2For more than 80 years, subtyping of Salmonella enterica has been routinely performed by serotyping, a method in which surface antigens are identified based on agglutination reactions with specific antibodies. The serotyping scheme, which is continuously updated as new serovars are discovered, has generated over time a data set of the utmost significance, allowing long-term epidemiological surveillance of Salmonella in the food chain and in public health control. Conceptually, serotyping provides no information regarding the phyletic relationships inside the different Salmonella enterica subspecies. In epidemiological investigations, identification and tracking of salmonellosis outbreaks require the use of methods that can fingerprint the causative strains at a taxonomic level far more specific than the one achieved by serotyping. During the last 2 decades, alternative methods that could successfully identify the serovar of a given strain by probing its DNA have emerged, and molecular biology-based methods have been made available to address phylogeny and fingerprinting issues. At the same time, accredited diagnostics have become increasingly generalized, imposing stringent methodological requirements in terms of traceability and measurability. In these new contexts, the hand-crafted character of classical serotyping is being challenged, although it is widely accepted that classification into serovars should be maintained. This review summarizes and discusses modern typing methods, with a particular focus on those having potential as alternatives for classical serotyping or for subtyping Salmonella strains at a deeper level.

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The Salmonella genus is currently divided into 2 species, Salmonella bongori and Salmonella enterica, the latter being classified into 6 subspecies, S. enterica subsp. enterica, salamae, arizonae, diarizonae, houtenae, and indica (1). Most salmonellae possess two flagellin genes, fliC and fljB, encoding the so-called phase 1 and phase 2 flagellins, respectively. Only one flagellin type is expressed at a time in a Salmonella cell. Isolates are termed biphasic, monophasic, or nonmotile if they can express both, one, or no flagellar phases, respectively. Expression of the two genes is under the control of a switch mechanism called "phase inversion." The switch mechanism consists of the DNA inversion of a sequence segment, the so-called invertible region, by recombination of the two flanking sites, hixL and hixR, mediated by an invertase encoded by the hin gene included in this segment (2). The promoter of the fljBA operon is born on this invertible region. The fljBA operon encodes the FljB flagellin and FljA, a repressor of the fliC gene. The orientation of the invertible region determines whether the fljBA operon or the fliC gene will be expressed (2).Characterization of Salmonella spp. below the subspecies level is traditionally obtained by serotyping based on the KauffmannWhite-Le Minor scheme (1). Serovars are determined following agglutination of entire bacteria with specific sera to identify variants of the somatic (O) and flagellar (H) antigens. Serotyping results in an antigenic formula, indicating the O, H1, and H2 antigens separated by colons. The different O and H antigens are named by numbers, alphabetical characters, or both. Some O antigens can be present or absent without interfering with serovar identification. These antigens are interesting only as epidemiological markers within a given serovar. These accessory O antigens are underlined when their presence is linked to the integration of a prophage. O or H antigens are written within square brackets if they may be present or absent without relation to phage conversion. The 1,4,[5],12:i:1,2 antigenic formula, corresponding to S. enterica serovar Typhimurium, therefore indicates that (i) the O4 and O12 antigens are always present in this serovar, (ii) the O1 antigen can be present or absent due to phage conversion, (iii) the O5 antigen can be present or absent without relation to phage conversion, and (iv) the H1 antigen "i" and the H2 antigens "1" and "2" are always present. If no H1 or H2 antigen can be detected,

Background Linezolid is a critically important antibiotic used to treat human infections caused by MRSA and VRE. While linezolid is not licensed for food-producing animals, linezolid-resistant (LR) isolates have been reported in European countries, including Belgium. Objectives To: (i) assess LR occurrence in staphylococci and enterococci isolated from different Belgian food-producing animals in 2019 through selective monitoring; and (ii) investigate the genomes and relatedness of these isolates. Methods Faecal samples (n = 1325) and nasal swab samples (n = 148) were analysed with a protocol designed to select LR bacteria, including a 44–48 h incubation period. The presence of LR chromosomal mutations, transferable LR genes and their genetic organizations and other resistance genes, as well as LR isolate relatedness (from this study and the NCBI database) were assessed through WGS. Results The LR rate differed widely between animal host species, with the highest rates occurring in nasal samples from pigs and sows (25.7% and 20.5%, respectively) and faecal samples from veal calves (16.4%). WGS results showed that LR determinants are present in a large diversity of isolates circulating in the agricultural sector, with some isolates closely related to human isolates, posing a human health risk. Conclusions LR dedicated monitoring with WGS analysis could help to better understand the spread of LR. Cross-selection of LR transferable genes through other antibiotic use should be considered in future action plans aimed at combatting antimicrobial resistance and in future objectives for the rational use of antibiotics in a One Health perspective.

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