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The recent implementation of the Revised Bathing Water Directive in the European Union has highlighted the need for development of effective methods to differentiate between sources of faecal contamination. It had previously been shown that amplification of 16S rRNA genes of host-specific Bacteriodales species using the HF183F and CF128F primers could be used as markers for human and bovine faecal contamination in the United States. This paper determined the sensitivity and specificity of these markers in four Atlantic Rim countries (France, Ireland, Portugal and the United Kingdom) to evaluate their usefulness in determining the origin of faecal contamination. It was shown that the HF183F marker displayed high sensitivity (80-100%) and specificity (91-100%), and is reliable as an indication of human faecal contamination. The CF128F marker displayed 100% sensitivity in all four countries. However, strong regional variations in specificity (41-96%) were observed, highlighting the need for local validation before this marker is employed in source tracking of faecal contamination.

Recent studies suggest that sand can serve as a vehicle for exposure of humans to pathogens at beach sites, resulting in increased health risks. Sampling for microorganisms in sand should therefore be considered for inclusion in regulatory programmes aimed at protecting recreational beach users from infectious disease. Here, we review the literature on pathogen levels in beach sand, and their potential for affecting human health. In an effort to provide specific recommendations for sand sampling programmes, we outline published guidelines for beach monitoring programmes, which are currently focused exclusively on measuring microbial levels in water. We also provide background on spatial distribution and temporal characteristics of microbes in sand, as these factors influence sampling programmes. First steps toward establishing a sand sampling programme include identifying appropriate beach sites and use of initial sanitary assessments to refine site selection. A tiered approach is recommended for monitoring. This approach would include the analysis of samples from many sites for faecal indicator organisms and other conventional analytes, while testing for specific pathogens and unconventional indicators is reserved for high-risk sites. Given the diversity of microbes found in sand, studies are urgently needed to identify the most significant aetiological agent of disease and to relate microbial measurements in sand to human health risk.

Whilst the potential impact on beach users from microorganisms in water has received considerable attention, there has been relatively little investigation into microbial contaminants in sand. 33 beaches across Portugal were analyzed during a five year period (2006)(2007)(2008)(2009)(2010) to determine the presence of yeasts, pathogenic fungi, dermatophytes, total coliforms, E. coli and intestinal enterococci in sand.Our results showed that 60.4% of the samples were positive for fungi and that 25.2% were positive for the bacterial parameters. The most frequent fungal species found were Candida sp.and Aspergillus sp., whereas intestinal enterococci were the most frequently isolated bacteria.Positive associations were detected among analyzed parameters and country-regions but none among those parameters and sampling period.Regarding threshold values, we propose 15 cfu/g for yeasts, 17 cfu/g for potential pathogenic fungi, 8 cfu/g for dermatophytes. 84 cfu/g for coliforms, 250 cfu/g for E. coli, and 100 cfu/g for intestinal enterococciHighlights:x 60.4% of the analyzed marine beach sand samples were positive for the studied fungi x Candida sp. and Aspergillus sp. were the most frequent fungal species found x 25.2% of the analyzed marine beach sand samples were positive to the studied bacteria

Beaches worldwide provide recreational opportunities to hundreds of millions of people and serve as important components of coastal economies. Beach water is often monitored for microbiological quality to detect the presence of indicators of human sewage contamination so as to prevent public health outbreaks associated with water contact. However, growing evidence suggests that beach sand can harbor microbes harmful to human health, often in concentrations greater than the beach water. Currently, there are no standards for monitoring, sampling, analyzing, or managing beach sand quality. In addition to indicator microbes, growing evidence has identified pathogenic bacteria, viruses, and fungi in a variety of beach sands worldwide. The public health threat associated with these populations through direct and indirect contact is unknown because so little research has been conducted relating to health outcomes associated with sand quality. In this manuscript, we present the consensus findings of a workshop of experts convened in Lisbon, Portugal to discuss the current state of knowledge on beach sand microbiological quality and to develop suggestions for standardizing the evaluation of sand at coastal beaches. The expert group at the "Microareias 2012" workshop recommends that 1) beach sand should be screened for a variety of pathogens harmful to human health, and sand monitoring should then be initiated alongside regular water monitoring; 2) sampling and analysis protocols should be standardized to allow proper comparisons among beach locations; and 3) further studies are needed to estimate human health risk with exposure to contaminated beach sand. Much of the manuscript is focused on research specific to Portugal, but similar results have been found elsewhere, and the findings have worldwide implications.

The action of maslinic acid (2alpha,3beta-dihydroxyolean-12-en-28-oic acid) (1), a pentacyclic derivative present in the pressed fruits of the olive (Olea europaea), has been studied against the tachyzoites of Toxoplasma gondii. The capability of tachyzoites to infect Vero cells treated with 1 was affected. The LD(50) values were 58.2 muM for the isolated tachyzoites and 236 muM for the noninfected Vero cells. Zymograms of the T. gondii proteases incubated with 1 showed a dosage-dependent inhibition of some of the proteases. The parasites treated with 1 showed gliding motility and ultrastructural alterations. The present findings suggest that protease activity of the parasite required for cell invasion is the action target for maslinic acid (1).

Current regulations and legislation require critical revision to determine safety for alternative water sources and water reuse as part of the solution to global water crisis. In order to fulfill those demands, Lisbon municipality decided to start water reuse as part of a sustainable hydric resources management, and there was a need to confirm safety and safeguard for public health for its use in this context. For this purpose, a study was designed that included a total of 88 samples collected from drinking, superficial, underground water, and wastewater at three different treatment stages. Quantitative Polimerase Chain Reaction (PCR) detection (qPCR) of enteric viruses Norovirus (NoV) genogroups I (GI) and II (GII) and Hepatitis A (HepA) was performed, and also FIB (E. coli, enterococci and fecal coliforms) concentrations were assessed. HepA virus was only detected in one untreated influent sample, whereas NoV GI/ NoV GI were detected in untreated wastewater (100/100%), secondary treated effluent (47/73%), and tertiary treated effluent (33/20%). Our study proposes that NoV GI and GII should be further studied to provide the support that they may be suitable indicators for water quality monitoring targeting wastewater treatment efficiency, regardless of the level of treatment.

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