Effect of time restricted eating versus current practice in dietetics on glycaemic control and cardio-metabolic outcomes in individuals at risk of developing type 2 diabetes: Protocol for a multi-centre, parallel group, non-inferiority, randomised controlled trial.

BACKGROUND: Time restricted eating (TRE) is a dietary strategy that may improve metabolic health. However, no studies have compared TRE with current practice (CP) in dietetics. HYPOTHESIS: TRE will not be inferior to CP to improve glycaemic control in individuals at risk of type 2 diabetes (T2D). METHODS: This parallel group, randomised, non-inferiority, controlled trial randomised 247 participants by site and glycated haemoglobin (HbA1c) into TRE or CP (1:1) for 12 months. Participants were aged 35-70 years, with a body mass index (BMI) >25 but <45 kg/m2, and score ≥ 15 on the Australian type 2 diabetes risk (AUSDRISK) assessment, without a diagnosis of T2D. Study visits were balanced between groups and all participants received five consultations at 0, 0.5, 1, 2 and 3 months. TRE followed a self-selected 9 h eating window (≥0600 and ≤ 1900), whereas CP followed Australian dietary guidelines. OUTCOMES: The primary endpoint is the estimate of group mean difference (TRE vs CP) of HbA1c at 4 months in a covariate linear regression adjusting for stratification factors and sex. Secondary efficacy outcomes at 4 and 12 months are changes in fasting glucose, fasting insulin, HOMA-IR and nocturnal glucose by continuous glucose monitor incremental area under the curve and change in HbA1c at 12 months. Other endpoints are exploratory and will not be adjusted for multiplicity. CONCLUSIONS: We will determine whether TRE is an alternate strategy to current practice in dietetics to improve glucose control. TRIAL REGISTRATION: NCT04762251; 21 Feb 2021.

Exploring Australian Dietitians' knowledge, experience and perspectives of time-restricted eating in private practice: A qualitative study.

Time-restricted eating is a novel nutrition intervention with evidence of beneficial effects on weight loss, blood glucose management, and other metabolic health outcomes. Adherence to time-restricted eating is higher than some traditional nutrition interventions to support individuals living with overweight/obesity and type 2 diabetes mellitus. However, there may be an evidence-practice gap of time-restricted eating in Australian dietetic practice. The present study aimed to explore dietitians' knowledge, experiences, and perspectives of time-restricted eating and timing of eating advice in practice. Semi-structured interviews with 10 private practice dietitians across Australia were conducted. Audio recordings were transcribed and analysed thematically. Six themes were identified: (i) distinction of time-restricted eating to other fasting protocols; (ii) knowledge of health benefits of time-restricted eating; (iii) patient-led advice frequently given: timing of breakfast and dinner; (iv) dietitian-led advice frequently given: eating cut-off time to avoid late night snacking; (v) barriers and facilitators to offering time-restricted eating or timing of eating advice; (vi) timing of eating advice within professional guidelines and resources. These findings suggest the need for development of professional resources and educational development tools for dietitians on time-restricted eating.

The Need to Prioritize Education and Resources to Support Exercise in Type 1 Diabetes: Results of an Australian Survey of Adults With Type 1 Diabetes and Health Providers.

OBJECTIVES: Regular exercise is recommended for people with type 1 diabetes (PWD) to improve their health, but many do not meet recommended exercise targets. Educational resources supporting PWD to exercise exist, but their value is unclear. To determine the need for improved exercise resources in Australia, we surveyed adult PWD and health providers (HPs) about their confidence in managing type 1 diabetes (T1D) around exercise, barriers to exercise, and the adequacy of current resources. METHODS: Australian adult PWD and HPs completed surveys to rate the importance of exercise in T1D management, confidence in managing T1D around exercise, barriers to giving and receiving education, resources used, and what form new resources should take. RESULTS: Responses were received from 128 PWD and 122 HPs. Both groups considered exercise to be important for diabetes management. PWD cited time constraints (57%) and concern about dysglycemia (43%) as barriers to exercise, and many lacked confidence in managing T1D around exercise. HPs were more confident, but experienced barriers to providing advice, and PWD did not tend to rely on this advice. Instead, 72% of PWD found continuous glucose monitoring most helpful. Both groups desired better resources to support exercise in T1D, with PWD preferring to obtain information through a structured education program and HPs through eLearning. CONCLUSIONS: Australian HPs and PWD appreciate the importance of exercise in T1D management and express a clear desire for improved educational resources. Our findings provide a basis for developing a comprehensive package of resources for both adult PWD and HPs, to support exercise in PWD.

Efficacy of commercial peroxyacetic acid on Vibrio parahaemolyticus planktonic cells and biofilms on stainless steel and Greenshell™ mussel (Perna canaliculus) surfaces.

The potential of using commercial peroxyacetic acid (PAA) for Vibrio parahaemolyticus sanitization was evaluated. Commercial PAA of 0.005 % (v/v, PAA: 2.24 mg/L, hydrogen peroxide: 11.79 mg/L) resulted in a planktonic cell reduction of >7.00 log10 CFU/mL when initial V. parahaemolyticus cells averaged 7.64 log10 CFU/mL. For cells on stainless steel coupons, treatment of 0.02 % PAA (v/v, PAA: 8.96 mg/L, hydrogen peroxide: 47.16 mg/L) achieved >5.00 log10 CFU/cm2 reductions in biofilm cells for eight strains but not for the two strongest biofilm formers. PAA of 0.05 % (v/v, PAA: 22.39 mg/L, hydrogen peroxide: 117.91 mg/L) was required to inactivate >5.00 log10 CFU/cm2 biofilm cells from mussel shell surfaces. The detection of PAA residues after biofilm treatment demonstrated that higher biofilm production resulted in higher PAA residues (p < 0.05), suggesting biofilm is acting as a barrier interfering with PAA diffusing into the matrices. Based on the comparative analysis of genomes, robust biofilm formation and metabolic heterogeneity within niches might have contributed to the variations in PAA resistance of V. parahaemolyticus biofilms.

Novel hyperthermoacidic archaeal enzymes for removal of thermophilic biofilms from stainless steel.

AIMS: To test the efficacy of novel hot/acid hyperthermoacidic enzyme treatments on the removal of thermophilic spore-forming biofilms from stainless steel surfaces. METHODS AND RESULTS: The present study measured the efficacy of hyperthermoacidic enzymes (protease, amylase, and endoglucanase) that are optimally active at low pH (≈3.0) and high temperatures (≈80°C) at removing thermophilic bacilli biofilms from stainless steel (SS) surfaces. Plate counts, spore counts, impedance microbiology, as well as epifluorescence microscopy, and scanning electron microscopy (SEM) were used to evaluate the cleaning and sanitation of biofilms grown in a continuous flow biofilm reactor. Previously unavailable hyperthermoacidic amylase, protease, and the combination of amylase and protease were tested on Anoxybacillus flavithermus and Bacillus licheniformis, and endoglucanase was tested on Geobacillus stearothermophilus. In all cases, the heated acidic enzymatic treatments significantly reduced biofilm cells and their sheltering extracellular polymeric substances (EPS). CONCLUSIONS: Hyperthermoacidic enzymes and the associated heated acid conditions are effective at removing biofilms of thermophilic bacteria from SS surfaces that contaminate dairy plants.

Comparative genome identification of accessory genes associated with strong biofilm formation in Vibrio parahaemolyticus.

Vibrio parahaemolyticus biofilms on the seafood processing plant surfaces are a potential source of seafood contamination and subsequent food poisoning. Strains differ in their ability to form biofilm, but little is known about the genetic characteristics responsible for biofilm development. In this study, pangenome and comparative genome analysis of V. parahaemolyticus strains reveals genetic attributes and gene repertoire that contribute to robust biofilm formation. The study identified 136 accessory genes that were exclusively present in strong biofilm forming strains and these were functionally assigned to the Gene Ontology (GO) pathways of cellulose biosynthesis, rhamnose metabolic and catabolic processes, UDP-glucose processes and O antigen biosynthesis (p < 0.05). Strategies of CRISPR-Cas defence and MSHA pilus-led attachment were implicated via Kyoto Encyclopedia of Genes and Genomes (KEGG) annotation. Higher levels of horizontal gene transfer (HGT) were inferred to confer more putatively novel properties on biofilm-forming V. parahaemolyticus. Furthermore, cellulose biosynthesis, a neglected potential virulence factor, was identified as being acquired from within the order Vibrionales. The cellulose synthase operons in V. parahaemolyticus were examined for their prevalence (22/138, 15.94 %) and were found to consist of the genes bcsG, bcsE, bcsQ, bcsA, bcsB, bcsZ, bcsC. This study provides insights into robust biofilm formation of V. parahaemolyticus at the genomic level and facilitates: identification of key attributes for robust biofilm formation, elucidation of biofilm formation mechanisms and development of potential targets for novel control strategies of persistent V. parahaemolyticus.

Non-Targeted Metabolomic Profiling Identifies Metabolites with Potential Antimicrobial Activity from an Anaerobic Bacterium Closely Related to Terrisporobacter Species.

This work focused on the metabolomic profiling of the conditioned medium (FS03CM) produced by an anaerobic bacterium closely related to Terrisporobacter spp. to identify potential antimicrobial metabolites. The metabolome of the conditioned medium was profiled by two-channel Chemical Isotope Labelling (CIL) LC-MS. The detected metabolites were identified or matched by conducting a library search using different confidence levels. Forty-eight significantly changed metabolites were identified with high confidence after the growth of isolate FS03 in cooked meat glucose starch (CMGS) medium. Some of the secondary metabolites identified with known antimicrobial activities were 4-hydroxyphenyllactate, 3-hydroxyphenylacetic acid, acetic acid, isobutyric acid, valeric acid, and tryptamine. Our findings revealed the presence of different secondary metabolites with previously reported antimicrobial activities and suggested the capability of producing antimicrobial metabolites by the anaerobic bacterium FS03.

Inactivation of Cronobacter sakazakii biofilms using high voltage atmospheric cold plasma on various food-contact surfaces-a preliminary study.

Cronobacter sakazakii is an opportunistic foodborne pathogen in powdered infant formula (PIF) associated with several foodborne outbreaks. Biofilms of C. sakazakii in the PIF manufacturing plant may be a source of contamination, but information on the treatment of these biofilms is limited. This study investigated the inactivation of C. sakazakii biofilms on three food contact surfaces (stainless steel, silicone, and PVC) using high voltage atmospheric cold plasma (HVACP) applied as a dielectric barrier discharge in a 10:90 air: helium modified atmosphere. After the 90 s  of cold plasma exposure at 40 kV, C. sakazakii was reduced by ∼3 log CFU/coupon comparing to without cold plasma treatments. HVACP treatment caused cell shrinkage, fragmentation of the cell membrane, and leakage of cytoplasm. The inactivation of the cells on the surface was confirmed by live/dead staining. These above-mentioned results indicate the antibiofilm efficacy of HVACP on C. sakazakii isolates on various food contact surfaces.

Biofilm formation, sodium hypochlorite susceptibility and genetic diversity of Vibrio parahaemolyticus.

Vibrio parahaemolyticus is a marine oriented pathogen; and biofilm formation enables its survival and persistence on seafood processing plant, complicating the hygienic practice. The objectives of this study are to assess the ability of V. parahaemolyticus isolated from seafood related environments to form biofilms, to determine the effective sodium hypochlorite concentrations required to inactivate planktonic and biofilm cells, and to evaluate the genetic diversity required for strong biofilm formation. Among nine isolates, PFR30J09 and PFR34B02 isolates were identified as strong biofilm forming strains, with biofilm cell counts of 7.20, 7.08 log10 CFU/cm2, respectively, on stainless steel coupons after incubation at 25 °C. Free available chlorine of 1176 mg/L and 4704 mg/L was required to eliminate biofilm cells of 1.74-2.28 log10 CFU/cm2 and > 7 log10 CFU/cm2, respectively, whereas 63 mg/L for planktonic cells, indicating the ineffectiveness of sodium hypochlorite in eliminating V. parahaemolyticus biofilm cells at recommended concentration in the food industry. These strong biofilm-forming isolates produced more polysaccharides and were less susceptible to sodium hypochlorite, implying a possible correlation between polysaccharide production and sodium hypochlorite susceptibility. Genetic diversity in mshA, mshC and mshD contributed to the observed variation in biofilm formation between isolates. This study identified strong biofilm-forming V. parahaemolyticus strains of new multilocus sequence typing (MLST) types, showed a relationship between polysaccharide production and sodium hypochlorite resistance.

Complete Genome Sequences of Eight Faecalibacterium sp. Strains Isolated from Healthy Human Stool.

Eight Faecalibacterium sp. strains were isolated from feces of healthy human volunteers. Here, we describe their genome sequences. The genome sizes ranged from 2.78 Mbp to 3.23 Mbp, with an average GC content of 56.6% and encoding 2,795 protein-coding genes on average.

Differential effects of magnesium, calcium, and sodium on Listeria monocytogenes biofilm formation.

Listeria monocytogenes is a gram-positive foodborne pathogen that causes outbreaks of listeriosis associated with a diverse range of foods. L. monocytogenes forms biofilms as a strategy to enhance its survival in the environment. These biofilms then provide a source of contamination in processing plant environments. Cations like magnesium, calcium, and sodium are commonly found in the environment and are important to bacteria to maintain their homeostasis. It is, therefore, valuable to understand the relationship between these cations and biofilm formation. In this study, four isolates of L. monocytogenes from seafood processing environments were used to investigate the influence of magnesium, calcium, and sodium (1, 10, and 50 mM) on biofilms. The isolates selected were defined as being either a low biofilm former, a high biofilm former, an outbreak isolate, and a persistent isolate from the seafood industry. The study showed that the divalent cations magnesium and calcium increased biofilm formation compared with the monovalent cation, sodium. Fifty mM concentrations of the divalent cations significantly enhanced biofilm formation. The cations did not have a significant effect on the initial stages of biofilm formation but appeared to influence the later stages of biofilm development.

Monascus spp. and citrinin: Identification, selection of Monascus spp. isolates, occurrence, detection and reduction of citrinin during the fermentation of red fermented rice.

Red fermented rice (RFR) is rice fermented using Monascus spp. This product contains monacolin K, providing health benefits including mitigation of diarrhoea and improving blood circulation. RFR can produce pigments that can act as natural colour and flavouring agents. However, Monascus spp. (a fungal starter to ferment RFR) can also produce the mycotoxin, citrinin (CIT) which is believed to have adverse effects on human health. CIT in RFR has been reported worldwide by using different methods of detection. This review focuses on the production of RFR by solid-state fermentation (SSF) and submerged fermentation (SmF), the occurrence of CIT in RFR, CIT quantification, the factors affecting the growth of Monascus spp., pigments and CIT production in RFR, and possible methods to reduce CIT in RFR. This review will help the food industries, researchers, and consumers understand the risk of consuming RFR, and the possibility of controlling CIT in RFR.

Coconut Cream Agar as a simple and rapid semiquantitative method to screen citrinin-producing Monascus spp. isolates isolated from red fermented rice.

Red fermented rice (RFR) is produced using Monascus spp. This product has some health benefits. However, RFR can also contain the mycotoxin, citrinin (CIT) and that has adverse effects on human health. The objective of the study was to develop a simple and rapid screening method for the detection of Monascus spp. isolates that can produce CIT by using Coconut Cream Agar (CCA). RFR was spread onto CCA and other media and incubated at 30 °C for 7 days. All the media were observed daily under ultraviolet (UV) light and any Monascus spp. colony that produced light blue fluorescence was recorded as a CIT-producer. Two different isolates (MF1 and MS1) isolated from CCA were selected for further analysis. All (100%; 10/10 plates) of CCA inoculated with MF1 produced light blue fluorescence after incubation for 4 days, meanwhile 30% (3/10 plates) of MS1 produced weak fluorescence on CCA after incubation for 7 days. Isolates MF1 and MS1 were identified as M. purpureus with the ability to produce CIT by having polyketide synthase (pksCT) and transcriptional regulator (ctnA) genes. CIT was quantified by high-performance liquid chromatography (HPLC). CCA is a simple and rapid method to detect CIT-producers of Monascus spp.

Investigation of UV light treatment (254 nm) on the reduction of aflatoxin M1 in skim milk and degradation products after treatment.

This study investigates the reduction of aflatoxin M1 (AFM1) in skim milk by using ultraviolet light at 254 nm and the effects of influencing factors on the efficacy including treatment time (min), depth of samples (mm), contamination level (µg L-1), stirring, temperature, and fat content in milk. The colour and pH of milk samples were measured to evaluate the influence of the treatment on these values. It was found that short-wave ultraviolet radiation (UVC) reduced up to 50% of AFM1 in milk after 20 min of treatment regardless of the initial AFM1 contamination level. Treatment time, depth of samples, and stirring were all found to significantly (P < 0.05) enhance the reduction of AFM1. The milk colour was affected but there was no influence on the pH of milk samples at any duration of UV exposure. It is concluded that UVC light treatment has the potential to reduce AFM1 in milk.

Trans-kingdom interactions in mixed biofilm communities.

The microbial world represents a phenomenal diversity of microorganisms from different kingdoms of life, which occupy an impressive set of ecological niches. Most, if not all, microorganisms once colonize a surface develop architecturally complex surface-adhered communities, which we refer to as biofilms. They are embedded in polymeric structural scaffolds and serve as a dynamic milieu for intercellular communication through physical and chemical signalling. Deciphering microbial ecology of biofilms in various natural or engineered settings has revealed coexistence of microorganisms from all domains of life, including Bacteria, Archaea, and Eukarya. The coexistence of these dynamic microbes is not arbitrary, as a highly coordinated architectural setup and physiological complexity show ecological interdependence and myriads of underlying interactions. In this review, we describe how species from different kingdoms interact in biofilms and discuss the functional consequences of such interactions. We highlight metabolic advances of collaboration among species from different kingdoms, and advocate that these interactions are of great importance and need to be addressed in future research. Since trans-kingdom biofilms impact diverse contexts, ranging from complicated infections to efficient growth of plants, future knowledge within this field will be beneficial for medical microbiology, biotechnology, and our general understanding of microbial life in nature.

Control of aflatoxin M1 in skim milk by high voltage atmospheric cold plasma.

Cold plasma has potential for the degradation of aflatoxins in corn and hazelnuts; however, this has not been demonstrated for aflatoxin in milk. In this study, the efficacy of high voltage atmospheric cold plasma (HVACP) on the reduction of aflatoxin M1 (AFM1) in skim milk improved with increasing treatment times (1-20 min), using gas containing 65% oxygen (MA65) rather than air, increasing voltage (60-80 kV) and reducing sample volume (30 mL-10 mL). Direct treatment was more effective than indirect treatment. AFM1 in milk was degraded by 65.0 % and 78.9 % by air and MA65 respectively in 20 min with no change in milk colour. The toxicity of AFM1 after treatment was assessed using a brine shrimp model. A five-minute HVACP treatment reduced the toxicity of AFM1 by 83.9 % based on the increase in brine shrimp survival. HVACP is a promising method to reduce AFM1 in milk.

Antibacterial efficacy and possible mechanism of action of 2-hydroxyisocaproic acid (HICA).

The exploitation of natural antimicrobial compounds that can be used in food preservation has been fast tracked by the development of antimicrobial resistance to existing antimicrobials and the increasing consumer demand for natural food preservatives. 2-hydroxyisocaproic acid (HICA) is a natural compound produced through the leucine degradation pathway and is produced in humans and by certain microorganisms such as lactic acid bacteria and Clostridium species. The present study investigated the antibacterial efficacy of HICA against some important bacteria associated with food quality and safety and provided some insights into its possible antimicrobial mechanisms against bacteria. The results revealed that HICA was effective in inhibiting the growth of tested Gram-positive and Gram-negative bacteria including a multi-drug resistant P. aeruginosa strain in this study. The underlying mechanism was investigated by measuring the cell membrane integrity, membrane permeability, membrane depolarisation, and morphological and ultrastructural changes after HICA treatment in bacterial cells. The evidence supports that HICA exerts its activity via penetration of the bacterial cell membranes, thereby causing depolarisation, rupture of membranes, subsequent leakage of cellular contents and cell death. The current study suggests that HICA has potential to be used as an antibacterial agent against food spoilage and food-borne pathogenic bacteria, targeting the bacterial cell envelope.

Culture and genome-based analysis of four soil Clostridium isolates reveal their potential for antimicrobial production.

BACKGROUND: Soil bacteria are a major source of specialized metabolites including antimicrobial compounds. Yet, one of the most diverse genera of bacteria ubiquitously present in soil, Clostridium, has been largely overlooked in bioactive compound discovery. As Clostridium spp. thrive in extreme environments with their metabolic mechanisms adapted to the harsh conditions, they are likely to synthesize molecules with unknown structures, properties, and functions. Therefore, their potential to synthesize small molecules with biological activities should be of great interest in the search for novel antimicrobial compounds. The current study focused on investigating the antimicrobial potential of four soil Clostridium isolates, FS01, FS2.2 FS03, and FS04, using a genome-led approach, validated by culture-based methods. RESULTS: Conditioned/spent media from all four Clostridium isolates showed varying levels of antimicrobial activity against indicator microorganism; all four isolates significantly inhibited the growth of Pseudomonas aeruginosa. FS01, FS2.2, and FS04 were active against Bacillus mycoides and FS03 reduced the growth of Bacillus cereus. Phylogenetic analysis together with DNA-DNA hybridization (dDDH), average nucleotide identity (ANI), and functional genome distribution (FGD) analyses confirmed that FS01, FS2.2, and FS04 belong to the species Paraclostridium bifermentans, Clostridium cadaveris, and Clostridium senegalense respectively, while FS03 may represent a novel species of the genus Clostridium. Bioinformatics analysis using antiSMASH 5.0 predicted the presence of eight biosynthetic gene clusters (BGCs) encoding for the synthesis of ribosomally synthesized post-translationally modified peptides (RiPPs) and non-ribosomal peptides (NRPs) in four genomes. All predicted BGCs showed no similarity with any known BGCs suggesting novelty of the molecules from those predicted gene clusters. In addition, the analysis of genomes for putative virulence factors revealed the presence of four putative Clostridium toxin related genes in FS01 and FS2.2 genomes. No genes associated with the main Clostridium toxins were identified in the FS03 and FS04 genomes. CONCLUSIONS: The presence of BGCs encoding for uncharacterized RiPPs and NRPSs in the genomes of antagonistic Clostridium spp. isolated from farm soil indicated their potential to produce novel secondary metabolites. This study serves as a basis for the identification and characterization of potent antimicrobials from these soil Clostridium spp. and expands the current knowledge base, encouraging future research into bioactive compound production in members of the genus Clostridium.

In silico analysis revealing CsrA roles in motility-sessility switching and tuning VBNC cells in Vibrio parahaemolyticus.

The formation of biofilms is a survival strategy employed by bacteria to help protect them from changing or unfavourable environments. In this research, 319 genes which govern biofilm formation in V. parahaemolyticus, as reported in 1,625 publications, were analysed using protein-protein-interaction (PPI) network analysis. CsrA was identified as a motility-sessility switch and biofilm formation regulator. Through robust rank aggregation (RRA) analysis of GSE65340, the generation of viable but non-culturable (VBNC) cells that may enhance cell tolerance to stress, was found to be associated with the TCA cycle and carbon metabolism biological pathways. The finding that CsrA is likely to play a role in the development of VBNC cells improves understanding of the molecular mechanisms of VBNC formation in V. parahaemolyticus and contributes to on-going efforts to reduce the hazard posed by this foodborne pathogen.