Respiratory sample staining is a standard tool used to diagnose Pneumocystis jirovecii pneumonia (PjP). Although molecular tests are more sensitive, their interpretation can be difficult due to the potential of colonization. We aimed to validate a Pneumocystis jirovecii (Pj) real-time PCR (qPCR) assay in bronchoscopic bronchoalveolar lavage (BAL) and oropharyngeal washes (OW). We included 158 immunosuppressed patients with pneumonia, 35 lung cancer patients who underwent BAL, and 20 healthy individuals. We used a SYBR green qPCR assay to look for a 103 bp fragment of the Pj mtLSU rRNA gene in BAL and OW. We calculated the qPCR cut-off as well as the analytical and diagnostic characteristics. The qPCR was positive in 67.8% of BAL samples from the immunocompromised patients. The established cut-off for discriminating between disease and colonization was Ct 24.53 for BAL samples. In the immunosuppressed group, qPCR detected all 25 microscopy-positive PjP cases, plus three additional cases. Pj colonization in the immunocompromised group was 66.2%, while in the cancer group, colonization rates were 48%. qPCR was ineffective at diagnosing PjP in the OW samples. This new qPCR allowed for reliable diagnosis of PjP, and differentiation between PjP disease and colonization in BAL of immunocompromised patients with pneumonia.

Objectives: Surveillance of acute respiratory infection (ARI) informs vaccination, preventive, and management decisions. In many countries, immunofluorescence is the cornerstone for ARI surveillance. We aimed to determine the effect of adding multiplex polymerase chain reaction (mPCR) to conventional surveillance in ARI.Methods: Respiratory samples from patients with influenza-like illness (ILI) and severe acute respiratory infection (SARI) were tested by a conventional approach [direct immunofluorescence (DIF) and SARS-CoV-2 PCR, and a subset of samples underwent routine testing]. Negative specimens were tested by multiplex PCR (mPCR), and remain negative samples were sequenced. Descriptive, multivariable regression analyses were conducted.Results: Between March and June 2022, 299 patients were enrolled. Pathogens were detected in 43.8% of samples (131/299) tested by the conventional approach. Of the 168 negatives after the conventional approach, 157 (93.4%) were positive by mPCR, increasing the detection rate to 96.3% (288/299). With the conventional approach, the most frequent pathogen was respiratory syncytial virus (50.3%, 66/131), whereas with mPCR it was Haemophilus influenzae (37.5%, 63/168). mPCR significantly improved pathogen detection in ARI surveillance (Adjusted Incidence Rate Ratios 4.22 95% IC 4.22–5.85).Conclusion: Adding mPCR to respiratory surveillance conventionally based on DIF significantly enhanced virus and bacteria detection. mPCR should be considered for routine ARI surveillance.

Background: Human Adenovirus (HAdV) is a common cause of acute respiratory infections, typically mild in healthy individuals. However, in late 2022, an outbreak of severe acute respiratory infection caused by HAdV emerged among children in Colombia and other countries. Methods: We described an HAdV outbreak between February 2022 and April 2023. Children with severe acute respiratory infection and HAdV infection confirmed by polymerase chain reaction were included in 4 institutions in Antioquia, Colombia. Our study investigated the clinical manifestations and circulating HAdV genotypes before, during and after this HAdV outbreak. Results: A total of 133 HAdV cases were analyzed, 37 (27.8%) cases were classified as the preoutbreak group, 88 (66.1%) as the outbreak and 8 (6.0%) as the postoutbreak group. Predominant symptoms were fever (87.0%), rhinorrhea (57.1%) and dyspnea (36.8%). The need for intensive care unit admission and supplemental oxygen increased during the outbreak and peaked in the postoutbreak period. Phylogenetic analysis revealed that 71.4% (10/14) of preoutbreak sequences belonged to genotype HAdV-C89, while during the outbreak, 75.6% (28/37) were HAdV-B3. Clinical symptoms did not significantly differ between HAdV-C89 and HAdV-B3 infections, but children infected with HAdV-B3 were significantly older. Conclusions: This study highlights the shifting dynamics of HAdV genotypes in children and their epidemiologic impact. The emergence of HAdV-B3 in the post-COVID-19 period contributed to a severe acute respiratory infection outbreak, emphasizing the need for ongoing surveillance.

Tuberculosis (TB) in the pediatric population is a major challenge. Our objective was to describe the clinical and microbiological characteristics, radiological patterns, and treatment outcomes of children and adolescents (from 1 month to 17 years) with community-acquired pneumonia (CAP) caused by TB. We performed a prospective cohort study of a pediatric population between 1 month and 17 years of age and hospitalized in Medellín, Colombia, with the diagnosis of radiologically confirmed CAP that had ≤ 15 days of symptoms. The mycobacterial culture of induced sputum was used for the bacteriological confirmation; the history of TB contact, a tuberculin skin test, and clinical improvement with treatment were used to identify microbiologically negative TB cases. Among 499 children with CAP, TB was diagnosed in 12 (2.4%), of which 10 had less than 8 days of a cough, 10 had alveolar opacities, 9 were younger than 5 years old, and 2 had close contact with a TB patient. Among the TB cases, 50% (6) had microbiological confirmation, 8 had viral and/or bacterial confirmation, one patient had multidrug-resistant TB, and 10/12 had non-severe pneumonia. In countries with an intermediate TB burden, Mycobacterium tuberculosis should be included in the etiological differential diagnosis (as a cause or coinfection) of both pneumonia and severe CAP in the pediatric population.

Human adenovirus (HAdV) is detected in fewer than 10% of hospitalized children with acute respiratory infections, yet several regions reported unusual increases in 2021-2022. We investigated an HAdV outbreak detected through surveillance in Curitiba, Brazil, and Antioquia, Colombia, comparing clinical and genomic characteristics before and during the outbreak. We conducted a multicenter observational study of HAdV-associated severe acute respiratory infection in hospitalized children. Respiratory samples collected between February 2022 and April 2023 underwent metagenomic sequencing and hexon gene analysis. Clinical and phylogenetic analyses evaluated viral dynamics and genetic diversity. A HAdV outbreak occurred in both regions during late 2022. HAdV-C predominated before the outbreak, whereas HAdV-B3 became the dominant genotype at both sites during the outbreak. During the outbreak, bronchiolitis/asthma exacerbation was most common in Antioquia, whereas pneumonia predominated in Curitiba. Phylogenetic analysis of the hexon gene identified two distinct HAdV-B3 clades that diverged from a shared ancestor but expanded independently in Brazil and Colombia. Following relaxation of non-pharmacological measures, both regions experienced a substantial rise in HAdV-B3-associated disease. Despite increased case numbers, clinical patterns remained stable within sites but differed between regions. Integrated genomic and clinical surveillance provides important insights into adenovirus lineage expansion and outbreak dynamics.