Candida spp. isolated from recreational coastal waters of Rio de Janeiro - Brazil: Focus on antifungal resistance and virulence attributes.

The use of recreational waters is a widespread activity worldwide, and one of the risks associated with this practice is the exposure of bathers to microorganisms that may arise due to pollution caused by inadequate infrastructure and sanitation. In the present work, we isolated Candida spp. (n = 24) from five recreational beaches in Rio de Janeiro, Brazil, in order to evaluate their susceptibility to antifungals, the production of virulence attributes and the in vivo virulence using Tenebrio molitor larvae as a model. The ITS1-5.8S-ITS2 gene sequencing identified thirteen isolates (54.1 %) as C. tropicalis, seven (29.1 %) as C. krusei (Pichia kudriavzevii), one (4.2 %) as C. rugosa (Diutina rugosa), one (4.2 %) as C. mesorugosa (Diutina mesorugosa), one (4.2 %) as C. utilis (Cyberlindnera jadinii) and one (4.2 %) as C. parapsilosis. C. tropicalis isolates showed resistance to azoles and susceptibility to amphotericin B, flucytosine and caspofungin. C. krusei isolates were resistant to fluconazole, caspofungin and itraconazole, with 42.8 % resistance to flucytosine, besides susceptibility to voriconazole and amphotericin B. The remaining species were susceptible to all tested antifungals. All Candida isolates adhered to abiotic surfaces and formed biofilm on polystyrene, albeit to varying degrees, and produced aspartic protease and hemolytic activity, which are considered fungal virulence attributes. C. tropicalis, C. krusei and C. utilis isolates produced phytase, while the only esterase producer was C. tropicalis. Regarding resistance to osmotic stress, all isolates of C. tropicalis, C. parapsilosis and C. mesorugosa grew up to 7.5 % NaCl; the remaining isolates grew up to 1.87-3.75 % NaCl. The mortality caused by fungal challenges in T. molitor larvae was variable, with C. tropicalis, C. utilis and C. parapsilosis being more virulent than C. krusei and C. rugosa complex. Collectively, the presence of these yeasts, particularly the virulent and resistant isolates, in recreational waters can pose a significant health risk to bathers.

Spatiotemporal distribution of thermophilic free-living amoebae in recreational waters: A 5-year survey in Guadeloupe (French West Indies).

Free-living amoebae (FLA) such as Acanthamoeba, Balamuthia mandrillaris, Naegleria fowleri and Sappinia pedata are naturally widespread in freshwater, causing rare but fatal and debilitating infections in humans. Although recent studies have shown an increase in infection rates, there is a paucity of epidemiological studies regarding the presence of these emerging pathogens in water. Herein, we studied the diversity and relative abundance of thermophilic FLA in different recreational baths in a tropical climate for 5 years. From 2018 to 2022, a total of 96 water samples were collected from 7 recreational baths (natural, tiled, regularly cleaned or not, and with temperatures ranging from 27 to 40 °C). DNA was extracted from FLA cultivated at 37 °C to detect thermophilic culturable FLA. Metabarcoding studies were conducted through FLA 18S rRNA gene amplicons sequencing; amplicon sequence variants (ASV) were extracted from each sample and taxonomy assigned against PR2 database using dada2 and phyloseq tools. We also searched for Naegleria sp. and N. fowleri using PCR targeting ITS and NFITS genes (respectively) and we quantified them using an optimized most probable number (MPN) method for FLA. Our results showed that differences in FLA diversity and abundance were observed amongst the 7 baths, but without a clear seasonal distribution. Naegleria, Vermamoeba and Stenamoeba were the most represented genera, while the genera Acanthamoeba and Vahlkampfia were mainly found in 2 baths. The MPN values for Naegleria sp. (NT/l) increased between 2018 and 2022, but the MPN values for N. fowleri (NF/l) seemed to decrease. Globally, our results showed that since we cannot establish a seasonal distribution of FLA, the regular presence of FLA (namely Naegleria and Acanthamoeba) in recreational waters can pose a potential threat in terms of neuroinfections as well as Acanthamoeba keratitis. It is thus imperious to perform the regular control of these baths as a preventive health measure.

Acanthamoeba and a bacterial endocytobiont isolated from recreational freshwaters.

The quality of many freshwater environments is impacted by human activities, so that many rivers may represent a vehicle for the transmission of health-related microorganisms. This work aimed to isolate and identify genetically free-living amoeba (FLA) of the genus Acanthamoeba from a recreational river in Salta, Argentina, and isolate, if possible, an endocytobiont. Sampling took place at four points (P1-P4) throughout the river in the winter and the summer seasons. Free-living amoebae and Acanthamoeba were recovered from 20-L water concentrated through an ultrafiltration system. Isolation was performed in agar plates, confirmation of Acanthamoeba genus by PCR, and fellow identification and classification based on their sequence analyses. High concentrations of indicator bacteria were found especially in P2, which is intensively used for recreation. Out of a total of 29 FLA isolations, 9 were identified as Acanthamoeba genotype T4 subtype A, the most frequent genotype found in nature and associated with causing human disease. From an axenic culture of Acanthamoeba spp. (KY751412), a bacterial endocytobiont was isolated, and identified as Stenotrophomonas maltophilia. The endocytobiont showed resistance and intermediate resistance to a wide range of widely used antibiotics. Results were in concordance with the cosmopolitan behavior of Acanthamoeba, and showed the importance of studying this group of amoebae and related microorganisms in recreational environments.

Prevalence of free-living amoebae in swimming pools and recreational waters, a systematic review and meta-analysis.

Free-living amoebae (FLA) are cosmopolitan microorganisms known to be pathogenic to humans who often have a history of contact with contaminated water. Swimming pools and recreational waters are among the environments where the greatest human exposure to FLA occurs. This study aimed to determine the prevalence of FLA in swimming pools and recreational waters, through a systematic review and meta-analysis that included studies published between 1977 and 2022. A total of 106 studies were included and an overall prevalence of FLA in swimming pools and recreational waters of 44.34% (95% CI = 38.57-50.18) was found. Considering the studies published up to 2010 (1977-2010), between 2010 and 2015, and those published after 2010 (> 2010-2022), the prevalence was 53.09% (95% CI = 43.33-62.73) and 37.07% (95% CI = 28.87-45.66) and 45.40% (95% CI = 35.48-55.51), respectively. The highest prevalence was found in the American continent (63.99%), in Mexico (98.35%), and in indoor hot swimming pools (52.27%). The prevalence varied with the variation of FLA detection methods, morphology (57.21%), PCR (25.78%), and simultaneously morphology and PCR (43.16%). The global prevalence by genera was Vahlkampfia spp. (54.20%), Acanthamoeba spp. (33.47%), Naegleria spp. (30.95%), Hartmannella spp./Vermamoeba spp. (20.73%), Stenamoeba spp. (12.05%), and Vannella spp. (10.75%). There is considerable risk of FLA infection in swimming pools and recreational waters. Recreational water safety needs to be routinely monitored and, in case of risk, locations need to be identified with warning signs and users need to be educated. Swimming pools and artificial recreational water should be properly disinfected. Photolysis of NaOCl or NaCl in water by UV-C radiation is a promising alternative to disinfect swimming pools and artificial recreational waters.

Fast screening of enteropathogens in marine water samples.

This study aimed to fast screen the microbiological contamination of recreational waters using a TaqMan Array Card (TAC), a multiplexed platform designed for the simultaneous detection of 35 enteropathogens. Surface and deep marine water samples were concentrated by skimmed milk flocculation and processed for nucleic acid extraction protocol using QIAamp Fast DNA Stool Mini Kit. Twelve microorganisms and parasites, including bacteria (n = 6), protozoa (4), and viruses (2), were detected in 85.7% (24/28) of samples. Campylobacter (82.1%), Cryptosporidium (39.3%), and adenovirus (14.3%) were the most detected pathogens. Neither fungi nor helminths were detected. A spatial pollution profile of microbiological contamination was observed in the area. Methodologies for simultaneous detection of multiple pathogens, such as TAC, can assist decision-makers by providing a quick assessment of the microbiological water quality in areas used for recreational purposes, which in many cases are in accordance with the bacteriological indicators.

Predictors of carbapenemase-producing bacteria occurrence in polluted coastal waters.

The spread of carbapenemase-producing bacteria is a worldwide concern as it challenges healthcare, especially considering the insufficient development of antimicrobials. These microorganisms have been described not only in hospitals, but also in several environmental settings including recreational waters. Community exposure to antimicrobial-resistant bacteria through recreation might be relevant for human health, but risk assessment studies are lacking. Absence of effective and feasible monitoring in recreational aquatic matrices contributes to such a knowledge gap. Here, we aimed at assessing predictors of occurrence of medically relevant carbapenemase-producing bacteria in coastal waters. We quantitatively assessed recovery of carbapenemase-producing Enterobacteriaceae, Pseudomonas spp., Acinetobacter spp. and Aeromonas spp. in superficial coastal waters showing distinct pollution history across one year, and registered data regarding tide regimen, 7-days pluviosity, salinity, pH, water temperature. We analyzed data using General Estimating Equation (GEE) to assess predictors of such occurrence. Our results suggest that the sampling site had the strongest effect over concentration of these antimicrobial-resistant microorganisms, followed by pollution indexes and tide regimen. Increased salinity, advanced sampling time, water temperature, rainfall and decrease of pH were related to decrease concentrations. We provide a list of factors that could be easily monitored and further included in models aiming at predicting occurrence of carbapenemase producers in coastal waters. Our study may encourage researchers to further improve this list and validate the model proposed, so that monitoring and future public policies can be developed to control the spread of antimicrobial resistance in the environment.

Assessment of health risks from recreational exposure to Giardia and Cryptosporidium in coastal bathing waters.

Some Brazilian beaches are impacted by raw or poorly treated sewage. Thus, users (beachgoers, sports people, and children) are exposed to pathogens, which pose health concerns. This study aimed to estimate the probability of infection and disease by Giardia and Cryptosporidium, using the quantitative microbial risk assessment (QMRA), for three groups of bathers: children, adults, and open water swimmers. The concentrations of (oo)cysts were taken from a study run by CETESB (Environmental Company of Sao Paulo State) throughout 2011 and 2012, in which 203 samples were collected monthly and analyzed for (oo)cysts of Giardia and Cryptosporidium. Giardia was present in 43% of the samples, while Cryptosporidium in 13%. Infection probability was higher in beaches with more positive samples for Giardia cysts for the group of open water swimmers. In some cases, the highest annual risk obtained for giardiasis and cryptosporidiosis was 2.4 × 10-1 and 8.9 × 10-3 for open water swimmers, respectively, exceeding the incidence results found in the epidemiological study run in summer of 1999 in São Paulo state coast. The results bring insights to improve environmental quality in order to protect tourists' and residents' wellbeing.

'Don't put your head under water': enteric viruses in Brazilian recreational waters.

Like in many other countries, virologic analyses are not routinely performed in Brazil in monitoring water quality for recreational purposes. We surveyed current research regarding viral contamination of recreational water environments in Brazil. Among the enteric viruses studied in Brazilian recreational waters, we highlight adenoviruses, rotaviruses, enteroviruses and noroviruses. Although there has been relatively little research on outbreaks related to bathing in recreational water environments in Brazil, noroviruses and adenoviruses are the viruses that are most often related to outbreaks. Better surveillance of the occurrence of enteric viruses in water could improve the assessment of risk to human health as well as indicate the sources of contamination and thus demonstrate the importance of adequate environmental sanitation.

Microbial risk assessment in recreational freshwaters from southern Brazil.

In this study, total coliforms (TC), Escherichia coli, enterovirus (EV), rotavirus (RV), and human mastadenovirus species C and F (HAdV-C and HAdV-F) were evaluated in water samples from Belo Stream. For HAdV-C and F, the infectivity was assessed by integrated cell culture quantitative real-time polymerase chain reaction (ICC-qPCR). Samples were collected monthly (May/2015 to April/2016) at four sites. Viral analyses were performed for both ultracentrifuge-concentrated and unconcentrated samples. For site P4 (used for recreational purposes), QMRA was applied to estimate health risks associated with exposure to E. coli and HAdV-C and F. TC and E. coli were present throughout the collection period. EV and RV were not detected. HAdV-C were present in 8.51% (1.89E + 06 to 2.28E + 07 GC (Genomic Copies)/L) and 21.27% (2.36E + 05 to 1.29E + 07 GC/L) for unconcentrated and concentrated samples, respectively. For HAdV-F were 12.76% (2.77E + 07 to 3.31E + 08 GC/L) and 48.93% (1.10E + 05 to 4.50E + 08 GC/L) for unconcentrated and concentrated samples, respectively. For unconcentrated samples, infectivity for HAdV-C was detected in 37.20% (1st ICC-qPCR) and 25.58% (2nd ICC-qPCR). For HAdV-F, infectivity was detected in 6.97% (1st ICC-qPCR) and 6.97% (2nd ICC-qPCR). For concentrated samples, HAdV-C infectious was observed in 17.02% (1st ICC-qPCR) and in 8.51% (2nd ICC-qPCR). For HAdV-F, were present in 8.51% for both 1st and 2nd ICC-qPCR. Statistical analyzes showed significant difference between the collection sites when analyzed the molecular data of HAdV-F, data of TC and E. coli. Correlation tests showed direct correlation between HAdV-F with E. coli and TC. E. coli concentrations translated to the lowest estimates of infection risks (8.58E-05 to 2.17E-03). HAdV-F concentrations were associated with the highest infection risks at 9.99E-01 and for group C, 1.29E-01 to 9.99E-01. These results show that commonly used bacterial indicators for water quality may not infer health risks associated with viruses in recreational freshwaters.

Are fecal indicator bacteria appropriate measures of recreational water risks in the tropics: A cohort study of beach goers in Brazil?

Regulating recreational water exposure to pathogens within the tropics is a major public health and economic concern. Although numerous epidemiological studies estimating the risk to recreational marine water exposure have been conducted since the 1950s, few studies have been done in the tropics. Furthermore, many have suggested that the use of fecal indicator bacteria for monitoring recreational water quality in temperate regions is not appropriate in the tropics. We analyzed a large cohort study of five beaches in Sao Paulo, Brazil, conducted during consecutive weekends in the summer of 1999 that estimated risk to water, sand, and food exposures. Enterococci and Escherichia coli concentrations were measured each day of the study. Elevated risks were estimated for both swimming (OR = 1.36 95% CI: 1.05-1.58) and sand contact (OR = 1.29 95% CI 1.05-1.58). A 1 log increase in enterococci concentration was associated with an 11% increase in risk (OR = 1.11 95% CI: 1.04-1.19). For E. coli a 1-log increase in concentration was associated with 19% increase in risk (OR = 1.19 95% CI: 1.14-1.28). Most countries with beaches in the tropics are lower or middle income countries (LMIC) and rely on tourism as a major source of income. We present data that suggests fecal indicator bacteria such as enterococci are an appropriate indicator of risk in tropical urban settings where contamination is coming from predominantly human sources. Additional studies in tropical settings could help inform and refine guidelines for safe use of recreational waters.

Avaliação sanitária das águas e areias de praias da Baixada Santista, São Paulo / Sanitary assessment of waters and sand of beaches at Baixada Santista, São Paulo

O Brasil possui em sua área costeira grandes metrópoles onde diversas atividades humanas são desenvolvidas, podendo trazer prejuízos à qualidade das águas costeiras. O litoral de São Paulo, constituído por 16 municípios incluindo-se Cubatão, apresenta peculiaridades diferenciadas. O tipo de poluição mais encontrado na costa paulista é a poluição fecal causada por despejo de esgotos domésticos, os quais transportam organismos patogênicos cuja presença pode levar a agravos à saúde humana. O objetivo deste estudo é o de avaliar a qualidade sanitária de águas e areias de praias da Baixada Santista. Assim amostras de água e areia (úmida e seca) de três praias, como Gonzaguinha (São Vicente), Boqueirão (Santos) e Pitangueiras (Guarujá) foram coletadas com frequência mensal pelo período de um ano nas quais foram analisados os seguintes parâmetros: coliformes termotolerantes, Escherichia coli, enterococos, e ainda Staphylococcus coagulase (+), Pseudomonas aeruginosa, Candida albicans e dermatófitos. Para a realização das análises em amostras de água utilizou-se da Técnica da Membrana Filtrante e para as amostras de areia a Técnica de Tubos Múltiplos, de acordo com Normas Técnicas CETESB e Standard Methods. Para a determinação de dermatófitos, as amostras de água foram concentradas pela Técnica de Membrana Filtrante e as amostras de areia (seca e úmida) foram analisadas pela Técnica de inoculação em superfície e, concomitantemente, pela Técnica da Isca de Cabelo. Os resultados obtidos revelaram que 95 por cento das amostras analisadas foram positivas para a presença dos indicadores de contaminação fecal, seguidos por P. aeruginosa, presente em 75 por cento das amostras. C. albicans foi o microrganismo menos frequente (6,7 por cento) e foi detectada somente nas amostras de água. O dermatófito Microsporum sp foi detectado somente nas amostras de areia e esteve presente nas três praias...

Brazilian seashore comprises many big cities and the diverse activities which are performed in these areas can bring damages to seashore water quality. São Paulo State seashore comprises 16 cities including Cubatão, where each one shows their own particularities. In the São Paulo State seashore is mostly observed fecal pollution due to launching sewage into these waters which transport pathogens with potential to cause diseases. The objective of this study is to assess the sanitary quality of recreational waters and wet and dry sand from three beaches located at Baixada Santista such as Gonzaguinha (São Vicente), Boqueirão (Santos) and Pitangueiras (Guarujá). The samples (water, wet and dry sand) were collected with monthly frequency throughout a year and the following parameters were analyzed: thermotolerant coliforms, Escherichia coli, enterococos, Staphylococcus coagulase +, Pseudomonas aeruginosa, Candida albicans and dermatophytes. The bacteriological and C. albicans analyzes were performed according to CETESB and Standard Methods 21st edition. Dermatophyte analysis from water samples were carried out by membrane filtering technique and by spread plate technique and hair-baiting technique for sand samples. The results revealed that 95 per cent of the water samples were positive for fecal indicators following by P. aeruginosa (75 per cent). C. albicans was detected only in water samples showing 6.7 per cent of the observed frequency. Microsporum sp specie was detected only in sand samples in all beaches studied...

Riesgo sanitario por pesticidas organoclorados en aguas de una laguna pampeana argentina / Health risk by organochlorine pesticides in waters of an Argentine pampean shallow lake

En la laguna El Carpincho (Junín, provincia de Buenos Aires, Argentina) se determinaron agroquímicos organoclorados, por lo que podrían existir efectos negativos para la salud en sus usuarios. El objetivo del trabajo fue estimar el riesgo sanitario (ARS) por los pesticidas hallados durante el baño recreativo. El ARS se basó en el modelo USEPA probabilístico para riesgo agregado y acumulativo a los pesticidas alfa y δ Hexaclorociclohexano (HCH), Endosulfán y Endosulfán sulfato. El ARS se estimó tanto para efectos no carcinogenéticos como carcinogenéticos, considerando la ingesta accidental de agua y el contacto dérmico. El individuo expuesto asumido fue un pescador deportivo que toma un baño en la laguna de una hora por día. Su tipología (frecuencia de excursiones, duración y experiencia) se justificó en resultados previos referidos a aspectos sociales de la pesquería allí instalada. El riesgo agregado y acumulativo para los efectos no carcinogenéticos y carcinogenéticos fue de 3,19E-05 y 1,76E-08 (5 y 3 órdenes de magnitud menores al nivel de peligrosidad para cada tipo de efecto), siendo los isómeros de HCH los principales contribuyentes en ambos casos. El uso de la laguna para baño no representaría un riesgo atendible para las condiciones de exposición consideradas.

Organochlorine pesticides were detected in El Carpincho shallow lake (Junín, Buenos Aires province, Argentina), and they could have negative effects on its users' health. The object of this study was to assess the human health risk (HR) associated with these pesticides during recreational bathing. The probabilistic HR assessment was based on aggregate and cumulative USEPA models and applied to alpha and δ HCH, Endosulfan and Endosulfan sulphate pesticides. The cancer and noncancer risks were estimated for accidental drinking and dermal contact. The exposure considered was one hour a day-1 by a sport fisherman. The fisherman characteristics (annual fish trip frequency, event length, experience) were based on previous studies on social aspects of local anglers. The aggregate and cumulative noncancer and cancer risks were 3.19E-05 and 1.76E-08 (5 and 3 orders of magnitude lower than the level of hazard for each type of effect). Isomers of HCH were the major contributors in both cases. The recreational use of this shallow lake would not represent an adverse health risk considering the assumed conditions of exposure.