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In this paper, we address the problem of recovering arbitrary-shaped data clusters from datasets while facing \emph{high space constraints}, as this is for instance the case in many real-world applications when analysis algorithms are directly deployed on resources-limited mobile devices collecting the data. We present DBMSTClu a new space-efficient density-based \emph{non-parametric} method working on a Minimum Spanning Tree (MST) recovered from a limited number of linear measurements i.e. a \emph{sketched} version of the dissimilarity graph $\mathcal{G}$ between the $N$ objects to cluster. Unlike $k$-means, $k$-medians or $k$-medoids algorithms, it does not fail at distinguishing clusters with particular forms thanks to the property of the MST for expressing the underlying structure of a graph. No input parameter is needed contrarily to DBSCAN or the Spectral Clustering method. An approximate MST is retrieved by following the dynamic \emph{semi-streaming} model in handling the dissimilarity graph $\mathcal{G}$ as a stream of edge weight updates which is sketched in one pass over the data into a compact structure requiring $O(N \operatorname{polylog}(N))$ space, far better than the theoretical memory cost $O(N^2)$ of $\mathcal{G}$. The recovered approximate MST $\mathcal{T}$ as input, DBMSTClu then successfully detects the right number of nonconvex clusters by performing relevant cuts on $\mathcal{T}$ in a time linear in $N$. We provide theoretical guarantees on the quality of the clustering partition and also demonstrate its advantage over the existing state-of-the-art on several datasets., Proceedings of the 2018 SIAM International Conference on Data Mining

Growth of the food-borne human pathogen Listeria monocytogenes to large numbers in ready-to-eat food products greatly increases the risk of disease for susceptible consumers. A better knowledge of the population structure of L. monocytogenes present in retailed food could allow better prevention strategies to be developed. We present the analysis of 450 L. monocytogenes isolates, 179 responsible for sporadic human cases of listeriosis and 271 isolated from foods collected from retailers. All isolates were investigated by multiplex PCR (food isolates), allowing serovar predictions, or serotyped (human isolates), and DNA macrorestriction patterns were determined. Isolates from different sources were significantly differently distributed into PCR groups. PCR group IIa, corresponding to serovars 1/2a and 3a, was predominant in food isolates (58%; OR = 3.19; P − 7 ). A larger proportion of human isolates belonged to PCR group IVb, corresponding to serovars 4b, 4d and 4e (44%; OR = 5.69; P − 7 ). DNA macrorestriction pattern analysis of PCR group IIa isolates showed that isolates from pork products had a very low diversity (ID = 0.905) whereas isolates from humans were more diverse (ID = 0.976). Furthermore, 78% of the pork product isolates belonging to PCR group IIa exhibited only two Asc I profiles, a 1 and a 2 , which were very similar (94%). DNA array analysis of representative isolates showed that isolates with a 1 and a 2 profiles constitute a homogeneous population, whereas isolates exhibiting non a 1 – a 2 profiles are more diverse. Six of the isolates with a 1 and a 2 profiles were selected and investigated for their gene content using a DNA array. With respect to 295 strains present in our data collection, a specific pattern of the presence and absence of 15 genes was identified. Five are predicted to encode internalins and cell surface proteins, and eight of the genes were missing in this group. They code for cell surface proteins, transcriptional regulators, an acylase, a sugar phosphorylase and proteins of unknown functions. The ability of strains to multiply in different niches may be determined by the presence or absence of these genes.

A DNA macroarray containing 465 intragenic amplicons was designed to identify Staphylococcus aureus at the species level and to type S. aureus isolates. The genes selected included those encoding (i) S. aureus -specific proteins, (ii) staphylococcal and enterococcal proteins mediating antibiotic resistance and factors involved in their expression, (iii) putative virulence proteins and factors controlling their expression, and (iv) proteins produced by mobile elements. The macroarray was hybridized with the cellular DNAs of 80 S. aureus clinical isolates that were previously typed by analyses of their antibiograms and SmaI patterns. The set selected contained unrelated, endemic, and outbreak-related isolates belonging to 45 SmaI genotypes. In a gene content dendrogram, the 80 isolates were distributed into 52 clusters. The outbreak-related isolates were linked in the same or a closely related cluster(s). Clustering based on gene content provided a better discrimination than SmaI pattern analysis for the tested mecA + isolates that were endemic to Europe. All of the antibiotic resistance genes detected could be correlated with their corresponding phenotypes, except for one isolate which carried a mecA gene without being resistant. The 16 isolates responsible for bone infections were distinguishable from the 12 isolates from uninfected nasal carriers by a significantly higher prevalence of the sdrD gene coding for a putative SD (serine-aspartate) adhesin (in 15 and 7 isolates, respectively). In conclusion, the macroarray designed for this study offers an attractive and rapid typing method which has the advantage of providing additional information concerning the gene content of the isolate of interest.

According to the French Society of Microbiology, Staphylococcus aureus isolates are suspected to have decreased susceptibility to glycopeptide(s) when at least one colony is able to grow from an inoculum of 10 microL of 2 McFarland bacterial suspension plated on Mueller-Hinton agar containing 5 mg/L teicoplanin and incubated for 48 h at 35-37 degrees C. We analysed 89 methicillin-resistant S. aureus isolates (MRSA), collected in 2000-2001 from 24 hospitals located in 18 French cities, which were able to grow on this selective medium. These isolates were distributed into six groups on the basis of their glycopeptide resistance phenotypes: (A) glycopeptide susceptible (GSSA, 21 isolates); (B) heterogeneous teicoplanin intermediately resistant (hetero-TISA, 24 isolates); (C) heterogeneous and intermediately resistant to both glycopeptides, teicoplanin and vancomycin (hetero-GISA, six isolates); (D) heterogeneous vancomycin intermediately resistant/teicoplanin intermediately resistant (hetero-VISA/TISA, 30 isolates); (E) GISA (four isolates); (F) TISA (four isolates). Despite the persistent decrease in gentamicin-resistant MRSA isolates in French hospitals since 1993, their prevalence is very high in groups D, E and F. Moreover, most of the group C, D and E isolates exhibiting decreased susceptibility to both glycopeptides belong to the same major SmaI genotype, which has been detected in Europe since at least 1989.

We analyzed 62 clinical isolates of streptogramin A-resistant (SGA r ) Staphylococcus aureus collected between 1981 and 2001 in 14 hospitals located in seven French cities. These isolates, including five with decreased susceptibility to glycopeptides, were distributed into 45 antibiotypes and 38 Sma I genotypes. Each of these genotypes included between 1 and 11 isolates, the Sma I patterns of which differed by no more than three bands. Although numerous clones were identified, we observed the spread of monoclonal isolates either within the same hospital or within hospitals in distinct cities and at large time intervals. Hybridization with probes directed against 10 SGA r genes ( vatA , vatB , vatC , vatD , vatE , vgaA , vgaB , vgaAv , vgbA , and vgbB ) revealed six patterns: vgaAv (21 isolates), vatA - vgbA (24 isolates), vgaAv - vatB - vgaB (14 isolates), vgaAv - vatA - vgbA (1 isolate), vgaAv - vatA - vgbA - vatB - vgaB (1 isolate), and vgaA (1 isolate). We detected at least one SGA r determinant in all of the tested isolates. vgaAv , which is part of the recently characterized transposon Tn 5406 , was found in 59.7% of the tested isolates. Of the 16 streptogramin B-susceptible isolates, 14 carried vgaAv alone and were susceptible to the mixtures of streptogramins, whereas the 2 isolates carrying vgaAv - vatB - vgaB were resistant to these mixtures. vatA - vgbA was found on plasmids of the same apparent size in 26 (42%) of the tested clinical isolates from 18 unrelated Sma I genotypes. The possible dissemination of some of the multiple clones characterized in the present study with an expected increased selective pressure of streptogramins following the recent licensing of Synercid (quinupristin-dalfopristin) must be carefully monitored.

Methicillin-resistant strains susceptible to gentamicin (Gm s MRSA) have emerged since 1993 in several French hospitals. To study whether particular clones have spread in various French cities and whether some clones are related to gentamicin-resistant (Gm r ) MRSA strains, various methods (antibiotyping, phage typing, determination of Sma I macrorestriction patterns before and after hybridization with IS 256 transposase and aacA-aphD probes) were used to compare 62 Gm s MRSA strains isolated from 1995 to 1997 in nine cities and 15 Gm r MRSA strains. Eighteen major Sma I genotypes were identified, of which 11 included only Gm s MRSA strains and 5 included only Gm r MRSA strains. Each of the Gm r MRSA strains contained 6 to 13 Sma I fragments hybridizing with the insertion sequence IS 256 , of which a single band also hybridized with the aacA-aphD gene. No such hybridizing sequences were detected in 60 of the 62 Gm s MRSA strains. Thus, the divergence between Gm r and Gm s MRSA strains is revealed, not only by their distributions in distinct Sma I genotypes but also by the differences in hybridization patterns. Two of the 62 Gm s MRSA strains had the uncommon feature of carrying several Sma I bands hybridizing with IS 256 , suggesting that they are possibly related to the Gm r MRSA strains grouped in the same Sma I genotype. Five of the 11 Sma I genotypes including only Gm s MRSA strains contained strains from diverse cities, isolated during different years and with different antibiograms, suggesting that some clones have spread beyond their cities of origin and persisted.

We studied the Sma I and Sst II macrorestriction patterns of 54 Staphylococcus epidermidis strains isolated from 14 patients infected following the implantation of joint prostheses. Multiple strains from pus and infected tissue specimens of each patient were selected on the basis of different colony morphologies and drug resistance patterns. The same criteria were used to select 23 S. epidermidis strains from hand swabs of eight healthy individuals. For 10 of the 14 patients, all the intrapatient strains appeared to be closely or possibly related, whereas related strains were detected in the skin flora of only one of the eight healthy individuals. This observation suggests that, in most cases, the patients were infected by a single S. epidermidis clone which subsequently underwent rearrangements that yielded derivatives with divergent phenotypes and, occasionally, divergent macrorestriction patterns. The four patients whose specimens contained unrelated S. epidermidis strains were probably infected with several polyclonal strains.

Twenty well-characterized isolates of methicillin-resistant Staphylococcus aureus were used to study the optimal resolution and interlaboratory reproducibility of pulsed-field gel electrophoresis (PFGE) of DNA macrorestriction fragments. Five identical isolates (one PFGE type), 5 isolates that produced related PFGE subtypes, and 10 isolates with unique PFGE patterns were analyzed blindly in 12 different laboratories by in-house protocols. In several laboratories a standardized PFGE protocol with a commercial kit was applied successfully as well. Eight of the centers correctly identified the genetic homogeneity of the identical isolates by both the in-house and standard protocols. Four of 12 laboratories failed to produce interpretable data by the standardized protocol, due to technical problems (primarily plug preparation). With the five related isolates, five of eight participants identified the same subtype interrelationships with both in-house and standard protocols. However, two participants identified multiple strain types in this group or classified some of the isolates as unrelated isolates rather than as subtypes. The remaining laboratory failed to distinguish differences between some of the related isolates by utilizing both the in-house and standardized protocols. There were large differences in the relative genome lengths of the isolates as calculated on the basis of the gel pictures. By visual inspection, the numbers of restriction fragments and overall banding pattern similarity in the three groups of isolates showed interlaboratory concordance, but centralized computer analysis of data from four laboratories yielded percent similarity values of only 85% for the group of identical isolates. The differences between the data sets obtained with in-house and standardized protocols could be the experimental parameters which differed with respect to the brand of equipment used, imaging software, running time (20 to 48 h), and pulsing conditions. In conclusion, it appears that the standardization of PFGE depends on controlling a variety of experimental intricacies, as is the case with other bacterial typing procedures.

Summary In an Algerian hospital where pristinamycin (Pt) was extensively used for the treatment of chronic osteomyelitis and for prophylaxis in bone surgery, the prevalence of pristinamycin-resistant (Pt R ) staphylococci during a five-month period (20%) was higher than that among staphylococci isolated elsewhere in Algeria (4.5%). Analysis of 13 Pt R staphylococci isolated in this hospital revealed a diversity of plasmids and genes conferring resistance to Pt and to related antibiotics. Most of the Pt R staphylococci were unrelated: they belonged to either different taxa or types. Nevertheless, some of the unrelated staphylococci harboured structurally related plasmids carrying streptogramin resistance genes. Thus, these plasmids may have contributed to the dispersion of these genes.

Summary Various methods were used to identify 145 staphylococcal strains isolated from 25 traditional types of cheeses. With the biochemical micromethod rapid gallery system. ATB 32 Staph, only 33% of the strains could be identified. All the strains unidentified by API system (98 strains) were subjected to 35 conventional biochemical and physiological tests according to the Devriese's identification scheme (1985). The results were then analyzed by a numerical method using the Sokal and Michener similarity coefficient and the 98 strains were distributed into 15 clusters at the similarity level of 60%. Of the 145 strains analyzed, 36% were identified using the association of two phenotypic methods and 26% were considered as atypical S. xylosus or S. equorum strains. An identification method based on analysis of the ribotypes was applied to 67 strains from the 15 taxa. The cellular DNA was cleaved by the Hind III and Eco RI restriction enzymes. The restriction fragments were separated by electrophoresis, blotted and probed with fluorescein labeled plasmid pBA2 containing a Bacillus subtilis gene encoding 16S rRNA. Sixty six of the 67 strains ribotyped (98.5%) gave typical known species patterns. In conclusion, 99.5% of the 145 staphylococcal cheese strains were identified. All chese strains belonged to the novobiocin-resistant CNS species and near 90% to S. equorum, S. vitulus and S. xylosus which are not widespread among human associated strains.

The levels of resistance to pristinamycin (Pt) and to its major constituents, pristinamycin IIA and IB (PIIA and PIB, respectively; classified as streptogramins A and B, respectively) were determined for 126 staphylococcal isolates. The results suggest tentative susceptibility breakpoints of < or = 2, < or = 8, and < or = 0.5 microgram/ml for PIIA, PIB, and Pt, respectively. Fifty-six isolates that were inhibited by > or = 4 micrograms of PIIA per ml were investigated for the presence of staphylococcal genes encoding resistance to PIIA (vga, vat, and vatB) and PIB (vgb). None of these genes was found in the 4 isolates inhibited by 4 micrograms of PIIA per ml or in 4 of the other 52 isolates tested. The remaining 48 isolates harbored plasmids carrying vatB and vga or combinations of genes (vga-vat-vgb or vga-vat). The absence of any known PIIA resistance gene from the four Staphylococcus aureus isolates inhibited by > or = 8 micrograms of PIIA per ml suggests that there is at least one PIIA resistance mechanism in staphylococci that has not yet been characterized.

During a 14-month period, from December 1984 to February 1986, 630 Staphylococcus aureus isolates were identified at Broussais Hospital. Thirty-eight isolates (6%), from 35 patients, were found to be pefloxacin-resistant S. aureus (PRSA) with minimal inhibitory concentrations greater than or equal to 8 mg l −1 . PRSA isolates were tested for susceptibility to 35 antibiotics, including nine quinolones, and heavy metal ions. Phage-type was determined. Out of the 38 PRSA isolates, 35 (92%) were methicillin- and multiply-resistant; however, all PRSA isolates were sensitive to vancomycin and coumermycin. Fifteen isolates (39%) had similar phage-type and identical antibiotic susceptibility pattern with high level resistance to pefloxacin (MICs equal to 64 mg l −1 ); they were isolated from the same surgical unit. The 23 remaining PRSA isolates differed by their phage and susceptibility patterns. Pefloxacin MICs ranged from 8 to 512 mg l −1 with a bimodal distribution; cross-resistance was observed with the eight other quinolones tested. Only nine PRSA isolates (24%), including four ‘epidemic' isolates, were obtained from patients who had been treated with quinolones. From these data there is apparently no direct relationship between quinolone administration and selection of PRSA in infected patients.

The performance of a multiplex PCR assay that separates the four major serovars of the pathogenic Listeria monocytogenes into four distinct PCR groups was evaluated through a multicenter typing study. Identical panels of 90 Listeria isolates were distributed to five participating laboratories that were blind to the nature of the isolates. Isolates were characterized using the previously standardized protocol. Overall concordance was 96.6 to 100%, sufficient for the assay to be used as an alternative to serotyping and confidently applied in laboratories involved in L. monocytogenes typing.

The Vga and Msr resistance determinants, encoded by mobile genetic elements in various staphylococcal strains, belong to a family of ATP-binding cassette (ABC) proteins whose functions and structures are ill defined. Their amino acid sequences are similar to those of proteins involved in the immunity of streptomycetes to the macrolide-lincosamide-streptogramin antibiotics that they produce. Sequence analysis of the genomes of the gram-positive bacteria with low G+C contents revealed that Lmo0919 from Listeria monocytogenes is more closely related to Vga variants than to Msr variants. In the present study we compared the antibiotic resistance profiles conferred by the Vga-like proteins in two staphylococcal hosts. It was shown that Vga(A), the Vga(A) variant [Vga(A)v], and Lmo0919 can confer resistance to lincosamides and streptogramin A compounds, while only Vga(B) is able to increase the level of resistance to pristinamycin, a mixture of streptogramin A and streptogramin B compounds. By using polyclonal antibodies, we found that the Vga(A) protein colocalized with the β subunit of the F 1 -F 0 ATPase in the membrane fractions of staphylococcal cells. In order to identify functional units in these atypical ABC proteins, such as regions that might be involved in substrate specificity and/or membrane targeting, we analyzed the resistance phenotypes conferred by various plasmids carrying parts or modified versions of the vga (A) gene and we determined the subcellular localization of the gene products. Only polypeptides composed of two ABC domains were detected in the cell membranes. No region of drug specificity was identified. Resistance properties were dependent on the integrities of both Walker B motifs.

International audience; A total of 101 staphylococcal strains were ribotyped using EcoRI and HindIII as restriction enzymes and plasmid pBA2 as the rDNA probe. Isolates from 10 newly described staphylococcal taxa were among those examined. All the ribotypes were added to our database, Staph DB, which now contains the sizes of the bands of 135 EcoRI and 120 HindIII ribotypes from 408 strains belonging to 42 staphylococcal taxa. The relatedness of ribotypes was evaluated by using the Dice coefficient. The ribotypes, and thus the strains, were clustered by the unweighted pair group method with averages (UPGMA). Separation into clusters correlated well with the delineation of the staphylococcal species but not with that of the different subspecies. No discrimination was possible between Staphylococcus vitulinus and Staphylococcus pulvereri. Ecovar-specific groups were evident within Staphylococcus intermedius and Staphylococcus hyicus. The data increase the usefulness of rRNA gene restriction site polymorphism analysis for staphylococcal taxonomy.

Ten Staphylococcus caprae strains isolated from four patients and responsible for bone infections following implantation of orthopaedic material were compared to four S. caprae strains collected from milk samples of healthy goats. The following characteristics were investigated: Smal patterns, hybridization patterns with pBA2 (ribotypes), slime production, adhesion to matrix proteins (fibrinogen, fibronectin, collagen) and the staphylococcal adhesion genes (fnbA, clfA, cna, atlE, ica, fbe). None of the characteristics enabled us to distinguish the human strains from the goat strains. Slime was occasionally produced by S. caprae strains but all of them carried nucleotide sequences hybridizing at low stringency with the following genes: atlE encoding a S. epidermidis autolysin binding vitronectin and responsible for the primary adhesion to polystyrene, ica operon involved in the biosynthesis of a S. epidermidis extracellular polysaccharide, and the part of clfA encoding the serine-aspartate repeated region of a S. aureus cell-wall fibrinogen-binding protein.

International audience; A new novobiocin-susceptible species of the genus Staphylococcus, Staphylococcus pasteuri, is described on the basis of the results of a study of seven strains isolated from human, animal, and food specimens. DNA relatedness experiments (S1 nuclease method) showed that these strains form a homogeneous genomic species related at DNA homology levels of 2 to 13% to 27 type strains representing known Staphylococcus species. The use of a method based on rRNA gene restriction site polymorphism provides clear-cut distinction between this new species and Staphylococcus warneri, which is the most similar species phenotypically. The type strain of the new species is strain BM9357 (= ATCC 51129).

International audience; The usefulness of the ID32 Staph System and a method based on rRNA gene restriction site polymorphism was evaluated by the study of 42 staphylococcal clinical isolates phenotypically difficult to identify. The ID32 Staph micromethod and the genomic method are adapted for recognition of 27 and 31 staphylococcal taxa, respectively. The genomic method is based on a Dice analysis of the hybridization patterns obtained by cutting the cellular DNA either with EcoRI or with HindIII and by probing with pBA2, containing the Bacillus subtilis gene encoding 16S rRNA, labeled either with [alpha-32P]dCTP or with acetylaminofluorene. This study showed that the nonradioactive labeling provided a better resolution of the hybridizing bands than radioactive labeling. Of the 42 isolates selected, only 22 could be assigned to a staphylococcal species by the ID32 Staph System, whereas 35 could be identified by the genomic method. This latter method also enabled the screening of three unclassified isolates having hybridization patterns more closely related to each other than to any of the 31 staphylococcal taxa investigated. These three isolates could belong to a staphylococcal taxon not yet described.

Summary. Fifty-nine epidemiologically unrelated methicillin-resistant Staphylococcus aureus (MRSA) isolates from different geographical areas and 23 phage-type 77 MRSA isolates from France were investigated. Cellular DNA, digested with restriction endonucleases EcoRI or HindIII, was probed with plasmids carrying the gene encoding 16S rRNA (pBA2), the gene aacA-aphD (pSF815A) and the gene aacA-aphD plus part of IS256 (pIP 1307). When probed with pBA2, most of the unrelated isolates displayed the same hybridisation pattern. A greater diversity in patterns was detected in gentamicin-resistant strains with the two other probes. The most accurate fingerprinting of these isolates was obtained with the probe pIP 1307. Moreover, this probe appeared to be useful for tracing the phage-type 77 epidemic MRSA isolates widespread in French hospitals.

SUMMARY: The rRNA gene restriction patterns of 110 strains belonging to 12 staphylococcal species have been determined. The strains, isolated from various sources, were epidemiologically unrelated. Total DNA was cleaved with restriction enzymes HindlII and Eco RI, electrophoretically separated and probed with radiolabelled 16S rDNA from Bacillus subtilisinserted in a plasmid vector, pBR322. Fourty-four distinct Hindlll patterns and 44 distinct EcoRl patterns were observed. Strains belonging to different species had different patterns. Although distinct patterns were also observed within some species, a core of common bands could be discerned within each species or subspecies. Analysis of the patterns revealed two taxa in Staphylococcusxylosus which were not evident using phenotypic characteristics. Of 18 strains which were difficult to identify using phenotypic schemes, 15 showed patterns typical of known species. The three remaining atypical strains showed unusual patterns and may belong either to a known species, not included in the study, or to a new species. Since various patterns were observed within some species (e.g. S. aureus and S. epidermidis), rRNA gene restriction patterns may have epidemiological, as well as taxonomic interest.

Listeria monocytogenes (Lm) is a major human and animal foodborne pathogen. Here we show that hypervirulent Lm clones, particularly CC1, are strongly associated with dairy products, whereas hypovirulent clones, CC9 and CC121, are associated with meat products. Clone adaptation to distinct ecological niches and/or different food products contamination routes may account for this uneven distribution. Indeed, hypervirulent clones colonize better the intestinal lumen and invade more intestinal tissues than hypovirulent ones, reflecting their adaption to host environment. Conversely, hypovirulent clones are adapted to food processing environments, with a higher prevalence of stress resistance and benzalkonium chloride tolerance genes and a higher survival and biofilm formation capacity in presence of sub-lethal benzalkonium chloride concentrations. Lm virulence heterogeneity therefore reflects the diversity of the ecological niches in which it evolves. These results also have important public health implications and may help in reducing food contamination and improving food consumption recommendations to at-risk populations., Here, Maury et al. show that hypervirulent Listeria monocytogenes (Lm) clones associated to dairy products exhibit higher adaptation to the mammalian gut environment, while hypovirulent clones persist in food processing environment, suggesting a relationship between Lm pathogenic potential and niche adaptation.

The pathogenesis of Listeria monocytogenes depends on the ability of this bacterium to escape from the phagosome of the host cells via the action of the pore-forming toxin listeriolysin O (LLO). Expression of the LLO-encoding gene ( hly ) requires the transcriptional activator PrfA, and both hly and prfA genes are essential for L. monocytogenes virulence. Here, we used the hemolytic activity of LLO as a phenotypic marker to screen for spontaneous virulence-attenuating mutations in L. monocytogenes . Sixty nonhemolytic isolates were identified among a collection of 57,820 confirmed L. monocytogenes strains isolated from a variety of sources (0.1%). In most cases (56/60; 93.3%), the nonhemolytic phenotype resulted from nonsense, missense, or frameshift mutations in prfA . Five strains carried hly mutations leading to a single amino acid substitution (G299V) or a premature stop codon causing strong virulence attenuation in mice. In one strain, both hly and gshF (encoding a glutathione synthase required for full PrfA activity) were missing due to genomic rearrangements likely caused by a transposable element. The PrfA/LLO loss-of-function (PrfA − /LLO − ) mutants belonged to phylogenetically diverse clades of L. monocytogenes , and most were identified among nonclinical strains (57/60). Consistent with the rare occurrence of loss-of-virulence mutations, we show that prfA and hly are under purifying selection. Although occurring at a low frequency, PrfA − /LLO − mutational events in L. monocytogenes lead to niche restriction and open an evolutionary path for obligate saprophytism in this facultative intracellular pathogen.

International audience; Enterotoxins produced by Staphylococcus aureus are responsible for staphylococcal food-poisoning outbreaks (SFPO). In France, SFPO are the second cause of food-borne diseases after Salmonella. However, very little is known about the strains involved. The objective of this study was to characterize the staphylococcal strains related to these SFPO through phenotypic and genotypic analyses. A total of 178 coagulase-positive staphylococcal isolates recovered from 31 SFPO (1981-2002) were screened through biotyping. Thirty-three strains representative of the different biotypes in each SFPO were further examined for SmaI macrorestriction-type, phage-type, resistance to various antimicrobial drugs, presence of staphylococcal enterotoxin (se) genes sea to sei, and production of enterotoxins SEA to SED. All these 33 strains were identified as S. aureus species: 27 were of human biotypes and six ovine or non-host-specific biotypes. Most (74.1%) strains reacted with group III phages. Eleven strains were resistant to at least two classes of antibiotics and among them, two were resistant to methicillin. Twenty-nine strains carried one or several of the eight se genes tested; the gene sea was most common (n=23), and often linked to sed (n=12) or seh (n=5). The novel se genes seg-i were in all cases associated with se genes sea to sed except for one strain which carried only seg and sei. Pulsed-Field Gel Electrophoresis (PFGE) of SmaI macrorestriction digests of the 33 strains discriminated 32 PFGE patterns grouped into nine biotype-specific clusters. All five strains carrying sea and seh were grouped together into the same sub-cluster. Three of the four se-gene-negative strains were in one PFGE cluster: all four should be tested for se genes not included in this study and, if negative, be further investigated for the presence of unidentified SEs.