Modelling the growth kinetics of Listeria monocytogenes in pasta salads at different storage temperatures and packaging conditions.

The aim of this study was to model Listeria monocytogenes growth kinetics in ready to eat full meal pasta salads, containing fresh and cooked ingredients. With this aim, laboratory prepared salads, representing two formulations of commercial pasta salads, were spiked with L. monocytogenes and tested under categorised packaging and storage temperature conditions. L. monocytogenes enumeration results collected in 15 different laboratory prepared salad datasets were analysed with primary and secondary models. The models showing the best fit to describe L. monocytogenes growth kinetics in the laboratory prepared salads were then validated within commercial pasta salads. Baranyi no-lag was the best primary model fitting datasets collected at 12 °C, whereas the exponential model gave the best results for datasets collected at 4 °C. The maximum microbial specific growth rate (µmax) mean values obtained at 4 and 12 °C for salads packaged under air packaging conditions were 0.008 ±â€¯0.003 and 0.036 ±â€¯0.006 log10 (cfu/g) h-1, respectively. At the same temperatures, the µmax mean values obtained under modified atmosphere were 0.005 ±â€¯0.005 and 0.026 ±â€¯0.005 log10 (cfu/g) h-1, respectively. The Gamma secondary model predicted the growth kinetics of L. monocytogenes at both temperatures and packaging conditions and the µmax at the optimum temperature and the optimum pH for Listeria growth (µopt) estimated by the model corresponded to 0.247 ±â€¯0.009 log10 (cfu/g) h-1. Baranyi model without lag phase was used to generate growth kinetics under different scenarios. In the comparison of the predicted log10 concentrations respect to the observed ones the residues rarely exceeded 1 Log10 cfu/g. The selected models can be applied to describe the growth kinetics of L. monocytogenes in similar types of pasta salads with comparable pH, shelf life and storage conditions.

Antimicrobial Resistance in the Terrestrial Environment of Agricultural Landscapes in Norway.

The abundance and diversity of antimicrobial-resistant bacteria (ARB) and antimicrobial resistance genes (ARGs) in agricultural landscapes may be important for the spread of antimicrobial resistance (AMR) in the environment. The aim of this study was to apply screening methods for ARB and ARGs to investigate the impact of farming on the prevalence of AMR in a country with low antibiotic usage. We have analyzed samples (n = 644) from soil and wild terrestrial animals and plants (slugs, snails, mice, shrews, earthworms, and red clover) collected over two years in agricultural fields accompanied by nearby control areas with low human activity. All samples were investigated for the occurrence of 35 different ARGs using high-throughput quantitative PCR (HT-qPCR) on a newly developed DNA array. In addition, samples from the first year (n = 415) were investigated with a culture-based approach combined with whole-genome sequencing (WGS) to identify antimicrobial-resistant E. coli (AREC). ARGs were detected in 59.5% of all samples (2019 + 2020). AREC, which was only investigated in the 2019 samples, was identified in 1.9% of these. Samples collected in the autumn showed more ARGs and AREC than spring samples, and this was more pronounced for organic fields than for conventional fields. Control areas with low human activity showed lower levels of ARGs and a lack of AREC. The use of livestock manure was correlated with a higher level of ARG load than other farming practices. None of the soil samples contained antibiotics, and no association was found between AMR and the levels of metals or pesticides. High qualitative similarity between HT-qPCR and WGS, together with the positive controls to the validation of our 35 ARG assays, show that the microfluid DNA array may be an efficient screening tool on environmental samples. In conclusion, even in a country with a very low consumption of antimicrobials by production animals, our results support the hypothesis of these animals being a source of AREC and ARGs in agricultural environments, primarily through the use of manure.

Bacteriocin production, antibiotic susceptibility and prevalence of haemolytic and gelatinase activity in faecal lactic acid bacteria isolated from healthy Ethiopian infants.

The objective of this study was to characterise lactic acid bacteria (LAB) isolated from faecal samples of healthy Ethiopian infants, with emphasis on bacteriocin production and antibiotic susceptibility. One hundred fifty LAB were obtained from 28 healthy Ethiopian infants. The isolates belonged to Lactobacillus (81/150), Enterococcus (54/150) and Streptococcus (15/150) genera. Lactobacillus species were more abundant in the breast-fed infants while Enterococcus dominated the mixed-fed population. Bacteriocin-producing LAB species were isolated from eight of the infants. Many different bacteriocins were identified, including one new bacteriocin from Streptococcus salivarius, avicin A (class IIa) from Enterococcus avium, one class IIa bacteriocin from Enterococcus faecalis strains, one unknown bacteriocin from E. faecalis and two unknown bacteriocins from Lactobacillus fermentum strains and the two-peptide gassericin T from Lactobacillus gasseri isolate. Susceptibility tests performed for nine antibiotics suggest that some lactobacilli might have acquired resistance to erythromycin (3 %) and tetracycline (4 %) only. The streptococci were generally antibiotic sensitive except for penicillin, to which they showed intermediate resistance. All enterococci were susceptible to ampicillin while 13 % showed penicillin resistance. Only one E. faecalis isolate was vancomycin-resistant. Tetracycline (51 %) and erythromycin (26 %) resistance was prevalent among the enterococci, but multidrug resistance was confined to E. faecalis (47 %) and Enterococcus faecium (33 %). Screening of enterococcal virulence traits revealed that 2 % were ß-haemolytic. The structural genes of cytolysin were detected in 28 % of the isolates in five enterococcal species, the majority being E. faecalis and Enterococcus raffinosus. This study shows that bacteriocin production and antibiotic resistance is a common trait of faecal LAB of Ethiopian infants while virulence factors occur at low levels.

Microarray-based transcriptome of Listeria monocytogenes adapted to sublethal concentrations of acetic acid, lactic acid, and hydrochloric acid.

Listeria monocytogenes , an important foodborne pathogen, commonly encounters organic acids in food-related environments. The transcriptome of L. monocytogenes L502 was analyzed after adaptation to pH 5 in the presence of acetic acid, lactic acid, or hydrochloric acid (HCl) at 25 °C, representing a condition encountered in mildly acidic ready-to-eat food kept at room temperature. The acid-treated cells were compared with a reference culture with a pH of 6.7 at the time of RNA harvesting. The number of genes and magnitude of transcriptional responses were higher for the organic acids than for HCl. Protein coding genes described for low pH stress, energy transport and metabolism, virulence determinates, and acid tolerance response were commonly regulated in the 3 acid-stressed cultures. Interestingly, the transcriptional levels of histidine and cell wall biosynthetic operons were upregulated, indicating possible universal response against low pH stress in L. monocytogenes. The opuCABCD operon, coding proteins for compatible solutes transport, and the transcriptional regulator sigL were significantly induced in the organic acids, strongly suggesting key roles during organic acid stress. The present study revealed the complex transcriptional responses of L. monocytogenes towards food-related acidulants and opens the roadmap for more specific and in-depth future studies.

Correction: Characteristics of pulmonary multidrug-resistant tuberculosis patients in Tigray Region, Ethiopia: A cross-sectional study.

[This corrects the article DOI: 10.1371/journal.pone.0236362.].

ccrABEnt serine recombinase genes are widely distributed in the Enterococcus faecium and Enterococcus casseliflavus species groups and are expressed in E. faecium.

The presence, distribution and expression of cassette chromosome recombinase (ccr) genes, which are homologous to the staphylococcal ccrAB genes and are designated ccrAB(Ent) genes, were examined in enterococcal isolates (n=421) representing 13 different species. A total of 118 (28 %) isolates were positive for ccrAB(Ent) genes by PCR, and a number of these were confirmed by Southern hybridization with a ccrA(Ent) probe (n=76) and partial DNA sequencing of ccrA(Ent) and ccrB(Ent) genes (n=38). ccrAB(Ent) genes were present in Enterococcus faecium (58/216, 27 %), Enterococcus durans (31/38, 82 %), Enterococcus hirae (27/52, 50 %), Enterococcus casseliflavus (1/4, 25 %) and Enterococcus gallinarum (1/2, 50 %). In the eight other species tested, including Enterococcus faecalis (n=94), ccrAB(Ent) genes were not found. Thirty-eight sequenced ccrAB(Ent) genes from five different enterococcal species showed 94-100 % nucleotide sequence identity and linkage PCRs showed heterogeneity in the ccrAB(Ent) flanking chromosomal genes. Expression analysis of ccrAB(Ent) genes from the E. faecium DO strain showed constitutive expression as a bicistronic mRNA. The ccrAB(Ent) mRNA levels were lower during log phase than stationary phase in relation to total mRNA. Multilocus sequence typing was performed on 39 isolates. ccrAB(Ent) genes were detected in both hospital-related (10/29, 34 %) and non-hospital (4/10, 40 %) strains of E. faecium. Various sequence types were represented by both ccrAB(Ent) positive and negative isolates, suggesting acquisition or loss of ccrAB(Ent) in E. faecium. In summary, ccrAB(Ent) genes, potentially involved in genome plasticity, are expressed in E. faecium and are widely distributed in the E. faecium and E. casseliflavus species groups.

Characteristics of pulmonary multidrug-resistant tuberculosis patients in Tigray Region, Ethiopia: A cross-sectional study.

BACKGROUND: Tuberculosis (TB) is among the top 10 causes of mortality and the first killer among infectious diseases worldwide. One of the factors fuelling the TB epidemic is the global rise of multidrug resistant TB (MDR-TB). The aim of this study was to determine the magnitude and factors associated with MDR-TB in the Tigray Region, Ethiopia. METHOD: This study employed a facility-based cross-sectional study design, which was conducted between July 2018 and August 2019. The inclusion criteria for the study participants were GeneXpert-positive who were not under treatment for TB, PTB patients' ≥15 years of age and who provided written informed consent. A total of 300 participants were enrolled in the study, with a structured questionnaire used to collect data on clinical, sociodemographic and behavioral factors. Sputum samples were collected and processed for acid-fast bacilli staining, culture and drug susceptibility testing. Drug susceptibility testing was performed using a line probe assay. Logistic regression was used to analyze associations between outcome and predictor variables. RESULTS: The overall proportion of MDR-TB was 16.7% (11.6% and 32.7% for new and previously treated patients, respectively). Of the total MDR-TB isolates, 5.3% were pre-XDR-TB. The proportion of MDR-TB/HIV co-infection was 21.1%. A previous history of TB treatment AOR 3.75; 95% CI (0.7-2.24), cigarette smoking AOR 6.09; CI (1.65-2.50) and patients who had an intermittent fever (AOR = 2.54, 95% CI = 1.21-5.4) were strongly associated with MDR-TB development. CONCLUSIONS: The magnitude of MDR-TB observed among new and previously treated patients is very alarming, which calls for an urgent need for intervention. The high proportion of MDR-TB among newly diagnosed cases indicates ongoing transmission, which suggests the need for enhanced TB control program performance to interrupt transmission. The increased proportion of MDR-TB among previously treated cases indicates a need for better patient management to prevent the evolution of drug resistance. Assessing the TB control program performance gaps and an optimal implementation of the WHO recommended priority actions for the management of drug-resistant TB, is imperative to help reduce the current high MDR-TB burden in the study region.

Complex phenotypic and genotypic responses of Listeria monocytogenes strains exposed to the class IIa bacteriocin sakacin P.

Sakacin P is a class IIa bacteriocin that is active against the food-borne pathogen Listeria monocytogenes, and use of this compound as a biopreservative in foods has been suggested. In the present study, we characterized 30 spontaneous sakacin P-resistant mutants of L. monocytogenes obtained after single exposure to sakacin P. The frequency of development of sakacin P resistance for all strains was in the range from 10(-8) to 10(-9). Using the 50% inhibitory concentration (IC(50)) of sakacin P, the strains could be grouped into strains with high levels of resistance (IC(50), > or =10(4) ng ml(-1)) and strains with low levels of resistance (IC(50), <10(4) ng ml(-1)). Resistant strains belonging to the same IC(50) group also had similar physiological and genetic characteristics. Generally, the resistant strains showed substantial variations in many parameters, such as differences in the stability of the acquired resistance to sakacin P, growth fitness, food-related stress tolerance, and biofilm-forming ability. Fourier transform infrared spectroscopy revealed differences between wild-type and resistant strains in polysaccharide, fatty acid, and, protein regions. A mannose-specific phosphotransferase (PTS) operon has been described for class IIa bacteriocin resistance, and the sakacin P-resistant strains displayed both up- and downregulation of the expression of the mptA gene encoding the PTS system. This is the first comprehensive study of the diversity of a large number of spontaneous resistant mutants obtained after one exposure to a class IIa bacteriocin, particularly to sakacin P. The great diversity among the resistant strains exposed to the same stress conditions suggests that there are different resistance mechanisms.

Corrigendum to "The STARTEC Decision Support Tool for Better Tradeoffs between Food Safety, Quality, Nutrition, and Costs in Production of Advanced Ready-to-Eat Foods".

[This corrects the article DOI: 10.1155/2017/6353510.].

The STARTEC Decision Support Tool for Better Tradeoffs between Food Safety, Quality, Nutrition, and Costs in Production of Advanced Ready-to-Eat Foods.

A prototype decision support IT-tool for the food industry was developed in the STARTEC project. Typical processes and decision steps were mapped using real life production scenarios of participating food companies manufacturing complex ready-to-eat foods. Companies looked for a more integrated approach when making food safety decisions that would align with existing HACCP systems. The tool was designed with shelf life assessments and data on safety, quality, and costs, using a pasta salad meal as a case product. The process flow chart was used as starting point, with simulation options at each process step. Key parameters like pH, water activity, costs of ingredients and salaries, and default models for calculations of Listeria monocytogenes, quality scores, and vitamin C, were placed in an interactive database. Customization of the models and settings was possible on the user-interface. The simulation module outputs were provided as detailed curves or categorized as "good"; "sufficient"; or "corrective action needed" based on threshold limit values set by the user. Possible corrective actions were suggested by the system. The tool was tested and approved by end-users based on selected ready-to-eat food products. Compared to other decision support tools, the STARTEC-tool is product-specific and multidisciplinary and includes interpretation and targeted recommendations for end-users.

Global transcriptional analysis of spontaneous sakacin P-resistant mutant strains of Listeria monocytogenes during growth on different sugars.

Subclass IIa bacteriocins have strong antilisterial activity and can control the growth of Listeria monocytogenes in food. However, L. monocytogenes may develop resistance towards such bacteriocins. In this follow-up study, the transcriptomes of a high level (L502-1) and a low level (L502-6) spontaneous sakacin P-resistant mutant strain of L. monocytogenes were compared to the wild-type (L502). The growth of the resistant strains was reduced on mannose but not affected on cellobiose and the transcriptomics was performed during growth on these sugars. The mannose phosphotransferase system (PTS) encoded by the mptACD operon (mpt) is known for transporting mannose and also act as a receptor to class IIa bacteriocins. The mpt was repressed in L502-1 and this is in accordance with abolition of the bacteriocin receptor with resistance to class IIa bacteriocins. In contrast, the mpt was induced in L502-6. Despite the induction of the mpt, L502-6 showed 1,000 times more resistance phenotype and reduced growth on mannose suggesting the mannose-PTS may not be functional in L502-6. The microarray data suggests the presence of other transcriptional responses that may be linked to the sakacin P resistance phenotype particularly in L502-6. Most of commonly regulated genes encode proteins involved in transport and energy metabolism. The resistant strains displayed shift in general carbon catabolite control possibly mediated by the mpt. Our data suggest that the resistant strains may have a reduced virulence potential. Growth sugar- and mutant-specific responses were also revealed. The two resistant strains also displayed difference in stability of the sakacin P resistance phenotype, growth in the presence of both the lytic bacteriophage P100 and activated charcoal. Taken together, the present study showed that a single time exposure to the class IIa bacteriocin sakacin P may elicit contrasting phenotypic and transcriptome responses in L. monocytogenes possibly through regulation of the mpt.