RESUMEN
Benzene is a widespread groundwater contaminant that persists under anoxic conditions. The aim of this study was to more accurately investigate anaerobic microbial degradation pathways to predict benzene fate and transport. Preliminary genomic analysis of Geotalea daltonii strain FRC-32, isolated from contaminated groundwater, revealed the presence of putative aromatic-degrading genes. G. daltonii was subsequently shown to conserve energy for growth on benzene as the sole electron donor and fumarate or nitrate as the electron acceptor. The hbs gene, encoding for 3-hydroxybenzylsuccinate synthase (Hbs), a homolog of the radical-forming, toluene-activating benzylsuccinate synthase (Bss), was upregulated during benzene oxidation in G. daltonii, while the bss gene was upregulated during toluene oxidation. Addition of benzene to the G. daltonii whole-cell lysate resulted in toluene formation, indicating that methylation of benzene was occurring. Complementation of σ54- (deficient) E. coli transformed with the bss operon restored its ability to grow in the presence of toluene, revealing bss to be regulated by σ54. Binding sites for σ70 and the transition state regulator AbrB were identified in the promoter region of the σ54-encoding gene rpoN, and binding was confirmed. Induced expression of abrB during benzene and toluene degradation caused G. daltonii cultures to transition to the death phase. Our results suggested that G. daltonii can anaerobically oxidize benzene by methylation, which is regulated by σ54 and AbrB. Our findings further indicated that the benzene, toluene, and benzoate degradation pathways converge into a single metabolic pathway, representing a uniquely efficient approach to anaerobic aromatic degradation in G. daltonii. IMPORTANCE: The contamination of anaerobic subsurface environments including groundwater with toxic aromatic hydrocarbons, specifically benzene, toluene, ethylbenzene, and xylene, has become a global issue. Subsurface groundwater is largely anoxic, and further study is needed to understand the natural attenuation of these compounds. This study elucidated a metabolic pathway utilized by the bacterium Geotalea daltonii capable of anaerobically degrading the recalcitrant molecule benzene using a unique activation mechanism involving methylation. The identification of aromatic-degrading genes and AbrB as a regulator of the anaerobic benzene and toluene degradation pathways provides insights into the mechanisms employed by G. daltonii to modulate metabolic pathways as necessary to thrive in anoxic contaminated groundwater. Our findings contribute to the understanding of novel anaerobic benzene degradation pathways that could potentially be harnessed to develop improved strategies for bioremediation of groundwater contaminants.
RESUMEN
We report the complete genome sequences of six S19 poliovirus reference strains for all three poliovirus serotypes, including three Sabin vaccine-derived and three wild-type-derived strains. The S19 strains are extensively attenuated and genetically stable when compared to the reference poliovirus strains, while maintaining the same antigenicity and immunogenicity.
RESUMEN
The emergence and international spread of neurovirulent circulating vaccine-derived polioviruses (cVDPVs) across multiple countries in Africa and Asia in recent years pose a major challenge to the goal of eradicating all forms of polioviruses. Approximately 90% of all cVDPV outbreaks are caused by the type 2 strain of the Sabin vaccine, an oral live, attenuated vaccine; cVDPV outbreaks typically occur in areas of persistently low immunization coverage (1). A novel type 2 oral poliovirus vaccine (nOPV2), produced by genetic modification of the type 2 Sabin vaccine virus genome (2), was developed and evaluated through phase I and phase II clinical trials during 2017-2019. nOPV2 was demonstrated to be safe and well-tolerated, have noninferior immunogenicity, and have superior genetic stability compared with Sabin monovalent type 2 (as measured by preservation of the primary attenuation site [domain V in the 5' noncoding region] and significantly lower neurovirulence of fecally shed vaccine virus in transgenic mice) (3-5). These findings indicate that nOPV2 could be an important tool in reducing the risk for generating vaccine-derived polioviruses (VDPVs) and the risk for vaccine-associated paralytic poliomyelitis cases. Based on the favorable preclinical and clinical data, and the public health emergency of international concern generated by ongoing endemic wild poliovirus transmission and cVDPV type 2 outbreaks, the World Health Organization authorized nOPV2 for use under the Emergency Use Listing (EUL) pathway in November 2020, allowing for its first use for outbreak response in March 2021 (6). As required by the EUL process, among other EUL obligations, an extensive plan was developed and deployed for obtaining and monitoring nOPV2 isolates detected during acute flaccid paralysis (AFP) surveillance, environmental surveillance, adverse events after immunization surveillance, and targeted surveillance for adverse events of special interest (i.e., prespecified events that have the potential to be causally associated with the vaccine product), during outbreak response, as well as through planned field studies. Under this monitoring framework, data generated from whole-genome sequencing of nOPV2 isolates, alongside other virologic data for isolates from AFP and environmental surveillance systems, are reviewed by the genetic characterization subgroup of an nOPV working group of the Global Polio Eradication Initiative. Global nOPV2 genomic surveillance during March-October 2021 confirmed genetic stability of the primary attenuating site. Sequence data generated through this unprecedented global effort confirm the genetic stability of nOPV2 relative to Sabin 2 and suggest that nOPV2 will be an important tool in the eradication of poliomyelitis. nOPV2 surveillance should continue for the duration of the EUL.