Approaches to empower the implementation of new tools to detect and prevent foodborne pathogens in food processing.

Foodborne pathogens cause an important public health burden, which is estimated in 600 million cases and more than 400,000 deaths, globally every year. The most susceptible populations, such as children under the age of five, the elderly and immunocompromised, account for the majority of the deaths. Food safety incidents, outbreaks, sporadic cases, and recalls have recognized economic impact, estimated at 7 billion every year in the US. Food safety has become a priority, and the implementation of preventive controls and monitoring systems has raised the development of new tools to detect and prevent pathogens in the food chain. Detection tools have evolved quickly, from rapid testing methods to application of genomics and metagenomics. Importantly, to reduce food safety hazards at food processing, the food chain needs to be seen from farm to fork. This review summarized the main findings discussed during the 2016 OECD-sponsored symposium on food safety. These include i) trends in food safety that embrace the need to implement new tools in detection and prevention, ii) the very rapid evolution of technologies to detect foodborne pathogens, iii) holistic approaches to prevent pathogens require a whole chain approach, and iv) key pillars to facilitate global implementations of new tools in food safety.

Short communication: Characterization of Salmonella phages from dairy calves on farms with history of diarrhea.

Salmonella enterica can cause disease and mortality in calves. This pathogen is also a zoonosis that can be transmitted by animal contact or by food. The prevalence of Salmonella in dairy farms has been reported to range from 0 to 64%, and, due to the diversity of Salmonella serovars that can be circulating, Salmonella is an important concern for dairy production. Bacteriophages that infect Salmonella have been documented to be abundant and widely distributed in the dairy environment. The current study investigated the diversity of Salmonella serovars and Salmonella phages in 8 dairy farms with a history of diarrhea in southern Chile. A total of 160 samples from sick calves, healthy calves, and the environment were analyzed for Salmonella and phage. Isolated phages were characterized and classified by their host range using a panel of 26 Salmonella isolates representing 23 serovars. Host ranges were classified according to lysis profiles (LP) and their spatial distribution was mapped. Salmonella-infecting phages were identified, but none of the 160 samples were positive for Salmonella. A total of 45 phage isolates were obtained from sick calves (11), healthy calves (16), or the environment (18). According to their host range, 19 LP were identified, with LP1 being the most common on all 8 farms; LP1 represents phages that only lyse serogroup D Salmonella. The identification of Salmonella phages but not Salmonella in the same samples could suggest that these phages are controlling Salmonella in these farms.

Expression of Shigella flexneri gluQ-rs gene is linked to dksA and controlled by a transcriptional terminator.

BACKGROUND: Glutamyl queuosine-tRNA(Asp) synthetase (GluQ-RS) is a paralog of the catalytic domain of glutamyl-tRNA synthetase and catalyzes the formation of glutamyl-queuosine on the wobble position of tRNA(Asp). Here we analyze the transcription of its gene in Shigella flexneri, where it is found downstream of dksA, which encodes a transcriptional regulator involved in stress responses. RESULTS: The genomic organization, dksA-gluQ-rs, is conserved in more than 40 bacterial species. RT-PCR assays show co-transcription of both genes without a significant change in transcript levels during growth of S. flexneri. However, mRNA levels of the intergenic region changed during growth, increasing at stationary phase, indicating an additional level of control over the expression of gluQ-rs gene. Transcriptional fusions with lacZ as a reporter gene only produced ß-galactosidase activity when the constructs included the dksA promoter, indicating that gluQ-rs do not have a separate promoter. Using bioinformatics, we identified a putative transcriptional terminator between dksA and gluQ-rs. Deletion or alteration of the predicted terminator resulted in increased expression of the lacZ reporter compared with cells containing the wild type terminator sequence. Analysis of the phenotype of a gluQ-rs mutant suggested that it may play a role in some stress responses, since growth of the mutant was impaired in the presence of osmolytes. CONCLUSIONS: The results presented here, show that the expression of gluQ-rs depends on the dksA promoter, and strongly suggest the presence and the functionality of a transcriptional terminator regulating its expression. Also, the results indicate a link between glutamyl-queuosine synthesis and stress response in Shigella flexneri.

IL-17A levels increase in the infarcted region of the left ventricle in a rat model of myocardial infarction.

Th17 cells, a recently described subtype of CD4+ effector lymphocytes, have been linked to cell-mediated autoimmune and inflammatory diseases as well as to cardiovascular diseases. However, the participation of IL-17A in myocardial ischemic injury has not been clearly defined. We therefore conducted the present study to evaluate IL-17A and Th17-related cytokine levels in a rat model of myocardial infarction (MI). MI was induced in male Sprague Dawley rats by coronary artery ligation. Controls were sham-operated (Sh) or non-operated (C). Blood and samples from the left ventricle (LV) were collected at weeks 1 and 4 post-MI. At week 1, MI animals exhibited increased IL-6, IL-23 and TGF-ß mRNA levels with no apparent change in IL-17 mRNA or protein levels in whole LV. Only TGF-ß mRNA remained elevated at week 4 post-MI. However, further analysis revealed that IL-17A mRNA and protein levels as well as IL-6 and IL-23 mRNA were indeed increased in the infarcted region, though not in the remote non infarcted region of the LV, except for IL-23 mRNA. The increased expression of IL-17A and Th17-related cytokines in the infarcted region of LV, suggests that this proinflammatory pathway might play a role in early stages of post MI cardiac remodelling.

Genomics of Salmonella contaminating backyard production systems reveals persistence and transmission of genetically related Salmonella on a farm basis.

Animals raised in backyard productive systems (BPS) have been frequently associated with Salmonella outbreaks. Several serovars have caused these events, showing that different BPSs can be contaminated by distinct Salmonella serovars. The aim of this study was to characterize the genomic diversity of Salmonella isolates obtained from BPSs in Central Chile to understand their genomic relatedness. A whole-genome SNP-based phylogenetic analysis of 22 Salmonella isolates from 12 locations revealed that S. Typhimurium isolates clustered based on the BPS that they were originally isolated from, and the same was established for S. Enteritidis isolates. Furthermore, our genomic analysis shows that animals from different species (i.e., a chicken, a duck and a pig) carried genetically related S. Typhimurium strains within the same BPS. Moreover, some of these genetically related isolates were obtained in different years (2013 and 2014), indicating that farm-specific Salmonella can persist in BPSs for multiple years and that interspecies transmission is plausible in this environment. Understanding the dynamics of interspecies transmission of Salmonella serovars within a contaminated BPS is fundamental to the design of mitigation strategies to reduce outbreaks of human Salmonella associated with backyard production systems.

Backyard Farms Represent a Source of Wide Host Range Salmonella Phages That Lysed the Most Common Salmonella Serovars.

The genus Salmonella has more than 2,600 serovars, and this trait is important when considering interventions for Salmonella control. Bacteriophages that are used for biocontrol must have an exclusively lytic cycle and the ability to lyse several Salmonella serovars under a wide range of environmental conditions. Salmonella phages were isolated and characterized from 34 backyard production systems (BPSs) with a history of Salmonella infections. BPSs were visited once, and cloacal or fecal samples were processed for phage isolation. Four hosts, Salmonella serovars Enteritidis, Heidelberg, Infantis, and Typhimurium, were used for phage isolation. The host range of the phages was later characterized with a panel of 23 Salmonella serovars (serovar diversity set) and 31 isolates obtained from the same farms (native set). Genetic relatedness for 10 phages with a wide host range was characterized by restriction fragment length polymorphism, and phages clustered based on the host range. We purified 63 phages, and 36 phage isolates were obtained on Salmonella Enteritidis, 16 on Salmonella Heidelberg, and 11 on Salmonella Infantis. Phages were classified in three clusters: (i) phages with a wide host range (cluster I), (ii) phages that lysed the most susceptible Salmonella serovars (serogroup D) and other isolates (cluster II), and (iii) phages that lysed only isolates of serogroup D (cluster III). The most susceptible Salmonella serovars were Enteritidis, Javiana, and Dublin. Seven of 34 farms yielded phages with a wide host range, and these phages had low levels of genetic relatedness. Our study showed an adaptation of the phages in the sampled BPSs to serogroup D Salmonella isolates and indicated that isolation of Salmonella phages with wide host range differs by farm. A better understanding of the factors driving the Salmonella phage host range could be useful when designing risk-based sampling strategies to obtain phages with a wide lytic host range for biocontrol purposes.

Genomic Diversity of Listeria monocytogenes Isolated from Clinical and Non-Clinical Samples in Chile.

Listeria monocytogenes is the causative agent of listeriosis, which is an uncommon but severe infection associated with high mortality rates in humans especially in high-risk groups. This bacterium survives a variety of stress conditions (e.g., high osmolality, low pH), which allows it to colonize different niches especially niches found in food processing environments. Additionally, a considerable heterogeneity in pathogenic potential has been observed in different strains. In this study, 38 isolates of L. monocytogenes collected in Chile from clinical samples (n = 22) and non-clinical samples (n = 16) were analyzed using whole genome sequencing (WGS) to determine their genomic diversity. A core genome Single Nucleotide Polymorphism (SNP) tree using 55 additional L. monocytogenes accessions classified the Chilean isolates in lineages I (n = 25) and II (n = 13). In silico, Multi-locus sequence typing (MLST) differentiated the isolates into 13 sequence types (ST) in which the most common were ST1 (15 isolates) and ST9 (6 isolates) and represented 55% of the isolates. Genomic elements associated with virulence (i.e., LIPI-1, LIPI-3, inlA, inlB, inlC, inlG, inlH, inlD, inlE, inlK, inlF, and inlJ) and stress survival (i.e., stress survival islet 1 and stress survival islet 2) were unevenly distributed among clinical and non-clinical isolates. In addition, one novel inlA premature stop codon (PMSC) was detected. Comparative analysis of L. monocytogenes circulating in Chile revealed the presence of globally distributed sequence types along with differences among the isolates analyzed at a genomic level specifically associated with virulence and stress survival.

[Infectious diseases in the genomic era]. / Infectología en la era de la genómica.

Next generation sequencing (NGS) technologies have arrived, changing research and infectious disease research into a new era, the "genomic era". Currently, the developed world is introducing NGS in a number of applications, including clinical diagnostics, epidemiology, and microbiology. In developing countries NGS is being progressively introduced. Technologies currently available allow to sequence the whole genome of bacterial and viral strains for an approximate cost of $100 USD, which is highly cost savings compared to old-technologies for genome sequencing. Here we review recent publication of whole genome sequencing used for, (i) tracking of foodborne outbreaks, with emphasis in Salmonella and Listeria monocytogenes, (ii) building genomic databases for Governments, (iii) investigating nosocomial infections, and (iv) clinical diagnosis. The genomic era is here to stay and researchers should use these "massive databases" generated by this technology to decrease infectious diseases and thus improve health of humans and animals.

IL-17A levels increase in the infarcted region of the left ventricle in a rat model of myocardial infarction

Th17 cells, a recently described subtype of CD4+ effector lymphocytes, have been linked to cell-mediated autoimmune and inflammatory diseases as well as to cardiovascular diseases. However, the participation of IL-17A in myocardial ischemic injury has not been clearly defined. We therefore conducted the present study to evaluate IL-17A and Th17-related cytokine levels in a rat model of myocardial infarction (MI). MI was induced in male Sprague Dawley rats by coronary artery ligation. Controls were sham-operated (Sh) or non-operated (C). Blood and samples from the left ventricle (LV) were collected at weeks 1 and 4 post-MI. At week 1, MI animals exhibited increased IL-6, IL-23 and TGF-β mRNA levels with no apparent change in IL-17 mRNA or protein levels in whole LV. Only TGF-β mRNA remained elevated at week 4 post-MI. However, further analysis revealed that IL-17A mRNA and protein levels as well as IL-6 and IL-23 mRNA were indeed increased in the infarcted region, though not in the remote non infarcted region of the LV, except for IL-23 mRNA. The increased expression of IL-17A and Th17-related cytokines in the infarcted region of LV, suggests that this proinflammatory pathway might play a role in early stages of post MI cardiac remodelling.