The SARS-CoV-2 spike (S) protein, the viral mediator for binding and entry into the host cell, has sparked great interest as a target for vaccine development and treatments with neutralizing antibodies. Initial data suggest that the virus has low mutation rates, but its large genome could facilitate recombination, insertions, and deletions, as has been described in other coronaviruses. Here, we deep-sequenced the complete SARS-CoV-2 S gene from 18 patients (10 with mild and 8 with severe COVID-19), and found that the virus accumulates deletions upstream and very close to the S1/S2 cleavage site (PRRAR/S), generating a frameshift with appearance of a stop codon. These deletions were found in a small percentage of the viral quasispecies (2.2%) in samples from all the mild and only half the severe COVID-19 patients. Our results suggest that the virus may generate free S1 protein released to the circulation. We suggest that natural selection has favored a “Don’t burn down the house” strategy, in which free S1 protein may compete with viral particles for the ACE2 receptor, thus reducing the severity of the infection and tissue damage without losing transmission capability.

Antifungal resistance is increasing by the emergence of intrinsically resistant species and by the development of secondary resistance in susceptible species. A previous study performed in Spain revealed levels of azole resistance in molds between 10 to 12.7% but secondary resistance in A. fumigatus was not detected. We used itraconazole supplemented media to select resistant strains. A total of 500 plates supplemented with 2 mg/L of ITZ were sent to 10 Spanish tertiary hospitals, molecular identification and antifungal susceptibility testing were performed. In addition, cyp51A gene was sequenced in those A. fumigatus strains showing azole resistance. A total of 493 isolates were included in the study. Sixteen strains were isolated from patients with an infection classified as proven, 104 as probable and 373 as colonization. Aspergillus was the most frequent genera isolated with 80.3% followed by Scedosporium/Lomentospora (7.9%), Penicillium/Talaromyces (4.5%), Fusarium (2.6 %) and the order Mucorales (1 %). Antifungal resistance was detected in Scedosporium/Lomentospora species, Fusarium, Talaromyces and Mucorales. Three strains of A. fumigatus senso stricto were resistant to azoles, two of them harbored TR₃₄+L98H mechanism of resistance and the other one had no mutations in cyp51A. The level of azole resistance in A. fumigatus remains low but cryptic species represent over 10% of the isolates and have broader but overall higher range of antifungal resistance

A remarkable percentage of acute gastroenteritis cases remain etiologically undiagnosed. The aim of the study was to determine the prevalence of common and emerging enteric viruses, such as novel human astroviruses, among undiagnosed samples from children with acute gastroenteritis. Epidemiological studies for novel human astroviruses are still scarce. Stool samples collected over two consecutive winter seasons (2016–2017) from children with gastroenteritis in Spain, which were negative for bacteria, rotavirus, and adenovirus by routine diagnostics were screened by real-time RT-PCR assays for the presence of classical and novel astrovirus, rotavirus, norovirus GI and GII, sapovirus, and adenovirus. Overall, 220/384 stool samples (57.3%) were positive for at least one virus. Co-infections were identified in 21% of cases. Among a total of 315 viruses identified, adenovirus was the most prevalent (n = 103), followed by rotavirus (n = 51), sapovirus (n = 50), classical astrovirus (n = 43), novel astroviruses (n = 42), and norovirus (n = 26). Novel astroviruses were present in 13.3% of virus-positive cases. Most novel astroviruses were found in children <2-year-old (30/39 children, 77%, p = 0.01) and were found in co-infection (66%). Only classical astroviruses demonstrated significant differences in the Cq values during mono-infections compared to co-infections. In conclusion, common enteric viruses may be frequently found in children with undiagnosed gastroenteritis, indicating the need to implement more sensitive diagnostic methods. Novel astroviruses circulate in the community and could be the cause of gastroenteritis among young children.

Antitrypanosomal treatment with Benznidazole (BZ) or Nifurtimox may be recommended for patients with chronic Chagas disease (CD) to reduce the onset or progression of symptoms. However, such treatment has limited efficacy and high level of toxic effects. In addition, the current cure biomarker (serology conversion) precludes any treatment assessment unless a prolonged follow-up is arranged. PCR is thus the most useful, alternative surrogate marker for evaluating responses to treatment. The aim of this study is to describe the usefulness of real-time PCR in monitoring BZ treatment within a large cohort of chronic CD cases in Barcelona. A total of 370 chronic CD patients were monitored with real-time PCR post-BZ treatment. The median follow-up was 4 years (IQR 2.2–5.3y), with a median of 3 clinical visits (IQR 2–4). Only 8 patients (2.2%) presented with at least one incident of positive real-time PCR after treatment and were therefore considered as treatment failure. Four of those failure patients had completed full course treatment, whereas the remaining cases had defaulted with a statistical difference between both groups (p = 0.02). Half of the failure patients had undergone less than 4 years of follow-up monitoring all presented with parasitemia before treatment. BZ treatment failure was highly infrequent in our cohort. BZ discontinuation was a risk factor for positive real-time PCR results during clinical follow-up. Regular testing with real-time PCR during follow-up allows for early detection of treatment failure in patients with chronic CD. Chagas disease has become a public health concern for health services in Spain, provided that the destination is the primary choice for Latin American migrants in Europe. Chronic phase is the most frequent clinical form of the disease outside of endemic areas. Most of these patients usually undergo treatment with Benznidazole, primarily childbearing women. Monitoring response to antiparasitic treatment without the presence of an early cure biomarker poses as one of the greatest challenges in managing this disease. In this study, almost 400 chronic Chagas disease patients treated with Benznidazole in two referral centers in Barcelona were included. Real-time PCR and serology were performed in all clinical visits after treatment; real-time PCR has been used as a surrogate biomarker to assess response to treatment. The failure rate of Benznidazole was very low in our cohort, with treatment discontinuation being a risk factor for such failure. In summary, our data supports that regular testing by real- time PCR for monitoring treatment response in chronic Chagas disease patients could be a useful surrogate marker for failure in such individuals.

Pneumocystis jirovecii pneumonia (PJP) is an important cause of pneumonia in the HIV-negative immunocompromised population, for whom the fungal load is low, the differential diagnosis is difficult, and a bronchoalveolar lavage (BAL) sample is often not readily available. Molecular techniques have improved the microbiological diagnosis in this scenario. The usefulness of two real-time PCR techniques targeting nuclear single-copy and mitochondrial multicopy genes, respectively, applied to oral wash specimens (OW) for PJP diagnosis was assessed, and its accuracy was compared to a BAL fluid-based diagnosis. Immunocompromised patients having PJP in the differential diagnosis of an acute respiratory episode, and from whom OW and BAL or lung biopsy specimens were obtained ≤48 h apart, were retrospectively included. PCRs targeting the dihydropteroate synthase gene (DHPS) and the mitochondrial small-subunit (mtSSU) rRNA gene were performed in paired OW-BAL specimens. Thirty-six patients were included (88.6% HIV negative). Fifteen patients (41.7%) were classified as PJP, and a further 8 were considered P. jirovecii colonized. Quantification of DHPS and mtSSU in BAL fluid showed an accuracy of 96.9% and 93.0%, respectively, for PJP diagnosis, whereas a qualitative approach performed better when applied to OW (accuracy, 91.7%) irrespective of the PCR target studied (kappa = 1). Qualitative molecular diagnosis applied to OW showed an excellent performance for PJP diagnosis regardless of the target studied, being easier to interpret than the quantitative approach needed for BAL fluid.