Listeria monocytogenes is an opportunistic pathogen that is widely distributed in the environment. Here we show the prevalence and transmission of L. monocytogenes in dairy farms in the Cantabria region, on the northern coast of Spain. A total of 424 samples was collected from 14 dairy farms (5 organic and 9 conventional) and 211 L. monocytogenes isolates were recovered following conventional microbiological methods. There were no statistically significant differences in antimicrobial resistance ratios between organic and conventional farms. A clonal relationship among the isolates was assessed by pulsed field gel electrophoresis (PFGE) analysis and 64 different pulsotypes were obtained. Most isolates (89%, n = 187) were classified as PCR serogroup IVb by using a multiplex PCR assay. In this case, 45 isolates of PCR serogroup IVb were whole genome-sequenced to perform a further analysis at genomic level. In silico MLST analysis showed the presence of 12 sequence types (ST), of which ST1, ST54 and ST666 were the most common. Our data indicate that the environment of cattle farms retains a high incidence of L. monocytogenes, including subtypes involved in human listeriosis reports and outbreaks. This pathogen is shed in the feces and could easily colonize dairy products, as a result of fecal contamination. Effective herd and manure management are needed in order to prevent possible outbreaks.

The aim of this study was the examination of the genetic diversity and antimicrobial susceptibility of C. jejuni isolates obtained from dairy farms situated in Cantabria. The presence of Campylobacter jejuni was scrutinized in dairy farms situated in the Cantabria region (Atlantic coast, North of Spain). A total of 520 samples were collected from 12 dairy farms and 62 C. jejuni isolates were achieved. Sixty-one (61) of the isolates proceeded from fresh feces and only one from the stable (soil). Characterization of the isolates was conducted by Pulsed Field Gel Electrophoresis (PFGE) analysis. Antimicrobial susceptibility testing was carried out by standardized disk diffusion test. The PFGE analysis showed a high genetic diversity. From the 62 C. jejuni isolates, 27 different PFGE types were obtained with 70% similarity. The results of the antimicrobial susceptibility tests showed that 21 out of 27 strains were resistant to ciprofloxacin (78%), and 15 of them were also resistant to tetracycline (55%), whereas none of the 27 strains analyzed were resistant to erythromycin. C. jejuni was capable of surviving in livestock waste for at least 20–25 days, whereas the maximum detectable survival time on crops was of six days. This study reveals the high genetic diversity and ciprofloxacin resistance of C. jejuni in dairy cattle farms in Northern Spain, a fact that highlights the urgent need for the surveillance and control of this foodborne pathogen.

Listeria monocytogenes causes listeriosis, a relatively rare but severe disease associated with high mortality rates, particularly among in high-risk populations. To reduce its presence in food production environments and increase the microbiological safety of food by identifying and developing new compounds that inhibit L. monocytogenes biofilm formation, this work joins the methodology of high-throughput screening (HTS) with the search for innovative applications of microbial natural products. Experimental conditions of an in vitro HTS assay have been set up to detect microbial extracts with antibiofilm-forming activity against a reference strain of L. monocytogenes (EGD-e, ATCC BAA-679) susceptible to disinfectants. Activity has been assessed by measuring fluorescence using resazurin (a redox indicator that allows quantifying the cell viability in biofilms). So far, 1520 microbial extracts have been tested, 127 of which have been selected based on their activity profile against the EGD-e strain. Subsequently, these extracts have been tested against a panel of five strains of L. monocytogenes resistant to disinfectants and persistently isolated from the meat industry, and their chromatographic profiles have been examined to prioritize their further study according to their activity and the presence of already known molecules. Twelve extracts have shown broad inhibition biofilm formation against the five resistant strains and three of them are candidates for further bioactivity-guided fractionation and structural elucidation. Further work is underway to characterize the chemical composition of these three extracts and explore their potential applications, with the aim of developing natural biocide formulations that may have a reduced environmental impact.

Listeria monocytogenes is one of the main foodborne pathogens in terms of mortality. To reduce its presence in food production environments and increase the microbiological safety of food by identifying and developing new biocides, this work joins the methodology of HTS (High Throughput Screening) with the search for innovative applications of microbial natural products. Experimental conditions of an in vitro HTS assay have been set up to detect microbial extracts with antibiofilm activity against a reference strain of L. monocytogenes (EGD-e, ATCC BAA-679) susceptible to disinfectants. Activity has been assessed by measuring fluorescence using resazurin (a redox indicator that allows quantifying the viability of the cells present in the biofilms). So far, 1520 microbial extracts have been tested, 127 of which have been selected based on their activity profile against the EGD-e strain. Subsequently, these extracts have been tested against a panel of five strains of L. monocytogenes resistant to disinfectants and persistently isolated from the meat industry, and their chromatographic profiles have been examined to prioritize their further study according to their activity and the presence of already known molecules. A total of 12 extracts has shown broad antibiofilm activity against the five resistant strains and three of them contain potentially active unknown molecules. Further work is underway to characterize the chemical composition and explore potential applications of these three extracts with the aim of developing natural biocide formulations with reduced environmental impact.