Nutraceutical Potential of Lentinula edodes' Spent Mushroom Substrate: A Comprehensive Study on Phenolic Composition, Antioxidant Activity, and Antibacterial Effects.

Lentinula edodes, commonly known as shiitake mushroom, is renowned for its potential health advantages. This research delves into the often-overlooked by-product of shiitake cultivation, namely spent mushroom substrate (SMS), to explore its nutraceutical properties. The SMS samples were collected and subjected to different extraction methods, namely short or long agitation, and ultrasound-assisted extractions using different temperatures and distilled water or a 50% (v/v) ethanol as solvents. The extracts were tested for phenolic content (total phenols, ortho-diphenols, and flavonoids), antioxidant capacity (DPPH, 2,2-diphenyl-1 picrylhydrazyl; ABTS, 2,2'-azino-bis-3-ethylbenzothiazoline-6-sulfonic acid; and FRAP, ferric reducing antioxidant power), and antibacterial activity. The different extraction methods revealed substantial variations (p < 0.05) in phenolic composition and antioxidant capacity. The highest phenolic content and antioxidant capacity were achieved using 24 h extraction, agitation, 50 °C, and ethanol as the solvent. Furthermore, the extracted compounds displayed antibacterial activity in specific tested bacterial strains. This study highlights the nutraceutical potential of L. edodes' SMS, positioning it as a valuable dietary supplement for animal nutrition, with emphasis on its prebiotic properties. Hence, this research unveils the promising health benefits of SMS in both human and animal nutrition.

Hydrocinnamic acid and perillyl alcohol are effective against Escherichia coli biofilms when used alone and combined with antibiotics.

AIMS: The use of phytochemicals to improve the effectiveness of antibiotics is a promising strategy for the development of novel antimicrobials. In this study, the antibiofilm activity of perillyl alcohol and hydrocinnamic acid, both phytochemicals present in several plants, and two antibiotics from different classes (amoxicillin and chloramphenicol) was tested, alone and in combination, against Escherichia coli. METHODS AND RESULTS: Each molecule was tested at the minimum inhibitory concentration (MIC), 5 × MIC, and 10 × MIC, and characterized concerning biomass removal, metabolic inactivation, and cellular culturability. The highest percentages of metabolic inactivation (88.5% for 10 × MIC) and biomass reduction (61.7% for 10 × MIC) were obtained with amoxicillin. Interestingly, for 5 × MIC and 10 × MIC, phytochemicals provided a total reduction of colony-forming units (CFUs). Dual and triple combinations of phytochemicals and antibiotics (at MIC and 5 × MIC) demonstrated high efficacy in metabolic inactivation, moderate efficacy in terms of biomass reduction, and total reduction of cellular culturability for 5 × MIC. CONCLUSIONS: The results demonstrated the antibiofilm potential of phytochemicals, highlighting the advantage of phytochemical/antibiotic combinations for biofilm control.

Escherichia coli Phylogenetic and Antimicrobial Pattern as an Indicator of Anthropogenic Impact on Threatened Freshwater Mussels.

Freshwater bivalves are widely used as accumulation indicators and monitoring tools for assessing contaminant effects on different levels of biological integration. This pilot study aimed to explore the phylogenetic diversity of Escherichia coli isolated from freshwater mussels (Margaritifera margaritifera and Potomida littoralis) and characterize their phenotypes and antibiotic resistance profiles. Samples were collected in the Rabaçal and Tua Rivers, in the Douro basin, Portugal-two sites representing different levels of anthropogenic contamination. Antimicrobial susceptibility testing was performed via the disk diffusion method with 21 antibiotics. Results showed that 31% of isolates were multidrug-resistant (MDR). Thus, freshwater mussels provide an effective and time-integrated approach for identifying/quantifying fecal indicators, including MDR bacteria. PCR-based assays were designed for assessing phylogenetic E. coli groups. Among the E. coli isolates, the highest prevalence (44%) was observed in group D or E, followed by group E or Clade I (25%), group A (19%), and group B1 (13%). E. coli isolated from M. margaritifera predominantly exhibited a higher prevalence of phylogroups D or E, whereas E. coli from P. littoralis showed associations with phylogroups E or clade I, B1, A, and D or E. Our results provide new insights into the phylogenetic diversity of E. coli in freshwater bivalves. Additionally, the findings highlight the possible linkage of phylogroups with the host species, the geographical location in the water stream, and human activity. Using E. coli as a bioindicator isolated from freshwater mussels helps us grasp how human activities affect the environment. This study has important implications for those interested in safeguarding water resources, especially in tackling antibiotic resistance in aquatic ecosystems.

An Overview of Cefiderocol's Therapeutic Potential and Underlying Resistance Mechanisms.

Antimicrobial resistance continues to increase globally and treatment of difficult-to-treat (DTT) infections, mostly associated with carbapenem-resistant (CR) Pseudomonas aeruginosa, CR Acinetobacter baumannii, and CR- and third-generation-cephalosporins-resistant Enterobacterales remains a challenge for the clinician. The recent approval of cefiderocol has broaden the armamentarium for the treatment of patients with DTT infections. Cefiderocol is a siderophore cephalosporin that has shown excellent antibacterial activity, in part due to its innovative way of cell permeation. It is relatively stable compared to most commonly found carbapenamases. However, some resistant mechanisms to cefiderocol have already been identified and reduced susceptibility has developed during patient treatment, highlighting that the clinical use of cefiderocol must be rational. In this review, we summarize the current available treatments against the former resistant bacteria, and we revise and discuss the mechanism of action of cefiderocol, underlying the biological function of siderophores, the therapeutic potential of cefiderocol, and the mechanisms of resistance reported so far.

Biofilm Production by Critical Antibiotic-Resistant Pathogens from an Equine Wound.

As in human medicine, in veterinary medicine, chronic wounds are often related to polymicrobial infections and the presence of a biofilm, which compromises the effectiveness of therapeutic approaches. In this study, a Lusitano mare presented a 21-day-old chronic wound that was only being treated with an antiseptic. A swab sample was collected, and three isolates of Staphylococcus aureus and one of Pseudomonas aeruginosa were isolated. S. aureus did not show resistance to a panel of antibiotics. However, the P. aeruginosa isolate showed a resistance profile to carbapenems and fluoroquinolones, which may suggest a cross-resistance between antiseptic and antibiotics, given that no antibiotic therapy was applied to the wound or the mare in the previous year. Further experiments were conducted to assess the ability of the isolates to form biofilms, and to ascertain their susceptibility to gentamicin. The results demonstrated that the isolates produced biofilms. Gentamicin at the minimum inhibitory concentration (MIC) and 10× MIC caused biofilm removal between 59.3% and 85.7%, with the highest removal percentage being obtained for the P. aeruginosa isolate (at 10× MIC concentration). This study reveals that an equine wound was colonized by antibiotic resistant bacteria, and that all the wound colonizers could form biofilms, demonstrating the relevance of an adequate diagnosis and treatment when there is a suspicion of a biofilm-infected wound. It also highlights the possibility of resistance transmission between animals, animals and humans, or animals and the environment.

Equine Gram-Negative Oral Microbiota: An Antimicrobial Resistances Watcher?

Horses are considered as reservoirs of multidrug resistant bacteria that can be spread through the environment and possibly to humans. The aim of this study was to characterize the oral Gram-negative microbiota of healthy horses and evaluate their antimicrobial susceptibility profile in a One Health approach. For this purpose, samples were collected from the gingival margin of healthy horses, free of antimicrobial therapy, cultured in selective mediums, identified, and tested for antimicrobial susceptibility. Fifty-five Gram-negative isolates were identified, with 89.5% being zoonotic and 62% affecting humans, which were also found commonly in the environment. Forty-eight isolates (96%) were MDR. The phenotypic resistance presented as higher to macrolides (81.8%), ß-lactams (55.4%), and quinolones (50%), and lower to sulfonamides (27.3%), tetracyclines, and amphenicols (both with 30.9%). In total, 51.5% of the isolates presented resistance to carbapenems. In addition to being the first report on the commensal oral microbiota of horses and respective susceptibility profile, this study highlights the horse as a valuable sentinel that can control the evolution and transmission of multidrug-resistant bacteria between the "One Health triad" since it is in contact with humans, other animals, and the environment, in different geographic locations.

Drinking-water isolated Delftia acidovorans selectively coaggregates with partner bacteria and facilitates multispecies biofilm development.

Coaggregation plays an important role in the development of multispecies biofilms in different environments, often serving as an active bridge between biofilm members and other organisms that, in their absence, would not integrate the sessile structure. The ability of bacteria to coaggregate has been reported for a limited number of species and strains. In this study, 38 bacterial strains isolated from drinking water (DW) were investigated for their ability to coaggregate, in a total of 115 pairs of combinations. Among these isolates, only Delftia acidovorans (strain 005P) showed coaggregating ability. Coaggregation inhibition studies have shown that the interactions mediating D. acidovorans 005P coaggregation were both polysaccharide-protein and protein-protein, depending on the interacting partner bacteria. Dual-species biofilms of D. acidovorans 005P and other DW bacteria were developed to understand the role of coaggregation on biofilm formation. Biofilm formation by Citrobacter freundii and Pseudomonas putida strains highly benefited from the presence of D. acidovorans 005P, apparently due to the production of extracellular molecules/public goods favouring microbial cooperation. This was the first time that the coaggregation capacity of D. acidovorans was demonstrated, highlighting its role in providing a metabolic opportunity for partner bacteria.

Hydrocinnamic Acid and Perillyl Alcohol Potentiate the Action of Antibiotics against Escherichia coli.

The treatment of bacterial infections has been troubled by the increased resistance to antibiotics, instigating the search for new antimicrobial therapies. Phytochemicals have demonstrated broad-spectrum and effective antibacterial effects as well as antibiotic resistance-modifying activity. In this study, perillyl alcohol and hydrocinnamic acid were characterized for their antimicrobial action against Escherichia coli. Furthermore, dual and triple combinations of these molecules with the antibiotics chloramphenicol and amoxicillin were investigated for the first time. Perillyl alcohol had a minimum inhibitory concentration (MIC) of 256 µg/mL and a minimum bactericidal concentration (MBC) of 512 µg/mL. Hydrocinnamic acid had a MIC of 2048 µg/mL and an MBC > 2048 µg/mL. Checkerboard and time-kill assays demonstrated synergism or additive effects for the dual combinations chloramphenicol/perillyl alcohol, chloramphenicol/hydrocinnamic acid, and amoxicillin/hydrocinnamic acid at low concentrations of both molecules. Combenefit analysis showed synergism for various concentrations of amoxicillin with each phytochemical. Combinations of chloramphenicol with perillyl alcohol and hydrocinnamic acid revealed synergism mainly at low concentrations of antibiotics (up to 2 µg/mL of chloramphenicol with perillyl alcohol; 0.5 µg/mL of chloramphenicol with hydrocinnamic acid). The results highlight the potential of combinatorial therapies for microbial growth control, where phytochemicals can play an important role as potentiators or resistance-modifying agents.

In Vitro Effect of Eucalyptus Essential Oils and Antiseptics (Chlorhexidine Gluconate and Povidone-Iodine) against Bacterial Isolates from Equine Wounds.

Considering the increasing antibiotics resistance, there has been a propensity to replace them with antiseptics when it comes to wound management and treatment. Nevertheless, in recent years, there have been reports regarding resistance to antiseptics by some bacterial strains. There is also concern about the environmental impact of these substances. The aim of this study was to compare the antimicrobial efficacy of antiseptics and eucalyptus essential oils on bacterial strains from horse's wounds. We used twelve Escherichia coli, eight Staphylococcus aureus, two Staphylococcus pseudintermedius, one Staphylococcus vitulinus and one Staphylococcus saprophyticus strains from equine wounds. The effect of Eucalyptus radiata essential oil, Eucalyptus globulus essential oil, povidone-iodine and chlorhexidine gluconate against the isolated strains was evaluated applying the Kirby-Baüer method. Regarding the Escherichia coli strains, E. radiata and the mixture of E. radiata and E. globulus had a better inhibitory effect than antiseptics. E. globulus had a better effect against most Staphylococcus spp. compared to E. radiata. For both Gram-negative and Gram-positive strains tested, chlorhexidine gluconate had a better inhibitory effect than povidone-iodine. The antibacterial efficacy of essential oils highlights their potential to substitute or complement the use of antiseptics and so reduce resistance to antiseptics.

Role of Goats in the Epidemiology of Coxiella burnetii.

Since its first description in the late 1930s, Q fever has raised many questions. Coxiella burnetii, the causative agent, is a zoonotic pathogen affecting a wide range of hosts. This airborne organism leads to an obligate, intracellular lifecycle, during which it multiplies in the mononuclear cells of the immune system and in the trophoblasts of the placenta in pregnant females. Although some issues about C. burnetii and its pathogenesis in animals remain unclear, over the years, some experimental studies on Q fever have been conducted in goats given their excretion pattern. Goats play an important role in the epidemiology and economics of C. burnetii infections, also being the focus of several epidemiological studies. Additionally, variants of the agent implicated in human long-term disease have been found circulating in goats. The purpose of this review is to summarize the latest research on C. burnetii infection and the role played by goats in the transmission of the infection to humans.

Exploring the Antibiotic Resistance Profile of Clinical Klebsiella pneumoniae Isolates in Portugal.

While antibiotic resistance is rising to dangerously high levels, resistance mechanisms are spreading globally among diverse bacterial species. The emergence of antibiotic-resistant Klebsiella pneumoniae, mainly due to the production of antibiotic-inactivating enzymes, is currently responsible for most treatment failures, threatening the effectiveness of classes of antibiotics used for decades. This study assessed the presence of genetic determinants of ß-lactam resistance in 102 multi-drug resistant (MDR) K. pneumoniae isolates from patients admitted to two central hospitals in northern Portugal from 2010 to 2020. Antimicrobial susceptibility testing revealed a high rate (>90%) of resistance to most ß-lactam antibiotics, except for carbapenems and cephamycins, which showed antimicrobial susceptibility rates in the range of 23.5−34.3% and 40.2−68.6%, respectively. A diverse pool of ß-lactam resistance genetic determinants, including carbapenemases- (i.e., blaKPC-like and blaOXA-48-like), extended-spectrum ß-lactamases (ESBL; i.e., blaTEM-like, blaCTX-M-like and blaSHV-like), and AmpC ß-lactamases-coding genes (i.e., blaCMY-2-like and blaDHA-like) were found in most K. pneumoniae isolates. blaKPC-like (72.5%) and ESBL genes (37.3−74.5%) were the most detected, with approximately 80% of K. pneumoniae isolates presenting two or more resistance genes. As the optimal treatment of ß-lactamase-producing K. pneumoniae infections remains problematic, the high co-occurrence of multiple ß-lactam resistance genes must be seen as a serious warning of the problem of antimicrobial resistance.

Occurrence and seasonality of Campylobacter spp. in Portuguese dairy farms.

The growing idea that natural products are better for consumption is creating behaviors that can lead to food safety problems and an increase of healthcare costs. One of the trending products is raw milk, which in some countries is sold by vending machines outside dairy farms. Campylobacteriosis is the most common gastrointestinal infection in humans in the European Union since 2005. Several outbreaks have been associated with the consumption of raw milk contaminated with Campylobacter spp. In the present study, the occurrence and seasonality of Campylobacter spp. in raw cow's milk were determined. Other samples from the dairy farm environment were also analyzed to understand the possible routes of contamination. The study was conducted from November 2020 to September 2021 in randomly selected dairy farms located in northern Portugal. One liter of milk was collected from bulk cooling tanks transported to the laboratory and analyzed within 24 h. Campylobacter spp. detection and identification were performed using real-time PCR methodology and confirmation followed ISO standards. From 100 dairy farms evaluated, the occurrence of Campylobacter spp. was estimated at 4.0 % in raw cow's milk samples. In the samples from the environment of the farms, only contaminated fecal samples were found, corresponding to an occurrence of 4.2 %. Positivity was observed in summer months. The results of this study indicate the potential risk of campylobacteriosis after the consumption of raw milk. Consumers who seek raw milk for health reasons should be aware of the risk, especially if they belong to vulnerable groups. Moreover, it raises the question of how climate change will impact food safety, suggesting that routine surveillance for zoonotic pathogens should be implemented in dairy farms.

Genomic epidemiological analysis of Klebsiella pneumoniae from Portuguese hospitals reveals insights into circulating antimicrobial resistance.

Klebsiella pneumoniae (Kp) bacteria are an increasing threat to public health and represent one of the most concerning pathogens involved in life-threatening infections and antimicrobial resistance (AMR). To understand the epidemiology of AMR of Kp in Portugal, we analysed whole genome sequencing, susceptibility testing and other meta data on 509 isolates collected nationwide from 16 hospitals and environmental settings between years 1980 and 2019. Predominant sequence types (STs) included ST15 (n = 161, 32%), ST147 (n = 36, 7%), ST14 (n = 26, 5%) or ST13 (n = 26, 5%), while 31% of isolates belonged to STs with fewer than 10 isolates. AMR testing revealed widespread resistance to aminoglycosides, fluoroquinolones, cephalosporins and carbapenems. The most common carbapenemase gene was blaKPC-3. Whilst the distribution of AMR linked plasmids appears uncorrelated with ST, their frequency has changed over time. Before year 2010, the dominant plasmid group was associated with the extended spectrum beta-lactamase gene blaCTX-M-15, but this group appears to have been displaced by another carrying the blaKPC-3 gene. Co-carriage of blaCTX-M and blaKPC-3 was uncommon. Our results from the largest genomics study of Kp in Portugal highlight the active transmission of strains with AMR genes and provide a baseline set of variants for future resistance monitoring and epidemiological studies.

Multiresistant bacteria: Invisible enemies of freshwater mussels.

Freshwater mussels are among the most endangered groups of fauna anywhere in world. The indiscriminate use of antibiotics has led to the emergence of resistant strains. These antibiotic-resistant bacteria play a key role in increasing the risk allied with the use of surface water and in spread of resistance genes. Two endangered freshwater mussel species, Margaritifera margaritifera and Potomida littoralis, were sampled at 4 sampling sites along a 50 km stretch of River Tua. Water samples were taken at same sites. Of the total of 135 isolates, 64.44% (39.26% from water and 25.19% from mussels) were coliform bacteria. Site T1, with the lowest concentration of coliform bacteria, and site T2 were the only ones where M. margaritifera was found. No E. coli isolates were found in this species and the pattern between water and mussels was similar. P. littoralis, which was present at T3/T4 sites, is the one that faces the highest concentration of bacterial toxins, which are found in treated wastewater effluents and around population centers. Sites T3/T4 have the isolates (water and mussels) with the highest resistance pattern, mainly to ß-lactams. Water and P. littoralis isolates (T3/T4) showed resistance to penicillins and their combination with clavulanic acid, and to cephalosporins, precisely to a fourth generation of cephalosporin antibiotics. The analysis provides important information on the risk to water systems, as well as the need to investigate possible management measures. It is suggested that future studies on the health status of freshwater bivalves should incorporate measures to indicate bacteriological water quality.

Biofilms in Diabetic Foot Ulcers: Impact, Risk Factors and Control Strategies.

Diabetic foot ulcers (DFUs) are a serious complication from diabetes mellitus, with a huge economic, social and psychological impact on the patients' life. One of the main reasons why DFUs are so difficult to heal is related to the presence of biofilms. Biofilms promote wound inflammation and a remarkable lack of response to host defences/treatment options, which can lead to disease progression and chronicity. In fact, appropriate treatment for the elimination of these microbial communities can prevent the disease evolution and, in some cases, even avoid more serious outcomes, such as amputation or death. However, the detection of biofilm-associated DFUs is difficult due to the lack of methods for diagnostics in clinical settings. In this review, the current knowledge on the involvement of biofilms in DFUs is discussed, as well as how the surrounding environment influences biofilm formation and regulation, along with its clinical implications. A special focus is also given to biofilm-associated DFU diagnosis and therapeutic strategies. An overview on promising alternative therapeutics is provided and an algorithm considering biofilm detection and treatment is proposed.

Bacterial coaggregation in aquatic systems.

The establishment of a sessile community is believed to occur in a sequence of steps where genetically distinct bacteria can become attached to partner cells via specific molecules, in a process known as coaggregation. The presence of bacteria with the ability to autoaggregate and coaggregate has been described for diverse aquatic systems, particularly freshwater, drinking water, wastewater, and marine water. In these aquatic systems, coaggregation already demonstrated a role in the development of complex multispecies sessile communities, including biofilms. While specific molecular aspects on coaggregation in aquatic systems remain to be understood, clear evidence exist on the impact of this mechanism in multispecies biofilm resilience and homeostasis. The identification of bridging bacteria among coaggregating consortia has potential to improve the performance of wastewater treatment plants and/or to contribute for the development of strategies to control undesirable biofilms. This study provides a comprehensive analysis on the occurrence and role of bacterial coaggregation in diverse aquatic systems. The potential of this mechanism in water-related biotechnology is further described, with particular emphasis on the role of bridging bacteria.

Antimicrobial, Antibiofilm, and Antioxidant Properties of Boletus edulis and Neoboletus luridiformis Against Multidrug-Resistant ESKAPE Pathogens.

Multidrug-resistant ESKAPE pathogens (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter species) has become the most recurrent global cause of skin and soft-tissue infections, belonging to the WHO priority pathogens list. Successful therapy remains challenging and entails the assessment of novel and successful antibiotics. In this study, mushrooms are considered a valuable and unique source of natural antimicrobial compounds. Therefore, this study aimed to evaluate the antimicrobial and antibiofilm properties of Boletus edulis (B. edulis) and Neoboletus luridiformis (N. luridiformis) aqueous and methanolic extracts against ESKAPE isolates from clinical wound infections. Disk diffusion and microdilution methods were used to assess the antimicrobial activity. Phytochemical characterization was achieved by analysis of total phenols, orthodiphenols content, and antioxidant activity as well as by high-performance liquid chromatography-diode array detector (HPLC-DAD). Human foreskin fibroblasts-1 (HFF-1) cell viability was performed by the MTT assay. Aqueous and methanolic extracts of B. edulis and N. luridiformis showed antimicrobial and antibiofilm properties against multidrug-resistant bacteria, although with different efficacy rates. The results showed that there is a convincing relation between the content of phenolic compounds, antioxidant activity, and antimicrobial activity suggesting that the presence of phenolic compounds may explain the biological effects. HPLC analysis revealed high levels of protocatechuic acid, homogentisic acid, pyrogallol, gallic acid, p-catechin, and dihydroxybenzoic acid in the aqueous extract of B. edulis, explaining the highest antimicrobial and antibiofilm properties. Importantly, the mushrooms extracts were non-cytotoxic at all the tested concentrations. Overall, the tested mushrooms extracts are good candidates to further explore its use in the prevention of wound infection, particularly by multidrug-resistant pathogens.

Multidrug-resistant Serratia rubidaea strains in the oral microbiota of healthy horses.

BACKGROUND: Many emergent pathogenic agents are cross-transmitted from animals to humans. Horses are considered as potential reservoirs of commensal, zoonotic, and multidrug-resistant bacteria. Equine bites could lead to infections caused by these agents, considering equine species as a public health concern. The more it is known about the equine oral microbiota the best secondary problems created by their commensal flora can be controlled. There are very few reports of Serratia rubidaea, a zoonotic and opportunistic bacterium, both in human and veterinary medicine. AIM: This study aimed to evaluate the Gram-negative microbiota of healthy equine oral cavities and their antimicrobial susceptibility. METHODS: During equine routine oral procedures, eight healthy horses were selected for this study, after discarding any abnormal dental conditions. Samples were collected from the subgingival space and gingival margin from the tooth 406 and both the identification and antimicrobial susceptibility test of Gram-negative bacteria were performed. RESULTS: This study reports the isolation of 32 Gram-negative agents, 27 of which were multidrug-resistant to the antimicrobial classes tested. High resistance rates were obtained to commonly used antimicrobial drugs, particularly macrolides and aminoglycosides as to carbapenems that are specific to human medicine. Two multi-drug resistance strains of S. rubidaea were found in the mouth of two healthy horses. CONCLUSION: Most Gram-negative isolates found in healthy horses were zoonotic and multi-drug resistant. This is a strong reason to consider the horse as an animal with a major place in the "One Health" concept. Equine clinicians should take precautions when working with horses' mouths. Antimicrobial sensitivity tests should be taken into consideration when finding the appropriate antimicrobial therapy protocol. To the authors' best knowledge, this is the first report about isolation of S. rubidaea from the mouth of the equine species.

Phenolic Profile and Bioactive Potential of Stems and Seed Kernels of Sweet Cherry Fruit.

Every year, large quantities of stems and pits are generated during sweet cherry processing, without any substantial use. Although stems are widely recognized by traditional medicine, detailed and feasible information about their bioactive composition or biological value is still scarce, as well as the characterization of kernels. Therefore, we conducted a study in which bioactivity potential of extracts from stems and kernels of four sweet cherry cultivars (Early Bigi (grown under net cover (C) and without net cover (NC)), Burlat, Lapins, and Van) were examined. The assays included antioxidant (by 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic) acid (ABTS), 2,2-diphenyl-1-picrylhydrazyl (DPPH) and ß-carotene-linoleic acid bleaching assays), and antibacterial activities against important Gram negative and Gram positive bacterial human isolates. Profile and individual phenolic composition of each extract were determined by High-performance liquid chromatography (HPLC) analysis. Extracts from stems of cv. Lapins and kernels of Early Bigi NC presented high levels of total phenolics, flavonoids, ortho-diphenols and saponins. Excepting for cv. Early Bigi NC, major phenolic compounds identified in stems and kernels were sakuranetin and catechin, respectively. In cv. Early Bigi NC the most abundant compounds were ellagic acid for stems and protocatechuic acid for kernels. In all extracts, antioxidant activities showed a positive correlation with the increments in phenolic compounds. Antimicrobial activity assays showed that only stem's extracts were capable of inhibiting the growth of Gram positive isolates. This new data is intended to provide new possibilities of valorization of these by-products and their valuable properties.

Prevalence and serotypes of Shiga toxin-producing Escherichia coli (STEC) in dairy cattle from Northern Portugal.

The prevalence of Shiga toxin (Stx)-producing Escherichia coli (STEC) was determined by evaluating its presence in faecal samples from 155 heifers, and 254 dairy cows in 21 farms at North of Portugal sampled between December 2017 and June 2019. The prevalence of STEC in heifers (45%) was significantly higher than in lactating cows (16%) (p<0.05, Fisher exact test statistic value is <0.00001). A total of 133 STEC were isolated, 24 (13.8%) carried Shiga-toxin 1 (stx1) genes, 69 (39.7%) carried Shiga-toxin 2 (stx2) genes, and 40 (23%) carried both stx1 and stx2. Intimin (eae) virulence gene was detected in 29 (21.8%) of the isolates. STEC isolates belonged to 72 different O:H serotypes, comprising 40 O serogroups and 23 H types. The most frequent serotypes were O29:H12 (15%) and O113:H21 (5.2%), found in a large number of farms. Two isolates belonged to the highly virulent serotypes associated with human disease O157:H7 and O26:H11. Many other bovine STEC serotypes founded in this work belonged to serotypes previously described as pathogenic to humans. Thus, this study highlights the need for control strategies that can reduce STEC prevalence at the farm level and, thus, prevent food and environmental contamination.