Essential Oils and Their Combination with Lactic Acid Bacteria and Bacteriocins to Improve the Safety and Shelf Life of Foods: A Review.

The use of plant extracts (e.g., essential oils and their active compounds) represents an interesting alternative to chemical additives and preservatives applied to delay the alteration and oxidation of foods during their storage. Essential oils (EO) are nowadays considered valuable sources of food preservatives as they provide a healthier alternative to synthetic chemicals while serving the same purpose without affecting food quality parameters. The natural antimicrobial molecules found in medicinal plants represent a possible solution against drug-resistant bacteria, which represent a global health problem, especially for foodborne infections. Several solutions related to their application on food have been described, such as incorporation in active packaging or edible film and direct encapsulation. However, the use of bioactive concentrations of plant derivatives may negatively impact the sensorial characteristics of the final product, and to solve this problem, their application has been proposed in combination with other hurdles, including biocontrol agents. Biocontrol agents are microbial cultures capable of producing natural antimicrobials, including bacteriocins, organic acids, volatile organic compounds, and hydrolytic enzymes. The major effect of bacteriocins or bacteriocin-producing LAB (lactic acid bacteria) on food is obtained when their use is combined with other preservation methods. The combined use of EOs and biocontrol agents in fruit and vegetables, meat, and dairy products is becoming more and more important due to growing concerns about potentially dangerous and toxic synthetic additives. The combination of these two hurdles can improve the safety and shelf life (inactivation of spoilage or pathogenic microorganisms) of the final products while maintaining or stabilizing their sensory and nutritional quality. This review critically describes and collects the most updated works regarding the application of EOs in different food sectors and their combination with biocontrol agents and bacteriocins.

Human Breast Milk: A Source of Potential Probiotic Candidates.

This study focuses on the isolation of lactobacilli/bifidobacteria from human breast milk and their first characterization, in the perspective to find new probiotic candidates to be included in food products. More specifically, breast-milk-isolated strains demonstrated a very good aptitude to adhere to intestinal cells, in comparison with L. rhamnosus GG strain, taken as reference. The same behavior has been found for hydrophobicity/auto-aggregation properties. A remarkable antagonistic activity was detected for these isolates not only against spoilage and pathogenic species of food interest, but also against the principal etiological agents of intestinal infections. Indeed, isolated strains impaired spoilage and pathogenic species growth, as well as biofilm formation by gut pathogens. In addition, breast milk strains were characterized for their antibiotic susceptibility, displaying species-specific and strain-specific susceptibility patterns. Finally, to assess their technological potential, the fermentation kinetics and viability of breast milk strains in pasteurized milk were investigated, also including the study of the volatile molecule profiles. In this regard, all the strains pointed out the release of aroma compounds frequently associated with the sensory quality of several dairy products such as acetic acid, diacetyl, acetoin, acetaldehyde. Data here reported point up the high potential of breast-milk-isolated strains as probiotics.

Prevalence of HEV infection in acute non-ABC hepatitis and prognostic role of extrahepatic manifestations.

Background: HEV-3 and HEV-4 are emerging cause of zoonotic acute hepatitis in high-income countries. In Europe the disease is underdiagnosed but hyperendemic areas have been identified. We describe a population with acute non-ABC (n-ABC) hepatitis in Abruzzo, the Italian region with the highest seroprevalence reported. The study was included in the surveillance of acute hepatitis E by the Italian Institute of Public Health started in 2004 and implemented in 2015. Methods: Patients with n-ABC hepatitis during 2004-2018 in all Abruzzo Infectious Disease Departments were tested for HEV-IgM (Wantai®) and HEV-RNA (ORF3). Positive samples were sequenced (Beckman Coulter®) and phylogenetic tree (MEGA 6.06 software) obtained. Clinical data were retrospectively collected and an alimentary risk factors-questionnaire was administered. Categorical and quantitative variables were compared (Chi square test or Fisher test and Wilcoxon test). Results: 97 hospitalized patients were tested, most cases (91.7%) after 2015. Overall, HEV-IgM resulted positive in 36% and HEV-RNA detectable in 33.3%. All 24 sequences obtained were HEV-3, with two small groups of closely related strands. L'Aquila was the Province with higher positivity rate (44%). Retrospective clinical data were acquired in 86.5% of patients, no one having liver failure. Higher ALT-levels (1282.34 vs 893.25, p=0.0139) and extrahepatic symptoms (OR 16.69, p=0.0018) were strongly associated with HEV-IgM presence. Two small outbreaks are described. Conclusions: More than one third of n-ABC hepatitis in all Abruzzo are HEV-related. Extrahepatic symptoms correlate with HEV aetiology. Implementing surveillance is mandatory to really understand the extent of the disease.

Effects of Sub-Lethal High Pressure Homogenization Treatment on the Adhesion Mechanisms and Stress Response Genes in Lactobacillus acidophilus 08.

Cell surface hydrophobicity (CSH) and adhesion are very important phenotypical traits for probiotics that confer them a competitive advantage for the resilience in the human gastrointestinal tract. This study was aimed to understand the effects over time of a 50 MPa hyperbaric treatment on the surface properties of Lactobacillus acidophilus 08 including CSH, autoaggregation, and in vitro adhesion (mucin layer and Caco-2 cells). Moreover, a link between the hurdle applied and the expression of genes involved in the general stress response (groEL and clpP) and adhesion processes (efTu and slpA) was evaluated. High pressure homogenization (HPH) at 50 MPa significantly increased the CSH percentage (H%), autoaggregation and in vitro adhesion on mucin of L. acidophilus 08 cells compared with the untreated cells. Moreover, the hyperbaric hurdle induced an upregulation of the stress response genes groEL and ef-TU together with a down regulation of the clpP and S-layer slpA genes. Looking at the protein profile, HPH-treatment showed an increase in the number or intensity of protein bands at high and low molecular weights.

Probiotic and Metabolic Characterization of Vaginal Lactobacilli for a Potential Use in Functional Foods.

The main aim of this work was to verify the metabolic and functional aptitude of 15 vaginal strains belonging to Lactobacillus crispatus, Lactobacillus gasseri, and Limosilactobacillus vaginalis (previously Lactobacillus vaginalis), already characterized for their technological and antimicrobial properties. In order to evaluate the metabolic profile of these vaginal strains, a phenotype microarray analysis was performed on them. Functional parameters such as hydrophobicity, auto-aggregation, deconjugation of bile salts, adhesion to an intestinal cell line (Caco-2), and a simulated digestion process were evaluated for these strains. A good number of these strains showed hydrophobicity values higher than 70%. Regarding the auto-aggregation assay, the most promising strains were L. crispatus BC9 and BC1, L. gasseri BC10 and BC14, and L. vaginalis BC17. Moreover, L. crispatus BC4, BC6, BC7, and BC8 were characterized by strong bile salts hydrolase activity (BHS). In addition, L. crispatus BC8 and L. vaginalis BC17 were characterized by a medium ability to adhere to Caco-2 cells. Data related to digestion process showed a strong ability of vaginal lactobacilli to withstand this stress. In conclusion, the data collected show the metabolic versatility and several exploitable functional properties of the investigated vaginal lactobacilli.

Unravelling the Potential of Lactococcus lactis Strains to Be Used in Cheesemaking Production as Biocontrol Agents.

This research, developed within an exchange program between Italy and Canada, represents the first step of a three-year project intended to evaluate the potential of nisin-producing Lactococcus lactis strains isolated from Italian and Canadian dairy products to select a consortium of strains to be used as biocontrol agents in Crescenza and Cheddar cheese production. In this framework, the acidification and the production of nisin in milk, and the volatile molecule profiles of the fermented milk, were recorded. The strains were further tested for their anti-Listeria monocytogenes activity in milk. The data obtained highlighted good potential for some of the tested strains, which showed production of nisin beginning within 12 h after the inoculation and reaching maximum levels between 24 and 48 h. The highest inactivation levels of L. monocytogenes in milk was reached in the presence of the strains 101877/1, LBG2, 9FS16, 11FS16, 3LC39, FBG1P, UL36, UL720, UL35. The strains generated in milk-specific volatile profiles and differences in the presence of fundamental aromatic molecules of dairy products, such as 2-butanone and diacetyl. The results highlight the interesting potential of some L. lactis strains, the producer of nisin, to be further used as biocontrol agents, although the strains need to be tested for interaction with traditional thermophilic starters and tested in real cheesemaking conditions.

Evaluation of the fate of Lactobacillus crispatus BC4, carried in Squacquerone cheese, throughout the simulator of the human intestinal microbial ecosystem (SHIME).

Lactobacillus crispatus strain BC4, isolated from the human healthy vaginal environment and characterised by a strong antimicrobial activity against urogenital pathogens and foodborne microorganisms, was employed as a probiotic culture in the cheesemaking of the soft cheese Squacquerone. Such cheese is intended as a "gender food", that could be used as a hedonistic dietary strategy to reduce the incidence of woman vaginal dysbiosis and infections, given the evidence that a probiotic strain able to survive to the entire digestive process once ingested, can pass from intestine to vagina. This work was aimed to evaluate the resistance of L. crispatus BC4, carried in Squacquerone cheese, to different challenges of the human gastrointestinal tract, including the colon stage. The digestion process was tested using a Simulator of the Human Intestinal Microbial Ecosystem (SHIME®). The viability and metabolic activity of L. crispatus BC4 during the colon simulation were monitored by qPCR and gas chromatography, respectively, also in the presence of a complex microbiota. The results showed that L. crispatus BC4 survival was not affected by the gastric condition, while it was significantly affected by bile salts and pancreatic juice in small intestine conditions, where it decreased of approx. 0.6 log (colony-forming units) CFU/g. Differently, during colon simulation L. crispatus BC4 was able to grow in sterile colon conditions and to maintain viability in the presence of a complex microbiota. Moreover, during colon simulation, L. crispatus BC4 was metabolically active as demonstrated by the higher production of short chain fatty acids (SCFA) and lactate. In the presence of a complex gut microbiota, a decrease of lactate was observed, due to its conversion into propionate (anti-cholesterol activity) and butyrate (anti-inflammatory activity) by cross-feeding. However, no differences in propionate and butyrate production could be observed between control cheese and cheese containing L. crispatus BC4. Despite this may appear as a negative outcome, it must be taken into account that, in this setup, only a single dose of the cheese was tested and the outcome of the colonization and impact of the gut microbiota might be different when daily repeated doses are tested.

Suitability of the Nisin Z-producer Lactococcus lactis subsp. lactis CBM 21 to be Used as an Adjunct Culture for Squacquerone Cheese Production.

This research investigated the technological and safety effects of the nisin Z producer Lactococcus lactis subsp. lactis CBM 21, tested as an adjunct culture for the making of Squacquerone cheese in a pilot-scale plant. The biocontrol agent remained at a high level throughout the cheese refrigerated storage, without having a negative influence on the viability of the conventional Streptococcus thermophilus starter. The inclusion of CBM 21 in Squacquerone cheesemaking proved to be more effective compared to the traditional one, to reduce total coliforms and Pseudomonas spp. Moreover, the novel/innovative adjunct culture tested did not negatively modify the proteolytic patterns of Squacquerone cheese, but it gave rise to products with specific volatile and texture profiles. The cheese produced with CBM 21 was more appreciated by the panelists with respect to the traditional one.

Delayed presentation and late testing for HIV: demographic and behavioral risk factors in a multicenter study in Italy.

Ensuring timely access to care for persons with HIV is an important public health goal. To identify factors associated with delayed presentation to medical care after testing HIV-positive or with late HIV testing, we studied 968 patients at their first HIV care visit, enrolled in a multicenter study in Italy from 1997-2000. Patients completed a questionnaire on HIV-testing history, sexual behavior, and drug use behavior. Delayed presenters were patients with >6 months between their first HIV-positive test and presentation for HIV care; late testers were patients with CD4 count < 200 /mm or clinically defined AIDS at their first HIV-positive test. Among the study patients, 255 (26.3%) were delayed presenters, and 280 (28.9%) were late testers. In multinomial logistic regression analysis, injection drug use significantly increased (odds ratio [OR]= 5.04) the probability of delayed presentation but reduced (OR = 0.55) the chance of late testing. A previous HIV-negative test was associated with a reduced risk of both delayed presentation (OR = 0.39) and late testing (OR = 0.36). Unemployment was positively associated with delayed presentation and increasing age with late testing, whereas HIV counseling at the time of first positive HIV test strongly (OR = 0.42) reduced the odds of delayed presentation. Interventions aimed at promoting timely access to care of HIV-infected persons should consider differentiated programs for delayed presentation and late testing.