Early SARS-CoV-2 Reinfections Involving the Same or Different Genomic Lineages, Spain.

Centers for Disease Control and Prevention guidelines consider SARS-CoV-2 reinfection when sequential COVID-19 episodes occur >90 days apart. However, genomic diversity acquired over recent COVID-19 waves could mean previous infection provides insufficient cross-protection. We used genomic analysis to assess the percentage of early reinfections in a sample of 26 patients with 2 COVID-19 episodes separated by 20-45 days. Among sampled patients, 11 (42%) had reinfections involving different SARS-CoV-2 variants or subvariants. Another 4 cases were probable reinfections; 3 involved different strains from the same lineage or sublineage. Host genomic analysis confirmed the 2 sequential specimens belonged to the same patient. Among all reinfections, 36.4% involved non-Omicron, then Omicron lineages. Early reinfections showed no specific clinical patterns; 45% were among unvaccinated or incompletely vaccinated persons, 27% were among persons <18 years of age, and 64% of patients had no risk factors. Time between sequential positive SARS-CoV-2 PCRs to consider reinfection should be re-evaluated.

Correction: Díaz-García et al. Candida Genotyping of Blood Culture Isolates from Patients Admitted to 16 Hospitals in Madrid: Genotype Spreading during the COVID-19 Pandemic Driven by Fluconazole-Resistant C. parapsilosis. J. Fungi 2022, 8, 1228.

In the original publication [...].

SARS-CoV-2 superinfection and reinfection with three different strains.

We report a corona virus disease (COVID-19) case with unprecedented viral complexity. In the first severe episode, two different severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) strains (superinfection) were identified within a week. Three months after discharge, the patient was readmitted and was infected in a nosocomial outbreak with a different strain, suffering a second milder COVID-19 episode.

Candida Genotyping of Blood Culture Isolates from Patients Admitted to 16 Hospitals in Madrid: Genotype Spreading during the COVID-19 Pandemic Driven by Fluconazole-Resistant C. parapsilosis.

BACKGROUND: Candidaemia and invasive candidiasis are typically hospital-acquired. Genotyping isolates from patients admitted to different hospitals may be helpful in tracking clones spreading across hospitals, especially those showing antifungal resistance. METHODS: We characterized Candida clusters by studying Candida isolates (C. albicans, n = 1041; C. parapsilosis, n = 354, and C. tropicalis, n = 125) from blood cultures (53.8%) and intra-abdominal samples (46.2%) collected as part of the CANDIMAD (Candida in Madrid) study in Madrid (2019-2021). Species-specific microsatellite markers were used to define the genotypes of Candida spp. found in a single patient (singleton) or several patients (cluster) from a single hospital (intra-hospital cluster) or different hospitals (widespread cluster). RESULTS: We found 83 clusters, of which 20 were intra-hospital, 49 were widespread, and 14 were intra-hospital and widespread. Some intra-hospital clusters were first detected before the onset of the COVID-19 pandemic, but the number of clusters increased during the pandemic, especially for C. parapsilosis. The proportion of widespread clusters was significantly higher for genotypes found in both compartments than those exclusively found in either the blood cultures or intra-abdominal samples. Most C. albicans- and C. tropicalis-resistant genotypes were singleton and presented exclusively in either blood cultures or intra-abdominal samples. Fluconazole-resistant C. parapsilosis isolates belonged to intra-hospital clusters harboring either the Y132F or G458S ERG11p substitutions; the dominant genotype was also widespread. CONCLUSIONS: the number of clusters-and patients involved-increased during the COVID-19 pandemic mainly due to the emergence of fluconazole-resistant C. parapsilosis genotypes.

Clostridioides difficile infection epidemiology and clinical characteristics in COVID-19 pandemic.

Information on Clostridioides difficile infection (CDI) in patients with COVID-19 is scarce and points to an overall decrease of episodes during the pandemic. This situation results paradoxical, as COVID-19 patients had long periods of hospital stay and high use of antibiotics. We conducted a retrospective study from January 1st 2019 to December 31st 2020 comparing the incidence of hospital-acquired episodes of CDI (HA-CDI) among patients with and without COVID-19 admitted to our institution. During the study period, there were 47,048 patient admissions in 2019, 35,662 admissions of patients without COVID-19 in 2020 and 6,763 of COVID-19 patients. There were 68 episodes of HA-CDI in COVID-19 patients (14.75/10,000 days), 159 in 2020-non-COVID-19 patients (5.54/10,000 days) and 238 in 2019 (6.80/10,000 days). Comparison of HA-CDI in COVID-19 and non-COVID-19 patients indicates it occurs more frequently, in terms of CDI disease severity, COVID-19 does not seem to have a negative impact.

First importations of SARS-CoV-2 P.1 and P.2 variants from Brazil to Spain and early community transmission.

INTRODUCTION: SARS-CoV-2variants of concern (VOC) have been described in the UK (B.1.1.7), South Africa (B.1.351) and Brazil (P.1). Among them, the most scarce information has been obtained from the P.1 variant and more data on its global presence and about its spreading dynamics are needed. METHODS: Whole genome sequencing was performed prospectively on travellers arriving from Brazil and on a random selection of SARS-CoV-2 positive cases from our population. RESULTS: In this study we report the first SARS-CoV-2 P.1 and P.2 variants exported from Brazil to Spain. The case infected with the P.1 variant, who had only stayed in Rio de Janeiro, required hospitalisation. The two P.2 cases remained asymptomatic. A wider distribution for P.1 variant beyond the Brazilian Amazonia should be considered. The exportation of the P.2 variant, carrying the E484K mutation, deserves attention. One month after the first description of P.1 and P.2 importations from Brazil to Madrid, these variants were identified circulating in the community, in cases without a travel history, and involved in household transmissions CONCLUSION: Whole genome sequencing of SARS-CoV-2 positive travellers arriving from Brazil allowed us to identify the first importations of P.1 and P.2 variants to Spain and their early community transmission.

First importations of SARS-CoV-2 P.1 and P.2 variants from Brazil to Spain and early community transmission Primeras importaciones de las variantes SARS-CoV-2 P.1 y P.2 de Brasil a Madrid y transmisión comunitaria

Introduction SARS-CoV-2variants of concern (VOC) have been described in the UK (B.1.1.7), South Africa (B.1.351) and Brazil (P.1). Among them, the most scarce information has been obtained from the P.1 variant and more data on its global presence and about its spreading dynamics are needed. Methods Whole genome sequencing was performed prospectively on travellers arriving from Brazil and on a random selection of SARS-CoV-2 positive cases from our population. Results In this study we report the first SARS-CoV-2 P.1 and P.2 variants exported from Brazil to Spain. The case infected with the P.1 variant, who had only stayed in Rio de Janeiro, required hospitalisation. The two P.2 cases remained asymptomatic. A wider distribution for P.1 variant beyond the Brazilian Amazonia should be considered. The exportation of the P.2 variant, carrying the E484K mutation, deserves attention. One month after the first description of P.1 and P.2 importations from Brazil to Madrid, these variants were identified circulating in the community, in cases without a travel history, and involved in household transmissions Conclusion Whole genome sequencing of SARS-CoV-2 positive travellers arriving from Brazil allowed us to identify the first importations of P.1 and P.2 variants to Spain and their early community transmission.

Systematic Genomic and Clinical Analysis of Severe Acute Respiratory Syndrome Coronavirus 2 Reinfections and Recurrences Involving the Same Strain.

Estimates of the burden of severe acute respiratory syndrome coronavirus 2 reinfections are limited by the scarcity of population-level studies incorporating genomic support. We conducted a systematic study of reinfections in Madrid, Spain, supported by genomic viral analysis and host genetic analysis, to cleanse laboratory errors and to discriminate between reinfections and recurrences involving the same strain. Among the 41,195 cases diagnosed (March 2020-March 2021), 93 (0.23%) had 2 positive reverse transcription PCR tests (55-346 days apart). After eliminating cases with specimens not stored, of suboptimal sequence quality, or belonging to different persons, we obtained valid data from 22 cases. Of those, 4 (0.01%) cases were recurrences involving the same strain; case-patients were 39-93 years of age, and 3 were immunosuppressed. Eighteen (0.04%) cases were reinfections; patients were 19-84 years of age, and most had no relevant clinical history. The second episode was more severe in 8 cases.

Different dynamics of mean SARS-CoV-2 RT-PCR Ct values between the first and second COVID-19 waves in the Madrid population.

SARS-CoV-2 RT-PCR cycle threshold values from 18,803 cases (2 March-4 October) in Madrid define three stages: (i) initial ten weeks with sustained reduction in viral load (Ct: 23.4-32.3), (ii) stability with low viral loads (Ct: 31.9-35.5) in the next nine weeks and (iii) sudden increase with progressive higher viral loads until reaching stability at high levels in the next twelve weeks, coinciding with an increased percentage of positive cases and reduced median age. These data indicate differential virological/epidemiological patterns between the first and second COVID-19 waves in Madrid.

SARS-CoV-2 B.1.1.7 Decline Is Not Driven by the Introduction of a More Successful Variant.

The SARS-CoV-2 variant of concern (VOC) Delta (B.617.2 lineage) displaced the predominant VOC Alpha (B.1.1.7 lineage) in the United Kingdom. In Madrid, recent start of the decline of predominant VOC Alpha suggested an equivalent phenomenon. However, 11 different variants, none overrepresented in frequency, occupied progressively over a period of 7 weeks the niche previously dominated by VOC Alpha. Only after these 7 weeks, VOC Delta started to emerge. Viral competition due to the entry of VOC Delta is not the major force driving the start of VOC Alpha decline in Madrid. IMPORTANCE Our data indicate that the dynamics of SARS-CoV-2 VOCs turnover in our setting differ from those proposed for other countries. A systematic genomic analysis, updated on a weekly basis, of representative randomly selected samples of SARS-CoV-2 circulating variants allowed us to define a lapse of 7 weeks between the start of VOC Alpha decline and the final emergence of VOC Delta. During this period, VOC Alpha showed a sustained decline, while 11 VOCs, variants of interest (VOIs), and other identified variants, none overrepresented, occupied the niche left by VOC Alpha. Only after these 7 weeks, emergence of VOC Delta occurred, indicating that viral competition involving VOC Delta was not the exclusive direct driving force behind the starting of VOC Alpha decline.

First importations of SARS-CoV-2 P.1 and P.2 variants from Brazil to Spain and early community transmission.

Introduction: SARS-CoV-2variants of concern (VOC) have been described in the UK (B.1.1.7), South Africa (B.1.351) and Brazil (P.1). Among them, the most scarce information has been obtained from the P.1 variant and more data on its global presence and about its spreading dynamics are needed. Methods: Whole genome sequencing was performed prospectively on travellers arriving from Brazil and on a random selection of SARS-CoV-2 positive cases from our population. Results: In this study we report the first SARS-CoV-2 P.1 and P.2 variants exported from Brazil to Spain. The case infected with the P.1 variant, who had only stayed in Rio de Janeiro, required hospitalisation. The two P.2 cases remained asymptomatic. A wider distribution for P.1 variant beyond the Brazilian Amazonia should be considered. The exportation of the P.2 variant, carrying the E484K mutation, deserves attention. One month after the first description of P.1 and P.2 importations from Brazil to Madrid, these variants were identified circulating in the community, in cases without a travel history, and involved in household transmissions. Conclusion: Whole genome sequencing of SARS-CoV-2 positive travellers arriving from Brazil allowed us to identify the first importations of P.1 and P.2 variants to Spain and their early community transmission.

Introducción: Se han descrito «variantes de preocupación¼ (VOC) de SARS-CoV-2 en el Reino Unido (B.1.1.7), Sudáfrica (B.1.351) y Brasil (P.1). Entre ellas, se dispone de información más escasa para la variante P.1 y se necesitan más datos sobre su presencia global y sobre su dinámica de expansión. Métodos: Se realizó secuenciación del genoma completo de forma prospectiva de SARS-CoV-2 en viajeros procedentes de Brasil y en una selección aleatoria de casos positivos de SARS-CoV-2 de nuestra población. Resultados: En este estudio reportamos las primeras variantes de SARS-CoV-2 P.1 y P.2 exportadas desde Brasil a España. El caso infectado por la variante P.1, que solo había permanecido en Río de Janeiro, requirió hospitalización. Los 2 casos de la variante P.2 permanecieron asintomáticos. Se debe considerar una distribución más amplia para la variante P.1 más allá de la Amazonía brasileña. La exportación de la variante P.2, que porta la mutación E484K, merece asimismo atención adicional. Un mes después de la primera descripción de las importaciones de P.1 y P.2 de Brasil a Madrid, se identificaron estas variantes circulando en la comunidad, en casos sin antecedentes de viaje, e implicadas en transmisiones domiciliarias. Conclusión: La secuenciación de genoma completo de viajeros positivos para SARS-CoV-2 procedentes de Brasil nos permitió identificar las primeras importaciones de variantes P.1 y P.2 a España y su transmisión comunitaria precoz.

The challenge of COVID-19 for a Clinical Microbiology Department.

OBJECTIVES: To quantify the workload and cost overload that the COVID-19 pandemic has meant for a Clinical Microbiology laboratory in a real-life scenario. METHODS: We compared the number of samples received, their distribution, the human resources, and the budget of a Microbiology laboratory in the COVID pandemic (March-December 2020) with the same months of the previous year. RESULTS: the total number of samples processed in the Clinical Microbiology laboratory in March to December 2020 increased 96.70% with respect to 2019 (from 246,060 to 483,993 samples), reflecting an increment of 127.50% when expressed as samples/1000 admissions (from 6057 to 13,780). The increase in workload was mainly at the expense of the virology (+2058%) and serology (+86%) areas. Despite additional personnel hiring, the samples processed per technician increased 12.5%. The extra cost attributed to Microbiology amounts to 6,616,511 euros (114.8%). CONCLUSIONS: This is the first study to provide quantitative figures about workload and cost increase caused by the COVID-19 in a Microbiology laboratory.

Sample pooling is efficient in PCR testing of SARS-CoV-2: a study in 7400 healthcare professionals.

SARS-CoV-2 pandemic shows the importance of having efficient virus diagnosis, especially in groups of particular relevance such as health care professionals, without involving a large economic expense. This is a prevalence study carried out in 7400 health care professionals in a 1350-bed hospital in Madrid, Spain. Pools of 10 samples were performed, using the Xpert® Xpress SARS-CoV-2 test for the diagnosis from clinical samples of nasopharyngeal exudate. A previous study was performed to evaluate the effect of the dilution in terms of sensitivity. The estimated sensitivity was over 95%. A total of 740 pools were performed, with a final result of 218 health care professionals being positive. Using the pooling system, the reagent cost reduction to the institution was 75.3%. It can be concluded that the described sample pooling system is a useful and efficient tool in the diagnosis of SARS-CoV-2 in certain groups, assuming a cost reduction without reducing the sensitivity.

Inference of Active Viral Replication in Cases with Sustained Positive Reverse Transcription-PCR Results for SARS-CoV-2.

The purpose of this study was to detect coronavirus disease 2019 (COVID-19) cases with persistent positive reverse transcription-PCR (RT-PCR) results for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), for which viable virus can be inferred due to the presence of subgenomic (SG) viral RNA, which is expressed only in replicating viruses. RNA remnants purified from diagnostic nasopharyngeal specimens were used as the templates for RT-PCR-specific detection of SG E gene RNA. As controls, we also detected viral genomic RNA for the E gene and/or a human housekeeping gene (RNase P). We assessed the samples of 60 RT-PCR-positive cases with prolonged viral SARS-CoV-2 shedding (24 to 101 days) since the first diagnostic RT-PCR. SG viral RNA was detected in 12/60 (20%) of the persistent cases, 28 to 79 days after the onset of symptoms. The age range of the cases with prolonged viral shedding and the presence of SG RNA was quite wide (40 to 100 years), and the cases were equally distributed between males (42%) and females (58%). No case was HIV positive, although seven were immunosuppressed. According to the severities of the COVID-19 episodes, they were mild (40%), intermediate (20%), and severe (40%). In a percentage of persistent SARS-CoV-2 PCR-positive cases, the presence of actively replicating virus may be inferred, far beyond diagnosis. We should not assume a universal lack of infectiousness for COVID-19 cases with prolonged viral shedding.

Detection of SARS-CoV-2 antibodies is insufficient for the diagnosis of active or cured COVID-19.

We assessed the performance of Abbott's SARS-CoV-2 IgG assay and the PanbioTM COVID-19 IgG/IgM rapid test device for the diagnosis of either active or cured COVID-19. Three cohorts of patients were chosen. Cohort 1, patients (n = 65) who attended the emergency department on March 30, 2020 with clinical suspicion of active COVID-19 (n = 56 with proven/probable COVID-19). Cohort 2, hospital workers (n = 92) who had either been (n = 40) or not (n = 52) diagnosed with proven/probable COVID-19 and were asymptomatic at the time of the sampling. Cohort 3, patients (n = 38) cared at the hospital before the start of the COVID-19 pandemic. Detection of serum antibodies was done using Abbott´s SARS-CoV-2 IgG assay and the PanbioTM COVID-19 IgG/IgM device. Both methods showed 98% agreement for IgG detection. No antibodies were detected in the 38 samples from hospitalized pre-COVID subjects. The diagnostic performance of IgGs detected by Abbott´s SARS-CoV-2 assay in Cohorts 1/2 was: sensitivity (60.7%/75%) and specificity (100%/84.6%). The diagnostic performance of IgM by PanbioTM COVID-19 in Cohorts 1/2 was: sensitivity (16%/17.5%) and specificity (100%/98.1%). We show that IgG detection alone is insufficient for the diagnosis of active or cured COVID-19. IgM detection has a limited diagnostic value.