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The transmission of SARS-CoV-2 virus through aerosols has become an outstanding issue, where plenty of spread aspects are being analyzed. Portable Air Cleaners (PAC) with high-efficiency particulate air (HEPA) filters have been discussed as an adjunctive means for indoor environments coronavirus decontamination. This study evaluates, first, the air and surfaces SARS-COV-2 RNA contamination due to positive patients in households, and second, the efficiency of a PAC with HEPA filter to eliminate virus. A total of 29 air and surface samples were collected inside 9 households, by using an air portable collector with gelatin filters and swabs. SARS-CoV-2 RNA detection was performed using real-time reverse transcription polymerase chain reaction (RT-PCR). Overall, all the air samples collected before using PAC and 75% of swab samples were positive for SARS-CoV-2. After the PAC usage, all samples except one were negative, displaying a 80% device effectiveness. Portable HEPA cleaners usage allowed the removal of SARS CoV-2 and, therefore, they could be recommended for places with inadequate ventilation, considering the limitations and functionality of the device., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021 The Authors. Published by Elsevier B.V. All rights reserved.)

A novel and multidisciplinary observational analysis of atmospheric components in the Central Iberian Peninsula is presented here. PM 2.5 concentrations and both populations of cultivable and non-cultivable microorganisms and concentrations of a wide range of trace elements associated have been simultaneously studied during multiple events along one year. The aim has been to characterize their potential relations and dependencies, and their seasonal, daily and hourly evolution. Tools that could explain the atmospheric mechanisms and sources from all these elements have been also evaluated. As it would be expected from a suburban environment, absolute levels obtained were not close to legislation limits. Anthropogenic and natural sources, such as heating home, soil resuspension, or Sahara dust intrusion; and atmospheric factors are responsible for higher PM 2.5 and metals concentrations in months with both low and high temperatures. Daily and hourly evolution depends on University Campus activity, especially on traffic flow and resuspended dust due to human transit. No statistical significant differences on daily or seasonal scales between cultivable counts of fungi and bacteria were displayed. However, using the q-PCR technique, the bacterial population was lower in winter. Positive correlations between PM 2.5 and relative humidity; and PM 2.5 and cultivable microorganism have been established. It was also the case among 7 of the 11 trace elements, indicating then common natural or anthropogenic sources. In summary, this work illustrates the interest of a combined inspection of elements, interactions and dependencies when studying the unique and continuous atmospheric environment, which are typically analysed separately., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Milk is a typical and satisfactory medium for the growth of lactic acid bacteria (LAB). These microorganisms are of vital importance in the quality of the milk since they contribute to its preservation and give differential organoleptic properties to the final product. Furthermore, LABs can act as biocontrol agents in the dairy industry by inhibiting the growth of undesirable bacteria present in milk and by improving the quality of dairy products such as cheese. In this context, knowing the transfer routes used by LABs from the livestock environment to the milk is of great importance within the dairy industry. Therefore, the objectives of the present study were to expand the knowledge of the LAB population present in the milk of Manchego ewe by means of DNA sequencing techniques and to evaluate the possible transfers of LAB species based on the management of each dairy farm. Samples of bulk tank milk, air (from the milking parlour and from the livestock housing), animal feed and teat surface (taken from 10 sheep per farm) were collected in 12 traditional livestock farms in Castilla-La Mancha (Spain), where each farm presented differences regarding their farming practices. A mixed-effects model was used to evaluate the effects of livestock practices on the distribution of LAB species. Results showed that the vast majority of species identified in the milk had an isolate that was also found in other matrices, which could indicate a microbial transference via the livestock environment to the milk. In addition, the mixed model showed that the factors that positively influence the LAB count were the low-line milking system and the daily use of acid detergent in cleaning the milking machine.

Staphylococci are one of the main microorganisms responsible for intramammary infections in sheep, causing important economic losses for farmers and eventually health problems in humans, especially by the consumption of dairy products made with raw milk containing toxic compounds, such as biogenic amines or antibiotic resistant bacteria. This study aimed to check the presence and safety of staphylococci in bulk tank ewe's milk from different farms, and to determine the relationship between the presence of these staphylococci and farming practices, by applying nonlinear canonical correlation models (OVERALS). Two-hundred and fifty-nine staphylococci from milk samples from eighteen farms were genotyped and representative isolates of the major clusters were identified as belonging to Staphylococcus (S.) aureus , S. epidermidis , S. arlettae , S. lentus , S. simulans , and S. chromogenes species. Identified isolates were assayed in terms of their safety, by evaluating resistance to antimicrobial drugs and the aminobiogenic capacity, using both phenotypic and genetic assays. Antibiotic resistance phenotypic assay revealed that 82.9% were resistant to some antibiotics, although in the genotypic assay only the genes tet M, erm B, erm C, and grl A were detected. Fifty-three percent were high biogenic amine (BA) producers, being putrescine the most produced amine. A lowered risk of finding antibiotic-resistant and BA-producing staphylococci is related to some farming methods such as enrolling in a breeding program, use of good farming practices, postdipping teat disinfection, hygienic livestock housing, or periodic check of the milking machine.

The aim of this study was to evaluate the air quality of an indoor swimming pool, analyzing diurnal and seasonal variations in microbiological counts and chemical parameters. The results indicated that yeast and bacteria counts, as well as carbon dioxide (CO 2 ), nitrogen oxides (NO x ) and O 3 concentrations, showed significant diurnal difference. On the other hand, temperature, relative humidity (R.H.), yeast counts and concentrations of CO 2 , particles, O 3 , toluene, and benzene showed seasonal differences. In addition, the relationship between indoor and outdoor air and the degree of correlation between the different parameters have been calculated, suggesting that CO 2 , fine particles and NO x would have indoor origin due to the human activity and secondary reactions favored by the chemical and environmental conditions of the swimming pool; while O 3 , benzene and toluene, would come from outside, mainly. The overall results indicated that indoor air quality (IAQ) in the swimming pool building was deficient by the high levels of CO 2 and microorganisms, low temperatures, and high R.H., because frequently the limits established by the legislation were exceeded. This fact could be due to the poor ventilation and the inadequate operation of heating, ventilation, and air-conditioning systems., (© 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

The aim of this study was to assess the species and the genetic diversity of the staphylococci population in raw milk from healthy goats. Isolates representative of all genotypes were screened for their potential pathogenicity by the occurrence of some relevant safety-related properties, such as antibiotic resistance, presence of virulence factor genes, biofilm formation ability and biogenic amine production. A total of 314 staphylococci were isolated, and randomly amplified polymorphic DNA-PCR analysis displayed 48 genotypes. Isolates were identified as belonging to S. epidermidis (87.5%), S. caprae (6.2%), S. aureus (4.2%) and S. simulans (2.1%) species. The antibiotic resistance varied strongly with strains, with S. epidermidis and S. aureus strains showing resistance to more number of antibiotics. A high occurrence of strains harbouring hemolysin genes was also found in both species. On the contrary, none of the strains assayed harboured enterotoxin or amino acid decarboxylase genes, and, although a moderate or high biofilm formation was observed in 29% of the strains, they did not harbour icaA or icaD genes. This study gives a first and extensive picture of safety-related properties within Staphylococcus species isolated from milk of healthy goats, displaying that these species can act as a reservoir for spreading genes related to safety., (Copyright © 2016 Elsevier B.V. All rights reserved.)

The Oenococcus oeni populations of Tempranillo wines from Castilla La Mancha and La Rioja winemaking regions were analysed from one to three years and up to ten wineries. The objective was to evaluate the genetic variability and the O. oeni population structure. For this purpose a MLST scheme based on four loci (gyrB, purK, pgm and recP genes) and PFGE with SfiI restriction enzyme were developed for later combination. The results showed an O. oeni population completely adapted to winemaking regions. A purifying selection influenced the genes evolution, especially recP that along with purK were the most interesting loci to analyse the genetic variability of the isolates. In this way linkage disequilibrium and intergenic and intragenic recombination were determined between isolates. PFGE typing with UPGMA data were not coincident with the phylograms assessed for MLST by Maximum likelihood and combination of both techniques differentiated all the isolates as strains. Those results led the research to conclude that O. oeni population from CM and LR was a panmictic population with a slight clonal evolution, so subpopulations could not be described. A broader study including more winemaking regions with different grape varieties and distinct ways of elaborating would be interesting to complete the knowledge about O. oeni populations., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

A study has been carried out in order to determine the effect of the lactic acid bacteria inoculation time on the kinetic of vinification and on chemical and sensory characteristics of Cabernet Franc wines. Traditional vinifications, with lactic acid bacteria inoculated after completion of alcoholic fermentation were compared with vinifications where yeast and bacteria were co-inoculated at the beginning of vinification. One commercial yeast strain and an autochthonous Oenococcus oeni strain (C22L9), previously identified and selected at our laboratory, were used. Monitoring of alcoholic and malolactic fermentations was carried out by yeast and lactic acid bacteria counts and by measuring l-malic acid concentration. Wines were chemically characterized and analysed for volatile compounds content. A sensory analysis, consisting of a descriptive and a triangular test, was also carried out. Results from this study showed that the concurrent yeast/bacteria inoculation of musts at the beginning of vinification produced a reduction in duration of the process without an excessive increase in volatile acidity. Differences in volatile compounds content and the corresponding impact on the sensorial profile of wines were also displayed. These results suggest that co-inoculation is a worthwhile alternative for winemaking of Cabernet Franc wines, if compared with traditional post-alcoholic fermentation lactic acid bacteria inoculation., (© The Author(s) 2014 Reprints and permissions: sagepub.co.uk/journalsPermissions.nav.)

The aim of this study was to assess, both quantitatively and qualitatively, the populations of lactic acid bacteria (LAB) and yeasts in air and wine of a winery, in order to evaluate the possible exchange of microorganisms between them. Samples were taken in a winery located in Castilla-La Mancha (Spain) during the winemaking period of two consecutive vintages (2011 and 2012). The microbial composition was determined by using both a culture-dependent method and a culture-independent method, PCR-denaturing gradient gel electrophoresis (PCR-DGGE). In addition, genetic characterization of isolates from plates was carried out. A high diversity of species was detected in air and wine samples from both vintages. Leuconostoc mesenteroides was the predominant lactic acid bacteria in air from both vintages while Oenococcus oeni was the predominant in wine. Saccharomyces cerevisiae was the most frequently isolated yeast in both air and wine. Typing of O. oeni and S. cerevisiae isolates from air and wine samples showed the presence of coincident genotypes in both samples, that would confirm the exchange of microorganisms between the two environments, air and wine, and furthermore some of these genotypes were also found at samples taken at different vintages, indicating that they would remain in the winery. The results display the influence of the activity taking place in the winery and the moment of fermentation of the wines in tanks, on the microorganisms present in the air and the role of the air for the dispersal of microorganisms within the winery., (Copyright © 2014 Elsevier B.V. All rights reserved.)

The aim of this study was the genetic characterisation and safety evaluation of 129 Enterococcus isolates obtained from wine undergoing malolactic fermentation. Genetic characterisation by randomly amplified polymorphic DNA-PCR displayed 23 genotypes. 25 isolates representative of all genotypes were identified as Enterococcus faecium by species-specific PCR and assayed for antibiotic resistance, presence of virulence genes and aminobiogenic capacity, both in decarboxylase medium and wine. The aminobiogenic capacity in wine was analysed in presence (assay 1) and absence (assay 2) of Oenococcus oeni CECT 7621. Resistance to tetracycline, cotrimoxazol, vancomycin and teicoplanin was exhibited by 96% of the strains, but none of them harboured the assayed virulence genes. All of the strains harboured the tyrosine decarboxylase (tdc) gene, while 44% were positive for tyramine in decarboxylase medium. Only five out of 25 strains survived in wine after seven days of incubation, and when concentrations of biogenic amines in wines were determined by HPLC, only those wines in which the five surviving strains occurred contained biogenic amines. Histamine, putrescine and cadaverine were detected in wines from both assays, although concentrations were higher in assay 2. Tyramine and phenylethylamine were detected only in absence of O. oeni. This research contributes for the knowledge of safety aspects of enterococci related to winemaking., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

The goal of this study was to examine the esterase activity of 243 lactic acid bacteria (LAB) strains from wines of different red grape varieties, belonging to the genera Oenococcus, Lactobacillus, Pediococcus and Enterococcus. p-Nitrophenyl octanoate was used as substrate. All strains presented esterase activity in the first screening, but only those showing higher activity were used in subsequent studies to determine the cellular location of this activity, the influence of pH, temperature and the presence of ethanol and the substrate specificity. For the thirteen selected strains, the highest activity was observed in the intracellular fraction. Responses to pH, temperature and ethanol were strain-dependent, but for all the strains, a marked decrease in activity in presence of ethanol was observed. When the influence of pH and ethanol acting together was studied at 25 °C and 37 °C, temperature-dependent differences were not observed for any of the strains except for Oen6. In the substrate specificity assay, the majority of strains of all genera displayed a trend to more readily hydrolyse ester substrates from C8 and longer., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Unlabelled: Different wine varieties, including some with low pH, were studied to determine the ability to grow and produce secondary metabolites of a previously selected autochthonous Oenococcus oeni strain (C22L9), compared with a commercial strain. Monitoring of malolactic fermentation (MLF) was carried out by microbiological and chemical analysis of wines. The concentration of some major volatile compounds and biogenic amines in wines before and after malolactic fermentation was also determined. The results showed major differences in MLF duration both between wines and strains, although the differences between strains were slight for most of the analyzed compounds. Statistically significant differences in citric acid degradation were found in all wine varieties and it was confirmed that O. oeni C22L9 is a poor degrader of citric acid; this means that MLF can be prolonged without the risk of producing high concentrations of diacetyl and acetoin. Sensory analysis of wines after MLF showed similar characteristics in wines from both strains. This study thus shows that O. oeni C22L9 possesses even better sensory and fermentation properties than the commercial strain and can be used in wines with different characteristics, which makes it highly valuable for industrial use., Practical Application: The increasingly use of grape varieties of low pH in winemaking and the higher alcohol content of wines, as a consequence of the climatic change, make interesting the study of the behavior during MLF of O. oeni strains in order to determine their ability to grow, when growth conditions are not optimal, and to produce secondary metabolites. A comparative study was conducted using an autochthonous O. oeni strain (C22L9) and a commercial O. oeni strain and 4 wine varieties., (© 2012 Institute of Food Technologists®)

The aim of this study was to evaluate the ability from a number of lactic acid bacteria isolated from different sources to produce glycosidase enzymes. Representative isolates (225) from clusters obtained after genotyping, using randomly amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) analysis, of 1,464 isolates, were screened for β-D-glucosidase activity. Thirty-five of them were selected for subsequent analysis. These strains were able to hydrolyze α-D-glucopyranoside, β-D-xylopyranoside and α-L-arabinofuranoside although β-D-glucosidase activity was the predominant activity for 22 of the selected strains. Only some of them did so with α-L-rhamnopyranoside. All of these were from wine samples and were identified as belonging to the Oenococcus oeni species using Amplification and Restriction Analysis of 16S-rRNA gene (16S-ARDRA). When the influence of pH, temperature and ethanol or sugars content on β-D-glucosidase activity was assayed, a strain-dependent response was observed. The β-D-glucosidase activity occurred in both whole and sonicated cells but not in the supernatants from cultures or obtained after cell sonication. Strains 10, 17, 21, and 23 retained the most β-D-glucosidase activity when they were assayed at the conditions of temperature, pH, ethanol and sugar content used in winemaking. These results suggest that these strains could be used as a source of glycosidase enzymes for use in winemaking.

The bacterial population during malolactic fermentation of Tempranillo wine was studied using the polymerase chain reaction-denaturing gradient gel electrophoresis, a culture-independent method successfully used for identification and monitoring of bacterial population in different habitats included food fermentations. The results showed that Oenococcus oeni was the predominant species in the malolactic fermentation of Tempranillo wines, although the presence of Gluconobacter oxydans, Asaia siamensis, Serratia sp., and Enterobacter sp. was also observed. These results were partly coincidental with those obtained from a culture-dependent method, using a selective medium. Therefore, it may be concluded that for a more complete knowledge of the bacterial community present during malolactic fermentation of Tempranillo wine, an approach that combines a culture-independent method and a culture-dependent method would be advisable.

The goal of this study is to carry out a characterization of 84 Oenococcus oeni strains isolated from Tempranillo wine samples taken at the cellars in Castilla-La Mancha, in order to select those showing the highest potential as oenological starter cultures. Various oenological properties were analyzed and the ability of some of these strains to grow and undergo MLF in simulated laboratory microvinifications was tested. Twenty-two strains were selected on the basis of fermentation assays and the eight that produced the best results in the chemical analysis of the wines were chosen for further assays. None of the eight strains was either able to produce biogenic amines or displayed tannase or anthocyanase activities. On the other hand all presented activity against p-NP-beta Glucopyranoside, p-NP-alpha Glucopyranoside and p-NP-beta xylopyranoside. Randomly Amplified Polymorphic DNA (RAPD)-PCR was used to determine the colonizing ability of the inoculated strains. C22L9 and D13L13 strains showed the highest implantation values. On the basis of this characterization, two strains have been selected which are suitable as starter cultures for MLF of Tempranillo wine. Use of these strains will ensure that MLF proceeds successfully and gives retention of the organoleptic characteristics of wines made in Castilla-La Mancha., ((c) 2009 Elsevier B.V. All rights reserved.)

The genetic diversity of 339 strains of lactic acid bacteria responsible for the spontaneous malolactic fermentation (MLF) in Tempranillo wines produced in six cellars located in different provinces of Castilla-La Mancha region of Spain was analyzed by using randomly amplified polymorphic DNA (RAPD). The results indicate that Oenococcus oeni was the predominant species in MLF of Tempranillo wines with a scant diversity of other lactic acid bacteria (LAB) species. The presence of a well-adapted cellar microbiota is confirmed, with MLF dominated by a limited number of O. oeni genotypes. Clustering of isolates from samples taken from different provinces showed the existence of coincident genotypes in different cellars of the region. This result suggests that a population of cosmopolitan O. oeni strains exists in the Castilla-La Mancha wine-growing region.

Three molecular techniques, randomly amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR), pulsed field gel electrophoresis (PFGE) and differential display-polymerase chain reaction (DD-PCR) have been used to assess the intraspecific diversity of the lactic microbiota responsible for spontaneous malolactic fermentation (MLF) in Cencibel wines made at a cellar in Castilla-La Mancha (Spain). The results indicate that RAPD-PCR and PFGE are of value in typing this microbiota. Better discrimination was achieved by RAPD-PCR. Reproducibility using DD-PCR was not good, which makes this method unsuitable. Combined numerical analysis of the patterns obtained from RAPD-PCR and PFGE allowed a better discrimination; this would therefore be a suitable tool to discriminate the diversity of bacterial communities like those found in MLF of wines. Genetic diversity data from combined numerical analysis suggest that there is considerable microbial diversity within MLF of Cencibel wines, with some genotypes coinciding in the two vinifications analysed.

Aim: Identification of the predominant lactic acid bacteria (LAB) involved in spontaneous fermentations of Almagro eggplants, and evaluation of the biodiversity by molecular typing., Methods and Results: Almagro eggplant fermentations in three factories (A, B and C) enjoying Protected Designation of Origin (PDO) status were monitored by chemical and microbiological analysis of brines. LAB isolates from brines were identified by phenotypic analysis and by species-specific PCR reactions and typed by randomly amplified polymorphic DNA (RAPD)-PCR. All isolates from factories A and C belonged to the genus Lactobacillus (Lact.), whereas isolates from factory B belonged to Lactobacillus (50%), Leuconostoc (Ln.) (25%) and Lactococcus (Lc.) (25%); 1.9% of this microbiota was considered cosmopolitan. The genera Leuconostoc and Lactococcus and the species Lact. acidophilus and Lact. paracasei had never previously been reported in Almagro eggplant fermentations., Conclusion: Considerable differences in the composition of the lactic acid microbiota participating in the Almagro eggplant fermentations exist. Brine NaCl concentration has a notable influence both in number and in the species participating., Significance and Impact of the Study: The original aspect of this work consists of an ecological study of the LAB taking part in spontaneous Almagro eggplant fermentations from different factories. Participation of Leuconostoc and Lactococcus species and of Lact. acidophilus and Lact. paracasei, which had never before been described for this pickle, and the evidence that a lactic fermentation does not always take place, were the most relevant results.