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Differentiation of Veillonella atypica, Veillonella dispar and Veillonella parvula using restricted fragment-length polymorphism analysis of 16S rDNA amplified by polymerase chain reaction.

Veillonella atypica, Veillonella dispar and Veillonella parvula cannot be reliably distinguished by conventional phenotypic tests, including the API ZYM test. In this study, restricted fragment-length polymorphism (RFLP) analysis of 16S rDNA amplified by polymerase chain reaction (PCR) was used to generate restriction profiles of the type strains of V. atypica, V. dispar and V. parvula and 20 Veillonella strains isolated from oral sites. 16S rRNA gene sequences from isolated genomic DNA samples were amplified by PCR. PCR products were purified and characterized by single digestion with 13 restriction endonucleases. Among them, Mn/I was found to discriminate the respective reference strains, and the clinical isolates were assigned to one of the three species on the basis of their restriction profiles by digestion with Mn/I. Thus, RFLP analysis of PCR-amplified 16S rDNA, using Mn/I, is a rapid and reliable method for the differentiation of V. atypica, V. dispar and V. parvula.

Base Sequence↗

Comparison of batch culture growth and fermentation of a poultry Veillonella isolate and selected Veillonella species grown in a defined medium.

The objective of this study was to develop a defined medium for quantitating nutritional requirements and fermentation products of a poultry cecal isolate of Veillonella and to compare these parameters with representative Veillonella species. The poultry isolate is one of 29 organisms from a continuous-flow culture that has been shown to be effective against Salmonella colonization in broilers. When the Veillonella species were grown in anaerobic batch culture, propionate and acetate were the only volatile fatty acids detected. Lactate was needed to provide energy for the growth of the Veillonella in the defined medium. The poultry isolate had significantly (p< 0.05) higher Y(lactate)(g of dry cell weight per mole of lactate utilized) and dry cell weight than the other Veillonella species when grown on amino acid supplemented defined media. Cultures of the Veillonella species in the defined medium grown with supplemented amino acids aspartate, threonine, arginine, and serine indicated that these amino acids were metabolized to acetate and propionate. Amino acid analysis on media inoculated with either V. atypica or the poultry isolate also indicated that these organisms may have different amino acid preferences. For nearly all of the amino acid supplemented media combinations the poultry isolate utilized significantly (p< 0.05) more threonine and serine whereas V. atypica utilized significantly (p< 0.05) more aspartate. The defined medium supported growth of all of the Veillonella species tested and should enable further in-depth physiological studies to be conducted on the poultry Veillonella studies.

Journal Article↗

Molecular identification of the first human isolate belonging to the Veillonella ratti-Veillonella criceti group based on 16S rDNA and dnaK gene sequencing.

Anaerobic gram-negative cocci belonging to the genus Veillonella are currently subdivided into eight species. Among them, four have been isolated either from human flora or from clinical samples during infectious processes: Veillonella atypica, V. dispar, V. parvula and V. montpellierensis. To date, the four other species have only been found in animals. In particular, V. ratti and V. criceti, since their characterization, have been exclusively reported from mouth and intestine of rodents. We report here for the first time a human isolate belonging to the V. ratti-V. criceti group recovered in mixed aerobic-anaerobic flora from a semen sample in a 24-year-old man attending the urology unit of our hospital for infertility. Identification of the isolate was based on 16S ribosomal DNA and dnaK gene sequencing. Since then, two strains of Veillonella sp. recovered from semen samples remained unidentified to the species level, and 16S rDNA-based phylogenetic analysis revealed that they might represent novel taxa within the genus Veillonella. Taken together, these observations suggest that host restriction may not exist in the genus Veillonella and that bacterial diversity remains underestimated both within this genus and in human semen.

Adult↗

Veillonella parvula meningitis: case report and review of Veillonella infections.

Veillonella parvula is a small, nonfermentative anaerobic gram-negative coccus that is part of the normal flora of the mouth, gastrointestinal tract, and vagina in humans. When isolated from clinical specimens, V. parvula is often regarded as a contaminant or commensal, but it has been implicated as a pathogen in infections of the sinuses, lungs, heart, bone, and central nervous system. Meningitis, however, is extremely rare; to our knowledge, only 2 cases have been previously described in the literature. We report a case of V. parvula meningitis and review the literature on Veillonella infections.

Female↗

Putrescine and cadaverine are constituents of peptidoglycan in Veillonella alcalescens and Veillonella parvula.

Veillonella alcalescens ATCC 17745, a strictly anaerobic, gram-negative small coccus, requires putrescine or cadaverine for growth (M. B. Ritchey, and E. A. Delwiche, J. Bacteriol. 124:1213-1219, 1975). Both putrescine and cadaverine were demonstrated to be incorporated exclusively into the peptidoglycan layer of V. alcalescens ATCC 17745. V. parvula GAI 0574 also proved to contain putrescine as a component of peptidoglycan. The primary chemical structure of the peptidoglycan common to the two Veillonella species is N-acetylglucosamine-N-acetylmuramic acid-L-alanine-D-glutamic acid gamma-meso-diaminopimelic acid-D-alanine. Putrescine or cadaverine links covalently to the alpha-carboxyl group of the D-glutamic acid residue of the peptidoglycan is necessary for normal cell growth. In V. alcalescens ATCC 17745, above 40% saturation at cadaverine linked to the alpha-carboxyl group of the D-glutamic acid residue of the peptidoglycan is necessary for normal growth.

Cadaverine↗

Purification and characterization of bacteriophage receptor on Veillonella rodentium cells. Phage-receptor on Veillonella.

Veillonellophage N2 adsorbed to polysaccharides (PSs) on Veillonella rodentium ATCC 17743 cell wall, and the bacteriophage receptor contained only glucosamine. D(+)-glucosamine hydrochloride (Sigma) also adsorbed the veillonellophage N2. These results therefore indicate that the receptor to the veillonellophage N2 is cell wall PSs. The PSs of the host cells as receptor have been characterized. Glucosamine accounted for approximately 100% of the weight of the PSs. The PSs which were partially resolved by Sephadex G-75 chromatography comprised approximately four glucosamine units. Their primary structure was determined by 400 MHz n.m.r. spectroscopy. One- and two-dimensional 1H-nmr experiments showed the PS to be a branched polymer. Glucosamine linkage was detected in one of the branches.

Bacteriophages↗

Ability of Veillonella and Neisseria species to attach to oral surfaces and their proportions present indigenously.

The present study describes the distribution of Veillonella and Neisseria species in the human oral cavity and indicates that their ability to attach to oral surfaces correlates with their proportions found in various sites of the mouth. The mean percentages of Veillonella and Neisseria of the total flora cultivable on anaerobic blood-agar plates was found to be: plaque, 0.75 and <0.13, respectively; lip, 0.38 and <0.05; cheek, 0.66 and <0.14; tongue dorsum, 9.4 and <0.12; saliva, 5.0 and <0.9. The ability of Veillonella and Neisseria species to attach to tooth surfaces was studied by cleaning the labial surfaces of incisors to render them relatively free of viable bacteria. Samples taken 1 hr later contained <0.27% Veillonella and <0.4% Neisseria, whereas saliva to which these teeth were exposed contained 20-fold higher proportions of Veillonella. These data indicate that Veillonella and Neisseria species possess a feeble ability to attach to cleaned teeth. The ability of these organisms to adhere to other oral surfaces was determined by introducing mixtures of streptomycin-resistant strains into the mouths of volunteers for 5 min. Labeled strains of Streptococcus sanguis and S. salivarius were included for comparative purposes. Analysis of samples obtained from oral surfaces after 45 min indicated that Veillonella and Neisseria adhere very poorly to preformed dental plaque as compared to S. sanguis. In contrast, Veillonella adhered to the tongue dorsum markedly better than Neisseria, S. sanguis, and S. salivarius. The greater ability of Veillonella to adhere to the tongue in relation to the other organisms studied correlates with the high proportions of Veillonella found on this site. The feeble ability of Neisseria to attach to surfaces in the oral cavity is reflected by their low proportions found on these surfaces.

Adult↗

The Genus Veillonella IV. Serological Groupings, and Genus and Species Emendations.

Rogosa, M. (National Institute of Dental Research, Bethesda, Md.). The genus Veillonella. IV. Serological groupings, and genus and species emendations. J. Bacteriol. 90:704-709. 1965.-Seven serological groups of Veillonella were found. The generic and species descriptions are enlarged and corrected. The type species is Veillonella parvula (Veillon and Zuber) Prévot, 1933. The proposed species, subspecies (infraspecific serogroups), and American Type Culture Collection (ATCC) type strains are: Veillonella parvula subsp. parvula subsp. nov. (antigenic group VI), ATCC 10790; Veillonella parvula subsp. rodentium subsp. nov. (antigenic group II), ATCC 17743; Veillonella parvula subsp. atypica subsp. nov. (antigenic group V), ATCC 17744; Veillonella alcalescens subsp. alcalescens subsp. nov. (antigenic group IV), ATCC 17745; Veillonella alcalescens subsp. ratti subsp. nov. (antigenic group III), ATCC 17746; Veillonella alcalescens subsp. criceti subsp. nov. (antigenic group I), ATCC 17747; and Veillonella alcalescens subsp. dispar subsp. nov. (antigenic group VII), ATCC 17748. The following species listed in Bergey's Manual (7th ed.) are rejected as members of the genus: V. discoides, V. reniformis, V. orbiculus, and V. vulvovaginitidis.

Journal Article↗

Intra-chromosomal heterogeneity between the four 16S rRNA gene copies in the genus Veillonella: implications for phylogeny and taxonomy.

Among the seven species characterized within the genus Veillonella, three (Veillonella dispar, Veillonella parvula and Veillonella atypica) have so far been isolated from human flora and during infectious processes. Sequencing and analysis of 16S rDNA (rrs) has been described as the best method for identification of Veillonella strains at the species level since phenotypic characteristics are unable to differentiate between species. rrs sequencing for the three species isolated from humans showed more than 98 % identity between them. Four rrs copies were found in the reference strains and in all the clinical isolates studied. The sequences of each rrs were determined for the clinical strain ADV 360.1, and they showed a relatively high level of heterogeneity (1.43 %). In the majority of cases, polymorphic positions corresponded to nucleotides allowing differentiation between the three species isolated from humans. Moreover, variability observed between rrs copies was higher than that between 16S rDNA sequences of V. parvula and V. dispar. Phylogenetic analysis showed that polymorphism between rrs copies affected the position of strain ADV 360.1 in the tree. Variable positions occurred in stems and loops belonging to variable and hypervariable regions of the 16S rRNA secondary structure but did not change the overall structure of the 16S rRNA. PCR-RFLP experiments performed on 27 clinical isolates of Veillonella sp. suggested that inter-rrs heterogeneity occurs widely among the members of the genus VEILLONELLA: These results, together with the lack of phenotypic criteria for species differentiation, give preliminary arguments for unification of V. dispar and V. parvula.

Base Sequence↗

[Recovery of Veillonella from saliva].

Veillonella spp. are anaerobic gram-negative cocci associated to oral health. Different types of cultures have been reported for the isolation of these microorganisms. Veillonella spp. colonies produce a red fluorescence, which is made visible through ultraviolet light and disappears in contact with oxygen. This feature would be very useful for rapid presumptive identification. The aims of this study were: 1. to compare the Rogosa selective medium for Veillonella with the cultures recommended by different authors in order to determine best saliva recovery, since this sample is generally used to determine the presence and predominance of this bacteria; 2. to detect red fluorescence production on these different culture media as a rapid method for identification. Selective medium for Veillonella, Schaedler agar for anaerobic bacteria with vitamin K, thioglycollate agar, brain heart infusion agar, Brucella agar, trypticase soy agar, and Columbia agar, all of them with and without the addition of vancomycin, and laked blood were used for this study. The tested sample was a saliva pool. Both, Veillonella colonies, and the total number of microorganisms were counted, and expressed as CFU/ml of saliva. The greatest Veillonella recovery in saliva was obtained with the selective medium for Veillonella with vancomycin and laked blood. The production of fluorescence was only observed in this medium.

Culture Media↗

Coaggregation properties of human oral Veillonella spp.: relationship to colonization site and oral ecology.

The primary habitats of oral veillonellae are the tongue, dental plaque, and the buccal mucosa. Isolates were obtained from each habitat and tested for coaggregation with a battery of other oral bacterial strains. All 59 tongue isolates tested for coaggregation were Veillonella atypica or Veillonella dispar. All but one of them coaggregated with strains of Streptococcus salivarius, a predominant inhabitant of the tongue surface but not subgingival dental plaque. These tongue isolates were unable to coaggregate with most normal members of the subgingival flora such as Actinomyces viscosus, Actinomyces naeslundii, Actinomyces israelii, and Streptococcus sanguis. In contrast, 24 of 29 Veillonella isolates, of which 20 were Veillonella parvula from subgingival dental plaque samples, coaggregated strongly with the three species of Actinomyces, S. sanguis, and other bacteria usually present in subgingival plaque, but they did not coaggregate with S. salivarius. The majority of isolates from the buccal mucosa (42 of 55) has coaggregation properties like those from the tongue. These results indicate that the three human oral Veillonella species are distributed on oral surfaces that are also occupied by their coaggregation partners and thus provide strong evidence that coaggregation plays a critical role in the bacterial ecology of the oral cavity.

Actinomyces↗

Synergism of lactate and succinate as metabolites utilized by Veillonella to inhibit the growth of Salmonella typhimurium and Salmonella enteritidis in vitro.

The inhibition of salmonellae growth by a Veillonella bacterium isolated from the cecal contents of adult chickens was examined. The Veillonella isolate was grown on an agar medium supplemented with 175 mumol of lactate or succinate/ml. Either 0, 100, 125, 150, or 175 mumol of succinate/ml was added to the lactate medium; either 0, 100, 125, 150, or 175 mumol of lactate/ml was added to the succinate medium; and the pH of all media was adjusted to 6.0. Agar overlays of Veillonella cultures grown on the media were inoculated with Salmonella typhimurium or S. enteritidis. The largest zones of inhibition of salmonellae growth were produced by Veillonella cultures grown on medium supplemented with 175 mumol/ml of both lactate and succinate. The widths of the zones of inhibition decreased as the concentration of lactate was reduced in the succinate medium and as the concentration of succinate was reduced in the lactate medium. Analyses of lactate broth and succinate broth inoculated with Veillonella indicated that inhibition of salmonellae growth on the agar media was related to the production of volatile fatty acids by Veillonella, the presence of residual succinate in the media, and the final pH of the media.

Analysis of Variance↗

Age-related frequency of penicillin resistance of oral Veillonella.

Veillonella spp. are early colonizing inhabitants in the mouth. As part of studies on penicillin resistance among oral indigenous anaerobic microbiota in childhood, the aim of the present longitudinal study was to examine the emergence of resistant strains in Veillonella populations. Altogether 305 Veillonella isolates from saliva of 49 healthy infants followed from 2 to 24 months of age were examined for their in vitro susceptibility to penicillin G and, further, 20 penicillin-resistant isolates representing 5 MIC categories to ampicillin, amoxicillin, amoxicillin/clavulanate, cefoxitin, and beta-lactamase production. In infants positive for oral Veillonella, the recovery rate of penicillin-resistant (MIC >/=2 microg/ml) strains increased with age up to 68%, however, most infants simultaneously harbored penicillin-susceptible strains. During the follow-up, the MIC(50) increased from 0.5 microg/ml to 2 microg/ml. In addition to penicillin G, 8/20 strains also showed reduced susceptibility to ampicillin and/or amoxicillin but none produced beta-lactamase. Our study suggests other mechanisms than enzymatic degradation of beta-lactam ring for resistance of oral Veillonella to penicillin.

Age Factors↗

Production and characterization of monoclonal antibodies to a Veillonella species from a continuous-flow culture system of chicken cecal bacteria.

Administering native intestinal flora to newly hatched chicks protects against cecal Salmonella colonization, and is known as competitive exclusion. Continuous-flow culture systems have been used to maintain defined competitive exclusion cultures. We have recently demonstrated that such a stable continuous-flow culture, CF3, contains 29 bacterial strains representing ten genera. Broiler chicks treated with CF3 are protected against Salmonella colonization of the ceca. Such protection is correlated with elevated concentrations of proprionic acid in the cecal contents of treated chicks. In this study we report on the preparation and characterization of monoclonal antibodies to one of the proprionic acid producing anaerobes contained in CF3, namely Veillonella CF3. Five different monoclonal antibodies were characterized with respect to: (1) isotype; (2)Veillonella specificity as judged by cross-reactivity profiles with other bacteria; (3) sensitivity as measured by the limit of detection of the number of colony forming units of Veillonella; and (4) antigen recognition of Veillonella by Western Blot analysis. These antibodies have been used to enumerate Veillonella in both the CF3 cultures and in the ceca of young chicks.

Journal Article↗

Research note: in vitro inhibition of Listeria monocytogenes growth by veillonellae cultures grown on tartrate media.

Veillonellae cultures were grown on agar media supplemented with tartrate and examined for inhibitory effects on the growth of Listeria monocytogenes. Veillonellae cultures grown on media supplemented with 0 or 50 mmol l-1 of tartrate did not inhibit the growth of L. monocytogenes; however, veillonellae grown on media supplemented with 100, 150 or 200 mmol l-1 of tartrate did inhibit the growth of L. monocytogenes. The inhibition of the growth of L. monocytogenes by the veillonellae was correlated with the increased production of acetate and propionate from tartrate by veillonellae and with the reduction of the pH of the media by L. monocytogenes.

Bacteriological Techniques↗

THE GENUS VEILLONELLA. I. GENERAL CULTURAL, ECOLOGICAL, AND BIOCHEMICAL CONSIDERATIONS.

Rogosa, M. (National Institutes of Health, Bethesda, Md.). The genus Veillonella. I. General cultural, ecological, and biochemical considerations. J. Bacteriol. 87:162-170. 1964.-Arguments are presented for excluding Veillonella discoides, V. reniformis, V. orbiculus, and V. vulvovaginitidis from the genus, and for restricting it to aerogenic organisms such as V. parvula and V. alcalescens. The genus Veillonella thus would comprise species which are anaerobic and nonmotile; are small, spherical, gram-negative cocci appearing as pairs, masses, and short chains; and are cytochrome-oxidase- and benzidine-negative. Veillonella would be further characterized in that glucose or any other carbohydrate is not fermented; indole is not produced; gelatin is not liquefied; nitrate is reduced; H(2)S is produced; propionic and acetic acids, CO(2), and H(2) are produced from lactate during growth; and pyruvic, oxaloacetic, malic, fumaric, and succinic acids are metabolized by resting cells, but citric, isocitric, and malonic acids are not. In addition to the above, a number of cultural, ecological, and biochemical characteristics are described. At present, V. parvula (the type species) and V. alcalescens would be retained as valid species. Errors in the descriptions of V. parvula and V. alcalescens are corrected by amended statements. These species are differentiated serologically. Also, V. alcalescens differs from V. parvula in having an absolute requirement for putrescine or cadaverine and in decomposing H(2)O(2).

Cadaverine↗