Shotgun metagenomics, when sufficient read depth exists for each taxon, enables capturing metagenome-assembled genomes (MAGs) directly from a microbial community. In 2021, an aromatherapy spray contaminated with Burkholderia pseudomallei caused an outbreak of melioidosis in the United States. Metagenome-assembled genome binning depends in part on different nucleotide compositions, and the contaminated aromatherapy spray contained other bacteria, including related species (e.g., Burkholderia cepacia, Burkholderia cenocepacia, Burkholderia multivorans, Burkholderia pseudomultivorans, Cupriavidus pauculus, and Pseudomonas aeruginosa with average nucleotide identity (ANI) to B. pseudomallei being 84.2%, 84.4%, 84.7%, 84.8%, 75.7%, and 72.3%, respectively, and AAI being 79.5%, 79.8%, 80.5%, 80.4%, 62.5%, and 52.9%, respectively). We performed metagenomic sequencing on the contaminated aromatherapy spray to determine if a public metagenomic pipeline (https://github.com/nf-core/mag) can form a MAG of B. pseudomallei. Upon completion of the pipeline, inter- and intracontig comparisons revealed few potential contaminants of related taxa. Conservative removal of those contigs was especially valuable, ultimately obtaining an ANI of 99.9% between the B. pseudomallei MAG and the genome of an isolate from the aromatherapy spray. This underscores the importance of quality checking recovered MAGs (e.g., for congeneric chimerism) for high-resolution objectives such as outbreak pathogenomics. Importantly, our analysis revealed that the identical conclusion was made possible with the B. pseudomallei MAG (as with its corresponding isolate genome), which was that the aromatherapy B. pseudomallei originated from South Asia (specifically India). Because rapid read-based (k-mer) taxonomic classification methods often report false positives, this operational framework could be valuable for rapid biothreat radar detection systems in public health surveillance.IMPORTANCEIn 2021, an imported aromatherapy spray caused a U.S. outbreak of melioidosis after contamination with Burkholderia pseudomallei. Using shotgun metagenomics, we reconstructed a near-complete genome of the pathogen directly from the product, despite the presence of other related microbes. The assembled genome showed 99.9% similarity to a cultured isolate. This work demonstrates that metagenomics can recover high-quality pathogen genomes from complex samples, supporting outbreak investigations and enhancing public health surveillance.

Endocarditis from melioidosis is rarely reported; only 14 cases have been reported in the literature. We present a fatal case of native aortic valve endocarditis due to Burkholderia pseudomallei complicated by embolic stroke and subdural empyema that occurred in a traveler who returned from Thailand to the United States.

Melioidosis, caused by Burkholderia pseudomallei, is an emerging disease in the United States and was declared endemic to the Gulf Coast region in 2022. Melioidosis has been sporadically reported in the American continents, the Caribbean, and more recently in Africa. We conducted lethal dose analyses in BALB/c and C57BL/6 mice exposed to small particle aerosols of B. pseudomallei strains from the Western Hemisphere and Africa. Those isolates exhibit a variety of virulence patterns, including rapidly-lethal disease and delayed onset of fatal disease. We found that the isolates we tested grew similarly in culture but displayed major differences in biofilm formation. Our data contribute to the growing knowledge of geographically distinct B. pseudomallei isolates and aid in ensuring that medical countermeasures in development are effective against a diverse collection of bacterial strains.

Infection with Burkholderia pseudomallei, the causative agent of melioidosis, is uncommon in the United States (U.S.), leading to delays in pathogen identification and clinical diagnosis which can often lead to laboratory exposures. The indirect hemagglutination assay (IHA) is the primary serological test for confirming exposure to B. pseudomallei. In the U.S., a titer of ≥1:40 suggests exposure to B. pseudomallei or a closely related species, and a 4-fold rise in IHA titer ≥1:40 with clinically compatible illness is considered diagnostically probable. A retrospective analysis of 160 voluntarily reported laboratory exposure events to B. pseudomallei across 29 U.S. jurisdictions and 5 countries between 2008–2024 was conducted. This analysis included post-exposure management data and IHA results for 855 exposed laboratory personnel who had serological monitoring performed at the U.S. Centers for Disease Control and Prevention (CDC). Among exposed laboratory personnel, 105 (12%) had a seropositive titer. Of these, ninety-one (87%) laboratory personnel remained seropositive (≥1:40) at their last IHA test. Five (1%) people had a 4-fold rise in titers, though none developed melioidosis. This report underscores the need for prospective studies to evaluate seropositive laboratory personnel and to update risk guidance for laboratory exposures in non-endemic areas.

Leptospirosis is a worldwide zoonosis caused by pathogenic Leptospira spp. with small rodents serving as the main reservoir. In Croatia, the serogroup Pomona has been detected most frequently, but its genomic diversity remains insufficiently characterized. This study presents the first whole genome sequencing analysis of 48 Croatian Leptospira spp. isolates collected from small rodents over a 14-year period. Serological typing confirmed that all the isolates belonged to the serogroup Pomona. Genomic analysis assigned them to L. kirschneri based on high genomic similarity using average nucleotide identity (ANI). The isolates were assigned to ST-98 using traditional multilocus sequence typing (MLST), while cgMLST identified seven genotype clusters, many of which showed geographic structuring. Phylogenetic analyses based on single nucleotide polymorphisms (SNPs) supported this structure and revealed a monophyletic clade of Croatian isolates distinct from other global L. kirschneri strains. Serological typing, MLST, and phylogenetic clustering support classification of the isolates as L. kirschneri, serogroup Pomona, most likely serovar Mozdok, although serovar Tsaratsovo cannot be excluded. These results indicate the existence of a geographically restricted and potentially host-adapted lineage of L. kirschneri in Croatia. The integration of ecological, serological, and genomic data in this study emphasizes the value of whole genome sequencing for understanding the population biology of Leptospira spp. serogroup Pomona. Moreover, it supports targeted, country-specific surveillance and control strategies for leptospirosis through the identification of circulating serovars and species in reservoir hosts, in line with a One Health approach.

We identified 4 cases of presumptive autochthonous melioidosis during 1983-2024 in Georgia, USA. Epidemiologic investigation identified no recent international travel before illness; all cases were geographically linked, and 3 patients became ill after a severe weather event. Bioinformatic analyses revealed Burkholderia pseudomallei genome sequences were highly related, suggesting a shared exposure.

Melioidosis is a fatal bacterial infection caused by a soil-borne bacterium Burkholderia pseudomallei. Pneumonia, localized abscess and acute septicemia are the most common clinical presentations for melioidosis. Septic arthritis is relatively uncommon clinical manifestation and may cause severe infection. Here, we report a melioidosis case of a 60-year old diabetic male presenting with fever and acute hip pain. B. pseudomallei was isolated from blood culture and it was sensitive to meropenem, amoxicillin-clavulanic acid, cotrimoxazole, ceftazidime and doxycycline. The genome sequence from the clinical isolate obtained from this patient clusters with other Bangladeshi strains and it is within the major clade of East and Southeast Asia strains. The patient received parental meropenem for four weeks followed by six months of oral cotrimoxazole and doxycycline. We followed up the patients every three months over the past seven months. After seven months of oral antibiotic therapy, the patient fully recovered without feeling pain in hip joint and no further complications. His blood culture was tested negative after seven months. The findings of the case report are valuable for raising clinical suspicion and prompting laboratory testing to detect melioidosis. The report also detailed the disease progression and the response to antibiotic therapy.

Bacillus anthracis causes anthrax through virulence factors encoded on two plasmids. However, non-B. anthracis organisms within the closely related, environmentally ubiquitous Bacillus cereus group (BCG) may cause an anthrax-like disease in humans through the partial adoption of anthrax-associated virulence genes, challenging the definition of anthrax disease. To elucidate these phenomena and their evolutionary past, we performed whole-genome sequencing on non-anthracis BCG isolates, including 93 archival (1967-2003) and 5 contemporary isolates (2019-2023). We produced annotated genomic assemblies and performed a pan-genome analysis to identify evidence of virulence gene homology and virulence gene acquisition by linear inheritance or horizontal gene transfer. At least one anthrax-associated virulence gene was annotated in ten isolates. Most homologous sequences in archival isolates showed evidence of pseudogenization and subsequent gene loss. The presence or absence of accessory genes, including anthrax-associated virulence genes, aligned with the phylogenetic structure of the BCG core genome. These findings support the hypothesis that anthrax-associated virulence genes were inherited from a common ancestor in the BCG and were retained or lost across different lineages, and contribute to a growing body of work informing public health strategies related to anthrax surveillance and identification.

In the United States in 2021, an outbreak of 4 cases of Burkholderia pseudomallei, the etiologic agent of melioidosis and a Tier One Select Agent (potential for deliberate misuse and subsequent harm), resulted in 2 deaths. The causative strain, B. pseudomallei ATS2021, was unintentionally imported into the United States in an aromatherapy spray manufactured in India. We established that ATS2021 represents a virulent strain of B. pseudomallei capable of robust formation of biofilm at physiologic temperatures that may contribute to virulence. By using mouse melioidosis models, we determined median lethal dose estimates and analyzed the bacteriologic and histopathologic characteristics of the organism, particularly the potential neurologic pathogenesis that is probably associated with the bimABm allele identified in B. pseudomallei strain ATS2021. Our data, combined with previous case reports and the identification of endemic B. pseudomallei strains in Mississippi, support the concept that melioidosis is emerging in the United States.

Burkholderia thailandensis, an opportunistic pathogen found in the environment, is a bacterium closely related to B. pseudomallei, the cause of melioidosis. Human B. thailandensis infections are uncommon. We isolated B. thailandensis from water in Texas and Puerto Rico and soil in Mississippi in the United States, demonstrating a potential public health risk.

Melioidosis, caused by Burkholderia pseudomallei, is a rare but potentially fatal bacterial disease endemic to tropical and subtropical regions worldwide. It is typically acquired through contact with contaminated soil or fresh water. Before this investigation, B. pseudomallei was not known to have been isolated from the environment in the continental United States. Here, we report on three patients living in the same Mississippi Gulf Coast county who presented with melioidosis within a 3-year period. They were infected by the same Western Hemisphere B. pseudomallei strain that was discovered in three environmental samples collected from the property of one of the patients. These findings indicate local acquisition of melioidosis from the environment in the Mississippi Gulf Coast region.

Melioidosis, endemic in Southeast Asia and Northern Australia, is an uncommon but frequently fatal opportunistic infection caused by the Gram-negative saprophyte Burkholderia pseudomallei. We describe the first reported case of activation of latent melioidosis concurrent with COVID-19-associated lymphopenia and neutropenia in the setting of poorly controlled diabetes. A 43-year-old HIV-positive, diabetic man presented to the emergency department with persistent chills and progressive dyspnea. He was admitted for hypoxia. Chest X-ray showed bilateral parenchymal infiltrates suspicious for COVID-19. Shortly after admission, he became acutely encephalopathic, had a generalized seizure, and was transferred to the intensive care unit after intubation. Further workup showed severe neutropenia and lymphopenia. The patient received empiric antimicrobial coverage and was found to be severe acute respiratory syndrome coronavirus 2 positive. He deteriorated rapidly with refractory shock and persistent hypoxemia, and died 40 hours after admission. Blood cultures and sputum cultures obtained via bronchoalveolar lavage returned positive for Burkholderia pseudomallei. Given confirmed compliance with antiretrovirals, stable CD4 counts, and no recent foreign travel, the patient likely contracted the B. pseudomallei infection from travel to Southeast Asia many years prior and only became symptomatic after succumbing to severe acute respiratory syndrome coronavirus 2 infection. This case highlights the importance of considering activation of latent opportunistic infections by COVID-19 in immunocompromised patients.

Bacillus cereus group bacteria containing the anthrax toxin genes can cause fatal anthrax pneumonia in welders. Two welder’s anthrax cases identified in 2020 were investigated to determine the source of each patient’s exposure. Environmental sampling was performed at locations where each patient had recent exposure to soil and dust. Samples were tested for the anthrax toxin genes by real-time PCR, and culture was performed on positive samples to identify whether any environmental isolates matched the patient’s clinical isolate. A total of 185 environmental samples were collected in investigation A for patient A and 108 samples in investigation B for patient B. All samples from investigation B were real-time PCR-negative, but 14 (8%) samples from investigation A were positive, including 10 from patient A’s worksite and 4 from his work-related clothing and gear. An isolate genetically matching the one recovered from patient A was successfully cultured from a worksite soil sample. All welder’s anthrax cases should be investigated to determine the source of exposure, which may be linked to their worksite. Welding and metalworking employers should consider conducting a workplace hazard assessment and implementing controls to reduce the risk of occupationally associated illnesses including welder’s anthrax.

Melioidosis, a potentially fatal infectious disease of humans and animals, including nonhuman primates (NHPs), is caused by the high-consequence pathogen Burkholderia pseudomallei. This environmental bacterium is found in the soil and water of tropical regions, such as Southeast Asia, where melioidosis is endemic. The global movement of humans and animals can introduce B. pseudomallei into nonendemic regions of the United States, where environmental conditions could allow establishment of the organism. Approximately 60% of NHPs imported into the United States originate in countries considered endemic for melioidosis. To prevent the introduction of infectious agents to the United States, the Centers for Disease Control and Prevention (CDC) requires newly imported NHPs to be quarantined for at least 31 d, during which time their health is closely monitored. Most diseases of public health concern that are transmissible from imported NHPs have relatively short incubation periods that fall within the 31-d quarantine period. However, animals infected with B. pseudomallei may appear healthy for months to years before showing signs of illness, during which time they can shed the organism into the environment. Melioidosis presents diagnostic challenges because it causes nonspecific clinical signs, serologic screening can produce unreliable results, and culture isolates are often misidentified on rapid commercial testing systems. Here, we present a case of melioidosis in a cynomolgus macaque (Macaca fascicularis) that developed a subcutaneous abscess after importation from Cambodia to the United States. The bacterial isolate from the abscess was initially misidentified on a commercial test. This case emphasizes the possibility of melioidosis in NHPs imported from endemic countries and its associated diagnostic challenges. If melioidosis is suspected, diagnostic samples and culture isolates should be submitted to a laboratory in the CDC Laboratory Response Network for conclusive identification and characterization of the pathogen.

Melioidosis is an underreported human disease of tropical and sub-tropical regions caused by the saprophyte Burkholderia pseudomallei. Although most global melioidosis cases are reported from tropical regions in Southeast Asia and northern Australia, there are multiple occurrences from sub-tropical regions, including the United States (U.S.). Most melioidosis cases reported from the continental U.S. are the result of acquiring the disease during travel to endemic regions or from contaminated imported materials. Only two human melioidosis cases from the continental U.S. have likely acquired B. pseudomallei directly from local environments and these cases lived only ~7 km from each other in rural Texas. In this study, we assessed the risk of acquiring melioidosis from the environment within the continental U.S. by surveying for B. pseudomallei in the environment in Texas where these two human melioidosis cases likely acquired their infections. We sampled the environment near the homes of the two cases and at additional sampling locations in surrounding counties in Texas that were selected based on ecological niche modeling. B. pseudomallei was not detected at the residences of these two cases or in the surrounding region. These negative data are important to demonstrate that B. pseudomallei is rare in the environment in the U.S. even at locations where locally acquired human cases likely have occurred, documenting the low risk of acquiring B. pseudomallei infection from the environment in the continental U.S.

Anthrax-causing members of Bacillus cereus sensu lato (s.l.) pose a serious threat to public health. While most anthrax-causing strains resemble B. anthracis phenotypically, rare cases of anthrax-like illness caused by strains resembling "B. cereus" have been reported. Here, whole-genome sequencing was used to characterize three B. cereus s.l. isolates associated with two 2020 welder anthrax cases in the United States, which resembled "B. cereus" phenotypically. Comparison of the three genomes sequenced here to all publicly available, high-quality B. cereus s.l. genomes (n = 2890 total genomes) demonstrated that genomes associated with each case effectively belonged to separate species at the conventional 95% average nucleotide identity prokaryotic species threshold. Two PubMLST sequence type 78 (ST78) genomes affiliated with a case in Louisiana were most closely related to B. tropicus and possessed genes encoding the Bps exopolysaccharide capsule, as well as hemolysin BL (Hbl) and cytotoxin K (CytK). Comparatively, a ST108 genome associated with a case in Texas was most closely related to B. anthracis; however, like other anthrax-causing strains most closely related to B. anthracis, this genome did not possess Bps-, Hbl-, or CytK-encoding genes. Overall, results presented here provide insights into the evolution of anthrax-causing B. cereus s.l.

3% We report an analysis of the genomic diversity of isolates of Burkholderia pseudomallei , the cause of melioidosis, recovered in Colombia from routine surveillance during 2016–2017. B. pseudomallei appears genetically di -verse, suggesting it is well established and has spread across the region.

Melioidosis, caused by the bacterium Burkholderia pseudomallei, is an uncommon infection that is typically associated with exposure to soil and water in tropical and subtropical environments. It is rarely diagnosed in the continental United States. Patients with melioidosis in the United States commonly report travel to regions where melioidosis is endemic. We report a cluster of four non-travel-associated cases of melioidosis in Georgia, Kansas, Minnesota, and Texas. These cases were caused by the same strain of B. pseudomallei that was linked to an aromatherapy spray product imported from a melioidosis-endemic area.

Nearly all cases of melioidosis in the continental United States are related to international travel to areas to which Burkholderia pseudomallei, the bacterium that causes melioidosis, is endemic. We report the diagnosis and clinical course of melioidosis in a patient from the United States who had no international travel history and the public health investigation to determine the source of exposure. We tested environmental samples collected from the patient's home for B. pseudomallei by PCR and culture. Whole-genome sequencing was conducted on PCR-positive environmental samples, and results were compared with sequences from the patient's clinical specimen. Three PCR-positive environmental samples, all collected from a freshwater home aquarium that had contained imported tropical fish, were a genetic match to the clinical isolate from the patient. This finding suggests a novel route of exposure and a potential for importation of B. pseudomallei, a select agent, into the United States from disease-endemic areas.