Quinolone-resistant Salmonella Infantis (n = 64) isolated from human stool samples, food and poultry during the years 2006-2011 were analysed for their resistance phenotypes, macrorestriction patterns and molecular mechanisms of decreased susceptibility to fluoroquinolones. Minimum inhibitory concentrations (MICs) of nalidixic acid (NAL) and ciprofloxacin (CIP) were determined by the agar dilution procedure, and the susceptibility to additional antimicrobial agents was determined by the disc diffusion method. To assess the influence of enhanced efflux activity, MICs were determined in the presence and absence of the inhibitor PAβN. The results of pulsed-field gel electrophoresis (PFGE) typing revealed that quinolone-resistant S. Infantis in Serbia had similar or indistinguishable PFGE profiles, suggesting a clonal spread. All S. Infantis showed combined resistance to NAL and tetracycline, whereas multiple drug resistance to three or more antibiotic classes was rare (2 isolates of human origin). The MICs ranged between 512 and 1024 μg/mL for NAL and 0.125-2 μg/mL for CIP. A single-point mutation in the gene gyrA leading to a Ser83→Tyr exchange was detected in all isolates, and a second exchange (Ser80→Arg) in the gene parC was only present in eight S. Infantis isolates exhibiting slightly higher MICs of CIP (2 μg/mL). The inhibitor PAβN decreased the MIC values of CIP by two dilution steps and of NAL by at minimum 3-6 dilution steps, indicating that enhanced efflux plays an important role in quinolone resistance in these isolates. The plasmid-mediated genes qnr, aac(6')-lb-cr and qepA were not detected by PCR assays.

ABSTRACT. The knowledge about virulence mechanisms, resistance to antimicrobial agents and the biofilm formation ability of Salmonella spp. in poultry industry has been expanded over the years. However, in spite of the research efforts and significant investments to improve management systems in poultry industry, it has become evident that none of the methods applied in all stages of food production chain are 100% effective in eliminating Salmonella spp. Different serovars are manifesting different mechanisms of invasiveness which depend on their ability to invade lower zones of the lamina propria, their ability to gain accesses to parenchymatous organs and survive in macrophages. The ubiquitous nature of Salmonella spp. due to their adaptation to animal and plant hosts, as well as their survival in hostile environments and their enhanced capacity to produce biofilms, contribute to a long lasting contamination of the environment, feed and animals. The emergency and spread of antimicrobial resistances in Salmonella spp. raise additional concerns.

Molecular typing and resistotyping coupled with gyrA single nucleotide polymorphism (SNP) of 60 Salmonella Enteritidis (SE) isolates originated from poultry, food, and humans in Serbia is described. Molecular fingerprinting was performed by randomly amplified polymorphic DNA (RAPD) using four primers, and the diversity index (D) was 0.688. In combination with resistotyping and gyrA SNP, D increased to 0.828. A total of 23 genetic groups were obtained. When four RAPD primers were combined, epidemic isolates from a fast-food restaurant outbreak were clustered in a distinctive genetic group. Among 60 SE strains, three had multiple resistances to three or more antibiotics. Nine strains were resistant to nalidixic acid (NAL; a non-fluorinated quinolone). The mutations in quinolone resistance-determining region (QRDR) found in NAL-resistant strains were attributed to Asp(87) → Asn in six strains, Asp(87) → Gly in one strain, and Ser(83) → Phe in one strain. One NAL-resistant strain had no mutations in QRDR, suggesting another mechanism of resistance.

Avian pathogenic Escherichia coli (APEC) causes colibacillosis within poultry flocks all around the world. There is a number of virulence mechanisms involved in the disease process in poultry and determination of some of the responsible genes is important for diagnosis of colibacillosis. In this work, research data regarding diagnostics of APEC and how certain clonal lineages could cause infection in different hosts is presented. In order to determine virulence genotype of APEC, multiplex polymerase chain reaction, based on a published sequence of seven pairs of primers (iroN, ompT, hlyF, iss, iutA, elitC and cvaC), was used in our laboratory. It was established in the research of other scientists that isolates with two or more of these genes can develop pathogenic phenotype, while isolates with one or none of the genes are mostly commensal E. coli. Additionally, virulence mechanisms in APEC were also briefly described. It was emphasized that resistance genes and virulence genes are sometimes co-located on the same plasmid and that such plasmids could be shared among related or unrelated bacteria species. Since APEC often confers resistance to antibiotics, the therapy is less effective in poultry with multidrug resistant strains. It was concluded that good management practice, treatment with probiotics and/or vaccination are necessary to reduce colibacillosis outbreaks. This approach is even more pronounced since APEC resides in intestine of healthy poultry and could cause disease if poultry is exposed to various stressors.

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Blastocystis is a common protist colonizing the gastrointestinal tract of humans and various animals. The first subtyping of Blastocystis isolates in pigs and humans in Serbia revealed unusual avian‐specific subtype ST6 in humans. In total, 48 pig faecal specimens collected on seven pig farms and 50 human faecal specimens positive to Blastocystis by microscopic examination were selected for the study. Eleven randomly selected PCR‐positive pig samples and 10 samples from human patients (with gastrointestinal complaints) were subjected to SSU rDNA sequencing. Three subtypes were identified (ST3, ST5 and ST6) by phylogenetic analysis. ST5 was found in all pig samples; while in human samples, we detected ST3 and ST6. The latter subtype is relatively uncommon in Europe and highly adapted to avian hosts; therefore, the possibility of sporadic zoonotic transmission to human patients should not be ignored.

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Methicillin-resistant Staphylococcus aureus is one of the most important human pathogens. Th ese bacteria have the ability to colonize animals andcross species barriers. Th ree major groups of MRSA strains have emerged. Healthcare associated MRSA and community acquired MRSA strains have potential to spread worldwide and oft en persist in hospitals and communities as clonal strains. Th e livestock-associated MRSA has been isolated from healthy human carriers but also from infected patients all around the world. Molecular typing of staphylococcal cassette chromosome mec (SCCmec), multilocus sequence typing in combination with pulse fi eld gel electrophoresis and spa typing are most frequently used for genetic characterization of MRSA strains. Th e community-acquired MRSA strains are capable of producing Panton Valentine leukocidin (PVL) cytotoxin which is their major virulence determinant. MRSA strains possess a number of virulence factors that are common in other bacteria and it is still not entirely explained which virulence factors or mechanisms of their regulation are important for the pathogenic potential, persistence in the environment or the ability to cause detrimental infection in patients. Recently, as a contribution to the progress of molecular biology, peculiar mechanisms of genetic regulation of virulence genes have been discovered and their role in pathogenesis of infection and epidemiology of MRSA has been studied.