Advances, applications, and limitations of portable and rapid detection technologies for routinely encountered foodborne pathogens.

Traditional foodborne pathogen detection methods are highly dependent on pre-treatment of samples and selective microbiological plating to reliably screen target microorganisms. Inherent limitations of conventional methods include longer turnaround time and high costs, use of bulky equipment, and the need for trained staff in centralized laboratory settings. Researchers have developed stable, reliable, sensitive, and selective, rapid foodborne pathogens detection assays to work around these limitations. Recent advances in rapid diagnostic technologies have shifted to on-site testing, which offers flexibility and ease-of-use, a significant improvement from traditional methods' rigid and cumbersome steps. This comprehensive review aims to thoroughly discuss the recent advances, applications, and limitations of portable and rapid biosensors for routinely encountered foodborne pathogens. It discusses the major differences between biosensing systems based on the molecular interactions of target analytes and biorecognition agents. Though detection limits and costs still need further improvement, reviewed technologies have high potential to assist the food industry in the on-site detection of biological hazards such as foodborne pathogens and toxins to maintain safe and healthy foods. Finally, this review offers targeted recommendations for future development and commercialization of diagnostic technologies specifically for emerging and re-emerging foodborne pathogens.

Whole-Genome Analysis of Escherichia Phage vB_EcoM-S1P5QW, Isolated from Manures Collected from Cattle Farms in Maine.

Here, we report a complete genome sequence of Escherichia phage vB_EcoM-S1P5QW, a T4-like bacteriophage that was isolated from manures collected from cattle farms in Maine. Escherichia phage vB_EcoM-S1P5QW can infect Escherichia coli O26:H11 strains and is devoid of virulence, antibiotic resistance, and lysogeny-associated genes, which may be meaningful for further biocontrol studies.

A review of epidemic investigation on cold-chain food-mediated SARS-CoV-2 transmission and food safety consideration during COVID-19 pandemic.

COVID-19 has brought speculations on potential transmission routes of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causal agent of the pandemic. It is reported that the main route of virus transmission to be person-to-person by respiratory droplets; however, people have raised concerns on the possible transmission of SARS-CoV-2 to humans via food and packaging and its potential effects on food safety. This review discusses food safety issues in the COVID-19 pandemic and reveals its possible transmission in cold-chain food. The first outbreak of COVID-19 in late 2019 was associated with a seafood market in Wuhan, China, while the second outbreak of COVID-19 in June 2020 was also related to a seafood market in Beijing, China. As of 2020, several frozen seafood products linked with SARS-CoV-2 have been reported in China. According to the current survey and scientific studies, the risk of infection by SARS-CoV-2 from cold-chain food, food products, and food packaging is thought to be very low. However, studies on food cold chain contamination have shown that SARS-CoV-2 remained highly stable under refrigerated (4°C) and even in freezing conditions (-10 to -80°C). Since one mode of SARS-CoV-2 transmission appears to be touching contaminated surfaces, it is important to clean and sanitize food contact surfaces properly. Understanding food safety hazard risks is essential to avoid potential negative health effects and SARS-CoV-2 transmission in the food supply chain during the COVID-19 pandemic.

Enhanced Detection of Major Pathogens and Toxins in Poultry and Livestock With Zoonotic Risks Using Nanomaterials-Based Diagnostics.

Nanotechnology has gained prominence over the recent years in multiple research and application fields, including infectious diseases in healthcare, agriculture, and veterinary science. It remains an attractive and viable option for preventing, diagnosing, and treating diseases in animals and humans. The apparent efficiency of nanomaterials is due to their unique physicochemical properties and biocompatibility. With the persistence of pathogens and toxins in the poultry and livestock industries, rapid diagnostic tools are of utmost importance. Though there are many promising nanomaterials-based diagnostic tests specific to animal disease-causing agents, many have not achieved balanced sensitivity, specificity, reproducibility, and cost-effectiveness. This mini-review explores several types of nanomaterials, which provided enhancement on the sensitivity and specificity of recently reported diagnostic tools related to animal diseases. Recommendations are also provided to facilitate more targeted animal populations into the development of future diagnostic tools specifically for emerging and re-emerging animal diseases posing zoonotic risks.

Food safety lessons learned from the COVID-19 pandemic.

The COVID-19 pandemic has ushered in a new era of food safety. To date, there is no evidence to suggest that consuming food is associated with COVID-19. Nevertheless, COVID-19's impact on food safety and security has been grave. The world is currently experiencing several supply chain issues as a direct result of extensive lockdowns and impacts on essential workers' safety. However, disruption in the food supply, while catastrophic in nature, has created opportunities for the advancement of medical science, data processing, security monitoring, foodborne pathogen detection, and food safety technology. This article will discuss the key components for food safety during the COVID-19 pandemic. The discussion will draw from lessons learned early in the outbreak and will analyze the etiology of the disease through a food safety perspective. From there, we will discuss personal protective equipment, detection of SARS-CoV-2, useful surrogates to study SARS-CoV-2, and the expanding field of data science, from the food safety point of view. In the future, scientists can apply the knowledge to the containment of COVID-19 and eventually to future pandemics.

A sandwich-type bacteriophage-based amperometric biosensor for the detection of Shiga toxin-producing Escherichia coli serogroups in complex matrices.

Immuno-based biosensors are a popular tool designed for pathogen screening and detection. The current antibody-based biosensors employ direct, indirect, or sandwich detection approaches; however, instability, cross-reactivity, and high-cost render them unreliable and impractical. To circumvent these drawbacks, here we report a portable sandwich-type bacteriophage-based amperometric biosensor, which is highly-specific to various Shiga toxin-producing Escherichia coli (STEC) serogroups. Environmentally isolated and biotinylated bacteriophages were directly immobilized onto a streptavidin-coated screen-printed carbon electrode (SPCE), which recognized and captured viable target cells. Samples (50 µL) were transferred to these bacteriophage-functionalized SPCEs (12 min, room temp) before sequentially adding a bacteriophage-gold nanoparticle solution (20 µL), H2O2 (40 mM), and 1,1'-ferrocenedicarboxylic acid for amperometric tests (100 mV s-1) and analysis (ANOVA and LSD, P < 0.05). The optimum biotin concentration (10 mM) retained 94.47% bacteriophage viability. Non-target bacteria (Listeria monocytogenes and Salmonella Typhimurium) had delta currents below the threshold of a positive detection. With less than 1 h turn-around time, the amperometric biosensor had a detection limit of 10-102 CFU mL-1 for STEC O157, O26, and O179 strains and R 2 values of 0.97, 0.99, and 0.87, respectively, and a similar detection limit was observed in complex matrices, 10-102 CFU g-1 or mL-1 with R 2 values of 0.98, 0.95, and 0.76, respectively. The newly developed portable amperometric biosensor was able to rapidly detect viable target cells at low inoculum levels, thus providing an inexpensive and improved alternative to the current immuno- and laboratory-based STEC screening methods.

Simultaneous Colorimetric Detection of a Variety of Salmonella spp. in Food and Environmental Samples by Optical Biosensing Using Oligonucleotide-Gold Nanoparticles.

Optical biosensors for rapid detection of significant foodborne pathogens are steadily gaining popularity due to its simplicity and sensitivity. While nanomaterials such as gold nanoparticles (AuNPs) are commonly used as signal amplifiers for optical biosensors, AuNPs can also be utilized as a robust biosensing platform. Many reported optical biosensors were designed for individual pathogen detection in a single assay and have high detection limit (DL). Salmonella spp. is one of the major causative agents of foodborne sickness, hospitalization and deaths. Unfortunately, there are around 2,000 serotypes of Salmonella worldwide, and rapid and simultaneous detection of multiple strains in a single assay is lacking. In this study, a comprehensive and highly sensitive simultaneous colorimetric detection of nineteen (19) environmental and outbreak Salmonella spp. strains was achieved by a novel optical biosensing platform using oligonucleotide-functionalized AuNPs. A pair of newly designed single stranded oligonucleotides (30-mer) was displayed onto the surface of AuNPs (13 nm) as detection probes to hybridize with a conserved genomic region (192-bases) of ttrRSBCA found on a broad range of Salmonella spp. strains. The sandwich hybridization (30 min, 55°C) resulted in a structural formation of highly stable oligonucleotide/AuNPs-DNA complexes which remained undisturbed even after subjecting to an increased salt concentration (2 M, final), thus allowing a direct discrimination via color change of target (red color) from non-target (purplish-blue color) reaction mixtures by direct observation using the naked eye. In food matrices (blueberries and chicken meat), nineteen different Salmonella spp. strains were concentrated using immunomagnetic separation and then simultaneously detected in a 96-well microplate by oligonucleotide-functionalized AuNPs after DNA preparation. Successful oligonucleotide/AuNPs-DNA hybridization was confirmed by gel electrophoresis while AuNPs aggregation in non-target and control reaction mixtures was verified by both spectrophotometric analysis and TEM images. Results showed that the optical AuNP biosensing platform can simultaneously screen nineteen (19) viable Salmonella spp. strains tested with 100% specificity and a superior detection limit of <10 CFU/mL or g for both pure culture and complex matrices setups. The highly sensitive colorimetric detection system can significantly improve the screening and detection of viable Salmonella spp. strains present in complex food and environmental matrices, therefore reducing the risks of contamination and incidence of foodborne diseases.

Discovery of Shiga Toxin-Producing Escherichia coli (STEC)-Specific Bacteriophages From Non-fecal Composts Using Genomic Characterization.

Composting is a complex biodegradable process that converts organic materials into nutrients to facilitate crop yields, and, if well managed, can render bactericidal effects. Majority of research focused on detection of enteric pathogens, such as Shiga toxin-producing Escherichia coli (STEC) in fecal composts. Recently, attention has been emphasized on bacteriophages, such as STEC-specific bacteriophages, associated with STEC from the fecal-contaminated environment because they are able to sustain adverse environmental condition during composting process. However, little is known regarding the isolation of STEC-specific bacteriophages in non-fecal composts. Thus, the objectives were to isolate and genomically characterize STEC-specific bacteriophages, and to evaluate its association with STEC in non-fecal composts. For bacteriophage isolation, the samples were enriched with non-pathogenic E. coli (3 strains) and STEC (14 strains), respectively. After purification, host range, plaque size, and phage morphology were examined. Furthermore, bacteriophage genomes were subjected to whole-genome sequencing using Illumina MiSeq and genomic analyses. Isolation of top six non-O157 and O157 STEC utilizing culture methods combined with PCR-based confirmation was also conducted. The results showed that various STEC-specific bacteriophages, including vB_EcoM-Ro111lw, vB_EcoM-Ro121lw, vB_EcoS-Ro145lw, and vB_EcoM-Ro157lw, with different but complementary host ranges were isolated. Genomic analysis showed the genome sizes varied from 42kb to 149kb, and most bacteriophages were unclassified at the genus level, except vB_EcoM-Ro111lw as FelixO1-like viruses. Prokka predicted less than 25% of the ORFs coded for known functions, including those essential for DNA replication, bacteriophage structure, and host cell lysis. Moreover, none of the bacteriophages harbored lysogenic genes or virulence genes, such as stx or eae. Additionally, the presence of these lytic bacteriophages was likely attributed to zero isolation of STEC and could also contribute to additional antimicrobial effects in composts, if the composting process was insufficient. Current findings indicate that various STEC-specific bacteriophages were found in the non-fecal composts. In addition, the genomic characterization provides in-depth information to complement the deficiency of biological features regarding lytic cycle of the new bacteriophages. Most importantly, these bacteriophages have great potential to control various serogroups of STEC.

Investigation of prevalence of free Shiga toxin-producing Escherichia coli (STEC)-specific bacteriophages and its correlation with STEC bacterial hosts in a produce-growing area in Salinas, California.

Shiga toxin-producing E. coli (STEC) causes approximately 265,000 illnesses and 3,600 hospitalizations annually and is highly associated with animal contamination due to the natural reservoir of ruminant gastrointestinal tracts. Free STEC-specific bacteriophages against STEC strains are also commonly isolated from fecal-contaminated environment. Previous studies have evaluated the correlation between the prevalence of STEC-specific bacteriophages and STEC strains to improve animal-associated environment. However, the similar information regarding free STEC-specific bacteriophages prevalence in produce growing area is lacking. Thus, the objectives of this research were to determine the prevalence of STEC-specific phages, analyze potential effects of environmental factors on the prevalence of the phages, and study correlations between STEC-specific bacteriophages and the bacterial hosts in pre-harvest produce environment. Surface water from 20 samples sites was subjected to free bacteriophage isolation using host strains of both generic E. coli and STEC (O157, six non-O157 and one O179 strains) cocktails, and isolation of O157 and non-O157 STEC strains by use of culture methods combined with PCR-based confirmation. The weather data were obtained from weather station website. Free O145- and O179-specific bacteriophages were the two most frequently isolated bacteriophages among all (O45, O145, O157 and O179) in this study. The results showed June and July had relatively high prevalence of overall STEC-specific bacteriophages with minimum isolation of STEC strains. In addition, the bacteriophages were likely isolated in the area-around or within city-with predominant human impact, whereas the STEC bacterial isolates were commonly found in agriculture impact environment. Furthermore, there was a trend that the sample sites with positive of free STEC bacteriophage did not have the specific STEC bacterial hosts. The findings of the study enable us to understand the ecology between free STEC-specific phages and STEC bacteria for further pre-harvest food safety management in produce environment.

Simultaneous direct detection of Shiga-toxin producing Escherichia coli (STEC) strains by optical biosensing with oligonucleotide-functionalized gold nanoparticles.

A simultaneous direct detection of Shiga-toxin producing strains of E. coli (STEC; "Big Six" - O26, O45, O103, O111, O121, and O145) as well as O157 strains by optical biosensing with oligonucleotide-functionalized gold nanoparticles (AuNPs) was developed. Initially, conserved regions of stx genes were amplified by asymmetric polymerase chain reaction (asPCR). Pairs of single stranded thiol-modified oligonucleotides (30-mer) were immobilized onto AuNPs and used as probes to capture regions of stx1 (119-bp) and/or stx2 (104-bp) genes from STEC strains. DNA samples from pure cultures and food samples were sandwich hybridized with AuNP-oligo probes at optimal conditions (50 °C, 30 min). A complex was formed from the hybridization of AuNP-probes and target DNA fragments that retained the initial red color of the reaction solutions. For non-target DNA, a color change from red to purplish-blue was observed following an increase in salt concentration, thus providing the basis of simultaneous direct colorimetric detection of target DNA in the samples. Enrichment and pooling systems were incorporated to efficiently process a large number of food samples (ground beef and blueberries) and detection of live targets. The detection limit was <1 log CFU g(-1), requiring less than 1 h to complete after DNA sample preparation with 100% specificity. Gel electrophoresis verified AuNP-DNA hybridization while spectrophotometric data and transmission electron microscope (TEM) images supported color discrimination based on the occurrence of molecular aggregation. In conclusion, the significant features of this approach took advantage of the unique colorimetric properties of AuNPs as a low-cost and simple approach yet with high specificity for simultaneous detection of STEC strains.