Personal impressions about PROMISE: Ama Szmolka

I was involved in the project working on Task 1.2 for Virulence and Antimicrobial resistance profiling of foodborne pathogens of illegal food imports, lead by Béla Nagy, VMRI, Hungary. Together with my colleagues from the Hungarian team, my major objective was to detect Salmonella from food samples derived from non-Schengen countries confiscated at the Hungarian borders and to isolate and characterize E. coli strains with multiple antimicrobial resistance.

Results from this study can be briefly summarized as follows (Kugler R, Szmolka A, Pászti J, Nagy B, 2014. Integrons and antimicrobial resistance genes of multidrug resistant Escherichia coli and coliform bacteria from foods of animal origin confiscated at the Hungarian borders. Abstract Book, European Symposium on Food Safety, P3-21, 7-9 May, Budapest. Poster):

„Introduction: The import of contaminated food may represent a food safety risk by the spread of pathogenic and/or multidrug resistant (MDR) bacteria and their determinants for virulence and antimicrobial resistance.

Purpose: Here we aimed to isolate and characterize MDR E. coli and coliform bacteria from food samples from non-Schengen countries confiscated at the Hungarian borders. 

Methods: E. coli and coliform colonies were isolated based on their phenotype on Chromocult® Coliform selective media. Furthermore, API®, PCR and 16S rDNS sequencing were used for species identification. Resistance phenotypes were determined by disc diffusion method for 18 antimicrobials with animal and human clinical relevance. Corresponding antimicrobial resistance and virulence gene patterns were identified using PCR microarray systems AMR05 and Ec03 respectively. The gene cassette arrangements of the integrons were defined by amplicon sequencing. 

Results: From the total of 207 confiscated food samples 833 coliform isolates were collected. Among them 17 (13 E. coli and 4 coliforms identified as Enterobacter spp.) showed resistance to at least three different antimicrobial classes thus were designated as MDR. The 17 strains represented 14 different food samples. Resistance genes strA, strB, sul2, blaTEM-1, tet(A) predominantly occurred, but in general the prevalence of the virulence genes was low. The identification of genes qnrB, aac(6’)-Ib, blaOXA-7 in some of the isolates indicated the presence of certain emerging antimicrobial resistance plasmids. Class 1 integrons were found in 10 of the 17 MDR isolates (9 E. coli, 1 coliform), and in the majority of them the sul1 gene was absent from their 3’ conserved segment (CS). Interestingly, in one of the pork samples we detected a non-typical class 1 integron carrying the sul3gene on its 3’CS. 

Significance: Above results showed that these illegal foods may frequently carry MDR E. coli and coliform bacteria with some unusual or new antimicrobial resistance traits.”

Besides I was responsible to perform microarray-based antimicrobial resistance and virulence genotyping of VTEC isolates provided by our WP1 partners, as reported by B.Nagy et al., (IAFP, Budapest, May, 2014). 

During the whole period of the project, I had the opportunity to attend two of the PROMISE Annual Meetings (Dublin, 2013 and Hydra, 2014), and one from the Regional ones, the latter we organized in Keszthely, Hungary. In my oppinion, all of them realized to reach the main goal: to bring together senior and early stage researchers from the field.

In addition to the above events, the program of the PROMISE Summer school (Brno, 2014) allowed us to gain deeper insight into the novel technics for microbe detection and identification and for studying host-pathogen inetractions at different levels.

Last, but not least, as a project member participating in PROMISE Pesearchers Mobility Program I also had the opportunity to collaborate directly, to share ideas and related methodology. As an Early Stage Researcher I was lucky to visit the lab of our VRI parner from Brno, I. Rychlik. In collaboration with his group we revealed some new aspects of the Salmonella-host interactions by discovering  expression of some  novel chicken genes.

Results from this study are close to the publication submission, and can be briefly summarized as follows (Ama Szmolka, Zoltán Wiener, Marta Elsheimer Matulova, Karolina Varmuzova, Ivan Rychlik. Gene expression profiles of chicken embryo fibroblasts in response to Salmonella Enteritidis infection):

The response of chicken to non-typhoidal Salmonella infection is becoming well characterised but the role of particular cell types in this response is still far from being understood. Therefore, in this study we characterised the response of chicken embryo fibroblasts (CEFs) to infection with two different S. Enteritidis strains by microarray analysis. The expression of chicken genes identified as significantly up- or down-regulated (≥3-fold) by microarray analysis was verified by real-time PCR followed by functional classification and interaction prediction of the genes using Gene Ontology and STRING Database. Finally the expression of the newly identified genes was tested in HD11 macrophages and in vivo in chicken caecum and spleen. Altogether 19 genes were induced in CEFs after S. Enteritidis infection. Twelve of them were also induced in HD11 macrophages and thirteen in the caecum of orally infected chickens. The majority of these genes were assigned different functions in the immune response, however five of them (LOC101750351, K123, BU460569, MOBKL2C and G0S2) have not been associated with the response of chicken to Salmonella infection so far. K123 and G0S2 were the only ’non-immune’ genes inducible by S. Enteritidis in fibroblasts, HD11 macrophages and in the caecum after oral infection. The function of K123 is unknown but G0S2 is involved in lipid metabolism and in the β-oxidation of fatty acids in mitochondria. Increased levels of G0S2 might decrease the availability of fatty acids to mitochondria. In non-professional phagocytes such as CEFs, this may lead to the dysfunction of mitochondria, apoptosis of CEFs and release of intracellular Salmonella. In phagocytes, G0S2 might be involved in the control of mitochondrial respiration, resulting in a decreased production of reactive oxygen species as respiration by-products and a lower damage to own tissues.

And finally my closing remark to the organizers: I was really happy to be part of the PROMISE Project !