Worldwide Distribution of Major Clones of Listeria monocytogenes

Listeria monocytogenes is worldwide a pathogen, but the geographic distribution of clones remains largely unknown. Genotyping of 300 isolates from the 5 continents and diverse sources showed the existence of few prevalent and globally distributed clones, some of which include previously described epidemic clones. Cosmopolitan distribution indicates the need for genotyping standardization

Spontaneous virulence loss in natural populations of Listeria monocytogenes

International audienceThe 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 Ϫ /LLO Ϫ) mutants belonged to phylogenetically diverse clades of L. monocyto-genes, 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 Ϫ /LLO Ϫ muta-tional events in L. monocytogenes lead to niche restriction and open an evolutionary path for obligate saprophytism in this facultative intracellular pathogen

Highly Rifampin-Resistant Listeria monocytogenes Isolated from a Patient with Prosthetic Bone Infection

International audienc

Clonogrouping, a Rapid Multiplex PCR Method for Identification of Major Clones of Listeria monocytogenes

A patent application describing the clonogrouping method was submitted (V. Chenal-Francisque, M. M. Maury, M. Lavina, M. Touchon, A. Leclercq, M. Lecuit, and S. Brisse, patent application EP 15 306 154.4, 10 July 2015, European Patent Office).International audienceThree multiplex PCR assays were developed to identify the 11 most common Listeria monocytogenes clones in clinical and food samples; 270 (95.7%) of 282 strains of serogroups IVb, IIb, IIa, and IIc were identified accurately. This novel tool is a rapid and efficient alternative to multilocus sequence typing for identification of L. monocytogenes clones

Relationship between bacterial virulence and nucleotide metabolism: a mutation in the adenylate kinase gene renders Yersinia pestis avirulent.

Nucleoside monophosphate kinases (NMPKs) are essential catalysts for bacterial growth and multiplication. These enzymes display high primary sequence identities among members of the family Enterobacteriaceae. Yersinia pestis, the causative agent of plague, belongs to this family. However, it was previously shown that its thymidylate kinase (TMPKyp) exhibits biochemical properties significantly different from those of its Escherichia coli counterpart [Chenal-Francisque, Tourneux, Carniel, Christova, Li de la Sierra, Barzu and Gilles (1999) Eur. J. Biochem. 265, 112-119]. In this work, the adenylate kinase (AK) of Y. pestis (AKyp) was characterized. As with TMPKyp, AKyp displayed a lower thermodynamic stability than other studied AKs. Two mutations in AK (Ser129Phe and Pro87Ser), previously shown to induce a thermosensitive growth defect in E. coli, were introduced into AKyp. The recombinant variants had a lower stability than wild-type AKyp and a higher susceptibility to proteolytic digestion. When the Pro87Ser substitution was introduced into the chromosomal adk gene of Y. pestis, growth of the mutant strain was altered at the non-permissive temperature of 37 degree C. In virulence testings, less than 50 colony forming units (CFU) of wild-type Y. pestis killed 100% of the mice upon subcutaneous infection, whereas bacterial loads as high as 1.5 x 10(4) CFU of the adk mutant were unable to kill any animals

“Epidemic Clones” of Listeria monocytogenes Are Widespread and Ancient Clonal Groups

International audienceThe food-borne pathogen Listeria monocytogenes is genetically heterogeneous. Although some clonal groups have been implicated in multiple outbreaks, there is currently no consensus on how "epidemic clones" should be defined. The objectives of this work were to compare the patterns of sequence diversity on two sets of genes that have been widely used to define L. monocyto-genes clonal groups: multilocus sequence typing (MLST) and multi-virulence-locus sequence typing (MvLST). Further, we evaluated the diversity within clonal groups by pulsed-field gel electrophoresis (PFGE). Based on 125 isolates of diverse temporal, geographical , and source origins, MLST and MvLST genes (i) had similar patterns of sequence polymorphisms, recombination, and selection, (ii) provided concordant phylogenetic clustering, and (iii) had similar discriminatory power, which was not improved when we combined both data sets. Inclusion of representative strains of previous outbreaks demonstrated the correspondence of epidemic clones with previously recognized MLST clonal complexes. PFGE analysis demonstrated heterogeneity within major clones, most of which were isolated decades before their involvement in outbreaks. We conclude that the "epidemic clone" denominations represent a redundant but largely incomplete nomenclature system for MLST-defined clones, which must be regarded as successful genetic groups that are widely distributed across time and space

RESEARCH Plague Outbreak in Libya, 2009, Unrelated to Plague in Algeria

After 25 years of no cases of plague, this diseas

Optimized Multilocus Variable-Number Tandem-Repeat Analysis Assay and Its Complementarity with Pulsed-Field Gel Electrophoresis and Multilocus Sequence Typing for Listeria monocytogenes Clone Identification and Surveillance

International audiencePopulations of the food-borne pathogen Listeria monocytogenes are genetically structured into a small number of major clonal groups, some of which have been implicated in multiple outbreaks. The goal of this study was to develop and evaluate an optimized multilocus variable number of tandem repeat (VNTR) analysis (MLVA) subtyping scheme for strain discrimination and clonal group identification. We evaluated 18 VNTR loci and combined the 11 best ones into two multiplexed PCR assays (MLVA-11). A collection of 255 isolates representing the diversity of clonal groups within phylogenetic lineages I and II, including representatives of epidemic clones, were analyzed by MLVA-11, multilocus sequence typing (MLST), and pulsed-field gel electropho-resis (PFGE). MLVA-11 had less discriminatory power than PFGE, except for some clones, and was unable to distinguish some epidemiologically unrelated isolates. Yet it distinguished all major MLST clones and therefore constitutes a rapid method to identify epidemiologically relevant clonal groups. Given its high reproducibility and high throughput, MLVA represents a very attractive first-line screening method to alleviate the PFGE workload in outbreak investigations and listeriosis surveillance. L isteriosis is a food-borne infection caused by the bacterium Listeria monocytogenes. Invasive forms of human listeriosis include septicemia, meningitis, and maternal-fetal infections (1). Listeriosis is associated with high hospitalization and fatality rates (almost 100% and 25 to 30%, respectively). Populations at risk include pregnant women, immunocompromised individuals, and the elderly. L. monocytogenes is widely present in the environment, including soil, water, vegetation, and silage, as well as in animals and animal-derived food, and can contaminate food in processing plants and retail establishments. L. monocytogenes is recognized as a public health issue and a serious challenge for the food industry, and this has led to the establishment of national surveillance systems in several countries. L. monocytogenes also stands out as a model system in the fields of microbiology, cell biology, and im-munology and for the study of host-pathogen interactions (2-5). L. monocytogenes strain characterization on the basis of sero-typing and molecular typing methods is used for surveillance, epidemiological tracking, and outbreak investigation purposes (6, 7). Genetic variants of L. monocytogenes have diversified into four major phylogenetic lineages, with lineages 1 and 2 each containing multiple clonal groups of public health importance (8-14). As these groups appear to differ in virulence and epidemic potential (6, 15), it will be interesting to better define their epidemiological, clinical, and microbiological specificities. For this purpose, tools for the easy identification of clonal groups are needed to recognize such groups and determine their presence in a large variety of sources. Several typing methods are currently available for L. monocytogenes strains. Conventional serotyping (16) and its molecular proxy PCR serogrouping (17) discriminate major categories of strains that correlate strongly (albeit not totally) with lin-eages and clones (11, 12, 14), but these methods do not have the necessary discriminatory power in the context of outbreak investigations. Pulsed-field gel electrophoresis (PFGE) is established as the gold standard for L. monocytogenes strain subtyping and is widely used for listeriosis surveillance and outbreak investigations (18). Yet, PFGE presents several practical disadvantages, as it is time-consuming and requires stringent standardization for inter-laboratory data comparison. Multilocus sequence typing (MLST) is a well-established reference method for global epidemiology and population biology (19, 20), as it renders interlaboratory genotype comparisons easy and unambiguous and as sequence data can be used to infer useful population genetic information such as amounts of genetic diversity, recombination rates, and strain phy-logeny. MLST also provides backward compatibility with genome sequencing (21). However, MLST is neither rapid nor cheap and has limited discriminatory power within L. monocytogenes (12, 22). Given the current limitations of available methods for L. monocytogenes strain typing, a potentially useful complementary approach is multilocus variable number of tandem repeat

ActA promotes Listeria monocytogenes aggregation, intestinal colonization and carriage.

International audienceListeria monocytogenes (Lm) is a ubiquitous bacterium able to survive and thrive within the environment and readily colonizes a wide range of substrates, often as a biofilm. It is also a facultative intracellular pathogen, which actively invades diverse hosts and induces listeriosis. So far, these two complementary facets of Lm biology have been studied independently. Here we demonstrate that the major Lm virulence determinant ActA, a PrfA-regulated gene product enabling actin polymerization and thereby promoting its intracellular motility and cell-to-cell spread, is critical for bacterial aggregation and biofilm formation. We show that ActA mediates Lm aggregation via direct ActA-ActA interactions and that the ActA C-terminal region, which is not involved in actin polymerization, is essential for aggregation in vitro. In mice permissive to orally-acquired listeriosis, ActA-mediated Lm aggregation is not observed in infected tissues but occurs in the gut lumen. Strikingly, ActA-dependent aggregating bacteria exhibit an increased ability to persist within the cecum and colon lumen of mice, and are shed in the feces three order of magnitude more efficiently and for twice as long than bacteria unable to aggregate. In conclusion, this study identifies a novel function for ActA and illustrates that in addition to contributing to its dissemination within the host, ActA plays a key role in Lm persistence within the host and in transmission from the host back to the environment