The bimolecular reactive transport in heterogeneous porous media: sub-diffusion in interpretation of laboratory experiment.

This present work consists of investigating the effects of particle size heterogeneity and flow rates on transport-reaction kinetics of CuSO4 and Na2EDTA2- in porous media, via the combination of a bimolecular reaction experiment and model simulations. In the early stages of transport, a peak is observed in the concentration breakthrough curve of the reactant CuSO4, related to the delayed mixing and reaction of the reactants. The numerical results show that an increase in flow rate promotes the mixing processes between the reactants, resulting in a larger peak concentration and a slighter tail of breakthrough curves, while an increase in medium heterogeneity leads to a more significant heavy tail. The apparent anomalous diffusion and heavy-tailing behavior can be effectively quantified by a novel truncated fractional derivative bimolecular reaction model. The truncated fractional-order model, taking into account the incomplete mixing, offers a satisfactory reproduction of the experimental data.

Negativicoccus succinicivorans gen. nov., sp. nov., isolated from human clinical samples, emended description of the family Veillonellaceae and description of Negativicutes classis nov., Selenomonadales ord. nov. and Acidaminococcaceae fam. nov. in the bacterial phylum Firmicutes.

Three strains of a hitherto unknown, Gram-negative, tiny, anaerobic coccus were collected from human clinical samples originating from skin and soft tissues. The three isolates displayed at least 99.9 % identity in their 16S rRNA gene sequences and more than 99.8 % identity in their dnaK gene sequences. The isolates were affiliated to the family Veillonellaceae, the coccobacillus Dialister micraerophilus being the most closely related species, but there was no more than 91.1 % identity in the 16S rRNA gene sequence between this species and the three isolates. Phylogeny based on the 16S rRNA gene confirmed that the three strains represent a novel and robust lineage within the current family Veillonellaceae. A similar genomic structure was demonstrated for the three isolates by PFGE-based analysis. Morphology and metabolic end products, as well as genotypic and phylogenetic data supported the proposal of the novel genus Negativicoccus gen. nov., with the novel species Negativicoccus succinicivorans sp. nov. [type strain ADV 07/08/06-B-1388(T) (=AIP 149.07(T)=CIP 109806(T)=DSM 21255(T)=CCUG 56017(T)) as type species]. Phylogenetic analyses based on the 16S rRNA gene sequences of members of the phylum Firmicutes and other phyla indicated that the family Veillonellaceae forms a robust lineage clearly separated from those of the classes 'Bacilli', 'Clostridia', Thermolithobacteria and 'Erysipelotrichi' in the phylum Firmicutes. Therefore, we propose that this family is a class-level taxon in the phylum Firmicutes, for which the name Negativicutes classis nov. is proposed, based on the Gram-negative type of cell wall of its members, with the type order Selenomonadales ord. nov. In this order, a novel family, Acidaminococcaceae fam. nov., is proposed and description of the family Veillonellaceae is emended.

Proposal to unify Clostridium orbiscindens Winter et al. 1991 and Eubacterium plautii (Séguin 1928) Hofstad and Aasjord 1982, with description of Flavonifractor plautii gen. nov., comb. nov., and reassignment of Bacteroides capillosus to Pseudoflavonifractor capillosus gen. nov., comb. nov.

We isolated several strains from various clinical samples (five samples of blood, four of intra-abdominal pus and one of infected soft tissue) that were anaerobic, motile or non-motile and Gram-positive rods. Some of the strains formed spores. Phylogenetic analysis of the 16S rRNA gene sequence showed that these organisms could be placed within clostridial cluster IV as defined by Collins et al. [(1994). Int J Syst Bacteriol 44, 812-826] and shared more than 99 % sequence similarity with Clostridium orbiscindens DSM 6740(T) and Eubacterium plautii DSM 4000(T). Together, they formed a distinct cluster, with Bacteroides capillosus ATCC 29799(T) branching off from this line of descent with sequence similarities of 97.1-97.4 %. The next nearest neighbours of these organisms were Clostridium viride, Oscillibacter valericigenes, Papillibacter cinnamivorans and Sporobacter termitidis, with sequence similarities to the respective type strains of 93.1-93.4, 91.2-91.4, 89.8-90 and 88.7-89.3 %. On the basis of biochemical properties, phylogenetic position, DNA G+C content and DNA-DNA hybridization, it is proposed to unify Clostridium orbiscindens and Eubacterium plautii in a new genus as Flavonifractor plautii gen. nov., comb. nov., with the type strain Prévot S1(T) (=ATCC 29863(T) =VPI 0310(T) =DSM 4000(T)), and to reassign Bacteroides capillosus to Pseudoflavonifractor capillosus gen. nov., comb. nov., with the type strain CCUG 15402A(T) (=ATCC 29799(T) =VPI R2-29-1(T)).

Jonquetella anthropi gen. nov., sp. nov., the first member of the candidate phylum 'Synergistetes' isolated from man.

Six clinical isolates of a hitherto unknown, strictly anaerobic, Gram-negative rod showing fastidious growth were subjected to a polyphasic taxonomic study, including phenotypic, genomic and phylogenetic feature analyses. 16S rRNA gene sequenced-based phylogeny revealed that the novel strains represent a homogeneous group distant from any recognized species in the candidate phylum 'Synergistetes'. The novel isolates were most closely related to species of the genus Dethiosulfovibrio, with 88.2-88.7 % 16S rRNA gene sequence similarity. Large-scale chromosome structure and DNA G+C content also differentiated the novel strains from members of the genus Dethiosulfovibrio. The novel strains were asaccharolytic. Major metabolic end products in trypticase/glucose/yeast extract broth were acetic, lactic, succinic and isovaleric acids and the major cellular fatty acids iso-C(15 : 0) and C(16 : 0). Based on the data presented here, a new genus, Jonquetella gen. nov., is proposed with one novel species, Jonquetella anthropi sp. nov. J. anthropi is the first characterized species of the candidate phylum 'Synergistetes' that includes human isolates. The G+C content of the DNA of the type strain of J. anthropi ADV 126(T) (=AIP 136.05(T)=CIP 109408(T)=CCUG 53819(T)) is 59.4 mol%.

Veillonella denticariosi sp. nov., isolated from human carious dentine.

Selective culture of human carious dentine for Veillonella strains resulted in the isolation of two strains of a Gram-negative, coccus-shaped bacterium that has not been described previously. Comparative 16S rRNA and dnaK gene sequence analysis indicated that the two strains were homogeneous and comprised a distinct lineage within the genus Veillonella, phylogenetically most closely related to Veillonella rodentium. This was supported by DNA-DNA hybridization, which showed clearly that the two strains were similar and distinct from other Veillonella species, and the production of major cellular fatty acids (C(13 : 0) and C(17 : 1)omega8), which is consistent with other members of the genus Veillonella. Based on these observations, strains RBV81 and RBV106(T) represent a novel species, for which the name Veillonella denticariosi sp. nov. is proposed, with the type strain RBV106(T) (=CIP 109448(T) =CCUG 54362(T) =DSM 19009(T)).

Acidaminococcus intestini sp. nov., isolated from human clinical samples.

Eleven strains of a hitherto unknown, Gram-negative, anaerobic coccus were recovered from various human clinical samples of patients hospitalized in two geographically distant French hospitals. These strains displayed the morphology and growth characteristics of those related to the genus Acidaminococcus. The clinical isolates shared at least 99.9 and 99.7 % of their nucleotide positions in the 16S and 23S rRNA gene sequences, respectively. They displayed 95.6 and 88.9 % 16S and 23S rRNA gene sequence similarities, respectively, with Acidaminococcus fermentans. The 16S rRNA-based phylogeny revealed that all the clinical isolates grouped in a statistically well supported cluster separate from A. fermentans. Enzymic activity profiles as well as metabolic end product patterns, including propionic acid production, differentiated the novel bacteria from A. fermentans. Finally, phenotypic, genotypic and phylogenetic data, including large-scale chromosome structure and DNA G+C content, supported the proposal of a novel species of the genus Acidaminococcus, for which the name Acidaminococcus intestini sp. nov. is proposed. The type strain is ADV 255.99(T) (=AIP 283.01(T)=CIP 108586(T)=CCUG 50930(T)).

Moryella indoligenes gen. nov., sp. nov., an anaerobic bacterium isolated from clinical specimens.

Three Gram-positive, anaerobic, non-spore-forming, rod-shaped bacteria with pointed ends were isolated from clinical specimens. The organisms were weakly saccharolytic and produced indole, acetate, butyrate and lactate as major metabolic end products. 16S rRNA gene sequence analysis indicated that the isolates had no known close relatives among recognized bacteria but that they exhibited a phylogenetic association with Clostridium rRNA cluster XIVa [as defined by Collins, M. D. et al. (1994). Int J Syst Bacteriol 44, 812-826]. The closest recognized relatives were the type strains of Clostridium clostridioforme, Clostridium bolteae and Clostridium asparagiforme (16S rRNA gene sequence similarity values of 90.2-91.4 %). These results suggest that these three clinical isolates represent a novel species of a new genus, for which the name Moryella indoligenes gen. nov., sp. nov. is proposed. The type strain of Moryella indoligenes is AIP 220.04(T) (=CIP 109174(T)=CCUG 52648(T)).

Surgical wound infection associated with Psychrobacter phenylpyruvicus-like organism.

A case of surgical wound infection caused by Psychrobacter phenylpyruvicus-like organism is described. The strain showed phenotypic characteristics typical of P. phenylpyruvicus, but 16S rRNA sequencing showed 98.2% relatedness to Moraxella phenylpyruvica strain 752/52 and only 94.8% with P. phenylpyruvicus type strain ATCC 23333(T). The results of molecular analysis suggest that the strain we isolated may represent a new species within the genus Psychrobacter.

Contribution of C. beijerinckii and C. sporogenes in association with C. tyrobutyricum to the butyric fermentation in Emmental type cheese.

The relationship between C. tyrobutyricum, C. sporogenes and C. beijerinckii in experimental cheese conditions, and their influences on late-blowing and butyric fermentation, have been investigated. A molecular approach using a PCR-TTGE method in combination with conventional methods, such as microbiological and physico-chemical analysis, was performed to monitor the evolution of these clostridial species, simultaneously with the occurrence of cheese defects. Sixteen Emmental type cheeses were produced from milk inoculated with different clostridial spore associations. In all cheeses inoculated with C. tyrobutyricum, obvious signs of late blowing were detected. In cheeses inoculated with C. beijerinckii or C. sporogenes, a formation of holes in cheese body was observed, with a concomitant slight amount of butyric acid production. Even though C. beijerinckii and C. sporogenes were less metabolically active and less numerically important than C. tyrobutyricum in cheese as shown by TTGE profiles, the association of these species to C. tyrobutyricum enhanced the butyric fermentation and the cheese defects. The level of butyric content in ripened cheese increased to 268 mg 100 g(-1) in presence of C. tyrobutyricum, and reached a maximum of 414 mg 100 g(-1) in presence of the C. beijerinckii-C. tyrobutyricum (1:10) association. The propionic fermentation was also higher in cheese inoculated with C. tyrobutyricum, and was slowed down in presence of C. beijerinckii and C. sporogenes. From 30 days of ripening, a strong correlation between the chemical contents and the intensity of cheese defects was demonstrated. A chemical analysis of cheese associated with a molecular method for microbial spoilage investigation allows the prediction of the level of late blowing at early stages of ripening, and the understanding of the origin of the defect.

First isolation of Clostridium amygdalinum from a patient with chronic osteitis.

We describe a case of osteitis caused by a new and unusual Clostridium species, Clostridium amygdalinum, an environmental, moderately thermophilic bacterium. This is the first documented report of human infection caused by this organism.

Isolation from canned foods of a novel Thermoanaerobacter species phylogenetically related to Thermoanaerobacter mathranii (Larsen 1997): emendation of the species description and proposal of Thermoanaerobacter mathranii subsp. Alimentarius subsp. Nov.

Several anaerobic, thermophilic, Gram-positive bacteria were isolated from dairy products and canned meats. While some isolates were identified as Thermoanaerobacter thermohydrosulfuricus, comparisons of 16S rDNA genes indicated that others were phylogenetically closely related to Thermoanaerobacter mathranii, and more distantly related to Thermoanaerobacter thermocopriae and Thermoanaerobacter italicus. Biochemical characteristics, phylogenetic analysis, G+C content, and DNA-DNA hybridization experiments demonstrated that the strains AIP 504.99, AIP 505.99T and AIP 431.03, notwithstanding their high sequence similarities differ from T. mathranii and represent a novel T. mathranii subspecies for which the name T. mathranii subsp. alimentarius is proposed. The type strain is strain AIP 505.99T = CIP 108280T = CCUG 49566T. Emendation of the species description for T. mathranii is proposed to include this subspecies.

Phenotypic and genotypic characterization of some Moorella sp. strains isolated from canned foods.

Six anaerobic thermophilic strains isolated from various spoiled cans including fish soups and cooked meats were characterized using a polyphasic approach. These strains were closely related to Moorella thermoacetica or Moorella thermoautotrophica species. Except the spacer region between the 16S and the 23S rRNA genes, which exhibited two PCR profiles distinguishing both species, the genotypic and phylogenetic analyses grouped these isolates, the type strains, and all sequences of Moorella thermoacetica and Moorella thermoautotrophica species contained in the GenBank database within a unique cluster. Moreover, all 16S rDNA sequences shared two characteristic DNA fragments, which were highly specific of Moorella thermoacetica/Moorella thermoautotrophica strains. However, taken together, the phenotypic, physiological and genotypic methods were conflicting, and did not enable affiliation of the isolates with one or the other species. To our knowledge, this study represents the first report of characterization of Moorella species isolated from spoiled cans. These results and previous work, very strongly argue in favor of questioning the taxonomic status of the two species.

Dialister micraerophilus sp. nov. and Dialister propionicifaciens sp. nov., isolated from human clinical samples.

Seventeen anaerobic, Gram-negative, tiny coccobacilli were collected in France from various human clinical samples. Biochemical analyses as well as molecular studies, including 16S rRNA and dnaK gene sequencing, affiliated all the isolates to the genus Dialister. However, 16S rRNA and dnaK gene sequence similarities were below 95.2 and 79.7 %, respectively, when comparisons were performed with the currently described species Dialister pneumosintes and Dialister invisus. Two clusters consisting of 13 and four isolates could be differentiated. 16S rRNA- and dnaK-based phylogeny confirmed that these two clusters represent two novel and distinct lineages within the genus Dialister. Finally, phenotypic, genotypic and phylogenetic data supported the proposal of the two novel species Dialister micraerophilus sp. nov. (type strain ADV 04.01T=AIP 25.04T=CIP 108278T=CCUG 48837T) and Dialister propionicifaciens sp. nov. (type strain ADV 1053.03T=AIP 26.04T=CIP 108336T=CCUG 49291T). The G+C content of the DNA of the D. micraerophilus type strain is 36.3 mol%. On the basis of 16S rRNA gene sequence analysis, 11 isolates originating from Canada could also be affiliated to D. micraerophilus sp. nov., and were included in the species description.

Bacteremia caused by a metronidazole-resistant Prevotella sp. strain.

Metronidazole resistance among Prevotella spp. is rare. We report here the first case of bacteremia due to a high-level metronidazole-resistant Prevotella sp. responsible for treatment failure.

Brain abscess due to Actinobacillus actinomycetemcomitans.

Actinobacillus actinomycetemcomitans, a constituent of the oral flora, is a rare cause of brain abscesses. We report the case of a 47-year-old male who presented with multiple brain abscesses due to this organism, presumably originating from his poor dentition. Problems met in isolating and identifying A. actinomycetemcomitans suggest that its true rate of isolation from non-oral samples may have been underestimated.

Development and validation of PCR primers to assess the diversity of Clostridium spp. in cheese by temporal temperature gradient gel electrophoresis.

A nested-PCR temporal temperature gradient gel electrophoresis (TTGE) approach was developed for the detection of bacteria belonging to phylogenetic cluster I of the genus Clostridium (the largest clostridial group, which represents 25% of the currently cultured clostridial species) in cheese suspected of late blowing. Primers were designed based on the 16S rRNA gene sequence, and the specificity was confirmed in PCRs performed with DNAs from cluster I and non-cluster I species as the templates. TTGE profiles of the PCR products, comprising the V5-V6 region of the 16S rRNA gene, allowed us to distinguish the majority of cluster I species. PCR-TTGE was applied to analyze commercial cheeses with defects. All cheeses gave a signal after nested PCR, and on the basis of band comigration with TTGE profiles of reference strains, all the bands could be assigned to a clostridial species. The direct identification of Clostridium spp. was confirmed by sequencing of excised bands. C. tyrobutyricum and C. beijerinckii contaminated 15 and 14 of the 20 cheese samples tested, respectively, and C. butyricum and C. sporogenes were detected in one cheese sample. Most-probable-number counts and volatile fatty acid were determined for comparison purposes. Results obtained were in agreement, but only two species, C. tyrobutyricum and C. sporogenes, could be isolated by the plating method. In all cheeses with a high amount of butyric acid (>100 mg/100 g), the presence of C. tyrobutyricum DNA was confirmed by PCR-TTGE, suggesting the involvement of this species in butyric acid fermentation. These results demonstrated the efficacy of the PCR-TTGE method to identify Clostridium in cheeses. The sensitivity of the method was estimated to be 100 CFU/g.

Oribacterium sinus gen. nov., sp. nov., within the family 'Lachnospiraceae' (phylum Firmicutes).

A hitherto unknown anaerobic bacillus isolated from sinus pus in a young child (strain AIP 354.02T) was characterized by using phenotypic and genotypic methods. 16S rRNA gene sequence analysis indicated that this strain was phylogenetically affiliated with several sequences of cloned 16S rRNA gene inserts previously deposited in the public databases. According to their 16S rRNA gene sequence similarities, these uncultivated bacteria, together with strain AIP 354.02T, formed a separate subgroup belonging to the family 'Lachnospiraceae' within the phylum Firmicutes. Oribacterium gen. nov. is proposed for this group of organisms and Oribacterium sinus gen. nov. sp. nov. for strain AIP 354.02T (= CIP 107991T = CCUG 48084T).

Veillonella montpellierensis sp. nov., a novel, anaerobic, Gram-negative coccus isolated from human clinical samples.

Three strains of a hitherto unknown, Gram-negative, anaerobic coccus were isolated from human samples. At the phenotypic level, the isolates displayed all the characteristics of bacteria belonging to the genus Veillonella. Sequence analysis revealed that the three strains shared >99.5% similarity in 16S rDNA sequence and >98.4% similarity in dnaK sequence. The three unknown strains formed a separate subclade that was clearly remote from Veillonella species of human and animal origin. Based on these results, the three strains were considered to represent a novel species within the genus Veillonella, for which the name Veillonella montpellierensis is proposed. The type strain of the species is ADV 281.99T (=CIP 107992T=CCUG 48299T).

Osteosynthesis-associated bone infection caused by a nonproteolytic, nontoxigenic Clostridium botulinum-like strain.

A nonproteolytic, nontoxigenic Clostridium botulinum strain identified by conventional and molecular techniques as type B-, E-, or F-like (BEF-like) was isolated from a human postsurgical wound. All previous reports of such strains have been from environmental sources. Since toxin production is the main taxonomic denominator for C. botulinum, a new name is needed for nonproteolytic, nontoxigenic BEF-like clinical isolates.

Insertional inactivation of the lpxA gene involved in the biosynthesis of lipid A in Neisseria meningitidis resulted in lpxA/lpxA::aph-3' heterodiploids.

The lpxA gene is known to be involved in the biosynthesis of lipid A in Gram-negative bacteria and thought to be an essential gene. However, viable meningococcal lpxA mutants devoid of detectable endotoxin (lipooligosaccharide) have been reported. We characterised such mutants in strains of Neisseria meningitidis belonging to serogroups B and C using molecular and biochemical analysis. While lpxA mutants with no detectable or a low level of lipooligosaccharide could be obtained in N. meningitidis, the simple insertional inactivation of lpxA was not possible. In all mutants, we obtained lpxA/lpxA::aph-3' heterodiploids harbouring one copy of the wild-type lpxA gene and one copy of the inactivated lpxA gene by insertion of the kanamycin resistance cassette, aph-3'. The absence of lipooligosaccharide in these mutants may result from a negative transdominance effect of a truncated LpxA protein on the wild-type LpxA protein.