance of self-collected samples was similar to that of physician-collected samples, suggesting that self-sampling is a reliable strategy for diagnosing mpox.

Purpose: We aimed to describe the outcomes, focusing on the hearing and neurological development, of children born to mothers with SARS-CoV-2 infection during pregnancy, and to evaluate the persistence of maternal antibodies in the first months of life. Methods:An observational, prospective study at a tertiary hospital in Madrid (Spain) on infants born to mothers infected with SARS CoV-2 during pregnancy between March and September 2020. A follow-up visit at 1-3 months of age with a physical and neurological examination, cranial ultrasound (cUS), SARS-CoV-2 RT-PCR on NPS and SARS-CoV-2 serology were performed. Hearing was evaluated at birth through the automated auditory brainstem response and at six months of age through the auditory steady-state response. A neurodevelopmental examination using the Bayley-III scale was performed at 12 months of age. Results:Of 95 infants studied, neurological examination was normal in all of them at the follow-up visit, as was the cUS in 81/85 (95%) infants, with only mild abnormalities in four of them. Serology was positive in 47/95 (50%) infants, which was not associated with symptoms or severity of maternal infection. No hearing loss was detected and neurodevelopment was normal in 96% of the children (median Z score: 0). Conclusions: In this cohort, the majority of infants born to mothers with SARS-CoV-2 infection during pregnancy were healthy children with a normal cUS, no hearing loss and normal neurodevelopment in the first year of life. Only half of the infants had a positive serological result during the follow-up.

We have investigated the evolution of the neutralizing response against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants at 8 months after Pfizer-BNT162b2 vaccination in coronavirus disease 2019 (COVID-19)–naive (n = 21) and COVID-19–convalescent (n = 21) individuals. Neutralizing levels declined for all variants (range 2- to 3.7-fold). Eight months after vaccination, a significant proportion (4/21) of naive individuals lacked detectable neutralizing activity against the highly transmissible SARS-CoV-2 delta variant. In the convalescent group, the impressive high initial humoral response resulted in detectable neutralizing antibody levels against all variants throughout this period.

We have measured the humoral response to messenger RNA (mRNA) vaccines in COVID-19 naïve and convalescent individuals. Third doses of mRNA COVID-19 vaccines induced a significant increase in potency and breadth of neutralization against SARS-CoV-2 variants of concern (VoC) including Omicron subvariants BA.1, BA.2, and BA.2.12.1, that were cross-neutralized at comparable levels and less for BA.4/5. This booster effect was especially important in naïve individuals that only after the third dose achieved a level that was comparable with that of vaccinated COVID-19 convalescents except for BA.4/5. Avidity of RBD-binding antibodies was also significantly increased in naïve individuals after the third dose, indicating an association between affinity maturation and cross neutralization of VoC. These results suggest that at least three antigenic stimuli by infection or vaccination with ancestral SARS-CoV-2 sequences are required to induce high avidity cross-neutralizing antibodies. Nevertheless, the circulation of new subvariants such as BA.4/5 with partial resistance to neutralization will have to be closely monitored and eventually consider for future vaccine developments.

We evaluated the accuracy of patient-collected skin lesions, pharyngeal, and rectal swabs amongst 50 individuals enrolled in a study of monkeypox viral dynamics. We found that the performance of self-collected samples was similar to that of physician-collected samples, suggesting that self-sampling is a reliable strategy for diagnosing monkeypox.

Background In May, 2022, several European countries reported autochthonous cases of monkeypox, which rapidly spread globally. Early reports suggest atypical presentations. We aimed to investigate clinical and virological characteristics of cases of human monkeypox in Spain. Methods This multicentre, prospective, observational cohort study was done in three sexual health clinics in Madrid and Barcelona, Spain. We enrolled all consecutive patients with laboratory-confirmed monkeypox from May 11 to June 29, 2022. Participants were offered lesion, anal, and oropharynx swabs for PCR testing. Participant data were collected by means of interviews conducted by dermatologists or specialists in sexually transmitted infections and were recorded using a standard case report form. Outcomes assessed in all participants with a confirmed diagnosis were demographics, smallpox vaccination, HIV status, exposure to someone with monkeypox, travel, mass gathering attendance, risk factors for sexually transmitted infections, sexual behaviour, signs and symptoms on first presentation, virological results at multiple body sites, co-infection with other sexually transmitted pathogens, and clinical outcomes 14 days after the initial presentation. Clinical outcomes were followed up until July 13, 2022. Findings 181 patients had a confirmed monkeypox diagnosis and were enrolled in the study. 166 (92%) identified as gay men, bisexual men, or other men who have sex with men (MSM) and 15 (8%) identified as heterosexual men or heterosexual women. Median age was 37.0 years (IQR 31.0-42.0). 32 (18%) patients reported previous smallpox vaccination, 72 (40%) were HIV-positive, eight (11%) had a CD4 cell count less than 500 cells per mu L, and 31 (17%) were diagnosed with a concurrent sexually transmitted infection. Median incubation was 7.0 days (IQR 5.0-10.0). All participants presented with skin lesions; 141 (78%) participants had lesions in the anogenital region, and 78 (43%) in the oral and perioral region. 70 (39%) participants had complications requiring treatment: 45 (25%) had a proctitis, 19 (10%) had tonsillitis, 15 (8%) had penile oedema, six (3%) an abscess, and eight (4%) had an exanthem. Three (2%) patients required hospital admission. 178 (99%) of 180 swabs from skin lesions collected tested positive, as did 82 (70%) of 117 throat swabs. Viral load was higher in lesion swabs than in pharyngeal specimens (mean cycle threshold value 23 [SD 4] vs 32 [6], absolute difference 9 [95% CI 8-10]; p

The standard RT-PCR assay for COVID-19 is laborious and time-consuming, limiting the availability of testing. Rapid antigen-detection tests are faster and less expensive; however, the reliability of these tests must be validated before they can be used widely. The objective of this study was to determine the reliability of the PanbioTM COVID-19 Ag Rapid Test Device (PanbioRT) (Abbott) for SARS-CoV-2 in nasopharyngeal swab specimens. This was a prospective multicenter study in ten Spanish university hospitals of patients from hospital units with clinical symptoms or epidemiological criteria for COVID-19. Patients whose onset of symptoms or exposure was more than 7 days earlier were excluded. Two nasopharyngeal exudate samples were taken to perform the PanbioRT and a diagnostic RT-PCR test. Among the 958 patients studied, 359 (37.5%) were positive by RT-PCR and 325 (33.9%) were also positive by the PanbioRT. Agreement was 95.7% (kappa score: 0.90). All 34 false-negative PanbioRT results were in symptomatic patients, with 23.5% of them at 6–7 days since the onset of symptoms and 58.8% presenting CT >30 values for RT-PCR, indicating a low viral load. Overall sensitivity and specificity for the PanbioRT were 90.5% and 98.8%, respectively. The PanbioRT provides good clinical performance as a point-of-care test, with even more reliable results for patients with a shorter clinical course of the disease or a higher viral load. While this study has had a direct impact on the national diagnostic strategy for COVID-19 in Spain, the results must be interpreted based on the local epidemiological context.

Background Cytomegalovirus (CMV) is the leading cause of congenital infection worldwide. Up to 15%-20% of infected newborns will develop long-term sequelae such as hearing loss and neurologic abnormalities. The aim of this study was to investigate the prevalence of congenital CMV infection (cCMV) and associated clinical abnormalities in Spain. Methods A prospective screening for cCMV by viral load in saliva was performed. Saliva samples were obtained within the first 72 hours of life in a maternity ward in Madrid (Spain), during a 1-year period. All positive screening tests were confirmed with viral load in urine. Clinical, laboratory, auditory, visual and cerebral imaging assessments were performed in all children with cCMV. Results Of the 4097 neonates born during the study period, 3190 (78%) were included. CMV viral load in saliva was detectable in 24/3190 (0.75%) children, and congenital infection was confirmed in 15/3190 (0.47%, CI 95%: 0.29%-0.77%). Positive predictive value was 62.5% (CI 95%: 46.5%-76.1%). Two infants presented symptoms at birth. Eight (53.3%) children showed abnormalities in magnetic resonance imaging; most of them isolated white matter abnormalities. Newborns with abnormalities in magnetic resonance imaging showed higher viral loads in blood and saliva (P = 0.04). Conclusions One in 200 neonates born in our hospital presented a cCMV infection. CMV viral load in saliva has been shown to be a simple and highly accepted screening method but should be confirmed by CMV detection in urine. In spite of the fact that half of infected children had abnormalities in cerebral imaging, diagnosis during the neonatal period would have been impossible without a screening program in most cases.

Monitoring for cytomegalovirus (CMV)-specific cell-mediated immunity (CMV-CMI) may be useful for individualizing valganciclovir (VGCV) prophylaxis after kidney transplantation (KT). We performed a commercial ELISA-based interferon (IFN)-γ release assay (QTF-CMV) from posttransplant months 2-5 (362 points) in 120 CMV-seropositive KT recipients that received antithymocyte globulin as induction therapy and VGCV prophylaxis (median of 92 days). Forty-seven patients (39.3%) had CMV infection after discontinuation of prophylaxis. The QTF-CMV assay was reactive, nonreactive, and indeterminate in 264 (72.9%), 90 (24.9%), and 8 points (2.2%). The QTF-CMV assay at prophylaxis discontinuation exhibited suboptimal accuracy for predicting protective CMV-CMI (sensitivity: 77.4%; specificity: 34.3%; positive predictive value [PPV]: 64.1%; negative predictive value [NPV]: 50.0%), with no differences in 1-year CMV infection rates between patients with negative (nonreactive or indeterminate) or reactive results (45.8% vs 36.1%; P = .244). Specificity and PPV to predict protective CMV-CMI improved by elevating the IFN-γ cutoff value to 1.13 IU/mL (65.7% and 71.4%) and 7.0 IU/mL (85.7% and 76.2%), although NPVs decreased. The QTF-CMV assay as per manufacturer's interpretative criteria performed poorly to predict protection from CMV infection following discontinuation of VGCV prophylaxis among ATG-treated CMV-seropositive KT recipients. This performance is slightly improved by modifying the IFN-γ positivity threshold.

The clinical characteristics, management, and outcome of coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) after solid organ transplant (SOT) remain unknown. We report our preliminary experience with 18 SOT (kidney [44.4%], liver [33.3%], and heart [22.2%]) recipients diagnosed with COVID-19 by March 23, 2020 at a tertiary-care center at Madrid. Median age at diagnosis was 71.0 ± 12.8 years, and the median interval since transplantation was 9.3 years. Fever (83.3%) and radiographic abnormalities in form of unilateral or bilateral/multifocal consolidations (72.2%) were the most common presentations. Lopinavir/ritonavir (usually associated with hydroxychloroquine) was used in 50.0% of patients and had to be prematurely discontinued in 2 of them. Other antiviral regimens included hydroxychloroquine monotherapy (27.8%) and interferon-s (16.7%). As of April 4, the case-fatality rate was 27.8% (5/18). After a median follow-up of 18 days from symptom onset, 30.8% (4/13) of survivors developed progressive respiratory failure, 7.7% (1/13) showed stable clinical condition or improvement, and 61.5% (8/13) had been discharged home. C-reactive protein levels at various points were significantly higher among recipients who experienced unfavorable outcome. In conclusion, this frontline report suggests that SARS-CoV-2 infection has a severe course in SOT recipients.

ObjectiveTo report 3 patients infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) who developed generalized myoclonus.MethodsPatient data were obtained from medical records from the University Hospital “12 de Octubre,” Madrid, Spain.ResultsThree patients (2 men and 1 woman, aged 63–88 years) presented with mild hypersomnia and generalized myoclonus following the onset of the so-called inflammatory phase of coronavirus disease 2019 (COVID-19). All of them had presented previously with anosmia. Myoclonus was generalized with both positive and negative jerks, predominantly involving the facial, trapezius, sternocleidomastoid, and upper extremities muscles. These myoclonic jerks occurred spontaneously and were extremely sensitive to multisensory stimuli (auditive and tactile) or voluntary movements, with an exaggerated startle response. Other causes of myoclonus were ruled out, and none of the patients had undergone respiratory arrest or significant prolonged hypoxia. All of them improved, at least partially, with immunotherapy.ConclusionsOur 3 cases highlight the occurrence of myoclonus during the COVID-19 pandemic as a post- or para-infectious immune-mediated disorder. However, we cannot rule out that SARS-CoV-2 may spread transneuronally to first- and second-order structures connected with the olfactory bulb. Further investigation is required to clarify the full clinical spectrum of neurologic symptoms and optimal treatment.

Previous reports hypothesized that cytomegalovirus (CMV) may predispose to non-CMV infection after kidney transplantation (KT). We analysed the incidence of non-CMV infection (overall, bacterial and opportunistic) in 291 KT recipients according to the previous development of any level or high-level (≥1,000 IU/ml) CMV viremia. Exposure to CMV replication was assessed throughout fixed intervals covering first the 30, 90, 180 and 360 post-transplant days (cumulative exposure) and non-overlapping preceding periods (recent exposure). Adjusted Cox models were constructed for each landmark analysis. Overall, 67.7 and 50.5% patients experienced non-CMV and CMV infection, respectively. Patients with cumulative CMV exposure had higher incidence of non-CMV infection beyond days 30 (p-value = 0.002) and 90 (p-value = 0.068), although these associations did not remain after multivariable adjustment. No significant associations were observed for the remaining landmark models (including those based on high-level viremia or recent CMV exposure), or when bacterial and opportunistic infection were separately analysed. There were no differences in viral kinetics (peak CMV viremia and area under curve of CMV viral load) either. Our findings do not support the existence of an independent association between previous CMV exposure and the overall risk of post-transplant infection, although results might be affected by power limitations.

[Objectives]: To describe the etiology of community-acquired pneumonia (CAP) in hospitalized children in Spain and analyze the predictors of the etiology., [Hypothesis]: The different etiological groups of pediatric CAP are associated with different clinical, radiographic, and analytical data., [Design]: Observational, multicenter, and prospective study., [Patient selection]: This study included children aged 1 month to 17 years with CAP, who were hospitalized between April 2012 and May 2019., [Methods]: An extensive microbiological workup was performed. The clinical, radiographic, and analytical parameters were analyzed for three etiological groups., [Results]: Among the 495 children included, at least one causative pathogen was identified in 262 (52.9%): pathogenic viruses in 155/262 (59.2%); atypical bacteria (AB), mainly Mycoplasma pneumonia, in 84/262 (32.1%); and typical bacteria (TyB) in 40/262 (15.3%). Consolidation was observed in 89/138 (64.5%) patients with viral CAP, 74/84 (88.1%) with CAP caused by AB, and 40/40 (100%) with CAP caused by TyB. Para-pneumonic pleural effusion (PPE) was observed in 112/495 (22.6%) patients, of which 61/112 (54.5%) presented a likely causative pathogen: viruses in 12/61 (19.7%); AB in 23/61 (37.7%); and TyB in 26/61 (42.6%). Viral etiology was significantly frequent in young patients and in those with low oxygen saturation, wheezing, no consolidation, and high lymphocyte counts. CAP patients with AB as the etiological agent had a significantly longer and less serious course as compared to those with other causative pathogens., [Conclusions]: Viruses and M. pneumoniae are the main causes of pediatric CAP in Spain. Wheezing, young age, and no consolidation on radiographs are indicative of viral etiology. Viruses and AB can also cause PPE. Since only a few cases can be directly attributed to TyB, the indications for antibiotics must be carefully considered in each patient., Instituto de Investigación Hospital 12 de Octubre (i+12), Grant/Award Number: AY191212‐1; Instituto de Salud Carlos III (Ministry of Economy, Industry and Competitiveness) and co‐funded by the European Regional Development Funds, Grant/Award Number: Project PI17/01458; Instituto Ramón y Cajal de Investigación Sanitaria (IRYCIS), Grant/Award Number: PCAPE 2011_0025 Register 320/11; Research Project of Universidad Europea de Madrid, Grant/Award Number: 2017/UEM03

An early analysis of circulating monocytes may be critical for predicting COVID-19 course and its sequelae. In 131 untreated, acute COVID-19 patients at emergency room arrival, monocytes showed decreased surface molecule expression, including low HLA-DR, in association with an inflammatory cytokine status and limited anti-SARS-CoV-2-specific T cell response. Most of these alterations had normalized in post-COVID-19 patients 6 months after discharge. Acute COVID-19 monocytes transcriptome showed upregulation of anti-inflammatory tissue repair genes such as BCL6, AREG and IL-10 and increased accessibility of chromatin. Some of these transcriptomic and epigenetic features still remained in post-COVID-19 monocytes. Importantly, a poorer expression of surface molecules and low IRF1 gene transcription in circulating monocytes at admission defined a COVID-19 patient group with impaired SARS-CoV-2-specific T cell response and increased risk of requiring intensive care or dying. An early analysis of monocytes may be useful for COVID-19 patient stratification and for designing innate immunity-focused therapies.

Background The objective of this study was to investigate the neutralizing response against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants of concern (VoC) during coronavirus disease 2019 (COVID-19) convalescence and after vaccination. Methods COVID-19-convalescent and -naïve individuals were tested for neutralizing activity against SARS-CoV-2 VoC Alpha, Beta, Gamma, and Delta at 1 and 7 months postinfection and 4–6 weeks after BNT162b2 vaccination. Results Vaccination induced a high neutralizing response in naïve individuals. Interestingly, vaccination of convalescent patients induced a boosted response that was able to neutralize all VoC at high titers. Conclusions Vaccination with BNT162b2 induced high levels of neutralization against SARS-CoV-2 VoC in most patients; this is especially beneficial in COVID-19-convalescent individuals.

Objetives. To perform a description of the etiology of hospitalized children with community-acquired pneumonia (CAP) in Spain and analyze predictors related to etiology. Hypothesis. The different etiological groups of pediatric CAP are associated to different clinical, radiographic and analytical data. Design. Observational, multi-center, prospective study. Patient selection. Patients from 1 month to 17 years admitted because of CAP from April 2012 to May 2019. Methods. An extensive microbiological workup was done. Clinical, radiographic and analytical parameters were analyzed in order to differentiate viral, atypical bacteria (AB) and typical bacteria (TyB) pneumonia. Results. 495 children were enrolled. At least one likely causative pathogen was identified in 262 (52.9%). Pathogenic viruses in 155/262 (59.2%), AB in 84/262 (32.1%) and TyB in 40/262 (15.3%). Consolidation was found in 89/138 (64.5%) CAP attributed to virus only, in 74/84 (88.1%) of CAP attributed to AB and 40/40 (100%) of CAP attributed to TyB. Para-pneumonic pleural effusion (PPE) was found in 112/495 (22.6%) patients, 61/112 (54.5%) with a likely causative pathogen: virus 12/61 (19.7%), AB 23/61 (37.7%) and TyB 26/61 (42.6%). Viral etiology was significantly more frequent in younger patients and those with lower oxygen saturation, wheezing, no-consolidation and higher lymphocyte counts. Patients with AB were significantly more likely to have more days of fever at admission and a higher rate of use of antibiotics before admission. Conclusions. Viruses and AB are the main cause of pediatric CAP in Spain. Wheezing, younger age and no-consolidation on the X-ray support viral etiology. Viruses and AB can also cause PPE. The use of antibiotic in pediatric CAP can be restricted.

Background and ObjectivesEstablishing the etiology of community-acquired pneumonia (CAP) in children at admission is challenging. As a result, most children receive antibiotics that do not need.This study aims to build and validate a diagnostic tool combining clinical, analytical and radiographical features to sequentially differentiate viral from bacterial CAP, and among bacterial CAP, typical from atypical bacteria, to improve choice of treatment.MethodsConsecutive hospitalized children between 1 month and 16 years of age with CAP were enrolled. An extensive microbiological workup was performed. A score was built with a training set of 70% patients, to first differentiate between viral and bacterial CAP and secondly, typical from atypical bacterial CAP. To select variables, a Ridge model was used. Optimal cut-off points were selected to maximize specificity setting a high sensitivity (80%). Weights of each variable were calculated with a multivariable logistic regression. The score was validated with the rest of the participants.ResultsIn total, 262 (53%) children (median age, 2 years, 52.3% male) had an etiological diagnosis.The step 1 discriminates viral from bacterial CAP. Bacterial CAPs were classified with a sensitivity=97%, a specificity=48%, and a ROC’s area under the curve (AUC)=0.81. If a CAP was classificated as bacterial, it was assessed with step 2. The step 2 differentiates typical vs. atypical bacterial CAP. Typical bacteria were classified with a sensitivity=100%, a specificity=64%, and AUC=0.90.ConclusionThis two-steps tool can facilitate the physician’s decision to prescribe antibiotics without compromising patient safety.Article summaryWe validated a clinical tool to predict the aetiology of CAP in children safely. This tool differentiates CAP into viral, atypical bacteria and typical bacteria.“What’s Known on This Subject”Establishing the aetiology of community-acquired pneumonia (CAP) in children at admission is challenging. As a result, most admitted children with CAP receive antibiotics.“What This Study Adds”We validated a clinical tool to predict the aetiology of pneumonia in children safely, differentiating among viral, atypical bacteria and typical bacteria.

OBJECTIVES This study focused on the role that BK polyomavirus (BKPyV) genotypes can play in the development of BKPyV-associated complications in renal transplant recipients. METHODS A retrospective observational study (January 2015 to April 2018) was conducted by analyzing BKPyV genotypes in 180 blood samples with detectable BKPyV viral load (VL) > 1000 copies/mL, from 63 renal transplant recipients. VL and BKPyV genotypes detections were based on real-time PCR (rt-PCR)-specific assays. RESULTS Forty-four patients (44/63 [69.8%]) were men, and the median age was 55.0 (interquartile range 49.0-66.0 years). Eleven patients had clinical manifestations (11/63 [17.5%]). The most frequently detected genotypes were I (14/63 [22.2%]) and II (13/63 [20.6%]). Half of the patients (33/63 [52.4%]) had a mixed genotype, most with genotypes I and II (25/33 [75.8%]). Patients with infection by mixed genotypes showed VLs that were detected earlier (in the first year after transplantation) than those with a single genotype (25/33 [75.8%] vs 13/30 [43.3%], P = .009) and demonstrated greater risk of developing clinical manifestations associated with BKPyV (odds ratio 12.609, 95% confidence interval 1.503-105.807). Moreover, patients with first BKPyV VL > 10 000 copies/mL more frequently presented mixed genotypes (12/16 [75.0%] vs 21/47 [44.7%], P = .036). CONCLUSIONS The probability of developing clinical manifestations is higher in infections by mixed genotypes. Therefore, the detection of BKPyV genotypes by rt-PCR can provide relevant information to stratify patients' risk of BKPyV-associated complications and guide the clinical management of BKPyV infection in kidney transplant recipients.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was first detected in Madrid, Spain, on 25 February 2020. It increased in frequency very fast and by the end of May more than 70,000 cases had been confirmed by reverse transcription-polymerase chain reaction (RT-PCR). To study the lineages and the diversity of the viral population during this first epidemic wave in Madrid we sequenced 224 SARS-CoV-2 viral genomes collected from three hospitals from February to May 2020. All the known major lineages were found in this set of samples, though B.1 and B.1.5 were the most frequent ones, accounting for more than 60% of the sequences. In parallel with the B lineages and sublineages, the D614G mutation in the Spike protein sequence was detected soon after the detection of the first coronavirus disease 19 (COVID-19) case in Madrid and in two weeks became dominant, being found in 80% of the samples and remaining at this level during all the study periods. The lineage composition of the viral population found in Madrid was more similar to the European population than to the publicly available Spanish data, underlining the role of Madrid as a national and international transport hub. In agreement with this, phylodynamic analysis suggested multiple independent entries before the national lockdown and air transportation restrictions., This research was partially supported through the European Commission Horizon 2020 Framework Programme (VIRUSCAN, FETPROACT-2016). E.D. was recipient of a Marie Skłodowska- Curie Individual Fellowship (Grant Agreement number 796084). E.V. is supported by the Subprograma Juan Rodés, Instituto de Salud Carlos III, Subdirección General de Redes y Centros de Investigación Cooperativa, Ministerio de Ciencia, Innovación y Universidades, Spain (JR18/00048). R.R. is supported by the Subprograma Río Hortega, Instituto de Salud Carlos III, Subdirección General de Redes y Centros de Investigación Cooperativa, Ministerio de Ciencia, Innovación y Universidades, Spain (CM19/00229).

Background The impact of late-onset cytomegalovirus (CMV) infection (LOCI) on cardiac allograft vasculopathy (CAV) has yet to be established. Methods A retrospective study was performed for patients who had undergone heart transplantation (HT) between January 1995 and October 2017 to analyze epidemiology of LOCI (any positive level of CMV pp65 antigenemia or DNAemia after 100 days, without previous CMV replication) and its association with CAV. Our main hypothesis was that LOCI causes less direct and indirect effects compared to early onset infection (EOCI). Results Late-onset cytomegalovirus infection developed in 57 of 410 patients (13.9%) in a median time of 4.7 months post-transplant. CAV at 10 years was diagnosed in 31.6% of patients with LOCI, 34.6% with EOCI, and in 19.3% of CMV-uninfected patients. In the multivariate analysis, EOCI was an independent variable for developing CAV (HR 1.8, 95% CI 1.13-2.82, P = .01). Patients with LOCI showed a trend toward a higher risk of CAV, but the difference was not statistically significant (HR 1.7, 95% CI 0.95-3.08, P = .07). In the complementary log-log model, LOCI and EOCI had a similar CAV-free survival, and a higher probability of developing CAV than CMV-uninfected patients (P = .02). Conclusions Cytomegalovirus infection after HT may result in the same long-term events regardless of its onset, with a higher risk of developing CAV at 10 years than patients without CMV.

Background: The impact of coronavirus disease 2019 (COVID-19) on hematological patients has not been comprehensively reported to date. Methods: We analyzed 39 hematological patients diagnosed with SARS-CoV-2 infection at our institution from March 7 to April 7, 2020. Clinical characteristics and outcomes were compared to a matched contemporary control group of 53 non-cancer patients hospitalized at our center. Univariate and multivariate analyses were carried out to assess the risk factors associated with poor outcome. Results: Median patient age was 64.7 years (range 36-88 years). The most frequent hematological diseases were lymphoma (30%), multiple myeloma (30%), and chronic lymphocytic leukemia (15%). On the WHO severity scale, 12.8% of the cancer patients were classified as mild, 41% as moderate, and 46.2% as severe. Most of the hematological patients were treated with hydroxychloroquine (89%) and lopinavir/ritonavir (79.5%); 51% of the patients received corticoids and 30.8% (n=12) received tocilizumab. In the multivariate analysis, we observed that only age >70 years and C reactive protein >10 mg/dl were associated with higher risk of death (odds ratio 34.86 (3.407-356.8) and 13.56 (1.28-143.45), respectively). An unfavorable impact of chemotherapy administration on COVID-19 outcome was not demonstrated. The non-hematological and hematological patients had similar clinical and laboratory characteristics; however, but mortality was higher in hematological patients (35.9% vs. 13.2%; Odds Ratio 6.5; 95% CI=1.868-23.688; P=0.003). Conclusion: Patients with hematological malignancies had a similar incidence of COVID-19, but the severity and mortality were much higher than in non-cancer patients even in the absence of therapy. Funding Statement: This work received funding from the research contract COV20/00181 from the Spanish Ministry of Science and Innovation, Instituto de Salud Carlos III. Our research group was supported by the Fundacion Cris contra el cancer. Declaration of Interests: The authors declare no competing interests. Ethics Approval Statement: This retrospective study was approved by the Hospital Universitario 12 de Octubre Institutional Review Board (n 20/182).

Background Which are the consequences of severe acute respiratory syndrome coronavirus 2 (SARS‐CoV2) infection in liver transplant (LT) recipients? Methods We attempted to address this question by reviewing our single‐center experience during the first two months of the pandemics at a high incidence area. Results Nineteen adult patients (5 females) were diagnosed by May 5th, 2020. Median age was 58 (range 55‐72), and median follow‐up since transplantation was 83 (range 20‐183) months. Cough (84.2%), fever (57.9%) and dyspnea (47.4%) were the most common symptoms. Thirteen patients (68.4%) had pneumonia in X‐ray/CT scan. Hydroxychloroquine was administered in 11 patients, associated with lopinavir/ritonavir and interferon β in 2 cases each. Immunomodulatory therapy with tocilizumab was used in 2 patients. Immunosuppression (IS) was halted in one patient, and modified in only other two due to potential drug interactions. Five (26.3%) patients were managed as outpatient. Two patients (10.5%) died, 10 (52.6%) were discharged home, and 2 (10.5%) were still hospitalized after a median follow‐up of 41 days from the onset of symptoms. Baseline IS regimen remained unchanged in all surviving recipients, with good liver function. Conclusions Our preliminary experience shows a broad spectrum of disease severity in LT patients with COVID‐19, with a favorable outcome in most of them without needing to modify baseline IS.

AIM Previous studies have suggested a more frequent and severe course of novel coronavirus Sars-coV-2 infection in cancer patients undergoing active oncologic treatment. Our aim was to describe the characteristics of the disease in this population and to determine predictive factors for poor outcome in terms of severe respiratory distress (ARDS) or death. PATIENTS AND METHODS Patients consecutively admitted for SARS-CoV-2 infection were prospectively collected and retrospective statistical analysis was performed. Univariate and multivariate analyses were performed to assess potential factors for poor outcomes defined as ARDS or death. RESULTS 63 patients were analyzed, 34 of them developed respiratory failure (70% as ARDS). Lymphocytes/mm3 (412 vs. 686; p=0.001), serum albumin (2.84 vs. 3.1); LDH (670vs. 359; p, Highlights • Cancer patients are supposed to be specially vulnerable for Sars-CoV-2 infection. • Active cancer treatment could be deleterious in case of simultaneous SARS-CoV-2 infection. • Outcomes of patients under treatment and admitted for SARS-CoV-2 infection are described. • Hospitalized SARS-CoV-2 cancer patients show similar death rate to non-cancer patients.

Background Information regarding the incidence and characteristics of COVID-19 pneumonia amongst pregnant women is scarce. Methods Single-centre experience with 32 pregnant women diagnosed with COVID-19 between March 5 to April 5, 2020 at Madrid, Spain. Findings COVID-19 pneumonia was diagnosed in 61·5% (32/52) women. Only 18·7% (6/32) had some underlying condition (mostly asthma). Supplemental oxygen therapy was required in 18 patients (56·3%), with high-flow requirements in six (18·7%). Eight patients (25·0%) fulfilled the criteria for acute distress respiratory syndrome. Invasive mechanical ventilation was required in two patients (6·2%). Tocilizumab was administered in five patients (15·6%). Delivery was precipitated due to COVID-19 in three women (9·4%). All the newborns had a favourable outcome, with no cases of neonatal SARS-CoV-2 transmission. Severe cases of pneumonia requiring supplemental oxygen were more likely to exhibit bilateral alveolar or interstitial infiltrates on chest X-ray (55·6% vs. 0·0%; P-value = 0·003) and serum C-reactive protein (CRP) levels >10 mg/dL (33·0% vs. 0·0%; P-value = 0·05) at admission than those with no oxygen requirements. Interpretation Pregnant women with COVID-19 have a high risk of developing pneumonia, with a severe course in more than half of cases. The presence of bilateral kung infiltrates and elevated serum CRP at admission may identify women at-risk of severe COVID-19 pneumonia. Funding Instituto de Salud Carlos III (COV20/00,181), Spanish Ministry of Science and Innovation.

BackgroundOn January 31st the first case of COVID-19 was detected in Spain, an imported case from Germany in Canary Islands, and thereafter on February 25th the first case was detected in Madrid. The first case of COVID-19 was confirmed at the Hospital Universitario 12 de Octubre on March 1st, a large public hospital with 1200 beds, covering an area over 400000 inhabitants in southern Madrid. During March 2020 highly active circulation of SARS-CoV-2 was experienced in Madrid with 24090 cases officially reported by March 29th.MethodsSince the beginning of the epidemics the Occupational Health and Safety Service (OHSS) organized the consulting and testing of the hospital personnel with confirmed exposure and also those presenting symptoms suggestive of viral respiratory infection. For molecular diagnosis of SARS-CoV-2 infection both nasopharyngeal and oropharyngeal swabs were obtained from suspected cases and processed at the Microbiology Laboratory by automatized specific PCR methods that was operative from February 25th as part of the preparedness.ResultsFrom a total of 6800 employees of the hospital, 2085 (30,6 %) were tested during the period 1-29 March 2020, some of them repeatedly (2286 total samples). The first HCW infected was confirmed on March 9th. A total of 791 HCW and personnel were confirmed to be infected by March 29th, representing 38% of those tested and 11,6 % of all the hospital workers. The proportion of infected individuals was estimated among the different groups of occupational exposure and the evolution of the cases during the expansive epidemic wave was compared between HCW and those patients attending at the Emergency Department (ER) during the same period and adjusted by the same age range. There were no statistically significant differences in the proportion of SARS- CoV-2 positive PCR detection between HCW from high risk areas involved in close contact with COVID-19 patients in comparison with clerical, administrative or laboratory personnel without direct contact with patients. The curves of evolution of accumulated cases between patients and HCW during March 2020 showed an almost parallel shape.DiscussionThe recommendation from our OHSS did not include testing of asymptomatic cases but was highly proactive in testing even patients with minor symptoms therefore, a high proportion of HCW and non-sanitary personnel was tested in March 2020 during the rapid period of expansion of the epidemics in Madrid, accounting for a total of 30,6 % of the hospital employees. Most of the COVID-19 cases among the hospital HCW and personnel were mild and managed at home under self-isolation measures, however 23 (3%) required hospitalization mostly due to severe bilateral interstitial pneumonia, two of those cases required mechanical ventilation at the ICU. No fatalities occurred during the study period.Although there were some cases of highly probable transmission from COVID-19 patients to HCWs, mainly at the first phase of the epidemics, there were no significant differences on the infection rates of HCW and hospital personnel that can be related to working in areas of high exposure risk. Furthermore, the evolution of cases during the same time period (March 2020) between patients attending the ER and hospital staff suggests that both groups were driven by the same dynamics. This experience is similar to the communicated from Wuhan verified by the WHO Joint Mission and also from recent experiences at hospital in the Netherlands, where most of the infections of HCW were related to household or community contacts.SignificanceSince the collective of hospital HCW are exhaustively screened in specific centers, their rate of infection for SARS-CoV-2 could be an indicator of the epidemic dynamics in the community. There appears to be a close connection between HCW infection and the driving forces of transmission in the community. Although we cannot exclude an additional risk factor of infection by SARS-CoV-2 due to the fact of the hospital environment, the similar proportions of positive cases among all the areas of the hospital and the evolutive wave of infection, as compared with the community, are clear arguments against a major factor of occupational risk. Exhaustive testing, such as the one carried out in our institution, covering over one third of all the workers, could be used as a reference of the population infected in the community. Since a significant proportion of COVID-19 cases can be asymptomatic and not all the hospital employees were actually tested, it is highly likely that this 11,6 % is a minimum estimation of the impact of SARS-CoV-2 circulation in Madrid during the first 4 weeks of the epidemics. This is in high and clear contrast with the official figures circulating at national and international levels. This has important implications to more precisely estimate the actual number of cases in the community and to develop public health policies for containment, treatment and recovery.

We report our experience with pregnant women with COVID-19 pneumonia diagnosed between March 5 to April 5, 2020 at a tertiary-care center at Madrid, Spain COVI

Background: Coronavirus Disease 2019 (COVID-19) pandemic has implacably stricken on the wellness of many countries and their health-care systems. The aim of the present study is analyze the clinical characteristics of the initial wave of patients with COVID-19 attended in our center, and to identify the key variables predicting the development of respiratory failure. Methods: We retrospectively recruited all hospitalized adult patients who tested positive for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) rRT-PCR assay performed on respiratory samples from March 2nd to 18th, 2020. Patients were followed up to May 1st, 2020 or death. Respiratory failure was defined as a PaO2/FiO2 ratio ≤200 mm Hg or the need for mechanical ventilation (either non-invasive positive pressure ventilation or invasive mechanical ventilation). Findings: We included 521 patients of whom 416 (81%) had abnormal Chest X-ray on admission. Median age was 64·6 ± 18·2 years, with 317 patients (60.8%) over 60 years. One hundred eighty-one (34·7%) developed respiratory failure after a median time from onset of symptoms of 9 days (IQR: 6-11) and the in-hospital mortality was 23·8% (124/521). The modelling process concluded into a logistic regression multivariable analysis and a predictive score at admission. Age, peripheral pulse oximetry, lymphocyte count, lactate dehydrogenase and C-reactive protein were the selected variables. The model has a good discriminative capacity with an area under the ROC curve of 0·85 (0·82-0·88). Interpretation: The application of a simple and reliable score at admission seems to be a useful tool to predict respiratory failure in hospitalized COVID-19 patients. Funding: This study was supported by the Spanish Ministry of Science and Innovation, Instituto de Salud Carlos III (COVID-19 research call COV20/00181) — co‐ financed by the European Development Regional Fund “A way to achieve Europe”. Mikel Mancheno-Losa was supported by a Research Contract “Rio Hortega” from the Spanish Ministry of Science and Innovation, Instituto de Salud Carlos III (CM19/00226). Mario Fernandez-Ruiz holds a research contract “Miguel Servet” from the Spanish Ministry of Science and Innovation, Instituto de Salud Carlos III (CP 18/00073). Declaration of Interests: The authors declared they do not have anything to disclose regarding conflict of interest with respect to this manuscript. Ethics Approval Statement: The protocol was approved by the Hospital 12 de Octubre Clinical Research Ethics Committee (reference 20/117) and granted a waiver of informed consent due to its retrospective observational design.

BACKGROUND Recent studies suggest that Epstein-Barr virus DNAemia (EBVd) may act as a surrogate marker of post-transplant immunosuppression. This hypothesis has not been tested so far in lung transplant (LT) recipients. METHODS We included 63 patients undergoing lung transplantation at our center between October 2008 and May 2013. Whole blood EBVd was systematically assessed by real-time polymerase chain reaction assay on a quarterly basis. The occurrence of late complications (overall and opportunistic infection [OI] and chronic lung allograft dysfunction [CLAD]) was analyzed according to the detection of EBVd within the first 6 months post transplantation. RESULTS Any EBVd was detected in 30 (47.6%) patients. Peak EBVd was higher in patients with late overall infection (2.23 vs. 1.73 log10 copies/mL; P = 0.026) and late OI (2.39 vs. 1.74 log10 copies/mL; P = 0.004). The areas under receiver operating characteristic curves for predicting both events were 0.806 and 0.871 respectively. The presence of an EBVd ≥2 log10 copies/mL during the first 6 months post transplantation was associated with a higher risk of late OI (adjusted hazard ratio [aHR] 7.92; 95% confidence interval [CI] 2.10-29.85; P = 0.002). Patients with detectable EBVd during the first 6 months also had lower CLAD-free survival (P = 0.035), although this association did not remain statistically significant in the multivariate analysis (aHR 1.26; 95% CI 0.87-5.29; P = 0.099). CONCLUSIONS Although preliminary in nature, our results suggest that the detection of EBVd within the first 6 months after transplantation is associated with the subsequent occurrence of late OI in LT recipients.

Objectives To evaluate the progression of the seroprevalence of SARS-CoV-2 in the pregnant population of the south of Madrid during the first wave of the COVID-19 pandemic. Secondarily we aimed to evaluate maternal and perinatal outcomes. Study design Retrospective cohort study conducted at Hospital Universitario 12 de Octubre during weeks 10 to 19 of 2020, coinciding with the Spanish lockdown. We tested 769 serum samples obtained from routine serological testing during the first and third trimesters of pregnancy for specific IgG anti SARS-CoV-2 RBD and S proteins. RT-PCR tests were performed in suspected cases according to clinical practice. We compared maternal and perinatal outcomes in those with delivered pregnancies (n = 578) according to the presence or absence of specific IgG antibodies. Those with positive IgG were subdivided by the presence or absence of Covid-19 related symptoms at any time and the results of RT-PCR testing if performed. Therefore, we had 4 study groups: G1 (IgG negative), G2 (IgG positive, asymptomatic, RT-PCR testing negative or not done), G3 (IgG positive, symptomatic, RT-PCR testing negative or not done), and G4 (IgG positive, symptomatic, RT-PCR positive). Results Seropositivity increased from 0% to 21.4% (95% CI 11.8–31.0) during the study period, of which 27.9% had an asymptomatic course. Overall outcomes were favorable with a significant increased rate of preterm birth in G4 vs G1 (21.4% vs 6.7%) and cesarean/operative delivery (50% vs 26.9%). Asymptomatic and mild cases did not have differences regarding pregnancy course when compared to seronegative women. There were no documented cases of vertical or horizontal transmission. Conclusion Seroprevalence in pregnant women in southern Madrid went up to 21.4% of which 27.9% had an asymptomatic course. Overall perinatal results were favorable, especially in those asymptomatic.

Background Data concerning the use of leflunomide-a drug approved for rheumatoid arthritis with in vitro anticytomegalovirus (CMV) activity-in lung transplant (LT) recipients are scarce. Aims To report the use of leflunomide in LT recipients diagnosed with CMV infection/disease. Material and methods We performed a single-center retrospective study including LT recipients who received leflunomide for CMV infection or as secondary prophylaxis after viremia clearance. We also conducted a full systematic PubMed search until June 30, 2017. Results We identified 5 LT recipients in our center plus 7 patients reported in the literature. All patients had previously received ganciclovir (GCV) and foscarnet (FOS), with drug-induced adverse effects described in 6 recipients (50%). Antiviral resistance mutations were observed in 8 patients (66.7%). Leflunomide was prescribed for CMV infection in 9 of 12 patients (75%) and as secondary prophylaxis in 3 patients (25%). Initial decrease of CMV viremia after starting leflunomide was observed in 7 of 9 recipients (77.7%), although this response was only transient in 2 patients. Long-term suppression of CMV viremia was reported in 7 of 12 patients (58.3%). In 3 recipients (25%), leflunomide was discontinued due to adverse effects. Discussion Our study has some limitations, such as the small number of patients included, its retrospective nature, and absence of leflunomide drug monitoring in serum. Notwithstanding, in our experience, leflunomide proved to be particularly effective as an anti-CMV secondary prophylaxis treatment and for clearing low-grade viremia. Moreover, leflunomide combined with a short course of GCV or intravitreal FOS also proved to be very effective in some patients. Conclusion Leflunomide, alone or in combination, could be an effective treatment in selected LT recipients with GCV-resistant CMV infection and as secondary prophylaxis. Further studies are necessary.

Viral infections are a major cause of morbidity and even mortality in solid organ transplant recipients. This article reviews key aspects of infections in solid organ transplant recipients from respiratory viruses, such as influenza, polyomavirus, erythrovirus B19 and measles.

Experiments were performed on 120 rabbits to compare the probability of infection after bone surgery without an implant, with polymethylmethacrylate, and with bone autografts. Staphylococcus aureus phage type 94/96. Isolated from a human osteomyelitis, was instilled into the intramedullar cavity after reaming of the femoral canal and before insertion of the implant. The different 50% infective doses were determined for each of the groups for comparative purposes. The bacterial concentrations required to produce infection in femora without an implant were two times less than those necessary in femora implanted with polymethylmethacrylate. The bone graft required bacterial concentrations nine times less than those necessary to infect femora containing polymethylmethacrylate and four times less than those required to infect femora without an implant. The results presented here confirm that the susceptibility to infection in orthopaedic surgery is not only material dependent but also bacteria dependent.

Objectives This study compares the infectivity of SARS-CoV-2 in respiratory samples from patients with mild COVID-19 with those from hospitalized patients with severe bilateral pneumonia. In severe COVID-19, we also analysed the presence of neutralizing activity in paired sera. Methods We performed cell cultures on 193 real-time reverse transcription polymerase chain reaction respiratory samples, positive for SARS-CoV-2, obtained from 189 patients at various times, from clinical diagnosis to follow-up. Eleven samples were obtained from asymptomatic individuals, 91 samples from 91 outpatients with mild forms of COVID-19 and 91 samples from 87 inpatients with severe pneumonia. In these patients, neutralizing activity was analysed in 30 paired sera collected after symptom onset >10 days. Results We detected a cytopathic effect (CPE) in 91/193 (47%) samples. Viral viability was maintained for up to 10 days in patients with mild COVID-19. In patients with severe COVID-19, the virus remained viable for up to 32 days after the onset of symptoms. Patients with severe COVID-19 presented infectious virus at a significantly higher rate in the samples with moderate to low viral load (cycle threshold value ≥ 26): 32/75 (43%) versus 14/63 (22%) for mild cases (p 1:1024 in 10% (3/30) of samples. Discussion Patients with severe COVID-19 might shed viable virus during prolonged periods of up to 4 weeks after symptom onset, even when presenting high cycle threshold values in their respiratory samples and despite having developed high neutralizing antibody titres.