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FEMS Microbiology Letters Nov 2019The aims of this study were to evaluate the occurrence of Listeria monocytogenes and Salmonella spp. in sliced cheese and ham from retail markets in southern Brazil, as...
The aims of this study were to evaluate the occurrence of Listeria monocytogenes and Salmonella spp. in sliced cheese and ham from retail markets in southern Brazil, as well as to perform molecular characterization and to assess the antimicrobial resistance profile of the isolates. Samples (n = 160) of sliced cheese and ham were collected at retail level from the city of Pelotas, Brazil. The isolation of L. monocytogenes and Salmonella spp. was performed and the isolates were confirmed by PCR, submitted to antimicrobial susceptibility testing and pulsed-field gel electrophoresis (PFGE). Listeria monocytogenes was found in 9.4% (15/160) of the samples. All L. monocytogenes isolates were positive for the prs, inlA, inlC and inlJ genes. Salmonella spp. was not isolated. Regarding the antimicrobial susceptibility, one (6.6%) L. monocytogenes isolate was resistant to streptomycin and four (26.6%) to clindamycin. Macrorestriction analysis with ApaI and AscI enzymes yielded two major PFGE groups I and II. All L. monocytogenes isolates showed virulence genes, and some of them were resistant to clinically used antimicrobials, representing a risk to public health. Moreover, PFGE patterns with high similarity were visualized in L. monocytogenes isolates at different times, demonstrating adaptability of the pathogen at retail level in the region.
Topics: Anti-Bacterial Agents; Brazil; Cheese; Cities; Drug Resistance, Bacterial; Electrophoresis, Gel, Pulsed-Field; Genes, Bacterial; Genotype; Genotyping Techniques; Listeria monocytogenes; Meat Products; Microbial Sensitivity Tests; Phenotype; Polymorphism, Restriction Fragment Length; Salmonella; Virulence Factors
PubMed: 31834356
DOI: 10.1093/femsle/fnz249 -
Food Microbiology Sep 2021Food business operators are responsible for food safety and assessment of shelf lives for their ready-to-eat products. For assisting them, a customized software based on...
Food business operators are responsible for food safety and assessment of shelf lives for their ready-to-eat products. For assisting them, a customized software based on predictive models, ListWare, is being developed. The aim of this study was to develop and validate a predictive model for the growth of Listeria monocytogenes in sliced roast beef. A challenge study was performed comprising 51 different combinations of variables. The growth curves followed the Baranyi and Roberts model with no clear lag phase and specific growth rates in the range <0.005-0.110 hr. A linear regression model was developed based on 528 observations and had an adjusted R-square of 0.80. The significant predictors were storage temperature, sodium lactate, interactions between sodium acetate and temperature, and MAP packaging and temperature. The model was validated in four laboratories in three countries. For conditions where the model predicted up to + log 2 cfu/g Listeria concentration, the observed concentrations were true or below the predicted concentration in 90% of the cases. For the remaining 10%, the roast beef was coated with spices and therefore different from the others. The model will be implemented in ListWare web-application for calculation of "Listeria shelf life".
Topics: Animals; Cattle; Fast Foods; Food Contamination; Food Safety; Food Storage; Kinetics; Listeria monocytogenes; Meat Products; Models, Biological; Regression Analysis; Temperature
PubMed: 33875206
DOI: 10.1016/j.fm.2021.103770 -
Applied and Environmental Microbiology Apr 2021is a deadly intracellular pathogen mostly associated with consumption of ready-to-eat foods. This study investigated the effectiveness of total beef fat (BF-T) from...
is a deadly intracellular pathogen mostly associated with consumption of ready-to-eat foods. This study investigated the effectiveness of total beef fat (BF-T) from flaxseed-fed cattle and its fractions enriched with monounsaturated fatty acids (BF-MUFA) and polyunsaturated fatty acids (BF-PUFA), along with commercially available long-chain fatty acids (LC-FA), as natural antimicrobials against BF-T was ineffective at concentrations up to 6 mg/ml, while was susceptible to BF-MUFA and BF-PUFA, with MICs at pH 7 of 0.33 ± 0.21 mg/ml and 0.06 ± 0.03 mg/ml, respectively. The MIC of C14:0 was significantly lower than those of C16:0 and C18:0 (0.05). Fatty acids 9-C16:1, C18:2n-6, and C18:3n-3 showed stronger inhibitory activity than 9-C18:1 and conjugated C18:2, with MICs of <1 mg/ml. Furthermore, global transcriptional analysis by transcriptome sequencing (RNA-seq) was performed to characterize the response of to selected fatty acids. Functional analysis indicated that antimicrobial LC-UFA repressed the expression of genes associated with nutrient transmembrane transport, energy generation, and oxidative stress resistance. On the other hand, upregulation of ribosome assembly and translation process is possibly associated with adaptive and repair mechanisms activated in response to LC-UFA. Virulence genes and genes involved in bile, acid, and osmotic stresses were largely downregulated, and more so for 9-C16:1, C18:2n-6, and C18:3n-3, likely through interaction with the master virulence regulator PrfA and the alternative sigma factor σ is a bacterial pathogen known for its ability to survive and thrive under adverse environments and, as such, its control poses a significant challenge, especially with the trend of minimally processed and ready-to-eat foods. This work investigated the effectiveness of fatty acids from various sources as natural antimicrobials against and evaluated their potential role in pathogenicity modulation, using the strain ATCC 19111. The findings show that long-chain unsaturated fatty acids (LC-UFA), including unsaturated beef fat fractions from flaxseed-fed cattle, could have the potential to be used as effective antimicrobials for through controlling growth as well as virulence attenuation. This not only advances our understanding of the mode of action of LC-UFA against but also suggests the potential for use of beef fat or its fractions as natural antimicrobials for controlling foodborne pathogens.
Topics: Animals; Anti-Bacterial Agents; Cattle; Fats; Fatty Acids; Gene Expression Profiling; Gene Expression Regulation, Bacterial; Listeria monocytogenes; Red Meat
PubMed: 33608290
DOI: 10.1128/AEM.03027-20 -
Journal of Food Protection Apr 2020d-Tryptophan (d-Trp) has a significant inhibitory effect on growth of gram-negative bacteria under osmotic stress. However, the inhibitory effect of d-Trp on the...
ABSTRACT
d-Tryptophan (d-Trp) has a significant inhibitory effect on growth of gram-negative bacteria under osmotic stress. However, the inhibitory effect of d-Trp on the gram-positive Listeria monocytogenes under chilled and thermal stresses has not been evaluated previously. The effect of d-Trp on L. monocytogenes growth under cold and/or heat stress in milk and cream was dependent on the magnitude of the temperature stress. Low temperatures (4, 7, and 10°C) and treatment with 40 mM d-Trp resulted in significant inhibition of L. monocytogenes growth during the 4-week storage period. Lower temperatures more effectively inhibited growth. When added before thermal processing, 40 mM d-Trp completely inactivated L. monocytogenes (>6-log reduction) heated at 60°C for 25 min or 65°C for 20 min. These results suggest that d-Trp can be used as a preservative for controlling the growth of L. monocytogenes in milk and cream at refrigeration temperatures and could be used to enhance the thermal inactivation of L. monocytogenes.
Topics: Animals; Colony Count, Microbial; Food Microbiology; Listeria monocytogenes; Milk; Temperature; Tryptophan
PubMed: 32221568
DOI: 10.4315/0362-028X.JFP-19-414 -
International Journal of Food... Jun 2021Listeria monocytogenes is a foodborne human pathogen that causes systemic infection, fetal-placental infection in pregnant women causing abortion and stillbirth and...
Listeria monocytogenes is a foodborne human pathogen that causes systemic infection, fetal-placental infection in pregnant women causing abortion and stillbirth and meningoencephalitis in elderly and immunocompromised individuals. This study aimed to analyse L. monocytogenes from different sources from New Zealand (NZ) and to compare them with international strains. We used pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (MLST) and whole-genome single nucleotide polymorphisms (SNP) to study the population structure of the NZ L. monocytogenes isolates and their relationship with the international strains. The NZ isolates formed unique clusters in PFGE, MLST and whole-genome SNP comparisons compared to the international isolates for which data were available. PFGE identified 31 AscI and 29 ApaI PFGE patterns with indistinguishable pulsotypes being present in seafood, horticultural products and environmental samples. Apart from the Asc0002:Apa0002 pulsotype which was distributed across different sources, other pulsotypes were site or factory associated. Whole-genome analysis of 200 randomly selected L. monocytogenes isolates revealed that lineage II dominated the NZ L. monocytogenes populations. MLST comparison of international and NZ isolates with lineage II accounted for 89% (177 of 200) of the total L. monocytogenes population, while the international representation was 45.3% (1674 of 3473). Rarefaction analysis showed that sequence type richness was greater in NZ isolates compared to international trend, however, it should be noted that NZ isolates predominantly came from seafood, horticulture and their respective processing environments or factories, unlike international isolates where there was a good mixture of clinical, food and environmental isolates.
Topics: Environmental Microbiology; Food Handling; Food Microbiology; Genetic Variation; Genome, Bacterial; Horticulture; Humans; Listeria monocytogenes; New Zealand; Seafood
PubMed: 33838478
DOI: 10.1016/j.ijfoodmicro.2021.109166 -
BMC Infectious Diseases Jun 2021Listeria monocytogenes (LM) has come to be a major public health issue of at-risk groups, causing high morbidity and mortality. Despite this data, studies are very...
BACKGROUND
Listeria monocytogenes (LM) has come to be a major public health issue of at-risk groups, causing high morbidity and mortality. Despite this data, studies are very limited in developing countries like Ethiopia. Thus, we aimed to isolate and characterize LM in terms of antibiogram and biofilm formation among pregnant women with fever, women with a history of spontaneous abortion, women with a history of fetal loss, and women with preterm delivery at Jimma University Medical Center (JUMC), southwest Ethiopia.
METHODS
A cross-sectional study was done among 144 women from June to August 2019. Isolates were tested for antibiotic susceptibility and biofilm formation using disc diffusion and microtiter plate method, respectively. Data were collected using a structured questionnaire, entered into Epidata 3.1 and logistic regression was done by SPSS v25.0.
RESULTS
LM was isolated in 8 (5.56%) of 144 screened women. The isolation rate of LM was relatively higher among women with a history of fetal loss (9.7%), followed by women with preterm delivery (6.25%). One of the six cord blood was positive for LM, indicating that the transplacental transmission rate at JUMC was 16.7%. More than 2% of women with an ongoing pregnancy were found to have LM septicemia, which could hurt their fetus. All of the isolates tested were susceptible to Ampicillin. However, all of the isolates were resistant to Penicillin and Meropenem and were biofilm producers.
CONCLUSIONS
The high magnitude of pregnancy-related listeriosis in the current study setting appears that implementation of educational programs targeting risk reduction and more studies to identify sources of LM are warranted. The choice of antibiotics should be after susceptibility testing.
Topics: Academic Medical Centers; Anti-Bacterial Agents; Cross-Sectional Studies; Ethiopia; Female; Humans; Listeria monocytogenes; Listeriosis; Microbial Sensitivity Tests; Pregnancy; Pregnancy Complications, Infectious
PubMed: 34118865
DOI: 10.1186/s12879-021-06254-w -
Molecular Microbiology Mar 2020The universe of Molecular Microbial Pathogenesis is filled with many female and male stars. But there are two particularly bright shining supernovae-like stars: the late...
The universe of Molecular Microbial Pathogenesis is filled with many female and male stars. But there are two particularly bright shining supernovae-like stars: the late Stanley Falkow and the very lively and creative Pascale Cossart. These two outstanding luminaries, surrounded by numerous planets, do not only belong to different scientific generations but their splendor also comes from very different scientific concepts. Stanley Falkow, often referred to as the 'Father of Molecular Microbial Pathogenesis', made many groundbreaking contributions to this field by addressing almost all important bacterial pathogens. Pascale Cossart, who could be called in analogy the 'Queen of Modern Molecular Microbial Pathogenesis' by combining the Microbiology and Cell Biology, concentrates in her similarly impressive scientific work essentially on a single bacterial species which she studied and still studies in great depth: the facultative intracellular bacterial pathogen Listeria monocytogenes-and the vast majority of her most prominent publications deals with this pathogen in almost all facets. It is certainly not an exaggeration to say that she together with her co-workers and collaborators developed this model bacterium into a paradigm among the intracellular bacterial pathogens.
Topics: Female; History, 20th Century; History, 21st Century; Host-Pathogen Interactions; Humans; Listeria monocytogenes; Listeriosis; Virulence; Virulence Factors
PubMed: 32185837
DOI: 10.1111/mmi.14450 -
Journal of Bacteriology Jan 2021Lysozyme is an important component of the innate immune system. It functions by hydrolyzing the peptidoglycan (PG) layer of bacteria. The human pathogen is...
Lysozyme is an important component of the innate immune system. It functions by hydrolyzing the peptidoglycan (PG) layer of bacteria. The human pathogen is intrinsically lysozyme resistant. The peptidoglycan -deacetylase PgdA and -acetyltransferase OatA are two known factors contributing to its lysozyme resistance. Furthermore, it was shown that the absence of components of an ABC transporter, referred to here as EslABC, leads to reduced lysozyme resistance. How its activity is linked to lysozyme resistance is still unknown. To investigate this further, a strain with a deletion in , coding for a membrane component of the ABC transporter, was constructed in strain 10403S. The mutant showed a 40-fold reduction in the MIC to lysozyme. Analysis of the PG structure revealed that the mutant produced PG with reduced levels of -acetylation. Using growth and autolysis assays, we showed that the absence of EslB manifests in a growth defect in media containing high concentrations of sugars and increased endogenous cell lysis. A thinner PG layer produced by the mutant under these growth conditions might explain these phenotypes. Furthermore, the mutant had a noticeable cell division defect and formed elongated cells. Microscopy analysis revealed that an early cell division protein still localized in the mutant, indicating that a downstream process is perturbed. Based on our results, we hypothesize that EslB affects the biosynthesis and modification of the cell wall in and is thus important for the maintenance of cell wall integrity. The ABC transporter EslABC is associated with the intrinsic lysozyme resistance of However, the exact role of the transporter in this process and in the physiology of is unknown. Using different assays to characterize an deletion strain, we found that the absence of EslB affects not only lysozyme resistance but also endogenous cell lysis, cell wall biosynthesis, cell division, and the ability of the bacterium to grow in media containing high concentrations of sugars. Our results indicate that EslB is, by means of a yet-unknown mechanism, an important determinant for cell wall integrity in .
Topics: Bacterial Proteins; Cell Wall; Gene Deletion; Gene Expression Regulation; Listeria monocytogenes; Muramidase; Peptidoglycan; Virulence
PubMed: 33229460
DOI: 10.1128/JB.00553-20 -
BMC Microbiology Nov 2019High temperature requirement A (HtrA) is a widely expressed chaperone and serine protease in bacteria. HtrA proteases assemble and hydrolyze misfolded proteins to...
BACKGROUND
High temperature requirement A (HtrA) is a widely expressed chaperone and serine protease in bacteria. HtrA proteases assemble and hydrolyze misfolded proteins to enhance bacterial survival under stress conditions. Listeria monocytogenes (L. monocytogenes) is a foodborne pathogen that induces listeriosis in humans. In previous studies, it was shown that deletion of htrA in the genome of L. monocytogenes increased the susceptibility to cellular stress and attenuated virulence. However, expression and protease activity of listerial HtrA (LmHtrA) were never analyzed in detail.
RESULTS
In this study, we cloned LmHtrA wildtype (LmHtrA) and generated a proteolytic inactive LmHtrA mutant. Recombinant LmHtrA and LmHtrA were purified and the proteolytic activity was analyzed in casein zymography and in vitro cleavage assays. LmHtrA activity could be efficiently blocked by a small molecule inhibitor targeting bacterial HtrA proteases. The expression of LmHtrA was enhanced in the stationary growth phase of L. monocytogenes and significantly contributed to bacterial survival at high temperatures.
CONCLUSIONS
Our data show that LmHtrA is a highly active caseinolytic protease and provide a deeper insight into the function and mechanism, which could lead to medical and biotechnological applications in the future.
Topics: Bacterial Proteins; Caseins; Food Microbiology; Gene Expression Regulation, Bacterial; Heat-Shock Proteins; Heat-Shock Response; Listeria monocytogenes; Microbial Viability; Protein Folding; Protein Multimerization; Proteolysis; Up-Regulation
PubMed: 31726993
DOI: 10.1186/s12866-019-1633-1 -
Toxins Jun 2020is among the best-characterized intracellular pathogens. Its virulence factors, and the way they interfere with host cells to hijack host functions and promote the...
is among the best-characterized intracellular pathogens. Its virulence factors, and the way they interfere with host cells to hijack host functions and promote the establishment and dissemination of the infection, have been the focus of multiple studies over the last 30 years. During cellular infection, was shown to induce host DNA damage and delay the host cell cycle to its own benefit. However, whether the cell cycle stage would interfere with the capacity of to infect human cultured cell lines was never assessed. We found here that preferentially infects cultured cells in G2/M phases. Inside G2/M cells, the bacteria lead to an increase in the overall mitosis duration by delaying the mitotic exit. We showed that infection causes a sustained activation of the spindle assembly checkpoint, which we correlated with the increase in the percentage of misaligned chromosomes detected in infected cells. Moreover, we demonstrated that chromosome misalignment in -infected cells required the function of two virulence factors, ActA and InlC. Our findings show the pleiotropic role of virulence factors and their cooperative action in successfully establishing the cellular infection.
Topics: Bacterial Proteins; Caco-2 Cells; Chromosome Segregation; G2 Phase Cell Cycle Checkpoints; Host-Pathogen Interactions; Humans; Listeria monocytogenes; Listeriosis; M Phase Cell Cycle Checkpoints; Membrane Proteins; Mitosis; Virulence; Virulence Factors
PubMed: 32575670
DOI: 10.3390/toxins12060411