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Clinicopathologic considerations of fusobacteria chorioamnionitis.

When sought by light microscopy in formalin-fixed tissue, fusobacteria are apparent in 7-18% of chorioamnionitis cases. Brown and Hopps bacterial staining characteristically visualizes their long, slender and filamentous forms. Fusobacteria-like organisms in placentae have previously been associated with prematurity. Our findings indicate that perinatal infection with fusobacteria may cause neonatal death from prematurity, rather than from sepsis. In a study of 586 placentae, 14 specimens with chorioamnionitis and fusobacteria-like organisms were found. On the five occasions when microbiologic tests were made, fusobacteria were isolated. Prematurity of the newborns was associated with twelve of those placentae. Immunofluorescent labelling of the organisms is a helpful but incomplete means of diagnosis. Because fusobacteria antibodies are highly specific, they do not cross-react with other members of the genus Fusobacterium. A typical case is reported, in detail.

Adult↗

Beta-lactamase production and susceptibilities to amoxicillin, amoxicillin-clavulanate, ticarcillin, ticarcillin-clavulanate, cefoxitin, imipenem, and metronidazole of 320 non-Bacteroides fragilis Bacteroides isolates and 129 fusobacteria from 28 U.S. centers.

beta-Lactamase production (nitrocefin disk method) and agar dilution susceptibility of amoxicillin, amoxicillin-clavulanate, ticarcillin, ticarcillin-clavulanate, cefoxitin, imipenem, and metronidazole were determined for 320 Bacteroides species (not Bacteroides fragilis group) and 129 fusobacteria from 28 U.S. centers. Overall, 64.7% of Bacteroides species and 41.1% of fusobacteria were beta-lactamase positive. Among the Bacteroides species, positivity rates were highest for B. bivius (85.0%), followed by B. splanchnicus (83.3%), B. eggerthii (77.8%), and B. oralis (77.1%); 54.5% of black-pigmented Bacteroides species were beta-lactamase positive. Among the fusobacteria, Fusobacterium mortiferum showed the highest rate of beta-lactamase positivity (76.9%). MICs of amoxicillin (128 micrograms/ml) and ticarcillin (64 micrograms/ml) for 90% of all beta-lactamase-positive strains were reduced to 4 and 2 micrograms/ml, respectively, with the addition of clavulanate. MICs of amoxicillin and ticarcillin for 90% of all beta-lactamase-negative strains were 1 and 4 micrograms/ml, respectively, and greater than or equal to 98.4% of the strains were susceptible to the beta-lactams tested. Of the beta-lactamase-producing strains, 45.9% were susceptible to amoxicillin at less than or equal to 4 micrograms/ml and 93.4% were susceptible to ticarcillin at less than or equal to 64 micrograms/ml; the addition of clavulanate raised the rates to 90.4 and 100%, respectively. All strains were susceptible to cefoxitin, imipenem, and metronidazole. The activity of amoxicillin against 29 beta-lactamase-producing strains (10 Bacteroides species and 19 fusobacteria) was not enhanced by the addition of clavulanate; however, 82.7% of these strains were susceptible to amoxicillin, and all were susceptible to ticarcillin. Although beta-lactamase positivity is on the increase in non-B. fragilis group Bacteroides species and fusobacteria, amoxicillin-clavulanate, ticarcillin, cefoxitin, imipenem, and metronidazole should be suitable for the treatment of infections with these strains. The addition of clavulanate does not appreciably improve the efficacy of ticarcillin against these organisms.

Amoxicillin↗

Severe Fusobacteria infections (Lemierre syndrome) in two boys.

UNLABELLED: Abscess formation is a rare cause of febrile illness in childhood but always has to be considered in such clinical presentations. Belonging to the resident flora of the oropharyngeal region, Fusobacteria are known to cause local infections; from here they may extend to other sites via the bloodstream or are aspirated into the lung (Lemierre disease). We report on two boys with Lemierre disease due to infection by Fusobacteria in monoculture causing two different clinical phenotypes. Case 1 presented with a large subphrenic abscess and pneumonic infiltration of the right middle lobe. Primary focus of infection was periodontal disease. Case 2 presented with a life-threatening septicaemia due to a retropharyngeal abscess and perforated otitis media followed by osteomyelitis of the atlas and thrombosis of the left sigmoid sinus and internal jugular vein. CONCLUSION: Fusobacteria should be considered in any abscess formation in children. A thorough examination of the oropharyngeal region as a possible site of primary manifestation is mandatory.

Adolescent↗

Fusobacteria: new taxonomy and related diseases.

Fusobacteria are anaerobic gram-negative bacilli. Since the first reports in the late nineteenth century, various names have been applied to these organisms, sometimes with the same name being applied to different species. More recently, not only have there been changes to the nomenclature, but also attempts to differentiate between species which are believed to be either pathogenic or commensal or both. Because of their asaccharolytic nature, and a general paucity of positive results in routine biochemical tests, laboratory identification of the fusobacteria has been difficult. However, the application of novel molecular biological techniques to taxonomy has established a number of new species, together with the subspeciation of Fusobacterium necrophorum and F. nucleatum, and provided new methods for identification. The involvement of fusobacteria in a wide spectrum of human infections causing tissue necrosis and septicaemia has long been recognised, and, more recently, their importance in intra-amniotic infections, premature labour and tropical ulcers has been reported.

Animals↗

Susceptibility of Bacteroides non-fragilis and fusobacteria to amoxicillin, amoxicillin/clavulanate, ticarcillin, ticarcillin/clavulanate, cefoxitin, imipenem and metronidazole.

The susceptibility of 234 Bacteroides non-fragilis strains and 56 fusobacteria from 12 European centers to amoxicillin, amoxicillin/clavulanate, ticarcillin, ticarcillin/clavulanate, cefoxitin, imipenem and metronidazole was tested and related to beta-lactamase production. Beta-lactamase production was detected in 42.3 % of the Bacteroides strains and 26.8% of the fusobacteria. The MIC90 of amoxicillin for beta-lactamase-negative strains was 0.5 microgram/ml and the MIC90 of ticarcillin 2.0 micrograms/ml. In the case of beta-lactamase-positive strains the MIC90 of amoxicillin (32 micrograms/ml) and ticarcillin (16 micrograms/ml) dropped to less than or equal to 1.0 microgram/ml upon addition of clavulanate; 65.8% of these strains were susceptible to amoxicillin and 98.2% to ticarcillin, but all were susceptible when clavulanate was added. All strains were susceptible to imipenem and metronidazole, and 99.3% to cefoxitin.

Amoxicillin↗

The isolation of fusobacteria from tropical ulcers.

Tropical ulcer is a disease found most commonly in the tropics, although the disease is not confined to those areas. Young children are affected most frequently and the disease is thought to be a polymicrobial infection with fusobacteria, aerobic microorganisms and spirochaetes each playing a role. The fusobacteria have been reported on smears but have not been cultured thus far. We report the isolation and culture of these organisms from 46 ulcers and have also demonstrated the same organisms in skin biopsies from these lesions.

Bacteria, Anaerobic↗

A pyrolysis mass spectrometry study of fusobacteria.

Strains of fusobacteria (143) were examined by pyrolysis mass spectrometry (Py-MS) with a Horizon Instruments PYMS 200X. Fusobacterium necrogenes, F. necrophorum, F. nucleatum, F. mortiferum, F. varium, F. gonidiaformans, F. naviforme, F. russii and Leptotrichia buccalis were discriminated. Strains of fusobacteria isolated from tropical ulcers, although similar to F. mortiferum in conventional tests, were discriminated from each of these species in Py-MS. Identification of 416 spectra to species level agreed with conventional bacteriological methods in 91.8% of cases, was equivocal in 3.4% and disagreed in 4.8%. Classification based on pyrolysis data resolved groups largely corresponding to the recognised species. However, F. nucleatum strains were divided between two distinct groups. The tropical ulcer strains were resolved as a distinct homogeneous group. Py-MS is a rapid, inexpensive and convenient procedure for characterisation of bacteria, with the capacity for a high throughput of samples, although the initial cost of the apparatus is high.

Fusobacterium↗

Isolation of fusobacteria from the oral cavities of malnourished Nigerian children living in agricultural and herding villages.

A previous study demonstrated the presence and possible involvement of Fusobacterium necrophorum in the pathogenesis of noma lesions of children living in agricultural and herding villages in northwestern Nigeria. In order to determine if F. necrophorum was part of the oral flora of malnourished children with no noma lesions, a study of the fusobacteria present in the oral cavities of 30 children, 2-6 years of age in Sokoto State, was undertaken. Swabs taken of the oral cavity were cultured on selective fusobacteria medium using conventional anaerobic microbiological techniques. F. nucleatum was recovered from each child and F. necrophorum was isolated from the oral cavity of only one child. The presence of F. nucleatum and the lack of F. necrophorum, except in one case, suggests that the latter is not normal flora in the children at risk for noma. F. necrophorum, a putative trigger organism for noma may gain a foothold only when certain staging conditions (i.e., lowered host resistance and/or oral lesion) are present.

Anaerobiosis↗

Cellular fatty acid and soluble protein profiles of oral fusobacteria.

We compared the cellular fatty acid and protein content of 43 strains of oral fusobacteria isolated from patients with chronic gingivitis, acute necrotizing ulcerative gingivitis, and juvenile and adult periodontitis, as well as from the stump-tailed macaque, to that of eight reference strains of oral and non-oral Fusobacterium species. A gas-liquid chromatographic examination of trimethylsilyl derivatives of the fatty acid methyl esters revealed n14:0, 3-OH-14:0, n16:0, 16:1, 3-OH-16:0, n18:0, and 18:1 in each of the 51 study strains, and a variable occurrence of 14 other fatty acids. The n16:0 to 3-OH-16:0 ratio distinguished between Fusobacterium nucleatum and the non-oral species Fusobacterium varium, Fusobacterium necrophorum, Fusobacterium russii, Fusobacterium necrogenes, Fusobacterium mortiferum, and Fusobacterium naviforme. The soluble protein content, as determined by polyacrylamide gel electrophoresis, varied considerably among the oral fusobacterial strains studied, and underscored the heterogenous nature of these organisms. "Fingerprinting" of oral fusobacteria may be readily accomplished by polypeptide analysis.

Animals↗

Genotypic and phenotypic characterization of fusobacteria from Chinese and European patients with inflammatory periodontal diseases.

Phylogenetic and antigenic studies were performed on 48 human oral Fusobacterium strains from Chinese patients with either necrotizing ulcerative gingivitis (NUG) or gingivitis and on 23 Fusobacterium nucleatum or Fusobacterium periodonticum strains from European periodontitis patients. Alignment of partial 16S rRNA gene sequences resulted in a phylogenetic tree that corresponded well with the current classification of oral fusobacteria into F. periodonticum and several subspecies of F. nucleatum, in spite of much minor genetic variability. F. periodonticum, F. nucleatum subsp. animalis and a previously undescribed phylogenetic cluster (C4), that may represent an additional F. nucleatum subspecies, constituted discrete clusters distinct from the remainder of F. nucleatum with high bootstrap values. Chinese and European strains differed markedly with regard to their respective classification patterns, suggesting a predominance of F. peridonticum and F. nucleatum susp. animalis over F. nucleatum subsp. nucleatum and F. nucleatum subsp. fusiforme/vincentii in samples from China. Antigenic typing enabled the association of many previously described serovars with distinct phylogenetic clusters and when applied directly to uncultured clinical samples confirmed the differential distribution of oral Fusobacterium taxa in Chinese and European samples. Bacteria from cluster C4 and F. nucleatum subsp. animalis were significantly more prevalent and accounted for higher cell numbers in NUG than in gingivitis samples, suggesting a possible association of these rarely observed taxa with NUG in Chinese patients.

China↗

Enrichment of fusobacteria from the rumen that can utilize lysine as an energy source for growth.

Ruminal lysine degradation is a wasteful process that deprives the animal of an essential amino acid. Mixed ruminal bacteria did not deaminate lysine (50 mM) at a rapid rate, but lysine degrading bacteria could be enriched if Trypticase (5 mg/mL) was also added. Lysine degrading isolates produced acetate, butyrate and ammonia, were non-motile, stained Gram-negative and could also utilize lactate, glucose, maltose or galactose as an energy source for growth. Lactate was converted to acetate and propionate, and 16S rDNA indicated that their closest relatives were Fusobacterium necrophorum. Growing cultures produced ammonia at rates as high as 2400 nmol/mg protein/mL/min. Washed cell suspensions took up (14)C lysine (3 microM) at an initial rate of 6 nmol/mg protein/min, and glucose addition did not affect the transport. Cells washed aerobically had the same transport rate as those handled anaerobically, but only if the transport buffer contained sodium. The affinity constant for sodium was 8 mM, and sodium could not be replaced by lithium. Cells treated with the sodium/proton antiporter, monensin (5 microM), did not take up lysine, but a protonophore that inhibited growth (tetrachlorosalicylanilide, 10 microM) had no effect. An artificial membrane potential created by potassium diffusion did not increase the rate of lysine transport, and an Eadie-Hofstee plot indicated the transport rate was directly proportional to the lysine concentration. Decreasing the pH from 6.7 to 5.5 caused an 85% decrease in the rate of lysine transport. The addition of F. necrophorum JB2 (130 microg protein/mL) to mixed ruminal bacteria increased lysine degradation 10-fold, but only if the pH was 6.7 and monensin was not present. Further work will be needed to see if dietary lysine enriches fusobacteria in vivo.

Journal Article↗

Identification of fusobacteria in a routine diagnostic laboratory.

A scheme for differentiating Fusobacterium spp. and Leptotrichia spp. from Bacteroides spp. was devised after examining 114 strains of fusobacteria and asaccharolytic bacteroides (17 reference strains and 97 clinical isolates). Sensitivity to a 300 micrograms/ml plate of phosphomycin and an acid reaction on a lysine plate were found to be reliable for differentiating Fusobacterium spp. and L. buccalis from Bacteroides. Using a short set of simple cultural and biochemical tests, isolates could be identified as F. necrophorum, F. necrogenes, F. nucleatum, F. varium or L. buccalis. These tests were: indole, lecithinase, phosphatase, DNase and gas production, aesculin and casein hydrolysis, greening of casein/methylene blue agar, nitrite reduction, bile tolerance and haemolysis on horse blood agar.

Anti-Bacterial Agents↗

Factors affecting lysine degradation by ruminal fusobacteria.

Fusobacterium necrophorum can readily be enriched from the rumen with lysine, and its deamination rate is very rapid. The addition of F. necrophorum JB2 to mixed ruminal bacteria significantly increased lysine degradation, but only if the ratio of ruminal fluid to basal medium was less than 25%. If more ruminal fluid (pH 6.1) was added, ammonia production decreased by as much as 80%. Clarified, autoclaved ruminal fluid was also inhibitory. When F. necrophorum JB2 was grown in a lysine-limited continuous culture (0.1 h(-1) dilution rate) and pH was decreased using HCl, optical density decreased linearly, and the culture washed out at pH 5.6. Batch cultures of F. necrophorum JB2 deaminated as much lysine at pH 6.1 as at pH 6.6, but only if fermentation acids were not present. Sodium acetate (100 mM) had little effect at pH 6.6, but the same concentration inhibited ammonia production by 80% at pH 6.1. The idea that fermentation acids could prevent the enrichment of fusobacteria in vivo was supported by the observation that dietary lysine supplementation did not enhance the lysine deamination rate of the mixed ruminal bacteria.

Ammonia↗

Pleomorphism of fusobacteria isolated from the cockroach hindgut.

Fusobacteria are commonly isolated from the hindgut of the cockroach Eublaberus posticus . Eleven strains isolated from E. posticus by us were keyed to four species, Fusobacterium necrophorum, F. varium , F. gonidiaformans , and F. prausnitzii , using current taxonomic criteria. With the exception of F. gonidiaformis , all species showed rods with swollen centers and large bodies. The pleomorphism of F. varium was examined by phase microscopy and scanning and transmission electron microscopy. The pleomorphic process begins with a gradual swelling at the center of the rod until a large round body is formed. Some of these round bodies then fragment, giving rise to rod-shaped cells. When 10% yeast extract was added to growth media, pleomorphism was not induced. A dialyzable factor was found to account for this observation. Fermentation of [1-14C]glutamic acid gives rise to butyrate labeled in the carboxyl carbon, indicating that butyrate is formed by the hydroxyglutarate pathway which may be characteristic for the genus Fusobacterium.

Animals↗

Identification and characterization of a novel adhesin unique to oral fusobacteria.

Fusobacterium nucleatum is a gram-negative anaerobe that is prevalent in periodontal disease and infections of different parts of the body. The organism has remarkable adherence properties, binding to partners ranging from eukaryotic and prokaryotic cells to extracellular macromolecules. Understanding its adherence is important for understanding the pathogenesis of F. nucleatum. In this study, a novel adhesin, FadA (Fusobacterium adhesin A), was demonstrated to bind to the surface proteins of the oral mucosal KB cells. FadA is composed of 129 amino acid (aa) residues, including an 18-aa signal peptide, with calculated molecular masses of 13.6 kDa for the intact form and 12.6 kDa for the secreted form. It is highly conserved among F. nucleatum, Fusobacterium periodonticum, and Fusobacterium simiae, the three most closely related oral species, but is absent in the nonoral species, including Fusobacterium gonidiaformans, Fusobacterium mortiferum, Fusobacterium naviforme, Fusobacterium russii, and Fusobacterium ulcerans. In addition to FadA, F. nucleatum ATCC 25586 and ATCC 49256 also encode two paralogues, FN1529 and FNV2159, each sharing 31% identity with FadA. A double-crossover fadA deletion mutant, F. nucleatum 12230-US1, was constructed by utilizing a novel sonoporation procedure. The mutant had a slightly slower growth rate, yet its binding to KB and Chinese hamster ovarian cells was reduced by 70 to 80% compared to that of the wild type, indicating that FadA plays an important role in fusobacterial colonization in the host. Furthermore, due to its uniqueness to oral Fusobacterium species, fadA may be used as a marker to detect orally related fusobacteria. F. nucleatum isolated from other parts of the body may originate from the oral cavity.

Adhesins, Bacterial↗

Relationships among the oral fusobacteria assessed by DNA-DNA hybridization.

DNA was purified from 16 strains of Fusobacterium nucleatum and from five strains representing other Fusobacterium species. The relationships among fusobacteria were examined by DNA-DNA hybridization and by determining the guanine plus cytosine content of the DNA. F. nucleatum was found to comprise a heterogenous group of organisms related to Fusobacterium periodonticum and Fusobacterium simiae, but unrelated to any of the other species of Fusobacterium tested.

Cytosine↗

Fusobacteria. An important cause of chorioamnionitis.

Prematurity is the main cause of human perinatal disease and chorioamnionitis is the main cause of prematurity. The diverse etiologic role of infection is incompletely known. Fusobacteria (FB) are herein reported to be delineated by the Warthin-Starry (WS) stain; an animal model was developed to reinforce evidence that FB cause chorioamnionitis. Isolates of these bacteria from three human placentas were inoculated into rabbits to develop antisera. Within an annual study of 297 human placentas, 138 were processed with WS stains. By WS stain, microbiologic culture, and immunofluorescent labelling of three placentas, FB were proved to cause chorioamnionitis. Sixty-two (35%) of 176 premature newborns had chorioamnionitis. Of these 62, 11 (18%) had morphologically evident FB; FB-like organisms were associated with chorioamnionitis and prematurity. Our findings support and expand clinical investigation of the amniotic infection syndrome.

Animals↗

Susceptibility of Bacteroides fragilis, Bacteroides thetaiotaomicron, and Fusobacteria in Antibacterial agents.

The minimum inhibitory concentrations of 45 antimicrobial agents were established by the agar dilution method for 37 strains of Bacteroidaceae: Bacteroides fragilis (18 strains), B. thetaiotaomicron (8 strains), Fusobacterium nucleatum (6 strains), and Fusobacterium spp. (5 strains). The fusobacteria tested showed good susceptibility to beta-lactam-antibiotics, tetracyclines, lincomycins, and nitroimidazole compounds. Bacteria of the B. fragilis group were resistant to most of the penicillins and cephalosporins and could not be suppressed with certainty by nitroimidazoles and older tetracyclines. Doxycycline and minocycline, however, showed sufficient activity, and all our Bacteroides strains tested were sensitive to clindamycin. Especially the good in vitro susceptibility of all Bacteroidaceae strains investigated to cefoxitin may offer new therapeutic possibilities.

Anti-Bacterial Agents↗