G-site residue S67 is involved in the fungicide-degrading activity of a tau class glutathione S-transferase from Carica papaya.

Thiram is a toxic fungicide extensively used for the management of pathogens in fruits. Although it is known that thiram degrades in plant tissues, the key enzymes involved in this process remain unexplored. In this study, we report that a tau class glutathione S-transferase (GST) from Carica papaya can degrade thiram. This enzyme was easily obtained by heterologous expression in Escherichia coli, showed low promiscuity toward other thiuram disulfides, and catalyzed thiram degradation under physiological reaction conditions. Site-directed mutagenesis indicated that G-site residue S67 shows a key influence for the enzymatic activity toward thiram, while mutation of residue S13, which reduced the GSH oxidase activity, did not significantly affect the thiram-degrading activity. The formation of dimethyl dithiocarbamate, which was subsequently converted into carbon disulfide, and dimethyl dithiocarbamoylsulfenic acid as the thiram degradation products suggested that thiram undergoes an alkaline hydrolysis that involves the rupture of the disulfide bond. Application of the GST selective inhibitor 4-chloro-7-nitro-2,1,3-benzoxadiazole reduced papaya peel thiram-degrading activity by 95%, indicating that this is the main degradation route of thiram in papaya. GST from Carica papaya also catalyzed the degradation of the fungicides chlorothalonil and thiabendazole, with residue S67 showing again a key influence for the enzymatic activity. These results fill an important knowledge gap in understanding the catalytic promiscuity of plant GSTs and reveal new insights into the fate and degradation products of thiram in fruits.

Cellulose Surface Nanoengineering for Visualizing Food Safety.

Food safety is vital to human health, necessitating the development of nondestructive, convenient, and highly sensitive methods for detecting harmful substances. This study integrates cellulose dissolution, aligned regeneration, in situ nanoparticle synthesis, and structural reconstitution to create flexible, transparent, customizable, and nanowrinkled cellulose/Ag nanoparticle membranes (NWCM-Ag). These three-dimensional nanowrinkled structures considerably improve the spatial-electromagnetic-coupling effect of metal nanoparticles on the membrane surface, providing a 2.3 × 108 enhancement factor for the surface-enhanced Raman scattering (SERS) effect for trace detection of pesticides in foods. Notably, the distribution of pesticides in the apple peel and pulp layers is visualized through Raman imaging, confirming that the pesticides penetrate the peel layer into the pulp layer (∼30 µm depth). Thus, the risk of pesticide ingestion from fruits cannot be avoided by simple washing other than peeling. This study provides a new idea for designing nanowrinkled structures and broadening cellulose utilization in food safety.

Lipidomics of homeoviscous adaptation to low temperatures in Staphylococcus aureus utilizing exogenous straight-chain unsaturated fatty acids.

It is well established that Staphylococcus aureus can incorporate exogenous straight-chain unsaturated fatty acids (SCUFAs) into membrane phospho- and glyco-lipids from various sources in supplemented culture media and when growing in vivo during infection. Given the enhancement of membrane fluidity when oleic acid (C18:1Δ9) is incorporated into lipids, we were prompted to examine the effect of medium supplementation with C18:1Δ9 on growth at low temperatures. C18:1Δ9 supported the growth of a cold-sensitive, branched-chain fatty acid (BCFA)-deficient mutant at 12°C. Interestingly, we found similar results in the BCFA-sufficient parental strain, supported by the fact that the incorporation of C18:1Δ9 into the membrane increased membrane fluidity in both strains. We show that the incorporation of C18:1Δ9 and its elongation product C20:1Δ11 into membrane lipids was required for growth stimulation and relied on a functional FakAB incorporation system. Lipidomics analysis of the phosphatidylglycerol and diglycosyldiacylglycerol lipid classes revealed major impacts of C18:1Δ9 and temperature on lipid species. Growth at 12°C in the presence of C18:1Δ9 also led to increased production of the carotenoid pigment staphyloxanthin. The enhancement of growth by C18:1Δ9 is an example of homeoviscous adaptation to low temperatures utilizing an exogenous fatty acid. This may be significant in the growth of S. aureus at low temperatures in foods that commonly contain C18:1Δ9 and other SCUFAs in various forms. IMPORTANCE: We show that Staphylococcus aureus can use its known ability to incorporate exogenous fatty acids to enhance its growth at low temperatures. Individual species of phosphatidylglycerols and diglycosyldiacylglycerols bearing one or two degrees of unsaturation derived from the incorporation of C18:1Δ9 at 12°C are described for the first time. In addition, enhanced production of the carotenoid staphyloxanthin occurs at low temperatures. The studies describe a biochemical reality underlying membrane biophysics. This is an example of homeoviscous adaptation to low temperatures utilizing exogenous fatty acids over the regulation of the biosynthesis of endogenous fatty acids. The studies have likely relevance to food safety in that unsaturated fatty acids may enhance the growth of S. aureus in the food environment.

Subdural Empyema from Streptococcus suis Infection, South Korea.

In Jeju Island, South Korea, a patient who consumed raw pig products had subdural empyema, which led to meningitis, sepsis, and status epilepticus. We identified Streptococcus suis from blood and the subdural empyema. This case illustrates the importance of considering dietary habits in similar clinical assessments to prevent misdiagnosis.

High Prevalence of Echinostoma mekongi Infection in Schoolchildren and Adults, Kandal Province, Cambodia.

A high prevalence of Echinostoma mekongi infection (13.9%; 260/1,876) was found among schoolchildren and adults in Kandal Province, Cambodia, by fecal examination, worm expulsion, and molecular analysis of cox1 and nd1 genes. The source of infection was consumption of Pila sp. snails, a finding confirmed morphologically and molecularly.

Predictive Mapping of Antimicrobial Resistance for Escherichia coli, Salmonella, and Campylobacter in Food-Producing Animals, Europe, 2000-2021.

In Europe, systematic national surveillance of antimicrobial resistance (AMR) in food-producing animals has been conducted for decades; however, geographic distribution within countries remains unknown. To determine distribution within Europe, we combined 33,802 country-level AMR prevalence estimates with 2,849 local AMR prevalence estimates from 209 point prevalence surveys across 31 countries. We produced geospatial models of AMR prevalence in Escherichia coli, nontyphoidal Salmonella, and Campylobacter for cattle, pigs, and poultry. We summarized AMR trends by using the proportion of tested antimicrobial compounds with resistance >50% and generated predictive maps at 10 × 10 km resolution that disaggregated AMR prevalence. For E. coli, predicted prevalence rates were highest in southern Romania and southern/eastern Italy; for Salmonella, southern Hungary and central Poland; and for Campylobacter, throughout Spain. Our findings suggest that AMR distribution is heterogeneous within countries and that surveillance data from below the country level could help with prioritizing resources to reduce AMR.

Acute Gastroenteritis Associated with Norovirus GII.8[P8], Thailand, 2023.

Acute gastroenteritis associated with human norovirus infection was reported in Phuket, Thailand, in June 2023. We amplified GII.8[P8] from the outbreak stool specimens. Retrospective sample analysis identified infrequent GII.8[P8] in the country beginning in 2018. In all, the 10 whole-genome GII.8[P8] sequences from Thailand we examined had no evidence of genotypic recombination.

Detection of Rat Hepatitis E Virus in Pigs, Spain, 2023.

We identified rat hepatitis E virus (HEV) RNA in farmed pigs from Spain. Our results indicate that pigs might be susceptible to rat HEV and could serve as viral intermediaries between rodents and humans. Europe should evaluate the prevalence of rat HEV in farmed pigs to assess the risk to public health.

Foodborne Disease Outbreaks Linked to Foods Eligible for Irradiation, United States, 2009-2020.

Food irradiation can reduce foodborne illnesses but is rarely used in the United States. We determined whether outbreaks related to Campylobacter, Salmonella, Escherichia coli, and Listeria monocytogenes were linked to irradiation-eligible foods. Of 482 outbreaks, 155 (32.2%) were linked to an irradiation-eligible food, none of which were known to be irradiated.

Emergence of Group B Streptococcus Disease in Pigs and Porcupines, Italy.

We describe group B Streptococcus linked to disease in farmed pigs and wild porcupines in Italy. Occurrence in pigs was attributed to transmission from nonpasteurized bovine milk whey. Antimicrobial-resistance profiles in isolates from porcupines suggest no common source of infection. Our findings expand the known host range for group B Streptococcus disease.

Uncommon Salmonella Infantis Variants with Incomplete Antigenic Formula in the Poultry Food Chain, Italy.

Uncommon Salmonella Infantis variants displaying only flagellar antigens phenotypically showed identical incomplete antigenic formula but differed by molecular serotyping. Although most formed rough colonies, all shared antimicrobial resistances and the presence of usg gene with wild-type Salmonella Infantis. Moreover, they were undistinguishable wild-type Salmonella Infantis by whole-genome sequencing.

Vibrio mimicus Lineage Carrying Cholera Toxin and Vibrio Pathogenicity Island, United States and China.

Vibrio mimicus bacteria have caused sporadic cases and outbreaks of cholera-like diarrhea throughout the world, but the association of lineages with such events is unexplored. Genomic analyses revealed V. mimicus lineages carrying the virulence factors cholera toxin and toxin coregulated pilus, one of which has persisted for decades in China and the United States.

Genomic Diversity and Zoonotic Potential of Brucella neotomae.

After reports in 2017 of Brucella neotomae infections among humans in Costa Rica, we sequenced 12 strains isolated from rodents during 1955-1964 from Utah, USA. We observed an exact strain match between the human isolates and 1 Utah isolate. Independent confirmation is required to clarify B. neotomae zoonotic potential.

Emergence of Poultry-Associated Human Salmonella enterica Serovar Abortusovis Infections, New South Wales, Australia.

Salmonella enterica serovar Abortusovis is a ovine-adapted pathogen that causes spontaneous abortion. Salmonella Abortusovis was reported in poultry in 2009 and has since been reported in human infections in New South Wales, Australia. Phylogenomic analysis revealed a clade of 51 closely related isolates from Australia originating in 2004. That clade was genetically distinct from ovine-associated isolates. The clade was widespread in New South Wales poultry production facilities but was only responsible for sporadic human infections. Some known virulence factors associated with human infections were only found in the poultry-associated clade, some of which were acquired through prophages and plasmids. Furthermore, the ovine-associated clade showed signs of genome decay, but the poultry-associated clade did not. Those genomic changes most likely led to differences in host range and disease type. Surveillance using the newly identified genetic markers will be vital for tracking Salmonella Abortusovis transmission in animals and to humans and preventing future outbreaks.

Enhanced pyrophosphate detection: Utilizing oPD-derived carbon dots and Fe3+ interactions in a paper strip biosensor.

The development of portable, cost-effective, and straightforward DNA biosensors holds immense importance in various fields, including healthcare, environmental monitoring, and food safety. This study contributes to the objective by introducing an innovative approach for synthesizing carbon dots (Cdots) with high quantum yield (QY) and remarkable selectivity for Fe3+ ions. Utilizing o-phenylenediamine as a precursor, the study achieved a straightforward and environmentally friendly synthesis method, enabling the efficient detachment of metal ions from the Cdot surface upon introducing pyrophosphate (PPi). The presence of surface hydroxyl and amino groups facilitated specific Fe3+ recognition. Employing D-optimal response surface methodology, the study optimized Cdot synthesis parameters, identifying temperature and heating time as critical factors influencing QY. Statistical analysis confirmed the model's reliability, predicting maximum QY of 48.8 % with minimal deviation from experimental results. Characterization studies revealed the amorphous nature of Cdots through HR-TEM, XRD, and FTIR analysis. Furthermore, the proposed LAMP/PPi biosensing technique demonstrated higher sensitivity, specificity, and repeatability, with negligible interference from common anions and efficacy across varying pH levels. The limit of detection (LOD) of 0.079 (±0.01) µM and the detection range of 0.1 µM-2 mM underscore the biosensor's practical utility. This study highlights a promising direction for developing paper-based LAMP/PPi biosensors with potential diagnostics and environmental monitoring applications. Significantly, the biosensing technique is applicable to any DNA amplification method generating pyrophosphate (PPi) as a by-product.

Multiomics and biotechnologies for understanding and influencing cadmium accumulation and stress response in plants.

Cadmium (Cd) is one of the most toxic heavy metals faced by plants and, additionally, via the food chain, threatens human health. It is principally dispersed through agro-ecosystems via anthropogenic activities and geogenic sources. Given its high mobility and persistence, Cd, although not required, can be readily assimilated by plants thereby posing a threat to plant growth and productivity as well as animal and human health. Thus, breeding crop plants in which the edible parts contain low to zero Cd as safe food stuffs and harvesting shoots of high Cd-containing plants as a route for decontaminating soils are vital strategies to cope with this problem. Recently, multiomics approaches have been employed to considerably enhance our understanding of the mechanisms underlying (i) Cd toxicity, (ii) Cd accumulation, (iii) Cd detoxification and (iv) Cd acquisition tolerance in plants. This information can be deployed in the development of the biotechnological tools for developing plants with modulated Cd tolerance and detoxification to safeguard cellular and genetic integrity as well as to minimize food chain contamination. The aim of this review is to provide a current update about the mechanisms involved in Cd uptake by plants and the recent developments in the area of multiomics approach in terms of Cd stress responses, as well as in the development of Cd tolerant and low Cd accumulating crops.

Microbiological survey and genomic analysis of Cronobacter sakazakii strains isolated from US households and retail foods.

Cronobacter species are opportunistic pathogens that are capable of causing morbidity and mortality, particularly in infants. Although the transmission dynamics involved in Cronobacter infections remain largely unknown, contaminated powdered infant formula (PIF) has been linked to 30% of Cronobacter sakazakii cases involving invasive illness in infants. As several lines of evidence have implicated the domestic environment in PIF contamination, we undertook a microbiological survey of homes (N = 263) across the US. Cronobacter spp. and C. sakazakii were isolated from 36.1% and 24.7% of US homes, respectively, with higher recovery rates observed for floor and kitchen surfaces. Multi-locus sequence typing indicated that the dominant strain was C. sakazakii ST4, the sequence type most commonly associated with neonatal meningitis. For comparison purposes, retail foods (N = 4,009) were also surveyed, with the highest contamination frequencies (10.1%-26.3%) seen for nut products, seeds, and grains/baked goods/flours. The sequence type profile of isolates recovered from homes mirrored that of isolates recovered from retail foods, with increased representation of ST1, ST4, ST13, ST17, and ST40. Analysis of 386 whole genomic sequences revealed significant diversity. Redundancies were only observed for isolates recovered from within the same domicile, and there were no identical matches with sequences archived at the NCBI pathogen database. Genes coding for putative virulence and antibiotic resistance factors did not segregate with clinically significant sequence types. Collectively, these findings support the possibility that contamination events occurring within the home should not be overlooked as a contributor to community-onset Cronobacter infections. IMPORTANCE: Cronobacter sakazakii is an opportunistic pathogen that can cause significant morbidity and mortality in neonates. Its transmission dynamics are poorly understood, though powered infant formula (PIF) is thought to be the major transmission vehicle. How the PIF becomes contaminated remains unknown. Our survey shows that roughly 1/4 of US homes are contaminated with Cronobacter sakazakii, particularly in the kitchen setting. Our analyses suggest that the domestic environment may contribute to contamination of PIF and provides insights into mitigating the risk of transmission.

Time of arrival during plant disease progression and humidity additively influence Salmonella enterica colonization of lettuce.

The interplay between plant hosts, phytopathogenic bacteria, and enteric human pathogens in the phyllosphere has consequences for human health. Salmonella enterica has been known to take advantage of phytobacterial infection to increase its success on plants, but there is little knowledge of additional factors that may influence the relationship between enteric pathogens and plant disease. In this study, we investigated the role of humidity and the extent of plant disease progression on S. enterica colonization of plants. We found that high humidity was necessary for the replication of S. enterica on diseased lettuce, but not required for S. enterica ingress into the UV-protected apoplast. Additionally, the Xanthomonas hortorum pv. vitians (hereafter, X. vitians)-infected lettuce host was found to be a relatively hostile environment for S. enterica when it arrived prior to the development of watersoaking or following necrosis onset, supporting the existence of an ideal window during X. vitians infection progress that maximizes S. enterica survival. In vitro growth studies in sucrose media suggest that X. vitians may allow S. enterica to benefit from cross-feeding during plant infection. Overall, this study emphasizes the role of phytobacterial disease as a driver of S. enterica success in the phyllosphere, demonstrates how the time of arrival during disease progress can influence S. enterica's fate in the apoplast, and highlights the potential for humidity to transform an infected apoplast into a growth-promoting environment for bacterial colonizers. IMPORTANCE: Bacterial leaf spot of lettuce caused by Xanthomonas hortorum pv. vitians is a common threat to leafy green production. The global impact caused by phytopathogens, including X. vitians, is likely to increase with climate change. We found that even under a scenario where increased humidity did not enhance plant disease, high humidity had a substantial effect on facilitating Salmonella enterica growth on Xanthomonas-infected plants. High humidity climates may directly contribute to the survival of human enteric pathogens in crop fields or indirectly affect bacterial survival via changes to the phyllosphere brought on by phytopathogen disease.

Identification of pathways required for Salmonella to colonize alfalfa using TraDIS-Xpress.

Enteropathogenic bacteria, such as Salmonella, have been linked to numerous fresh produce outbreaks, posing a significant public health threat. The ability of Salmonella to persist on fresh produce for extended periods is partly attributed to its capacity to form biofilms, which pose a challenge to food decontamination and can increase pathogenic bacterial load in the food chain. Preventing Salmonella colonization of food products and food processing environments is crucial for reducing the incidence of foodborne outbreaks. Understanding the mechanisms of establishment on fresh produce will inform the development of decontamination approaches. We used Transposon-Directed Insertion site Sequencing (TraDIS-Xpress) to investigate the mechanisms used by Salmonella enterica serovar Typhimurium to colonize and establish on fresh produce over time. We established an alfalfa colonization model and compared the findings to those obtained from glass surfaces. Our research identified distinct mechanisms required for Salmonella establishment on alfalfa compared with glass surfaces over time. These include the type III secretion system (sirC), Fe-S cluster assembly (iscA), curcumin degradation (curA), and copper tolerance (cueR). Shared pathways across surfaces included NADH hydrogenase synthesis (nuoA and nuoB), fimbrial regulation (fimA and fimZ), stress response (rpoS), LPS O-antigen synthesis (rfbJ), iron acquisition (ybaN), and ethanolamine utilization (eutT and eutQ). Notably, flagellum biosynthesis differentially impacted the colonization of biotic and abiotic environments over time. Understanding the genetic underpinnings of Salmonella establishment on both biotic and abiotic surfaces over time offers valuable insights that can inform the development of targeted antibacterial therapeutics, ultimately enhancing food safety throughout the food processing chain. IMPORTANCE: Salmonella is the second most costly foodborne illness in the United Kingdom, accounting for £0.2 billion annually, with numerous outbreaks linked to fresh produce, such as leafy greens, cucumbers, tomatoes, and alfalfa sprouts. The ability of Salmonella to colonize and establish itself in fresh produce poses a significant challenge, hindering decontamination efforts and increasing the risk of illness. Understanding the key mechanisms of Salmonella to colonize plants over time is key to finding new ways to prevent and control contamination of fresh produce. This study identified genes and pathways important for Salmonella colonization of alfalfa and compared those with colonization of glass using a genome-wide screen. Genes with roles in flagellum biosynthesis, lipopolysaccharide production, and stringent response regulation varied in their significance between plants and glass. This work deepens our understanding of the requirements for plant colonization by Salmonella, revealing how gene essentiality changes over time and in different environments. This knowledge is key to developing effective strategies to reduce the risk of foodborne disease.

The OsZIP2 transporter is involved in root-to-shoot translocation and intervascular transfer of cadmium in rice.

Cadmium (Cd) is a toxic metal that poses serious threats to human health. Rice is a major source of dietary Cd but how rice plants transport Cd to the grain is not fully understood. Here, we characterize the function of the ZIP (ZRT, IRT-like protein) family protein, OsZIP2, in the root-to-shoot translocation of Cd and intervascular transfer of Cd in nodes. OsZIP2 is localized at the plasma membrane and exhibited Cd2+ transport activity when heterologously expressed in yeast. OsZIP2 is strongly expressed in xylem parenchyma cells in roots and in enlarged vascular bundles in nodes. Knockout of OsZIP2 significantly enhanced root-to-shoot translocation of Cd and alleviated the inhibition of root elongation by excess Cd stress; whereas overexpression of OsZIP2 decreased Cd translocation to shoots and resulted in Cd sensitivity. Knockout of OsZIP2 increased Cd allocation to the flag leaf but decreased Cd allocation to the panicle and grain. We further reveal that the variation of OsZIP2 expression level contributes to grain Cd concentration among rice germplasms. Our results demonstrate that OsZIP2 functions in root-to-shoot translocation of Cd in roots and intervascular transfer of Cd in nodes, which can be used for breeding low Cd rice varieties.