Food-related Environments Research Articles

Red Grape By-Products from the Demarcated Douro Region: Chemical Analysis, Antioxidant Potential and Antimicrobial Activity against Food-Borne Pathogens.

Wine production is one of the most important agricultural activities. The winemaking process generates a considerable volume of different residues characterized as by-products, such as pomace, seeds, stems, and skins. By-products are rich in polyphenols with antioxidant and antibacterial properties and may act as bacteriostatic or bactericidal agents against food-borne pathogens, improving food safety by enhancing antibiotic efficacy and reducing bacterial resistance. The aim of this study was to evaluate the phenolic composition and antioxidant activity of grape components (skins, seeds, and stems) from three red grape varieties (Periquita, Gamay, and Donzelinho Tinto) and determine their antibacterial activity against antibiotic-resistant bacteria, including Escherichia coli in food-producing animals and Listeria monocytogenes from food products and food-related environments. Ten phenolic compounds were quantified in these red grape varieties, with specific compounds found in different parts of the grape, including phenolic acids and flavonoids. Flavonoids are abundant in seeds and stems, malvidin-3-O-glucoside being the main anthocyanin in skins. The ethanolic extract from the seeds showed in vitro concentration-dependent activity against reactive species like •NO and O2•-. Gamay extract was the most effective, followed by Donzelinho Tinto and Periquita. Extracts showed varying antibacterial activity against Gram-positive and Gram-negative bacteria, with stronger effects on Gram-positive bacteria. L. monocytogenes was more susceptible, while E. coli was limited to three strains. Seeds exhibited the strongest antibacterial activity, followed by stems. The results of our study provide evidence of the potential of grape by-products, particularly seeds, as sources of bioactive compounds with antioxidant and antibacterial properties, offering promising avenues for enhancing food safety and combating antibiotic resistance in food production and related environments.

Inactivation of listerial biofilm on the food-contact surface using thymol and bio-modulated yeast microcarriers

Decontamination of bacterial biofilms formed on food-contact surfaces is a serious challenge in the food industry. In this study, a food-grade antimicrobial system was developed using yeast-based microcarriers and essential oil (EOs; thymol) to inactivate bacterial biofilms on food-contact surfaces. A bio-inspired method that involves pre-treatment of yeast microcarriers with simulated intestinal fluid (SIF) was employed to modulate the release of the encapsulated EOs. The effectiveness of the developed antimicrobial delivery system was evaluated based on physicochemical characterization, antimicrobial activities against planktonic bacterial cells in the presence of organic matter, affinity to bind bacterial biofilms, and co-incubation and residual antimicrobial activities against bacterial biofilms. The results illustrate that pre-treatment of yeast-based microcarriers with SIF significantly (p < 0.05) enhanced the release of the encapsulated EOs without influencing the encapsulation yield and binding of yeast-based microcarriers with bacterial biofilms. Strong antibiofilm activities were exhibited by the thymol encapsulated in the pre-digested yeast cells (pdYC@Thymol) compared to free thymol or thymol encapsulated in the undigested yeast microcarriers. Populations of Listeria innocua in biofilms (ca. 7.54 log CFU/cm2) decreased to ca. 2.63 log CFU/cm2 after 1 h of co-incubation with the pdYC@Thymol suspensions and further decreased to ca. 0.85 log CFU/cm2 after removal of the loosely bound pdYC@Thymol particles. Overall, this study suggests that the combined effects of the targeted delivery of the EOs via encapsulation in yeast-based microcarriers and facilitated release of the encapsulated EOs upon binding with bacterial biofilms can effectively inactivate the bacterial biofilms formed on food-contact surfaces, thereby enhancing the microbiological safety of the food-related environments.

Intestinal pathogens detected in cockroach species within different food-related environment in Pudong, China

Cockroaches are considered mechanical transmitters of infectious diseases, posing a threat to human health. This study assessed the potential of cockroaches in food-related environments to mechanically transmit intestinal pathogens. Cockroaches captured with traps were placed together into a low temperature refrigerator at − 80° for 2 h. Standard taxonomic keys and Fluorescent quantitative PCR techniques were applied for species identification and digestive tract etiological examination. A total of 360 cockroach traps were placed, with a positive rate of 20.8%, and 266 cockroaches were captured. In general, compared with other places and areas, the degree of infestation of cockroaches was more serious in catering places and kitchens. Blattella germanica were most found in catering places (40.2%), followed by Periplaneta fuliginosa in schools (22.2%). According to the life stage, among the 128 cockroach samples, 23 were positive for nymphs and 13 were positive for adults. There were statistically significant differences in the intestinal pathogen detection rates between nymphs and adults (P < 0.05). A total of eight intestinal pathogens were detected, and enterovirus infections were the main ones, with sapovirus being the most detected in Blattella germanica or nymph. Shiga toxin-producing Escherichia coli (STEC) was the most frequently isolated bacterium. Blastocystis hominis had the highest isolation rate. In contrast, 12 diarrhoeal disease pathogens were isolated, and the viruses and bacteria with the highest frequencies were norovirus and E. coli, respectively; no parasites were found. Blattella germanica and Periplaneta fuliginosa in food-related environments can act as potential vectors for the spread of intestinal pathogens and may pose a significant threat to public health.

Application of coliphage as biocontrol agent in combination with gamma irradiation to eliminate multi-drug-resistant E. coli in minimally processed vegetables

Biofilm formation is a rising concern in the food industry. Escherichia coli (E. coli) is one of the most important food-borne pathogens that can survive in food and food-related environments and eventually produce biofilms. This study suggested that both coliphages used were successful in preventing the creation of new biofilms as well as removing existing ones. Confocal laser scanning microscopy verified these findings. According to the findings, neither coliphage survived at 37 °C, but both remained stable at 4 °C and − 20 °C for extended periods of time. The study revealed that both coliphages demonstrated a greater degree of gamma irradiation resistance when compared to E. coli. The study’s results indicate that the implementation of a dual method, which incorporates gamma irradiation (1.5 kGy) and coliphage treatment, on various kinds of vegetables that were infected with E. coli, resulted in a significant reduction in bacterial count (surpassing 99.99%) following a 24-h incubation period. Combining gamma irradiation and the coliphage approach was significantly effective at lowering polysaccharide concentrations and proteins in the biofilm matrix. The results revealed that the pairing of gamma irradiation and coliphages acted in conjunction to cause disruptions in the matrix of biofilm, thereby promoting cell removal compared with either of the individual treatments. Ca+ ions strengthen the weak virion interaction with the relevant bacterial host cell receptors during the adsorption process. In conclusion, use of coliphage in combination with gamma irradiation treatment can be applied to improve fresh produce’s microbial safety and enhance its storability in supermarkets.Graphical

From farm to fork: Spread of a multidrug resistant Salmonella Infantis clone encoding blaCTX-M-1 on pESI-like plasmids in Central Italy

Salmonella enterica subsp. enterica serovar Infantis (S. Infantis) is one of the “top five Salmonella serovars” of clinical significance in the European Union (EU). Antimicrobial resistant and extended spectrum β-lactamase (ESBL) AmpC-producing S. Infantis have been described in food production systems and human clinical samples in Italy. Recently, an increase of MDR S. Infantis carrying blaCTX-M genes involved in 3rd generation cephalosporin resistance was noticed in the EU, including Italy, mainly due to the spread of S. Infantis harboring a pESI-like plasmid.The aim of this study was to investigate the occurrence of the S. Infantis pESI-like plasmid among antibiotic resistant S. Infantis strains isolated at different points of the food chain, and to provide a phylogenetic analysis to gain further insight on their transmission pathways from ‘farm to fork’.MDR S. Infantis strains (n. 35) isolated from 2016 to 2021 at different stages of the food chain (animals, food, food-related environments, and humans) were investigated with in depth molecular characterization using real-time PCR, S1 nuclease pulsed-field gel electrophoresis (S1-PFGE) and whole genome sequencing (WGS).Our study reported the occurrence of S. Infantis strains harboring pESI-like plasmids, carrying blaCTX-M-1 genes, in Central Italy, at different sampling points along the food chain. Results confirmed the presence of a plasmid with a molecular size around 224–310 kb, thus consistent with the pESI-like, in 97 % of the 35 samples investigated. Two variants of S. Infantis pESI-like IncFIB(K)_1_Kpn3 were detected, one associated with the European clone carrying blaCTX-M-1 (21 isolates) and the other associated with U.S. isolates carrying blaCTX-M-65 (2 isolates, pESI-like U.S. variant). The majority was resistant to 3rd generation cephalosporins but none of the strains tested positive for the carbapenemase encoding genes. A total of 118 virulence genes were identified in isolates harboring the pESI-like plasmid. cgMLST and SNP-based analysis revealed the presence of one main cluster, composed by strains isolated from the environment, animals, food and humans.The results of this investigation underline the importance of phylogenetic studies to monitor and understand pathogen and AMR spread in a One Health approach.

Sensitive detection of viable Cronobacter sakazakii by bioluminescent reporter phage emitting stable signals with truncated holin

As outbreaks of foodborne illness caused by the opportunistic pathogen Cronobacter sakazakii (Cs) continue to occur, particularly in infants consuming powdered infant formula (PIF), the need for sensitive, rapid, and easy-to-use detection of Cs from food and food processing environments is increasing. Here, we developed bioluminescent reporter bacteriophages for viable Cs-specific, substrate-free, rapid detection by introducing luciferase and its corresponding substrate-providing enzyme complex into the virulent phage ΦC01. Although the reporter phage ΦC01_lux, constructed by replacing non-essential genes for phage infectivity with a luxCDABE reporter operon, produced bioluminescence upon Cs infection, the emitted signal was quickly decayed due to the superior bacteriolytic activity of ΦC01. By truncating the membrane pore-forming protein holin and thus limiting its function, the bacterial lysis was delayed and the resultant engineered reporter phage ΦC01_lux_Δhol could produce a more stable and reliable bioluminescent signal. Accordingly, ΦC01_lux_Δhol was able to detect at least an average of 2 CFU/ml of Cs artificially contaminated PIF and Sunsik and food contact surface models within a total of 7 h of assays, including 5 h of pre-enrichment for Cs amplification. The sensitive, easy-to-use, and specific detection of live Cs with the developed reporter phage could be applied as a novel complementary tool for monitoring Cs in food and food-related environments for food safety and public health.

Co-culture of Cronobacter sakazakii and Staphylococcus aureus: Explore the influence of mixed biofilm formation and regulation of Cronobacter sakazakii biofilm formation genes

Bacterial biofilm is a protective matrix composed of metabolites secreted by bacteria that envelop bacteria. By forming a biofilm, bacteria can considerably improve their environmental tolerance. In food-related processing environment, different types of microorganisms are often present in biofilms. The main contaminating strain in the powdered infant formula (PIF) processing environment, Cronobacter sakazakii and Staphylococcus aureus continues to pollute the PIF processing environment after biofilm production. This study selected Cronobacter sakazakii with a weak biofilm-forming ability as one of the test organisms. The coexistence of Cronobacter sakazakii and Staphylococcus aureus on the surface of production equipment was simulated to analyze the interaction. Biofilm formation in the co-culture group was significantly higher than the others. In-depth study of the effect of Staphylococcus aureus on the biofilm formation genes of Cronobacter sakazakii. Results show two bacteria can coexist on the surface of a metal device, forming a more compact hybrid biofilm structure. Under co-culture conditions, S. aureus increased bcsA and fliD expression in Cronobacter sakazakii, whereas decreased bcsC expression. Signaling molecules produced by Staphylococcus aureus (Autoinducer 2) significantly promoted the biofilm formation of Cronobacter sakazakii at the concentration of 0–500 ng/mL (0.099–0.177) and up-regulated the expression of bcsA, filD and flhD genes.

Sublethal chlorine stress promotes the biofilm-forming ability of Salmonella enterica serovars enteritidis and expression of the related genes

Chlorine treatment is the most common disinfection method in food-related environments. In addition to being simple and inexpensive, this method is very effective if used properly. However, insufficient chlorine concentrations only cause a sublethal oxidative stress in the bacterial population and may alter the growth behavior of stressed cells. In the present study, the effect of sublethal chlorine stress on the biofilm formation characteristics of Salmonella Enteritidis was evaluated. Our results demonstrated that, sublethal chlorine stress (350 ppm total chlorine) activates the biofilm (csgD, agfA, adrA and bapA) and quorum-sensing (sdiA and luxS) related genes in planktonic cells of S. Enteritidis. The higher expression of these genes illustrated that the chlorine stress induced the initiation of the biofilm formation process in S. Enteritidis. Results of the initial attachment assay confirmed this finding. In addition, the number of chlorine-stressed biofilm cells was significantly higher than non-stressed biofilm cells after 48 h incubation at 37 °C. In S. Enteritidis ATCC 13076 and S. Enteritidis KL19, the number of chlorine-stressed biofilm cells were 6.93 ± 0.48 and 7.49 ± 0.57 log CFU/cm2, while the number of non-stressed biofilm cells were 5.12 ± 0.39 and 5.63 ± 0.51 log CFU/cm2, respectively. These findings were confirmed by measurements of the major components of biofilm, i.e., eDNA, protein and carbohydrate. The amount of these components in 48-h biofilms was higher when the cells were initially subjected to sublethal chlorine stress. However, the up-regulation of the biofilm and quorum sensing genes was not observed in 48-h biofilm cells, indicating that the effect of chlorine stress had vanished in the subsequent generations of Salmonella. In total, these results revealed that sublethal chlorine concentrations can promote the biofilm-forming ability of S Enteritidis.

Isolation of bacteriophages specific to Pseudomonas mosselii for controlling milk spoilage

Bacteriophages controlling heat-stable protease producing Pseudomonas mosselii were characterised and applied. Two Podoviridae family phages, designated ΦC106 and Φ21A, were isolated and shown to exhibit a broad host range. The optimal multiplicity of infection of both phages generating maximum titres was 10, latent periods were at 20 min and burst sizes were 1862 and 331 plaque-forming units per infected cell for ΦC106 and Φ21A, respectively. ΦC106 and Φ21A genomes did not encode known toxins, antibiotic-resistant genes, virulent factors of bacterial origin, or lysogenic markers such as integrase, recombinase, repressor/anti-repressor protein, and excisionase, guaranteeing their safety for addition into milk and dairy products. In vitro lytic activity and proteolytic activity of ΦC106 and Φ21A revealed a remarkable decrease in bacterial count and enzyme activity. These data suggest that ΦC106 and Φ21A are efficient in controlling the prevalence of spoilage-causing P. mosselii strains in dairy- and food-related environments.

Metataxonomic insights in the distribution of Lactobacillaceae in foods and food environments

Members of the family Lactobacillaceae, which now includes species formerly belonging to the genera Lactobacillus and Pediococcus, but also Leuconostocaceae, are of foremost importance in food fermentations and spoilage, but also as components of animal and human microbiota and as potentially pathogenic microorganisms. Knowledge of the ecological distribution of a given species and genus is important, among other things, for the inclusion in lists of microorganisms with a Qualified Presumption of Safety or with beneficial use. The objective of this work is to use the data in FoodMicrobionet database to obtain quantitative insights (in terms of both abundance and prevalence) on the distribution of these bacteria in foods and food environments.We first explored the reliability of taxonomic assignments using the SILVA v138.1 reference database with full length and partial sequences of the 16S rRNA gene for type strain sequences. Full length 16S rRNA gene sequences allow a reasonably good classification at the genus and species level in phylogenetic trees but shorter sequences (V1-V3, V3-V4, V4) perform much worse, with type strains of many species sharing identical V4 and V3-V4 sequences. Taxonomic assignment at the genus level of 16S rRNA genes sequences and the SILVA v138.1 reference database can be done for almost all genera of the family Lactobacillaceae with a high degree of confidence for full length sequences, and with a satisfactory level of accuracy for the V1-V3 regions. Results for the V3-V4 and V4 region are still acceptable but significantly worse. Taxonomic assignment at the species level for sequences for the V1-V3, V3-V4, V4 regions of the 16S rRNA gene of members of the family Lactobacillaceae is hardly possible and, even for full length sequences, and only 49.9 % of the type strain sequences can be unambiguously assigned to species.We then used the FoodMicrobionet database to evaluate the prevalence and abundance of Lactobacillaceae in food samples and in food related environments. Generalist and specialist genera were clearly evident. The ecological distribution of several genera was confirmed and insights on the distribution and potential origin of rare genera (Dellaglioa, Holzapfelia, Schleiferilactobacillus) were obtained.We also found that combining Amplicon Sequence Variants from different studies is indeed possible, but provides little additional information, even when strict criteria are used for the filtering of sequences.

A Meta-Analysis and Systematic Review of Listeria monocytogenes Response to Sanitizer Treatments.

Listeria monocytogenes is a ubiquitous organism that can be found in food-related environments, and sanitizers commonly prevent and control it. The aim of this study is to perform a meta-analysis of L. monocytogenes response to sanitizer treatments. According to the principle of systematic review, we extracted 896 records on the mean log-reduction of L. monocytogenes from 84 publications as the dataset for this study. We applied a mixed-effects model to describe L. monocytogenes response to sanitizer treatment by considering sanitizer type, matrix type, biofilm status, sanitizer concentration, treatment time, and temperature. Based on the established model, we compared the response of L. monocytogenes under different hypothetical conditions using forest plots. The results showed that environmental factors (i.e., sanitizer concentration, temperature, and treatment time) affected the average log-reduction of L. monocytogenes (p < 0.05). L. monocytogenes generally exhibited strong resistance to citric acid and sodium hypochlorite but had low resistance to electrolyzed water. The planktonic cells of L. monocytogenes were less resistant to peracetic acid and sodium hypochlorite than the adherent and biofilm cells. Additionally, the physical and chemical properties of the contaminated or inoculated matrix or surface also influenced the sanitizer effectiveness. This review may contribute to increasing our knowledge of L. monocytogenes resistance to sanitizers and raising awareness of appropriate safety precautions.

Development of Desiccation-Tolerant Probiotic Biofilms Inhibitory for Growth of Foodborne Pathogens on Stainless Steel Surfaces.

Lactic acid bacteria biofilms can be used to reduce foodborne pathogen contamination in the food industry. However, studies on growth inhibition of foodborne pathogens by inducing biofilm formation of antagonistic microorganisms on abiotic surfaces are rare. We developed a desiccation-tolerant antimicrobial probiotic biofilm. Lactobacillus sakei M129-1 and Pediococcus pentosaceus M132-2 isolated from fermented Korean foods were found to exhibit broad-spectrum antibacterial activity against Bacillus cereus, Escherichia coli O157:H7, Staphylococcus aureus, Listeria monocytogenes, and Salmonella enterica. Their biofilm levels were significantly (p < 0.05) higher on stainless steel than on polyethylene or ceramic. Biofilms of both isolates showed significantly (p < 0.05) enhanced resistance against desiccation (exposure to 43% atmospheric relative humidity) as compared with the isolates not in the biofilm form. The antimicrobial activity of the isolates was sustained in dried biofilms on stainless steel surface; the initial number of foodborne pathogens (average 7.0 log CFU/mL), inoculated on stainless steel chips containing L. sakei M129-1 or P. pentosaceus M132-2 biofilm decreased to less than 1.0 log CFU within 48 h. The lactic acid bacteria antibacterial biofilms developed in this study may be applied to desiccated environmental surfaces in food-related environments to improve microbiological food safety.

Identification of New Halomonas Strains from Food-related Environments.

Halomonas species, which are aerobic, alkaliphilic, and moderately halophilic bacteria, produce diverse biochemicals. To identify food-related Halomonas strains for bioremediation and the industrial production of biochemicals, 20 strains were isolated from edible seashells, shrimp, and umeboshi (pickled Japanese plum) factory effluents. All isolates were phylogenetically classified into a large clade of Halomonas species. Most isolates, which grew in wide pH (6–13) and salt concentration (0–14%) ranges, exhibited the intracellular accumulation of poly(3-hydroxybutyrate) granules. The characteristics of these isolates varied. A020 isolated from umeboshi factory effluents exhibited enhanced stress tolerance and proliferation and comprised two plasmids. IMZ03 and A020 grew to more than 200 OD600, while IMZ03 produced 3.5% 3-hydroxybutyrate in inorganic medium supplemented with 10% sucrose. The mucus of TK1-1 cultured on agar medium comprised approximately 64‍ ‍mM of ectoine. Whole-genome sequencing of A020 was performed to elucidate its origin and genomic characteristics. The genome ana­lysis revealed a region exhibiting synteny with a large virus genome isolated from the ocean, but did not identify any predictable pathogenic genes. Therefore, saline foods and related materials may be suitable resources for isolating Halomonas strains exhibiting unique, useful, and innocuous features.

Uses of Bacteriophages as Bacterial Control Tools and Environmental Safety Indicators.

Bacteriophages are bacterial-specific viruses and the most abundant biological form on Earth. Each bacterial species possesses one or multiple bacteriophages and the specificity of infection makes them a promising alternative for bacterial control and environmental safety, as a biotechnological tool against pathogenic bacteria, including those resistant to antibiotics. This application can be either directly into foods and food-related environments as biocontrol agents of biofilm formation. In addition, bacteriophages are used for microbial source-tracking and as fecal indicators. The present review will focus on the uses of bacteriophages like bacterial control tools, environmental safety indicators as well as on their contribution to bacterial control in human, animal, and environmental health.

Novel methods of microbiome analysis in the food industry.

The study of the food microbiome has gained considerable interest in recent years, mainly due to the wide range of applications that can be derived from the analysis of metagenomes. Among these applications, it is worth mentioning the possibility of using metagenomic analyses to determine food authenticity, to assess the microbiological safety of foods thanks to the detection and tracking of pathogens, antibiotic resistance genes and other undesirable traits, as well to identify the microorganisms responsible for food processing defects. Metataxonomics and metagenomics are currently the gold standard methodologies to explore the full potential of metagenomes in the food industry. However, there are still a number of challenges that must be solved in order to implement these methods routinely in food chain monitoring, and for the regulatory agencies to take them into account in their opinions. These challenges include the difficulties of analysing foods and food-related environments with a low microbial load, the lack of validated bioinformatics pipelines adapted to food microbiomes and the difficulty of assessing the viability of the detected microorganisms. This review summarizes the methods of microbiome analysis that have been used, so far, in foods and food-related environments, with a specific focus on those involving Next-Generation Sequencing technologies.

Chlorine-stressed Salmonella cells are significantly more engulfed by Acanthamoeba trophozoites and have a longer intracystic survival than the non-stressed cells

In the present study, the effect of sublethal chlorine-induced oxidative stress on the subsequent interaction of Salmonella enterica serovars Enteritidis and Typhimurium with Acanthamoeba castellanii and A. polyphaga was evaluated. Sublethal chlorine concentration was determined using the lag phase extension information and used to prepare chlorine-stressed Salmonella cells. Coculture experiments of Acanthamoeba and Salmonella cells were performed in Page's amoeba saline (PAS) at 25 °C for 2 h. The results showed that the chlorine-stressed Salmonella cells were significantly more engulfed by A. castellanii and A. polyphaga trophozoites than the non-stressed cells. The uptake rates of the chlorine-stressed and non-stressed Salmonella cells were in the range of 14.17–27.34 and 6.51–11.52% for A. castellanii, and in the range of 8.32–17.76 and 2.28–6.12% for A. polyphaga trophozoites, respectively. Moreover, intracystic survival time of chlorine-stressed cells of S. Enteritidis and S. Typhimurium was significantly longer than that of non-stressed cells. While, non-stressed Salmonella cells survived within A. castellanii and A. polyphaga cysts for 13–20 and 8–15 days, chlorine-stressed cells were recovered from A. castellanii and A. polyphaga cysts after 22–32 and 15–24 days, respectively. These results underscore the importance of bacterial exposure to sublethal chlorine concentrations in their interaction with free-living amoebae, and may lead to a better understanding of the parameters affecting the persistence of Salmonella enterica serovars in food-related environments.

Staphylococcus saprophyticus From Clinical and Environmental Origins Have Distinct Biofilm Composition.

Biofilm formation has been shown to be critical to the success of uropathogens. Although Staphylococcus saprophyticus is a common cause of urinary tract infections, its biofilm production capacity, composition, genetic basis, and origin are poorly understood. We investigated biofilm formation in a large and diverse collection of S. saprophyticus (n = 422). Biofilm matrix composition was assessed in representative strains (n = 63) belonging to two main S. saprophyticus lineages (G and S) recovered from human infection, colonization, and food-related environment using biofilm detachment approach. To identify factors that could be associated with biofilm formation and structure variation, we used a pangenome-wide association study approach. Almost all the isolates (91%; n = 384/422) produced biofilm. Among the 63 representative strains, we identified eight biofilm matrix phenotypes, but the most common were composed of protein or protein–extracellular DNA (eDNA)–polysaccharides (38%, 24/63 each). Biofilms containing protein–eDNA–polysaccharides were linked to lineage G and environmental isolates, whereas protein-based biofilms were produced by lineage S and infection isolates (p < 0.05). Putative biofilm-associated genes, namely, aas, atl, ebpS, uafA, sasF, sasD, sdrH, splE, sdrE, sdrC, sraP, and ica genes, were found with different frequencies (3–100%), but there was no correlation between their presence and biofilm production or matrix types. Notably, icaC_1 was ubiquitous in the collection, while icaR was lineage G-associated, and only four strains carried a complete ica gene cluster (icaADBCR) except one that was without icaR. We provided evidence, using a comparative genomic approach, that the complete icaADBCR cluster was acquired multiple times by S. saprophyticus and originated from other coagulase-negative staphylococci. Overall, the composition of S. saprophyticus biofilms was distinct in environmental and clinical isolates, suggesting that modulation of biofilm structure could be a key step in the pathogenicity of these bacteria. Moreover, biofilm production in S. saprophyticus is ica-independent, and the complete icaADBCR was acquired from other staphylococci.

Development of the Food Related Environments at School and Home Questionnaire (FRESH-Q)

ObjectivesDelineating determinants of food choice within the school and home environments of adolescents might help to translate advocacy efforts to effective interventions. However, this will require the development of valid instruments that measure school and home food environmental factors systematically. We conducted a cross-sectional study to develop a valid questionnaire that evaluates the food-related environments at schools and homes (FRESH- Q) of adolescents aged 10–15 years in Mumbai, India. MethodsA preliminary draft was developed based on a review of literature, eleven focus group discussions (FGDs) with adolescents (n = 36) and parents (n = 27), and eight in-depth interviews (IDIs) with school supervisors followed by the identification of school food (SFE) and home food environment (HFE) constructs and an item pool generation. The content validity was evaluated by an expert panel and the item-wise content validity indices (I-CVI) for clarity and relevance were calculated. The questionnaire was pretested in 42 adolescents for face validity and further administered among 315 adolescents (mean age 13.2 (2.1) years) for psychometric evaluation. The principal axis method of exploratory factor analysis (EFA) established the construct validity and the internal consistency was determined using Cronbach alpha values > 0.7. A sub-sample (n = 108) completed FRESH-Q twice to estimate test-retest reliability. ResultsContent analysis of FGDs and IDIs revealed six SFE and eleven HFE constructs. The I-CVI for the majority of items were satisfactory (0.72–0.96) and the Cronbach alpha values were 0.88 for SFE and 0.92 for HFE items. The intra-class correlation coefficients ranged from 0.78–0.96 indicating good to excellent reliability. Four factors – availability, and affordability of school foods, adolescents’ perceptions of SFE, and cafeteria policies explained 76.2% of the variance in SFE items. The EFA model identified five underlying constructs for HFE items – availability, accessibility and visibility of foods at home, family dietary habits, food purchasing behaviors, parents’ perceptions and consumption patterns. ConclusionsA valid 36 item self-reported questionnaire was developed to evaluate the school and home food environments of urban adolescents in India. Further investigations in different settings are recommended. Funding SourcesNone.

Antibacterial and antibiofilm activity of essential oil of clove against Listeria monocytogenes and Salmonella Enteritidis.

The antibacterial and antibiofilm activity of essential oil of clove against Listeria monocytogenes and Salmonella Enteritidis were investigated. The chemical composition of the oil was characterized by gas chromatography-mass spectrometry. Stock solution of the essential oil of clove was prepared in 95% (v/v) ethanol (EOC). The antibacterial assays were performed by disk diffusion assay and minimal inhibitory concentration (MIC). The biomass of adhered cells and preformed biofilms after incubation with different concentrations of EOC was assessed by crystal violet. Eugenol was the major bioactive compound of clove essential oil, accounting for 78.85% of the total composition. The MIC values for L. monocytogenes and S. Enteritidis were 0.05 mg/ml and 0.1 mg/ml, respectively. The initial cell adhesion at MIC was inhibited by 61.8% for L. monocytogenes and 49.8% for S. Enteritidis. However, the effect of EOC was less marked on biofilm eradication than on cell adhesion. At MIC and within 1 hour of incubation with the EOC, the preformed biofilms were reduced by 30.2% and 20.3% for L. monocytogenes and S. Enteritidis, respectively. These results suggest that sanitizers based on clove essential oil could be a potential strategy to control biofilms in food-related environments.

Biodiversity and technological features of Weissella isolates obtained from Brazilian artisanal cheese-producing regions

Weissella spp. strains (n = 57) were isolated from different Brazilian food-related environments and characterized based on their genetic profiles and technological potential. PFGE and Rep-PCR revealed the high level of biodiversity among isolates: PFGE grouped the isolates in three major profiles (similarity from 60 to 80%), while rep-PCR characterized the isolates as belonging to a single profile. Based on PFGE, isolates with identical pulsotypes were found within a same geographical region (Campo das Vertentes, Minas Gerais state). Based on these profiles, 26 isolates were selected and characterized based on their inhibitory and bacteriocinogenic activity against Listeria monocytogenes ATCC 15313, Staphylococcus aureus subsp. aureus ATCC 6538, Salmonella enterica subsp. enterica serovar Typhimurium ATCC 14028 and Escherichia coli ATCC 11229. Most isolates (n = 20) were able to inhibit the targets through organic acids. Most of the isolates (n = 12) were able to produce diacetyl, proteases, and/or coagulating milk, but none were able to produce exopolysaccharides. Isolate 16, in particular, was characterized as possessing high acidification ability, diacetyl and protease production, with promising technological potential in the dairy industry. The obtained results allowed the understanding of the Weissella spp. strains role as starter cultures in the dairy industry.