Efficacy of Food Safety Training in Commercial Food Service.

Proper food safety training is essential to decrease incidences and overall rates of foodborne illnesses and outbreaks. Though many commercial restaurants should provide proper food safety training to food handler employees, this training is not always offered or effective. Here, we summarize the results of a primary literature study concerning the effectiveness of food safety training in commercial settings. The literature chosen for review contained only studies with experimental food safety training, with before and after training data. Through evaluation of these studies, the best practice for ensuring effective training and follow-through were the use of food safety training programs, which incorporated both knowledge and behavior-based training. PRACTICAL APPLICATION: Food safety managers in food service establishments may consider reevaluating their current food safety training program to incorporate behavioral-based food safety training in addition to knowledge-based training.

Characterization of the total and viable bacterial and fungal communities associated with the International Space Station surfaces.

BACKGROUND: The International Space Station (ISS) is a closed system inhabited by microorganisms originating from life support systems, cargo, and crew that are exposed to unique selective pressures such as microgravity. To date, mandatory microbial monitoring and observational studies of spacecraft and space stations have been conducted by traditional culture methods, although it is known that many microbes cannot be cultured with standard techniques. To fully appreciate the true number and diversity of microbes that survive in the ISS, molecular and culture-based methods were used to assess microbial communities on ISS surfaces. Samples were taken at eight pre-defined locations during three flight missions spanning 14 months and analyzed upon return to Earth. RESULTS: The cultivable bacterial and fungal population ranged from 104 to 109 CFU/m2 depending on location and consisted of various bacterial (Actinobacteria, Firmicutes, and Proteobacteria) and fungal (Ascomycota and Basidiomycota) phyla. Amplicon sequencing detected more bacterial phyla when compared to the culture-based analyses, but both methods identified similar numbers of fungal phyla. Changes in bacterial and fungal load (by culture and qPCR) were observed over time but not across locations. Bacterial community composition changed over time, but not across locations, while fungal community remained the same between samplings and locations. There were no significant differences in community composition and richness after propidium monoazide sample treatment, suggesting that the analyzed DNA was extracted from intact/viable organisms. Moreover, approximately 46% of intact/viable bacteria and 40% of intact/viable fungi could be cultured. CONCLUSIONS: The results reveal a diverse population of bacteria and fungi on ISS environmental surfaces that changed over time but remained similar between locations. The dominant organisms are associated with the human microbiome and may include opportunistic pathogens. This study provides the first comprehensive catalog of both total and intact/viable bacteria and fungi found on surfaces in closed space systems and can be used to help develop safety measures that meet NASA requirements for deep space human habitation. The results of this study can have significant impact on our understanding of other confined built environments on the Earth such as clean rooms used in the pharmaceutical and medical industries.

Multi-drug resistant Enterobacter bugandensis species isolated from the International Space Station and comparative genomic analyses with human pathogenic strains.

BACKGROUND: The antimicrobial resistance (AMR) phenotypic properties, multiple drug resistance (MDR) gene profiles, and genes related to potential virulence and pathogenic properties of five Enterobacter bugandensis strains isolated from the International Space Station (ISS) were carried out and compared with genomes of three clinical strains. Whole genome sequences of ISS strains were characterized using the hybrid de novo assembly of Nanopore and Illumina reads. In addition to traditional microbial taxonomic approaches, multilocus sequence typing (MLST) analysis was performed to classify the phylogenetic lineage. Agar diffusion discs assay was performed to test antibiotics susceptibility. The draft genomes after assembly and scaffolding were annotated with the Rapid Annotations using Subsystems Technology and RNAmmer servers for downstream analysis. RESULTS: Molecular phylogeny and whole genome analysis of the ISS strains with all publicly available Enterobacter genomes revealed that ISS strains were E. bugandensis and similar to the type strain EB-247T and two clinical isolates (153_ECLO and MBRL 1077). Comparative genomic analyses of all eight E. bungandensis strains showed, a total of 4733 genes were associated with carbohydrate metabolism (635 genes), amino acid and derivatives (496 genes), protein metabolism (291 genes), cofactors, vitamins, prosthetic groups, pigments (275 genes), membrane transport (247 genes), and RNA metabolism (239 genes). In addition, 112 genes identified in the ISS strains were involved in virulence, disease, and defense. Genes associated with resistance to antibiotics and toxic compounds, including the MDR tripartite system were also identified in the ISS strains. A multiple antibiotic resistance (MAR) locus or MAR operon encoding MarA, MarB, MarC, and MarR, which regulate more than 60 genes, including upregulation of drug efflux systems that have been reported in Escherichia coli K12, was also observed in the ISS strains. CONCLUSION: Given the MDR results for these ISS Enterobacter genomes and increased chance of pathogenicity (PathogenFinder algorithm with > 79% probability), these species pose important health considerations for future missions. Thorough genomic characterization of the strains isolated from ISS can help to understand the pathogenic potential, and inform future missions, but analyzing them in in-vivo systems is required to discern the influence of microgravity on their pathogenicity.

Draft Genome Sequences from a Novel Clade of Bacillus cereus Sensu Lato Strains, Isolated from the International Space Station.

The draft genome sequences of six Bacillus strains, isolated from the International Space Station and belonging to the Bacillus anthracis-B. cereus-B. thuringiensis group, are presented here. These strains were isolated from the Japanese Experiment Module (one strain), U.S. Harmony Node 2 (three strains), and Russian Segment Zvezda Module (two strains).

Non-Toxin-Producing Bacillus cereus Strains Belonging to the B. anthracis Clade Isolated from the International Space Station.

In an ongoing Microbial Observatory investigation of the International Space Station (ISS), 11 Bacillus strains (2 from the Kibo Japanese experimental module, 4 from the U.S. segment, and 5 from the Russian module) were isolated and their whole genomes were sequenced. A comparative analysis of the 16S rRNA gene sequences of these isolates showed the highest similarity (>99%) to the Bacillus anthracis-B. cereus-B. thuringiensis group. The fatty acid composition, polar lipid profile, peptidoglycan type, and matrix-assisted laser desorption ionization-time of flight profiles were consistent with the B. cereus sensu lato group. The phenotypic traits such as motile rods, enterotoxin production, lack of capsule, and resistance to gamma phage/penicillin observed in ISS isolates were not characteristics of B. anthracis. Whole-genome sequence characterizations showed that ISS strains had the plcR non-B. anthracis ancestral "C" allele and lacked anthrax toxin-encoding plasmids pXO1 and pXO2, excluding their identification as B. anthracis. The genetic identities of all 11 ISS isolates characterized via gyrB analyses arbitrarily identified them as members of the B. cereus group, but traditional DNA-DNA hybridization (DDH) showed that the ISS isolates are similar to B. anthracis (88% to 90%) but distant from the B. cereus (42%) and B. thuringiensis (48%) type strains. The DDH results were supported by average nucleotide identity (>98.5%) and digital DDH (>86%) analyses. However, the collective phenotypic traits and genomic evidence were the reasons to exclude the ISS isolates from B. anthracis. Nevertheless, multilocus sequence typing and whole-genome single nucleotide polymorphism analyses placed these isolates in a clade that is distinct from previously described members of the B. cereus sensu lato group but closely related to B. anthracis. IMPORTANCE The International Space Station Microbial Observatory (Microbial Tracking-1) study is generating a microbial census of the space station's surfaces and atmosphere by using advanced molecular microbial community analysis techniques supported by traditional culture-based methods and modern bioinformatic computational modeling. This approach will lead to long-term, multigenerational studies of microbial population dynamics in a closed environment and address key questions, including whether microgravity influences the evolution and genetic modification of microorganisms. The spore-forming Bacillus cereus sensu lato group consists of pathogenic (B. anthracis), food poisoning (B. cereus), and biotechnologically useful (B. thuringiensis) microorganisms; their presence in a closed system such as the ISS might be a concern for the health of crew members. A detailed characterization of these potential pathogens would lead to the development of suitable countermeasures that are needed for long-term future missions and a better understanding of microorganisms associated with space missions.

Whole metagenome profiles of particulates collected from the International Space Station.

BACKGROUND: The built environment of the International Space Station (ISS) is a highly specialized space in terms of both physical characteristics and habitation requirements. It is unique with respect to conditions of microgravity, exposure to space radiation, and increased carbon dioxide concentrations. Additionally, astronauts inhabit a large proportion of this environment. The microbial composition of ISS particulates has been reported; however, its functional genomics, which are pertinent due to potential impact of its constituents on human health and operational mission success, are not yet characterized. METHODS: This study examined the whole metagenome of ISS microbes at both species- and gene-level resolution. Air filter and dust samples from the ISS were analyzed and compared to samples collected in a terrestrial cleanroom environment. Furthermore, metagenome mining was carried out to characterize dominant, virulent, and novel microorganisms. The whole genome sequences of select cultivable strains isolated from these samples were extracted from the metagenome and compared. RESULTS: Species-level composition in the ISS was found to be largely dominated by Corynebacterium ihumii GD7, with overall microbial diversity being lower in the ISS relative to the cleanroom samples. When examining detection of microbial genes relevant to human health such as antimicrobial resistance and virulence genes, it was found that a larger number of relevant gene categories were observed in the ISS relative to the cleanroom. Strain-level cross-sample comparisons were made for Corynebacterium, Bacillus, and Aspergillus showing possible distinctions in the dominant strain between samples. CONCLUSION: Species-level analyses demonstrated distinct differences between the ISS and cleanroom samples, indicating that the cleanroom population is not necessarily reflective of space habitation environments. The overall population of viable microorganisms and the functional diversity inherent to this unique closed environment are of critical interest with respect to future space habitation. Observations and studies such as these will be important to evaluating the conditions required for long-term health of human occupants in such environments.

Solibacillus kalamii sp. nov., isolated from a high-efficiency particulate arrestance filter system used in the International Space Station.

A Gram-stain-positive, rod-shaped, endospore-forming, aerobic bacterial strain, designated ISSFR-015T, was isolated from a high-efficiency particulate arrestance filter in the International Space Station and was characterized by polyphasic taxonomy. A comparative analysis of the 16S rRNA gene sequence (1494 bp) of strain ISSFR-015T showed highest similarity to Solibacillus isronensis B3W22T (98.9 %), followed by Solibacillus silvestris HR3-23T (98.6 %) and Bacillus cecembensis PN5T (96.7 %). DNA-DNA hybridization analysis revealed that the DNA relatedness values of strain ISSFR-015T with other closely related species were in the range of 41-47 % [S. silvestrisMTCC 10789T (47 %), S. isronensis MTCC 7902T (41 %) and B. cecembensis MTCC 9127T (43 %)]. The DNA G+C content of strain ISSFR-015T was 45.4 mol%. The major fatty acids were iso-C15 : 0 (45.2 %) and C17 : 1ω10c (12.1 %). The polar lipid profile contained diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, phosphatidylserine and one unknown phospholipid. The isoprenoid quinones present in strain ISSFR-015T were MK-7 (86.8 %), MK-6 (11.6 %) and MK-8 (1.0 %). The peptidoglycan type of the cell wall was A4α l-Lys-d-Glu. Based on the phylogenetic analysis, strain ISSFR-015T belongs to the genus Solibacillus. The polyphasic taxonomic data, including low DNA-DNA hybridization values, and the chemotaxonomic analysis confirmed that strain ISSFR-015T represents a novel species, for which the name Solibacillus kalamii sp. nov. is proposed. The type strain for this proposed species is ISSFR-015T (=NRRL B-65388T=DSM 101595T).

Draft Genome Sequences of Biosafety Level 2 Opportunistic Pathogens Isolated from the Environmental Surfaces of the International Space Station.

The draft genome sequences of 20 biosafety level 2 (BSL-2) opportunistic pathogens isolated from the environmental surfaces of the International Space Station (ISS) were presented. These genomic sequences will help in understanding the influence of microgravity on the pathogenicity and virulence of these strains when compared with Earth strains.