Advancing plastic pollution hotspotting at the subnational level: Brazil as a case study in the Global South.

Identifying sources is crucial for proposing effective actions to combat marine litter pollution. Here, we used an innovative approach to identify hotspots of mismanaged plastic waste (MPW) within Brazil and subsequent leakage to the ocean, based on population density, socio-economic conditions, municipal solid waste management and environmental parameters. We estimated plastic waste generation and MPW for each of the 5570 Brazilian municipalities, which totaled 3.44 million metric tons per year. Then, we estimated the probability of litter mobilization and transport (P) and the relative risk of leakage to the ocean (MPW × P). The Guanabara Bay and La Plata River comprised the main oceanic entry hotspots of litter produced in Brazil. The use of national databases allowed us to increase spatial and temporal granularity, offering a detailed baseline for the application of prevention and mitigation actions. However, overcoming data limitations is still a challenge in Brazil as in other Global South countries.

How far are we from robust estimates of plastic litter leakage to the environment?

Litter pollution is a global concern, and identifying sources and pathways is crucial for proposing preventative actions. Existing models of plastic litter leakage to the environment have provided worldwide estimates at a country-based level, but only a few initiatives address subnational scales. Adding relevant parameters and improving models is needed to reduce the limitations of global estimates. However, availability of information, which varies among countries and is critical in the Global South, may preclude such improvements. To understand the potentialities and limitations of subnational estimates of plastic litter leakage to the environment, we reviewed the parameters used in the literature and addressed data usability, considering Brazil as a case study. We gathered data on parameters identified for all 5570 Brazilian municipalities and evaluated their usability considering reliability and temporal and geographic granularity. We identified 51 parameters that are either currently used in models or could improve estimates, including parameters regarding territory, population density, socioeconomic condition, and solid waste generation, composition, collection, and final destination, selective waste collection, recycling, and hydrology. Only 29.4% of parameters were linked to data sources with good or very good usability, while most of them presented average usability (45.1%) and 7.8% were not linked to any data source. This panorama of low data usability reveals uncertainties and explicit difficulties of estimating plastic litter leakage to the environment, including mobilization from the terrestrial environment to the ocean. The Brazilian scenario reflects current data availability conditions and the difficulties of countries in the Global South to robustly understand plastic litter leakage and face land-based sources of marine litter.

Analysis of Cryptococcal Extracellular Vesicles: Experimental Approaches for Studying Their Diversity Among Multiple Isolates, Kinetics of Production, Methods of Separation, and Detection in Cultures of Titan Cells.

Extracellular vesicles (EVs) produced by members of the Cryptococcus genus are associated with fundamental processes of fungal physiology and virulence. However, several questions about the properties of cryptococcal EVs remain unanswered, mostly because of technical limitations. We recently described a fast and efficient protocol of high-yield EV isolation from solid medium. In this study, we aimed at using the solid medium protocol to address some of the open questions about EVs, including the kinetics of EV production, the diversity of EVs produced by multiple isolates under different culture conditions, the separation of vesicles in a density gradient followed by the recovery of functional EVs, the direct detection of EVs in culture supernatants, and the production of vesicles in solid cultures of Titan cells. Our results indicate that the production of EVs is directly impacted by the culture medium and time of growth, resulting in variable detection of EVs per cell and a peak of EV detection at 24 h of growth. Nanoparticle tracking analysis (NTA) of EV samples revealed that multiple isolates produce vesicles with variable properties, including particles of diverging dimensions. EVs were produced in the solid medium in amounts that were separated on a centrifugation density gradient, resulting in the recovery of functional EVs containing the major cryptococcal capsular antigen. We also optimized the solid medium protocol for induction of the formation of Titan cells, and analyzed the production of EVs by NTA and transmission electron microscopy. This analysis confirmed that EVs were isolated from solid cultures of cryptococcal enlarged cells. With these approaches, we expect to implement simple methods that will facilitate the analysis of EVs produced by fungal cells. IMPORTANCE Fungal extracellular vesicles (EVs) are considered to be important players in the biology of fungal pathogens. However, the limitations in the methodological approaches to studying fungal EVs impair the expansion of knowledge in this field. In the present study, we used the Cryptococcus genus as a model for the study of EVs. We explored the simplification of protocols for EV analysis, which helped us to address some important, but still unanswered, questions about fungal EVs.

Population genomic analysis of Cryptococcus Brazilian isolates reveals an African type subclade distribution.

The genomes of a large number of Cryptococcus neoformans isolates have been sequenced and analyzed in recent years. These genomes have been used to understand the global population structure of this opportunistic pathogen. However, only a small number of South American isolates have been considered in these studies, and the population structure of C. neoformans in this part of the world remains elusive. Here, we analyzed the genomic sequences of 53 Brazilian Cryptococcus isolates and deciphered the C. neoformans population structure in this country. Our data reveal an African-like structure that suggested repeated intercontinental transports from Africa to South America. We also identified a mutator phenotype in one VNBII Brazilian isolate, exemplifying how fast-evolving isolates can shape the Cryptococcus population structure. Finally, phenotypic analyses revealed wide diversity but not lineage specificity in the expression of classical virulence traits within the set of isolates.