This study evaluated the microbial quality, safety, and ecology of a retail delicatessen Galotyri-like fresh acid-curd cheese traditionally produced by mixing fresh natural Greek yogurt with ...‘Myzithrenio’, a naturally fermented and ripened whey cheese variety. Five retail cheese batches (mean pH 4.1) were analyzed for total and selective microbial counts, and 150 presumptive isolates of lactic acid bacteria (LAB) were characterized biochemically. Additionally, the most and the least diversified batches were subjected to a culture-independent 16S rRNA gene sequencing analysis. LAB prevailed in all cheeses followed by yeasts. Enterobacteria, pseudomonads, and staphylococci were present as <100 viable cells/g of cheese. The yogurt starters Streptococcus thermophilus and Lactobacillus delbrueckii were the most abundant LAB isolates, followed by nonstarter strains of Lactiplantibacillus, Lacticaseibacillus, Enterococcus faecium, E. faecalis, and Leuconostoc mesenteroides, whose isolation frequency was batch-dependent. Lactococcus lactis isolates were sporadic, except for one cheese batch. However, Lactococcus lactis, Enterobacteriaceae, Vibrionaceae, Salinivibrio, and Shewanellaceae were detected at fairly high relative abundances culture-independently, despite the fact that their viable counts in the cheeses were low or undetectable. Metagenomics confirmed the prevalence of S. thermophilus and Lb. delbrueckii. Overall, this delicatessen Galotyri-like cheese product was shown to be a rich pool of indigenous nonstarter LAB strains, which deserve further biotechnological investigation.
Nowadays, modification of surfaces by nanoparticulate coatings is a simple process that may have applications in reducing the prevalence of bacterial cells both on medical devices and food processing ...surfaces. To this direction, biofilm biological cycle of
Typhimurium,
O157:H7,
, and
on stainless steel and glass surfaces, with or without nanocoating was monitored. To achieve this, four different commercial nanoparticle compounds (two for each surface) based on organo-functionalized silanes were selected. In total 10 strains of above species (two for each species) were selected to form biofilms on modified or not, stainless steel or glass surfaces, incubated at 37°C for 72 h. Biofilm population was enumerated by bead vortexing-plate counting method at four time intervals (3, 24, 48, and 72 h). Organosilane based products seemed to affect bacterial attachment on the inert surfaces and/or subsequent biofilm formation, but it was highly dependent on the species and material of surfaces involved. Specifically, reduced bacterial adhesion (at 3 h) of
and
was observed (
< 0.05) in nanocoating glass surfaces in comparison with the control ones. Moreover, fewer
and
biofilm cells were enumerated on stainless steel coupons coated with organosilanes, than on non
coated surfaces at 24 h (
< 0.05). This study gives an insight to the efficacy of organosilanes based coatings against biofilm formation of foodborne pathogens, however, further studies are needed to better understand the impact of surface modification and the underlying mechanisms which are involved in this phenomenon.
Current information from conventional microbiological methods on the microbial diversity of table olives is insufficient. Next-generation sequencing (NGS) technologies allow comprehensive analysis of ...their microbial community, providing microbial identity of table olive varieties and their designation of origin. The purpose of this study was to evaluate the bacterial and yeast diversity of fermented olives of two main Greek varieties collected from different regions—green olives, cv. Halkidiki, from Kavala and Halkidiki and black olives, cv. Konservolia, from Magnesia and Fthiotida—via conventional microbiological methods and NGS. Total viable counts (TVC), lactic acid bacteria (LAB), yeast and molds, and Enterobacteriaceae were enumerated. Microbial genomic DNA was directly extracted from the olives’ surface and subjected to NGS for the identification of bacteria and yeast communities. Lactobacillaceae was the most abundant family in all samples. In relation to yeast diversity, Phaffomycetaceae was the most abundant yeast family in Konservolia olives from the Magnesia region, while Pichiaceae dominated the yeast microbiota in Konservolia olives from Fthiotida and in Halkidiki olives from both regions. Further analysis of the data employing multivariate analysis allowed for the first time the discrimination of cv. Konservolia and cv. Halkidiki table olives according to their geographical origin.
A geographical and seasonal assessment of indigenous lactic acid bacteria (LAB) in Maltese cow milk was conducted in this study. To investigate this, milk was collected from different regions of ...Malta during winter and summer seasons. Total viable counts (TVC) and LAB population were enumerated. Afterwards, LAB were isolated and identified by molecular methods. According to the results, similar TVC were enumerated on winter and summer samples, while highest LAB population was detected on summer samples. LAB isolates were grouped in seven different clusters which were assigned to Lactobacillus casei, Pediococcus pentosaceus, Lactobacillus plantarum, Weissella paramesenteroides, Lactobacillus rhamnosus, Lactococcus lactis, and Lactococcus garvieae. In addition, Enterococcus and Streptococcus species were also isolated. Season seemed to affect the genus/species of LAB since Lactobacillus were mainly isolated from winter samples, while Lactococcus and Enterococcus species were the main genera identified in summer samples. Regarding the geographical distribution, the majority of the Lactobacillus spp. were isolated from the South-eastern region in both seasons. In conclusion, through this study the diversity of indigenous LAB in the Maltese cow milk was monitored for the first time and highlighted that the microbial communities are affected by seasonality and geographical distribution of the farms.
Salmonella is recognized as one of the most significant enteric foodborne bacterial pathogens. In recent years, the resistance of pathogens to biocides and other environmental stresses, especially ...when they are embedded in biofilm structures, has led to the search for and development of novel antimicrobial strategies capable of displaying both high efficiency and safety. In this direction, the aims of the present work were to evaluate the antimicrobial activity of hydrosol of the Mediterranean spice Thymbra capitata against both planktonic and biofilm cells of Salmonella enterica serovar Typhimurium and to compare its action with that of benzalkonium chloride (BC), a commonly used industrial biocide. In order to achieve this, the disinfectant activity following 6-min treatments was comparatively evaluated for both disinfectants by calculating the concentrations needed to achieve the same log reductions against both types of cells. Their bactericidal effect against biofilm cells was also comparatively determined by in situ and real-time visualization of cell inactivation through the use of time-lapse confocal laser scanning microscopy (CLSM). Interestingly, results revealed that hydrosol was almost equally effective against biofilms and planktonic cells, whereas a 200-times-higher concentration of BC was needed to achieve the same effect against biofilm compared to planktonic cells. Similarly, time-lapse CLSM revealed the significant advantage of the hydrosol to easily penetrate within the biofilm structure and quickly kill the cells, despite the three-dimensional (3D) structure of Salmonella biofilm.
The results of this paper highlight the significant antimicrobial action of a natural compound, hydrosol of Thymbra capitata, against both planktonic and biofilm cells of a common foodborne pathogen. Hydrosol has numerous advantages as a disinfectant of food-contact surfaces. It is an aqueous solution which can easily be rinsed out from surfaces, it does not have the strong smell of the essential oil (EO) and it is a byproduct of the EO distillation procedure without any industrial application until now. Consequently, hydrosol obviously could be of great value to combat biofilms and thus to improve product safety not only for the food industries but probably also for many other industries which experience biofilm-related problems.
A total of 266 lactic acid bacteria (LAB) have been isolated from minced beef stored at 0, 5, 10 and 15 °C aerobically and under modified atmosphere packaging consisting of 40% CO
2–30% O
2–30% N
2 ...in the presence MAP (+) and absence MAP (−) of oregano essential oil. Sequencing of their 16S rRNA gene along with presence of the
katA gene demonstrated dominance of the LAB microbiota by
Leuconostoc spp. during aerobic storage at 5, 10 and 15 °C, as well as during MAP (−) and MAP (+) storage at 10 and 15 °C;
Lactobacillus sakei prevailed during aerobic storage at 0 °C, as well as at MAP (−) and MAP (+) storage at 0 and 5 °C. The sporadic presence of other species such as
Leuconostoc mesenteroides,
Weisella viridescens,
Lactobacillus casei and
Lactobacillus curvatus has also been determined. Pulsed-Field Gel Electrophoresis of high molecular weight genomic DNA revealed the dynamics of the isolated LAB strains. Prevalence of
Leuconostoc spp. was attributed to one strain only. On the other hand, packaging conditions affected
Lb. sakei strain spoilage dynamics.
The yeast and lactic acid bacteria (LAB) community structure and dynamics was assessed during storage of Conservolea natural black olives in different modified atmospheres. Black olives were packed ...without brine in polyethylene pouches under air and modified atmospheres (20%CO2–80%N2 and 40%CO2–30%O2–30%N2) and stored at 4 °C for a period of ca. 4 months. Yeast and LAB species diversity was evaluated before packaging, at the early (4 days), middle (77 days) and final (128 days) stages of storage. Yeast identification was based on restriction fragment length polymorphism and sequence analyses of the 5.8S internal transcribed spacer and the D1/D2 ribosomal DNA (rDNA) regions of isolates. Multiplex PCR amplification with recA gene derived primers was performed for the bacilli isolates for the differentiation of Lactobacillus plantarum group, while the remaining isolates were subjected to species identification by sequencing the V1-V3 variable region of the 16S rRNA gene. LAB heterogeneity at strain level was performed with Pulsed Field Gel Electrophoresis (PFGE). Analysis revealed a LAB community that composed of 5 species and 41 strains. The highest heterogeneity regarding strain frequency was observed for Lactobacillus pentosus with 20 strains, followed by L. plantarum with 9 strains. For yeasts, Pichia membranifaciens dominated in all pouches regardless of gas composition with frequency higher than 80% throughout storage. The composition of the modified atmosphere 20%CO2–80%N2 remained stable during storage. However, the presence of O2 in the aerobic and 40%CO2–30%O2–30%N2 pouches favoured the respiratory activity of yeasts resulting in a change in the initial gas composition with O2 depletion and CO2 accumulation.
► Lactic acid bacteria heterogeneity was affected by gas composition in the pouches. ► Lactic acid bacteria community consisted of 5 species and 41 strains. ► The highest strain frequency was observed for L. pentosus and L. plantarum. ► Yeast community was dominated by P. membranifaciens.
Anigrides Nymphes of Lake Kaiafas is a thermal spring that is well known for its therapeutical properties, as the hot water (32-34 °C) is rich in sulfur compounds and minerals. Nowadays, efforts are ...made from the Hellenic Republic to modernize the existing facilities and infrastructure networks of the area. To study the complex ecosystem of the thermal spring, we collected water from four sampling points (Lake, and Caves 1, 2, and 3). Filtration method was used for microbial enumeration. In parallel, total bacterial DNA was extracted and subjected to next-generation sequencing (NGS). A total of 166 different bacterial families were detected. Differences in families, genera, and species abundances were detected between the different sampling points. Specifically,
was the most common family detected in Lake and Cave 3. Similarly, in Caves 1 and 2,
was detected at a higher percentage compared to the rest of the families. Moreover, the detection of sequences assigned to waterborne or opportunistic pathogens, i.e.,
,
,
, and
, as well as
and
, is of great importance. Although the presence of pathogens was not examined by quantitative PCR, the detection of their sequences strengthens the need of the planned rehabilitation actions of this natural environment in order to allow human swimming.
The potential of biofilm formation of multifunctional starters Lactobacillus pentosus B281 and Pichia membranifaciens M3A during inoculated fermentation of Conservolea natural black olives according ...to Greek-style processing was investigated. Olives were directly brined in 8% (w/v) NaCl following three fermentation procedures namely, i) spontaneous fermentation, ii) inoculated fermentation with L. pentosus B281, and iii) co-inoculated fermentation with L. pentosus B281 and P. membranifaciens M3A. Lactic acid bacteria (LAB) and yeasts were monitored on olives by plate counting for a period of 153 days, whereas the survival of the inoculated strains was confirmed by Pulsed Field Gel Electrophoresis (PFGE) and Restriction Fragment Length Polymorphism (RFLP) analysis. Inoculated fermentation with L. pentosus B281 with/without the presence of the yeast resulted in higher acidification of the brine compared to the spontaneous process where no indigenous LAB could be enumerated. The population of LAB on olives ranged between 5.5 and 6.5 log CFU/g and it was maintained at higher levels compared to yeasts (3.5–4.5 log CFU/g) throughout the process. PFGE analysis revealed that L. pentosus B281 could successfully colonize the surface of black olives presenting high recovery rate (100%) at the end of fermentation in contrast to P. membranifaciens M3A that was successfully recovered (42%) only after 72 days of the process. The obtained results provide interesting perspectives for the production of natural black olives with functional properties.
•Lactobacillus pentosus and Pichia membranifaciens were used as starters in black olive fermentation.•Their in situ imposition on olives was studied in order to form biofilms.•L. pentosus could dominate the process and recovered 100% at the end of fermentation.•P. membranifaciens was recovered in the middle but not in the end of the process.•L. pentosus could be used as a multifunctional starter in black olive fermentation.
In the present study, the succession of the biota grown on a selective medium for pseudomonads (pseudomonas agar based medium – PAB) during the storage of meat under different conditions was ...monitored. Thus, minced beef was stored aerobically and under modified atmosphere packaging in the presence (MAP+) and absence (MAP−) of oregano essential oil at 0, 5, 10 and 15 °C. A total of 267 pure cultures were recovered from PAB throughout the storage period and subjected to PCR-Denaturing gradient gel electrophoresis (PCR-DGGE) for their differentiation. In parallel, the direct analysis of the whole cultivable community (WCC) from the same medium was applied. These two approaches were used in order to indicate the lack of selectivity. Fifteen different DGGE fingerprints were obtained after PCR – DGGE analysis of the isolates, which were assigned to Pseudomonas putida (3 fingerprints), Pseudomonas fragi and Pseudomonas fluorescens, Pseudomonas spp., Serratia liquefaciens (2), Citrobacter freundii, Serratia grimesii, Hafnia alvei (3), Rahnella spp. and Morganella morganii. Twelve of them occurred during the direct analysis of the WCC. The biota succession found to be affected from the different storage conditions. However, the outcome of the two strategies was quite different, which is leading to the use of different appropriated molecular approaches in order to widen the knowledge of bacterial succession of meat.
► Succession of the biota of a selective growth medium for pseudomonads was studied. ► PCR-denaturing gradient gel electrophoresis was applied. ► Biota succession found to be affected from the different storage conditions. ► Pseudomonas fragi was detected in the most conditions adopted. ► Pseudomonas putida was found unable to grow under elevated temperatures.
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