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Circulating Microbial Cell-Free DNA in Health and Disease
Human blood contains low biomass of circulating microbial cell-free DNA (cfmDNA) that predominantly originates from bacteria. Numerous studies have detected circulating cfmDNA in patients with infectious and non-infectious diseases, and in healthy individuals. Remarkable differences were found in the microbial composition of healthy subjects and patients compared to cohorts with various diseases or even patients with diversified prognoses, implying that these alterations may be associated with disease development. Although the function of circulating cfmDNA needs to be elucidated (whether it acts as a bystander of dysbiosis or a key player in disease development), several studies have demonstrated its potential as a non-invasive biomarker that may improve diagnosis and treatment efficacy. The origin of circulating cfmDNA is still the subject of much deliberation, but studies have identified members of various microbiome niches, including the gut, oral cavity, airways, and skin. Further studies investigating the origin and function of circulating cfmDNA are needed. Moreover, low-biomass microbiome studies are prone to contamination, therefore stringent negative experimental control reactions and decontamination frameworks are advised in order to detect genuine circulating cfmDNA.
Genoserotyping of Listeria monocytogenes strains originating from meat products and meat processing environments
Background. Listeria monocytogenes is a foodborne human pathogen and a causative factor of listeriosis, which is an illness with a high mortality rate. Serotyping is a method for differentiating L. monocytogenes isolates based on unique combinations of somatic (O) and flagellar (H) antigens on the surface of their cells. Standard serotyping involves agglutination methods, which require using antisera. However, there are also genoserotyping methods which allow to categorise L. monocytogenes isolates into particular groups of serotypes (referred to as serogroups) based on genetic analyses. Differentiating L. monocytogenes isolates is an important issue in terms of food safety, surveillance and traceability of contamination sources. In this work, we present results of the genoserotyping of 153 L. monocytogenes isolates originating from meat products and meat processing environments at Polish processing plants. Two protocols were used for genoserotyping analyses: the first one allows to differentiate between four most common serotypes (1/2a, 1/2b, 1/2c and 4b) and the second one allows to distinguish hipervirulent serovar 4h from other serotypes. Results and conclusion. Results achieved using both methods were consistent and all isolates were categorised into corresponding serogroups within the two methodologies. Most of the isolates (73.9 %) were characterised as members of the IIa serogroup (representing the 1/2a, 3a serovars). The IVb (4b, 4d, 4e) serogroup was the second most common (and comprised 18.3 % of isolates), followed by IIb (1/2b, 3b, 7) and IIc (1/2c, 3c), however, the last two groups were equally numerous (and each of them comprised 3.9 % of all isolates). None of the collected isolates belonged to the serogroup representing the 4a, 4c, 4ab and 4h serotypes.
Gene emrC Associated with Resistance to Quaternary Ammonium Compounds Is Common among Listeria monocytogenes from Meat Products and Meat Processing Plants in Poland
(1) Background: L. monocytogenes is a food pathogen of great importance, characterized by a high mortality rate. Quaternary ammonium compounds (QACs), such as benzalkonium chloride (BC), are often used as disinfectants in food processing facilities. The effectiveness of disinfection procedures is crucial to food safety. (2) Methods: A collection of 153 isolates of L. monocytogenes from meat processing industry was analyzed for their sensitivity to BC using the agar diffusion method. Genes of interest were detected with PCR. (3) Results: Genes emrC, bcrABC, and qacH were found in 64 (41.8%), 6 (3.9%), and 1 isolate (0.7%), respectively, and 79 isolates (51.6%) were classified as having reduced sensitivity to BC. A strong correlation between carrying QACs resistance-related genes and phenotype was found (p-value < 0.0001). Among 51 isolates originating from bacon (collected over 13 months), 48 had the emrC gene, which could explain their persistent presence in a processing facility. Isolates with the ilsA gene (from LIPI-3) were significantly (p-value 0.006) less likely to carry QACs resistance-related genes. (4) Conclusions: Reduced sensitivity to QACs is common among L. monocytogenes from the meat processing industry. Persistent presence of these bacteria in a processing facility is presumably caused by emrC-induced QACs resistance.
High Prevalence of Virulence-Associated Genes and Length Polymorphism in actA and inlB Genes Identified in Listeria monocytogenes Isolates from Meat Products and Meat-Processing Environments in Poland
Listeria monocytogenes is a human pathogen that has the ability to cause listeriosis, a disease with possible fatal outcomes. The typical route of infection is ingestion of the bacteria with contaminated food. In this study, 13 virulence-associated genes were examined with PCR in the genomes of 153 L. monocytogenes isolates collected from meat products and processing environments in Poland. All isolates possessed genes from LIPI-1—hly, actA, plcA, plcB and mpl—as well as four internalins: inlA, inlB, inlC, inlJ. Invasion-associated protein iap, as well as genes prfA and sigB, encoding regulatory proteins, were also detected in all isolates. Gene flaA, encoding flagellin, was detected in 113 (74%) isolates. This was the only gene that was not detected in all isolates, as its presence is serotype-dependent. Gene actA showed polymorphism with longer and shorter variants in PCR amplicons. Two isolates were characterized by truncated inlB genes, lacking 141 bp in their sequence, which was confirmed by gene sequencing. All isolates were positive in hemolysis assays, proving the synthesis of functional PrfA and Hly proteins. Four genotypes of L. monocytogenes based on actA polymorphism and two genotypes based on inlB polymorphism were distinguished within the isolates’ collection.