Legionellosis in Argentina: evolution of the laboratory diagnosis strategy

Legionnaires’ disease (LD) is severe acute pneumonia that occurs in sporadic or epidemic form and generally requires hospitalization. The bacterium L. pneumophila was first identified in 1977 as the cause of an outbreak of severe pneumonia in a convention centre in the USA in 1976. It has since been associated with outbreaks linked to poorly maintained artificial water systems. Legionella pneumophila serogroup 1 is the principal etiological agent responsible for the most prevalent and virulent legionellosis disease.

In August 2022, a cluster of six cases of bilateral pneumonia with no etiological cause was identified in San Miguel de Tucumán city, Tucuman Province, Argentina and notified to WHO . One private health facility linked all the cases and there was a very tight period covering symptom onset. In addition to bilateral pneumonia, all cases presented with fever, and muscle and abdominal pain. By early September, 11 cases had been identified, and four of them had died. Three of the four deaths were among healthcare workers. Ten of these cases (including the four who died) had underlying conditions and/or risk factors for severe disease.

The local public health laboratory returned negative tests on samples from these individuals for respiratory viruses, and other viral, bacterial, and fungal agents. Samples from the initial cases were sent to the National Reference Laboratory – Administración Nacional de Laboratorios e Institutos de Salud (ANLIS) for additional testing. The results were negative for COVID-19 (RT-PCR), Influenza, detection of antibodies for Coxiella, urinary antigen for Legionella spp., panel of 12 respiratory viruses, hantavirus (Elisa IgM), histoplasma (RT-PCR), Yersinia pestis (PCR) and micro agglutination for leptospirosis.

A metagenomics approach, which involved a highly sensitive total DNA sequencing approach on two bronchoalveolar lavage samples, found readings compatible with Legionella spp. ANLIS subsequently reported that their results were compatible with Legionella pneumophila (after amplifying 16S ribosomal gene products from the samples and using multiple bioinformatics approaches). In summary, despite neither urinary antigen nor validated syndromic PCR panels returning positive results in all the patients in the Tucuman outbreak, targeted metagenomics allowed the rapid identification of Legionella sp. in clinical samples, leading to a prompt environmental risk assessment and intervention.

Culture-independent metagenomics and targeted metagenomics detection of microbial species has the potential to provide rapid and precise real-time diagnostics results. However, it has limitations, including sequencing and taxonomic classification errors and lack of quality standards. Therefore is critical before using clinical metagenomics as a response to a potential outbreak or public health emergency to demonstrate the test validation, reproducibility, and quality assurance.