The Detection Limits of Legionella According to the EU Directive 2020/2184. Could That Be Too Permissive?

INTRODUCTION: The problem of detecting legionella after a case of legionellosis from the source of environmental contamination has been known since a long time ago. Legionella is a bacterium present in various natural and artificial habitats and especially in surface fresh waters. It is found in greater concentration in warm waters, at temperatures between 20 °C and 42 °C. The greatest risk factor for humans is represented by the presence of Legionella in water distribution systems in hospitals, medical equipment (e.g. respirators, dialyzers, inhalers, humidifiers, water, massage equipment used in balneotherapy) and turbines used in dental practices, especially for hospitalized individuals. In the EU directive 2020/2184, issued by the European parliament on 16/12/2020, the concentration of Legionella was added to the parameters to be determined in assessing the quality of drinking water intended for human consumption. The objectives were to improve the quality standard of drinking water, reduce the consumption of bottled water and consequently reduce plastic waste. The WHO notes that Legionella causes the greatest burden from a health point of view and it is included among the parameters that require careful monitoring with a limit of less than 1000 CFU/L. The aim of this report was to evaluate the new EU directive 2020/2184 on the light of our laboratory experience. MATERIALS AND METHODS: A total of 459 samples were processed at our Hygiene of food Laboratory - Department of Medical Sciences and Public Health. All statistical analyses were conducted using the SPSS statistical package (version 23 for Windows. SPSS, Inc. Chicago, Ill). RESULTS AND DISCUSSION: Of the 67 structures examined where the cases occurred, 35 showed samples with at least one over-threshold value considering the reference value of 100 CFU/L, whereas using the new limit of 1000 CFU/L, only 25 structures resulted as having at least one sample above the threshold. In our experience as a regional reference laboratory for Legionella research, the increase from 100 CFU/L to 1,000 CFU/L could lead to a lower alert level. In fact, in the period between October 2017 and October 2021, the median value of CFU/L in presence of a case was 0 (0-100). Despite the large amount of studies on Legionella only a few relate the withdrawals and the consequent CFU/L with the confirmed cases of legionellosis, as in our analysis. The 75° percentile values of the Legionella concentration equal to 100 CFU/L in all samples associated with cases and clusters leads us to hypothesize that the limit equal to 1000 CFU/L that will be introduced for environmental monitoring as per recent European regulations may not be sufficiently protective for minimizing risk in the population, especially in healthcare facilities where fragile patients are assisted.

Growth of Listeria monocytogenes in ready to eat salads at different storage temperatures and valuation of virulence genes expression.

BACKGROUND: Vegetables are major components of a healthy and balanced diet. However, 25% of foodborne diseases are linked to the consumption of vegetables. STUDY DESIGN: The aim of this work was to assess the microbiological risks associated with consumption of ready to eat salads (RTE). METHODS: Microbiological challenge tests were carried out for the evaluation of the L. monocytogenes growth potential in RTE salads stored at different temperatures. RESULTS: The results indicate that L.monocytogenes was able to grow (δ ≥ 0.5) in all storage conditions considered at the end of shelf life. In order to evaluate the virulence role of L. monocytogenes, the temperature-dependent transcription of major virulence genes was also investigated by RT-PCR. CONCLUSIONS: The microbiological challenge test allowed us to confirm, as also demonstrated by other authors, that RTE salads are able to support the growth of L. monocytogenes strains (d δ≥ 0.5) stored under different temperatures.

Microbiological environmental monitoring in high-risk departments during building activities in a hospital site.

BACKGROUND: This study examines the microbial and fungal contamination associated with the presence of renovation works in a hospital site in Sardinia (Italy). METHODS: Microbiological environmental monitoring was carried out before, during and at the conclusion of the works in the Ophthalmology Department in view of a risk assessment procedure. RESULTS: Although the median values of microbial and fungal counts were found raised during the works, protective measures set out by the internal procedures limited the contamination level. CONCLUSIONS: This study emphasizes the benefits of environmental surveillance for airborne contamination to help prevent outbreaks of nosocomial mycosis associated with construction work.

Microbiological Quality of the Water used in Agriculture in Sardinia.

BACKGROUND: The microbiological quality of the water used in irrigation is crucial for the safety of products, such as fruit and vegetables, especially when destined to be consumed raw. However, the microbiological quality of this water is not defined at a community regulatory level or at a national level. METHODS: With our present work, we wanted to investigate the microbiological quality of the water used for crop irrigation in various Sardinian provinces. Since in most fields the irrigation water is filtered to remove any impurities, the sample was processed twice - both before and after the filtering process. Furthermore, with the purpose of hypothesising the potential health risks attributable to the consumption of crops from the tested fields, samples of horticultural product were collect. Any eventual seasonal differences in the values of microbial concentration were assessed. Microorganism faecal contamination indicators (Escherichia coli, total coliform and faecal streptococci), but even the presence of the opportunistic pathogen such as Pseudomonas aeruginosa were researched in irrigation water. Total mesophilic counts (TMC) were assessed at 36°C and 22°C. On horticultural products we researched both the indicators of process parameters, such as Escherichia coli, Total mesophilic counts at 30°C, Enterobacteriaceae, Total Psychrophilic counts and Pseudomonas aeruginosa, and pathogens, such as Salmonella spp, Listeria monocytogenes and Yersinia enterocolitica. RESULTS: The number of target microorganisms, when present in irrigation water, was very limited: Escherichia coli, total coliform and faecal streptococci, were detected respectively in 48% and 67% of the water samples tested with average concentration values of 0.9, 1.2 and 1.4 log respectively. In fresh vegetable products, the total mesophilic counts (TMC) were found to have average values of 6.6x107 CFU/g. The average values of Enterobacteriaceae totalled 6.1x105 CFU/g; Escherichia coli was detected in only one sample (curly endive) with a value of 180 CFU/g. CONCLUSION: The data highlights the high quality of the water and how this contributed to achieving satisfactory quality on prime material. However the use of filters, to eliminate impurities, and reservoirs, may represent a crucial issue, if not managed correctly.

Hygiene and health risks associated with the consumption of edible lamellibranch molluscs.

Edible lamellibranch molluscs are frequently the cause of very serious outbreaks of food poisoning. Microbiological suitability for human consumption, according to EC Reg. n. 1441/2007, is only based on two faecal bacteriological contamination parameters (Escherichia coli and Salmonella) and does not consider other microorganisms which are naturally present within the marine environment and are potentially pathogenic such as those belonging to the genus Vibrio. In this study, a total of 540 samples of edible lamellibranch molluscs (mussels, clams and oysters) bred in Sardinia were analyzed for E. coli and Salmonella, according to EC Reg n. 1441/2005, and for the presence of Vibrios of major public health interest. Environmental parameters (seawater temperature and salinity) were also recorded. The 40% (n=216) of samples were positive for Vibrio while the levels of E. coli were above the legal European limit only in 6.4% of samples. A correlation between the presence of micro-organisms of faecal origin and potentially harmful Vibrios to human health was not observed. Vibrio alginolyticus was the most common in 68% of isolated species, followed by Vibrio parahaemolyticus in 22.6%, Vibrio cholerae in 6% and Vibrio vulnificus in 1.1%. Virulence gene (trh sequence) was present in 1.6% of the V. parahaemolyticus-positive samples. The V. cholerae, V. vulnificus and V. alginolyticus strains detected were non toxigenic. The absence of genes encoding major virulence factors in the studied strains does not exclude the possibility of exposure to pathogenic strains. This study indicates the potential health risk associated with the presence of pathogenic Vibrios in edible lamellibranch molluscs bred in Sardinia, where shellfish production is very important to the local economy.

Evaluation of the level of domestic hygiene in household kitchens.

BACKGROUND: Each year in industrialized countries, 30-40% or more cases of food poisoning occur in the household. The aims of this study are to describe the aspects related to food safety in households by carrying out a microbiological characterization of the kitchens in residential dwellings and to increase consumers' awareness concerning the importance of good hygienic practices, which are required for preventing foodborne diseases at household level. METHODS: The collection of data involved taking 760 analytical samples of kitchen-counter tops and food from 80 kitchens of private dwellings; the manner in which food was treated was evaluated from the moment of purchase to its transformation and storage by means of a checklist; the questionnaire enabled us to determine the extent of consumers' knowledge concerning the food sector. RESULTS: Overall, the results showed a good level of hygiene concerning both food and kitchen-counter top food surfaces. The respondents of the questionnaire did not appear to be aware of various health risks especially concerning the management of the temperature and compartments of refrigerators as well as food storage times and the cooking or heating of food. CONCLUSIONS: The data indicate that final consumers should take more care when cleaning kitchen-counter tops and washing salad; special training programmes should be included in school curricular in order to increase citizens' awareness and knowledge concerning food risks within the household.

Comparative analysis of polyphenolic profiles and antioxidant and antimicrobial activities of tunisian pome fruit pulp and peel aqueous acetone extracts.

Pome trees, apple, pear, and quince, are classified into the subfamily Pomoideae, belonging to the Rosaceae family. Their autumnal fruits are consumed worldwide in different forms, that is, fresh or transformed into jams, jelly, juices, etc. Their well-established beneficial properties to human health were found mainly related to their phenolic content. Pulp and peel aqueous acetone extracts obtained from Tunisian fruits at commercial maturity were comparatively evaluated for their phenolic profiles and antioxidant and antimicrobial potentials. The phenolic compounds present in the extracts were identified and quantified using RP-HPLC-DAD and ESI-MS techniques. Significant differences in the chromatographic profiles among these fruits, as well as between pulp and peel extracts of each fruit, were observed. Quince, followed by 'Red Delicious', peel extracts showed the highest phenolic content (160.33 and 110.90 mg/100 g of fresh weight). The stronger inhibitory effect on DPPH radicals corresponded to those obtained from peel materials. A comparative analysis of the antimicrobial potential against a range of microorganism strains was also carried out. Staphylococcus aureus, Pseudomonas aeruginosa, and Bacillus cereus were the most sensitive to the active extracts. Among the examined phenolic extracts, 'Red Delicious' and quince peels showed the highest effects for inhibiting bacteria growth. Minimum inhibitory and bactericide concentrations ranged from 10(2) to 10(4) microg of polyphenol/mL. Red skin apple and quince peels could be of great interest as important antioxidant and antimicrobial polyphenol sources.