Chantal Bizet

The Intestinal Microbiota Modulates the Anticancer Immune Effects of Cyclophosphamide

The Microbiota Makes for Good Therapy The gut microbiota has been implicated in the development of some cancers, such as colorectal cancer, but—given the important role our intestinal habitants play in metabolism—they may also modulate the efficacy of certain cancer therapeutics. Iida et al. (p. 967) evaluated the impact of the microbiota on the efficacy of an immunotherapy [CpG (the cytosine, guanosine, phosphodiester link) oligonucleotides] and oxaliplatin, a platinum compound used as a chemotherapeutic. Both therapies were reduced in efficacy in tumor-bearing mice that lacked microbiota, with the microbiota important for activating the innate immune response against the tumors. Viaud et al. (p. 971) found a similar effect of the microbiota on tumor-bearing mice treated with cyclophosphamide, but in this case it appeared that the microbiota promoted an adaptive immune response against the tumors. The gut microbiota promote the efficacy of several antineoplastic agents in mice. Cyclophosphamide is one of several clinically important cancer drugs whose therapeutic efficacy is due in part to their ability to stimulate antitumor immune responses. Studying mouse models, we demonstrate that cyclophosphamide alters the composition of microbiota in the small intestine and induces the translocation of selected species of Gram-positive bacteria into secondary lymphoid organs. There, these bacteria stimulate the generation of a specific subset of “pathogenic” T helper 17 (pTH17) cells and memory TH1 immune responses. Tumor-bearing mice that were germ-free or that had been treated with antibiotics to kill Gram-positive bacteria showed a reduction in pTH17 responses, and their tumors were resistant to cyclophosphamide. Adoptive transfer of pTH17 cells partially restored the antitumor efficacy of cyclophosphamide. These results suggest that the gut microbiota help shape the anticancer immune response.

Listeria rocourtiae sp. nov.

A Listeria-like strain isolated in Austria from pre-cut lettuce fitted the description of the genus Listeria although it could not be assigned to any of the known species. Comparison of the rrs gene (encoding 16S rRNA) sequence and gene content by DNA-array indicated affiliation to the genus Listeria. Phylogenetic distance from known species of the genus Listeria indicated that it represents a novel species. Since it can be differentiated from all other known species of the genus Listeria by using phenotypic tests, the name Listeria rocourtiae sp. nov. is proposed for the novel species. The type strain is CIP 109804(T) (=DSM 22097(T) =Allerberger 700284/02(T)). The type strain is avirulent as assessed by cell culture assays and inoculation of mice.

High Interlaboratory Reproducibility of Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry-Based Species Identification of Nonfermenting Bacteria

ABSTRACT Matrix-assisted laser desorption ionization-time of flight mass spectrometry has emerged as a rapid, cost-effective alternative for bacterial species identification. Identifying 60 blind-coded nonfermenting bacteria samples, this international study (using eight laboratories) achieved 98.75% interlaboratory reproducibility. Only 6 of the 480 samples were misidentified due to interchanges (4 samples) or contamination (1 sample) or not identified because of insufficient signal intensity (1 sample).

Identification and Functional Characterization of Arylamine N-Acetyltransferases in Eubacteria: Evidence for Highly Selective Acetylation of 5-Aminosalicylic Acid

Arylamine N-acetyltransferase activity has been described in various bacterial species. Bacterial N-acetyltransferases, including those from bacteria of the gut flora, may be involved in the metabolism of xenobiotics, thereby exerting physiopathological effects. We characterized these enzymes further by steady-state kinetics, time-dependent inhibition, and DNA hybridization in 40 species, mostly from the human intestinal microflora. We report for the first time N-acetyltransferase activity in 11 species of Proteobacteriaceae from seven genera: Citrobacter amalonaticus, Citrobacter farmeri, Citrobacter freundii, Klebsiella ozaenae, Klebsiella oxytoca, Klebsiella rhinoscleromatis, Morganella morganii, Serratia marcescens, Shigella flexneri, Plesiomonas shigelloides, and Vibrio cholerae. We estimated apparent kinetic parameters and found that 5-aminosalicylic acid, a compound efficient in the treatment of inflammatory bowel diseases, was acetylated with a catalytic efficiency 27 to 645 times higher than that for its isomer, 4-aminosalicylic acid. In contrast, para-aminobenzoic acid, a folate precursor in bacteria, was poorly acetylated. Of the wild-type strains studied, Pseudomonas aeruginosa was the best acetylator in terms of both substrate spectrum and catalytic efficiency. DNA hybridization with a Salmonella enterica serovar Typhimurium-derived probe suggested the presence of this enzyme in eight proteobacterial and four gram-positive species. Molecular aspects together with the kinetic data suggest distinct functional features for this class of microbial enzymes.

Molecular Characterization of Chromosomal Class C β-Lactamase and Its Regulatory Gene in Ochrobactrum anthropi

ABSTRACT Ochrobactrum anthropi, formerly known as CDC group Vd, is an oxidase-producing, gram-negative, obligately aerobic, non-lactose-fermenting bacillus of low virulence that occasionally causes human infections. It is highly resistant to all β-lactams except imipenem. A clinical isolate, SLO74, and six reference strains were tested. MICs of penicillins, aztreonam, and most cephalosporins tested, including cefotaxime and ceftazidime, were >128 μg/ml and of cefepime were 64 to >128 μg/ml. Clavulanic acid was ineffective and tazobactam had a weak effect in association with piperacillin. Two genes, ampR and ampC, were cloned by inserting restriction fragments of genomic DNA from the clinical strain O. anthropi SLO74 into pBK-CMV to give the recombinant plasmid pBK-OA1. The pattern of resistance to β-lactams of this clone was similar to that of the parental strain, except for its resistance to cefepime (MIC, 0.5 μg/ml). The deduced amino acid sequence of the AmpC β-lactamase (pI, 8.9) was only 41 to 52% identical to the sequence of other chromosomally encoded and plasmid-encoded class C β-lactamases. The kinetic properties of this β-lactamase were typical for this class of β-lactamases. Upstream from the ampC gene, the ampR gene encodes a protein with a sequence that is 46 to 62% identical to those of other AmpR proteins and with an amino-terminal DNA-binding domain typical of transcriptional activators of the Lys-R family. The deduced amino acid sequences of theampC genes of the six reference strains were 96 to 99% identical to the sequence of the clinical strain. The β-lactamase characterized from strain SLO74 was named OCH-1 (gene, blaOCH-I).

Electrophoretic pattern of peptidoglycan hydrolases, a new tool for bacterial species identification : application to 10 Lactobacillus species

Lactobacilli have been used as industrial starters for a long time, but in many cases their phenotypic identification is still neither easy nor reliable. Previously we observed that the cell wall peptidoglycan hydrolases of Lactobacillus helveticus were highly conserved enzymes; the aim of the present work was to determine whether peptidoglycan hydrolase patterns obtained by renaturing SDS-PAGE could be of interest in the identification of lactobacilli species. For that purpose, the peptidoglycan hydrolase patterns of 94 strains of lactobacilli belonging to 10 different species were determined; most of the species studied are used either in dairy, meat, bakery or vegetable fermentations: L. helveticus, L. acidophilus, L. delbrueckii, L. brevis, L. fermentum, L. jensenii, L. plantarum, L. sake, L. curvatus and L. reuteri. Within a species, the strains exhibited highly similar patterns: the apparent molecular weights of the lytic bands were identical, with only slight variations of intensity. Moreover, each species, including phylogenetically close species such as L. sake and L. curvatus, or L. acidophilus and L. helveticus, gave a different pattern. Interestingly, the closer the species were phylogenetically, the more related were their patterns. The sensitivity of the method was checked using various quantities of L. acidophilus cells: a peptidoglycan hydrolase extract of 5 x 10(6) cells was sufficient to obtain an informative pattern, as was a single colony. Finally, the method was also successfully applied to distinguish two Carnobacterium species. In conclusion, the electrophoretic pattern of peptidoglycan hydrolases is proposed as a new tool for lactobacilli identification: it is rapid, sensitive and effective even for phylogenetically close species. Furthermore, this work provides the first evidence of the potential overall taxonomic value of bacterial peptidoglycan hydrolases.

Marinobacter salarius sp. nov. and Marinobacter similis sp. nov., isolated from sea water.

Two non-pigmented, motile, Gram-negative marine bacteria designated R9SW1T and A3d10T were isolated from sea water samples collected from Chazhma Bay, Gulf of Peter the Great, Sea of Japan, Pacific Ocean, Russia and St. Kilda Beach, Port Phillip Bay, the Tasman Sea, Pacific Ocean, respectively. Both organisms were found to grow between 4°C and 40°C, between pH 6 to 9, and are moderately halophilic, tolerating up to 20% (w/v) NaCl. Both strains were found to be able to degrade Tween 40 and 80, but only strain R9SW1T was found to be able to degrade starch. The major fatty acids were characteristic for the genus Marinobacter including C16:0, C16:1 ω7c, C18:1 ω9c and C18:1 ω7c. The G+C content of the DNA for strains R9SW1T and A3d10T were determined to be 57.1 mol% and 57.6 mol%, respectively. The two new strains share 97.6% of their 16S rRNA gene sequences, with 82.3% similarity in the average nucleotide identity (ANI), 19.8% similarity in the in silico genome-to-genome distance (GGD), 68.1% similarity in the average amino acid identity (AAI) of all conserved protein-coding genes, and 31 of the Karlins genomic signature dissimilarity. A phylogenetic analysis showed that R9SW1T clusters with M. algicola DG893T sharing 99.40%, and A3d10T clusters with M. sediminum R65T sharing 99.53% of 16S rRNA gene sequence similarities. The results of the genomic and polyphasic taxonomic study, including genomic, genetic, phenotypic, chemotaxonomic and phylogenetic analyses based on the 16S rRNA, gyrB and rpoD gene sequence similarities, the analysis of the protein profiles generated using MALDI-TOF mass spectrometry, and DNA-DNA relatedness data, indicated that strains R9SW1T and A3d10T represent two novel species of the genus Marinobacter. The names Marinobacter salarius sp. nov., with the type strain R9SW1T ( =  LMG 27497T  =  JCM 19399T  =  CIP 110588T  =  KMM 7502T) and Marinobacter similis sp. nov., with the type strain A3d10T ( =  JCM 19398T  =  CIP 110589T  =  KMM 7501T), are proposed.

Global phylogeography and evolutionary history of Shigella dysenteriae type 1.

Together with plague, smallpox and typhus, epidemics of dysentery have been a major scourge of human populations for centuries1. A previous genomic study concluded that Shigella dysenteriae type 1 (Sd1), the epidemic dysentery bacillus, emerged and spread worldwide after the First World War, with no clear pattern of transmission2. This is not consistent with the massive cyclic dysentery epidemics reported in Europe during the eighteenth and nineteenth centuries1,3,4 and the first isolation of Sd1 in Japan in 18975. Here, we report a whole-genome analysis of 331 Sd1 isolates from around the world, collected between 1915 and 2011, providing us with unprecedented insight into the historical spread of this pathogen. We show here that Sd1 has existed since at least the eighteenth century and that it swept the globe at the end of the nineteenth century, diversifying into distinct lineages associated with the First World War, Second World War and various conflicts or natural disasters across Africa, Asia and Central America. We also provide a unique historical perspective on the evolution of antibiotic resistance over a 100-year period, beginning decades before the antibiotic era, and identify a prevalent multiple antibiotic-resistant lineage in South Asia that was transmitted in several waves to Africa, where it caused severe outbreaks of disease.

Global and regional dissemination and evolution of Burkholderia pseudomallei

The environmental bacterium Burkholderia pseudomallei causes an estimated 165,000 cases of human melioidosis per year worldwide and is also classified as a biothreat agent. We used whole genome sequences of 469 B. pseudomallei isolates from 30 countries collected over 79 years to explore its geographic transmission. Our data point to Australia as an early reservoir, with transmission to Southeast Asia followed by onward transmission to South Asia and East Asia. Repeated reintroductions were observed within the Malay Peninsula and between countries bordered by the Mekong River. Our data support an African origin of the Central and South American isolates with introduction of B. pseudomallei into the Americas between 1650 and 1850, providing a temporal link with the slave trade. We also identified geographically distinct genes/variants in Australasian or Southeast Asian isolates alone, with virulence-associated genes being among those over-represented. This provides a potential explanation for clinical manifestations of melioidosis that are geographically restricted.

Microbacterium binotii sp. nov., isolated from human blood

Two strains of non-spore-forming, rod-shaped, Gram-positive bacteria, CIP 101303(T) and CIP 102116, were isolated from human blood in 1976 and 1977, respectively. These strains had chemotaxonomic markers that were consistent with classification in the genus Microbacterium, i.e. MK-10, MK-11 and MK-12 as the major menaquinones, predominant iso- and anteiso-branched cellular fatty acids, galactose, mannose and rhamnose as the cell-wall sugars and ornithine as the diamino acid in the cell-wall peptidoglycan. The DNA G+C content was 70-72 mol%. Comparative 16S rRNA gene sequence studies revealed that strains CIP 101303(T) and CIP 102116 belonged to the genus Microbacterium and that they were related closely to Microbacterium halotolerans. The level of DNA-DNA relatedness showed that the two isolates represented a separate genomic species. Based on phenotypic and genotypic results, it is proposed that strains CIP 101303(T) and CIP 102116 be assigned to a novel species, Microbacterium binotii sp. nov. The type strain is CIP 101303(T) (=DSM 19164(T)).