Appl Environ Microbiol, 1996 Jun, 62(6), 1935 - 43 Molecular microbial diversity of an agricultural soil in Wisconsin; Borneman J et al.; A culture-independent survey of the soil microbial diversity in a clover-grass pasture in southern Wisconsin was conducted by sequence analysis of a universal clone library of genes coding for small-subunit rRNA (rDNA) . A rapid and efficient method for extraction of DNA from soils which resulted in highly purified DNA with minimal shearing was developed . Universal small-subunit-rRNA primers were used to amplify DNA extracted from the pasture soil . The PCR products were cloned into pGEM-T, and either hypervariable or conserved regions were sequenced . The relationships of 124 sequences to those of cultured organisms of known phylogeny were determined . Of the 124 clones sequenced, 98.4% were from the domain Bacteria . Two of the rDNA sequences were derived from eukaryotic organelles . Two of the 124 sequences were of nuclear origin, one being fungal and the other a plant sequence . No sequences of the domain Archaea were found . Within the domain, Bacteria, three kingdoms were highly represented: the Proteobacteria (16.1%), the Cytophaga-Flexibacter-Bacteroides group (21.8%), and the low G+C-content gram-positive group (21.8%) . Some kingdoms, such as the Thermotogales, the green nonsulfur group, Fusobacteria, and the Spirochaetes, were absent . A large number of the sequences (39.4%) were distributed among several clades that are not among the major taxa described by Olsen et al . (G.J . Olsen, C.R . Woese, and R . Overbeek, J . Bacteriol., 176:1-6, 1994) . From the alignments of the sequence data, distance matrices were calculated to display the enormous microbial diversity found in this soil in two ways, as phylogenetic trees and as multidimensional-scaling plots. Zentralbl Veterinarmed B, 1996 Jun, 43(4), 193 - 9 Isolation and antimicrobial susceptibility of obligate anaerobic bacteria recovered from the uteri of dairy cows with retained fetal membranes and postparturient endometritis; Cohen RO et al.; The uteri of 77 postparturient dairy cows were sampled . Samples were cultured aerobically and anaerobically, and the nature of bacterial growth was identified . A mixed aerobic and anaerobic bacterial infection was found in 55% of the samples . Actinomyces pyogenes was the predominant aerobic species; it was found in 70% of the samples, whereas Bacteroides melaninogenicus was the most frequent anaerobic species isolated . Altogether, 16 species belonging to the genus Bacteroides were identified with variable frequencies . It appears that more than one Bacteroides species colonizes the uterus of a given cow postpartum . The minimal inhibitory concentrations (MICs) of clindamycin, metronidazole, tetracycline and ciprofloxacin for 83 anaerobic isolates were determined . All isolates were susceptible to clindamycin (MIC90 of 0.064 microgram/ml) and all but two to metronidazole . Susceptibility to ciprofloxacin was variable, with a bimodal distribution of MIC values . The MIC of tetracycline for 90% of the isolates was > 256 micrograms/ml. Yakugaku Zasshi, 1996 Jun, 116(6), 491 - 6 {Synthesis and antibacterial properties of quinoxaline 1,4-dioxide derivatives}; Takabatake T et al.; Novel quinoxaline 1,4-dioxide derivatives were synthesized from benzofuroxans and the enolic form of 1,3-diketones or 3-oxoalkanoic esters or 3-oxoalkanamides or butanedioic esters catalyzed by silica gel or molecular sieves and their antibacterial activities were evaluated . As the results of antibacterial screening tests in vitro, quinoxaline 1,4-dioxides revealed strong activities against Bacteroides fragilis. Leuk Lymphoma, 1996 Jun, 22(1-2), 177 - 9 Hemophagocytic syndrome associated with Bacteroides fragilis infection in a patient with acute monoblastic leukemia; Bourquelot P et al.; The HS has been associated with malignant hemopathies . We report here a case of HS related to Bacteroides fragilis during the course of acute monoblastic leukemia . Evolution was fatal despite remission of the leukemia process and response of the infection to appropriate antibiotic therapy. J Bacteriol, 1996 Jun, 178(12), 3601 - 7 NBU1, a mobilizable site-specific integrated element from Bacteroides spp., can integrate nonspecifically in Escherichia coli; Shoemaker NB et al.; NBU1 is an integrated Bacteroides element that can he mobilized from Bacteroides donors to Bacteroides recipients . Previous studies have shown that a plasmid carrying the internal mobilization region of NBU1 could be transferred by conjugation from Bacteroides thetaiotaomicron to Escherichia coli . In this report, we show that NBU1 can integrate in E . coli . Whereas integration of NBU1 in B . thetaiotaomicron is site specific, integration of NBU1 in E . coli was relatively random, and the insertion frequency of NBU1 into the E . coli chromosome was 100 to 1,000 times lower than the frequency of integration in B . thetaiotaomicron . The frequency of NBU1 integration in E . coli could be increased about 10- to 70-fold, to a value close to that seen with B . thetaiotaomicron, if the primary integration site from B . thetaiotaomicron, BT1-1, was provided on a plasmid in the E . coli recipient or the NBU1 integrase gene, intN1, was provided on a high-copy-number plasmid to increase the amount of integrase available in the recipient . When the primary integration site was available in the recipient, NBU1 integrated site specifically in E . coli . Our results show that NBUs have a very broad host range and are capable of moving from Bacteroides spp . to distantly related species such as E . coli . Moreover, sequence analysis of NBU1 integration sites provided by integration events in E . coli has helped to identify some regions of the NBU1 attachment site that may play a role in the integration process. J Bacteriol, 1996 Jun, 178(12), 3594 - 600 The Bacteroides mobilizable insertion element, NBU1, integrates into the 3' end of a Leu-tRNA gene and has an integrase that is a member of the lambda integrase family; Shoemaker NB et al.; NBU1 is a 10.3-kbp integrated Bacteroides element that can be induced to excise from the chromosome and can be mobilized to a recipient by trans-acting functions provided by certain Bacteroides conjugative transposons . The NBU1 transfer intermediate is a covalently closed circle, which is presumed to be the form that integrates into the recipient genome . We report here that a 2.4-kbp segment of NBU1 was all that was required for site-specific integration into the chromosome of Bacteroides thetaiotaomicron 5482 . This 2.4-kbp region included the joined ends of the NBU1 circular form (attN1) and a single open reading frame, intN1, which encoded the integrase . Previously, we had found that NBU1 integrates preferentially into a single site in B . thetaiotaomicron 5482 . We have now shown that the NBU1 target site is located at the 3' end of a Leu-tRNA gene . The NBU1 integrase gene, intN1, was sequenced . The predicted protein had little overall amino acid sequence similarity to any proteins in the databases but had limited carboxy-terminal similarity to the integrases of lambdoid phages and to the integrases of the gram-positive conjugative transposons Tn916 and Tn1545 . We also report that the intN1 gene is expressed constitutively. Mol Microbiol, 1996 May, 20(4), 741 - 50 A gene product related to Tral is required for the mobilization of Bacteroides mobilizable transposons and plasmids; Smith CJ et al.; The antibiotic-resistance transposon Tn4555 from Bacteroides can be transferred between strains by conjugation . The transposon is not self-transmissible and must be mobilized by resident chromosomal tetracycline-resistance elements . In the present report, the mechanism of transfer was examined at the genetic level by deletion analysis and nucleotide sequencing of clones that conferred a transmissible phenotype on a non-mobilizable plasmid . The results suggested that the product of mobATn was required for mobilization and it worked in concert with a cis-acting oriT-like sequence . This mechanism was compared with the mobilization system of a cryptic Bacteroides plasmid, pBI143, and the two systems were found to share a common transfer strategy . The mobA gene products from both genetic elements were related and they had limited homology to the broad group of mobilization proteins (relaxases) typified by Tral of RP4 . Phylogenetic analysis of MobA and several other mobilization proteins from commensal gastro-intestinal tract organisms suggested that they formed a new subgroup of the Tral superfamily . The mobilization regions of both Tn4555 and pBI143 were located on discrete segments of DNA within the parent genetic element . These segments were delineated by regions of secondary structure, suggesting that they could be defined mobilization cassettes. J Periodontol, 1996 May, 67(5), 523 - 7 Long-term stability of Class II furcation defects treated with barrier membranes; Machtei EE et al.; The present longitudinal study was designed to explore the long-term efficacy of guided tissue regeneration (GTR) in Class II furcation defects and establish the factors that might be responsible for modifying this response . Subjects with two or more mandibular molars, one of which had Class II furcation defects, received the hygienic phase of therapy followed by baseline clinical measurements and subgingival plaque sampling . GTR procedure was performed in furcation defect sites using expanded polytetrafluoroethylene (ePTFE) membranes, while the other non-furcated molars received only scaling and root planning . Twenty-eight subjects (13 females, 15 males) aged 27 to 66 were included in this longitudinal analysis . Post-surgical treatment included routine home care supplemented with daily chlohexidine rinse and systemic tetracycline . Membranes were retrieved 4 to 6 weeks after surgery . During the first year, patients were initially seen bi-weekly and subsequently monthly for professional prophylaxis . At the end of this year, clinical measurements and samples were obtained . For the next 2 years, patients were seen bi-annually for maintenance visits . Clinical measurements and microbiological samples were then repeated . Next, a tighter maintenance protocol was established and patients were seen quarterly for scaling and oral hygiene reinforcement . Final measurements and samples were taken again 1 year later (4 years postoperative) . Significant probing reduction (3.00 mm) and gain in horizontal attachment (2.59 mm) were obtained 1 year postsurgery for the GTR sites . These changes were maintained over 4 years with a slight decline at the end of year 3 . Changes in probing depth (PD) from year 1 to 4 served to dichotomize the sites into stable (delta PD < or = 0.9 mm), and unstable (PD increase > or = 1 mm) . Of the 54 sites available for this analysis only 5 (9.3%) were unstable while 49 (90.7%) were stable or even further improved . Sites which exhibited minimal or no plaque (plaque index {PI} < or = 1) over the tight maintenance period had a further decrease in mean probing depth (0.43 mm) compared with a slight increase (-0.06 mm) in mean probing depth in sites with PI > or = 2 mm (P = 0.0235) . The same phenomenon was observed for changes in relative attachment level (RAL): mean gain in RAL was 0.61 mm compared to 0.25 mm for the 2 groups, respectively (P = 0.07) . Actinobacillus actinomycetemcomitans was only isolated from 2 sites at year 3, and none at year 4, compared to 21.45% of the sites at baseline . Porphyromonas gingivalis positive sites showed a continual decline over the years: 14.28% at baseline, 10.71% at year 1, and 5.1% at year 4 . On the contrary, Prevotella intermedia (Pi) and Bacteroides forsythus (Bf) infected sites remained at approximately the same rate throughout the 4 years of the study (40% to 50% and 30% to 40% for Pi and Bf, respectively) . Of these, Pi-infected sites exhibited less favorable clinical results compared to sites which were not infected with this microorganism . In summary, furcation defects treated with membrane barriers can be maintained in health for at least 4 years; however, good oral hygiene and frequent recall visits as part of a complete anti-infective therapy are essential . Finally, once treated, these teeth are comparable to similar molar teeth with no previous history of furcation pathosis. Lett Appl Microbiol, 1996 May, 22(5), 361 - 5 Detection of Bacteroides fragilis by PCR assay targeting the neuraminidase-encoding gene; Kuwahara T et al.; Oligonucleotide primers were designed on the basis of the sequence of the neuraminidase-encoding gene (nanH) of Bacteroides fragilis and used for the specific detection of this anaerobe by the nested PCR assay . Fifty-nine of 60 representative strains of Bact . fragilis were detected, while none of 45 strains of other species generated visible PCR products . The detection limits of Bact . fragilis cells and DNA by the nested PCR were 10 colony-forming units and 10 fg of chromosomal DNA, respectively . The PCR assay targeting the nanH gene has the potential for the detection of Bact . fragilis. J Bone Joint Surg Am, 1996 May, 78(5), 749 - 54 Aspiration of the hip joint before revision total hip arthroplasty . Clinical and laboratory factors influencing attainment of a positive culture; Lachiewicz PF et al.; The value of routine aspiration of the hip joint before revision of a hip arthroplasty remains controversial . We reviewed the results of such aspirations in an attempt to determine clinical or laboratory factors that could help the surgeon to identify hips that are infected and that should be aspirated preoperatively . One hundred and fifty consecutive revision total hip arthroplasties were performed by one of us . Preoperative aspiration was not performed or data were excluded for eight hips; no fluid was obtained from one of these hips (0.7 percent of the 150) . Of the remaining 142 hips, 128 had preoperative aspiration once and fourteen, twice . Twenty-one (15 percent) of the 142 hips were infected, as demonstrated by the intraoperative culture . The intraoperative culture for two of these hips, however, was considered to be false-positive . The initial aspiration was considered to be positive only if an organism grew on the solid medium or if grossly purulent fluid was obtained . The initial aspiration was positive for nineteen hips; on culture of specimens from one hip, Bacteroides thetaiotaomicron grew in the liquid medium only; and purulent fluid was obtained from one hip but no organisms grew on culture . Fourteen aspirations were repeated for various reasons, most commonly to confirm the presence of an unusual organism . The repeat aspiration did not change the diagnosis for these hips . When the two hips with a false-positive intraoperative culture were excluded, preoperative aspiration had a sensitivity of 92 percent, a specificity of 97 percent, and an accuracy of 96 percent . Seventeen of the nineteen truly infected hips were associated with an abnormally elevated erythrocyte-sedimentation rate (mean, 80.8 millimeters per hour) . However, fifty-eight (50 percent) of the 116 hips that were not infected, and for which the results were available, also had an abnormally elevated erythrocyte-sedimentation rate (mean, 32.0 millimeters per hour) . This difference was significant (p = 0.001, Fischer exact test) . The peripheral leukocyte count was not helpful in predicting infection . Hips in which the implants had been in situ for more than five years were less likely to be infected (p = 0.008, Fisher exact test) than those in which the implants had been in situ for five years or less . None of the infected hips in which the implants had been in situ for more than five years were associated with a normal erythrocyte-sedimentation rate . In this study, preoperative aspiration of the hip joint had an excellent sensitivity and specificity with regard to the prediction of infection, On the basis of our findings, we now favor a selective approach to aspiration, as determined by the erythrocyte sedimentation rate and the amount of time that the implant has been in situ. J Surg Res, 1996 May, 62(2), 273 - 7 Peritoneal interleukin-8 in acute appendicitis; Zeillemaker AM et al.; Interleukin-8 (IL-8) is a chemoattractant that is highly selective for neutrophils . This study was designed to investigate the presence of IL-8 in peritoneal fluid of patients with acute appendicitis . The clinical circumstances underlying the secretion of IL-8 by mesothelium and its mechanism of activation have not been defined . In an in vitro model for bacterial peritonitis the role of bacteria in activating human mesothelial cells to secrete IL-8 was studied . Cultured human mesothelium was incubated with various species of pathogenic bacteria, isolated from peritoneal exudate fluids of patients with appendicitis . The amount of IL-8 secreted by the cultured mesothelial cells was determined in an IL-8 ELISA, as IL-8 was present in the original peritoneal fluid of these patients . Peritoneal fluids from patients with a perforated appendix were found to contain a significantly higher concentration of IL-8 compared to peritoneal fluids from patients with nonperforating appendicitis (121.6 (57.8) ng/ml versus 0.2 (0.07) ng/ml, respectively; mean (SEM), P < or = 0.01) . Species of Bacteroides and Fusobacterium necrophorum induced IL-8 secretion from cultured mesothelial monolayers to levels comparable to those found in peritoneal fluids in vivo . Heat-killed bacteria and bacterial supernatant were also able to stimulate mesothelium to secrete IL-8 . The results suggest that in the early phase of bacterial peritonitis the influx of PMN is regulated by bacteria-induced IL-8 secretion by the mesothelium lining the peritoneal cavity. Oral Microbiol Immunol, 1996 Apr, 11(2), 79 - 87 Characteristics of systemic antibody responses of nonhuman primates following active immunization with Porphyromonas gingivalis, Prevotella intermedia and Bacteroides fragilis; Giardino A et al.; Periodontal disease is an infectious disease manifested by the progressive change of a healthy resident commensal microbiota to a pathogenic one characterized by a specific microbiota . Thus, the prospect for the use of selected bacteria or their antigens as a vaccine to interfere with the microbial changes and resulting progression of periodontal tissue destruction has been proposed . As a first step in examining the use of bacterial antigens as immunogens in periodontitis, this study characterized the humoral immune response in Macaca fascicularis after systemic immunization with intact Porphyromonas gingivalis, Prevotella intermedia and Bacteroides fragilis . Parental immunization of the nonhuman primate with the intact bacteria resulted in the production of specific and significantly elevated levels of antibodies to P . gingivalis and P . intermedia, with the predominant isotype being immunoglobulin G (IgG) . In contrast, the principal response to the nonoral, intestinal bacterium, B . fragilis, was of the IgM isotype . Immunization increased IgG, IgM, and IgA antibody by 14-227 fold to P . gingivalis and 8-108 fold to P . intermedia . The level of serum IgA antibody increased (77-227 fold) . The kinetics of the antibody response post-immunization and post-ligation differed with respect to each of the bacteria tested . IgG antibody to P . gingivalis increased through week 16 of the experiment and remained elevated above baseline through week 32 . The IgG antibody level to P . intermedia peaked at 4 weeks following the third immunization and decreased post-ligation to near baseline levels by week 16 . Characterization of the immune response after active immunization in the nonhuman primate has demonstrated a substantial and specific increase in antibody response which was sustained for several weeks . The insights obtained from these studies should help optimize the potential for immunologic interference with progressing periodontitis. Appl Environ Microbiol, 1996 Apr, 62(4), 1434 - 6 Inactivation of tryptic activity by a human-derived strain of Bacteroides distasonis in the large intestines of gnotobiotic rats and mice; Ramare F et al.; Tryptic activity disappeared and trypsin was no longer detected with an antitrypsin antiserum in the large intestines of gnotobiotic rats and mice monoassociated with a human-derived strain of Bacteroides distasonis, whereas tryptic activity was not modified in the small intestines . This function was shown to be strain specific. Antimicrob Agents Chemother, 1996 Apr, 40(4), 1014 - 6 Oligonucleotide probes for detection of cephalosporinases among Bacteroides strains; Mastrantonio P et al.; Two oligonucleotide probes selected from the sequences of cepA and cfxA genes, respectively, were used to detect beta-lactamase production among strains of the Bacteroides fragilis group . By using these probes, colony hybridization was shown to be a specific and rapid method for identifying the more prevalent beta-lactamase, CepA, and the rarer CfxA enzyme among B . fragilis strains. Microb Pathog, 1996 Apr, 20(4), 191 - 202 A comparison of the haemagglutinating and enzymic activities of Bacteroides fragilis whole cells and outer membrane vesicles; Patrick S et al.; The haemagglutinating and enzymic activities of the obligately anaerobic pathogenic bacterium Bacteroides fragilis were examined . Outer membrane vesicles are released from the surface of B . fragilis . They can be detected by electron microscopy in ultrathin sections and bacterial suspensions after negative staining . Electron microscopy and immunogold labelling with a MAb specific for surface polysaccharide of B . fragilis confirmed that the vesicles carried outer membrane associated epitopes . The haemagglutinating activity of whole cells from populations of B . fragilis strains NCTC9343, BE3 and LS66 enriched by Percoll density gradient centrifugation for a large capsule (LC), electron dense layer (EDL); non-capsulate by light microscopy) and outer membrane vesicles (OMV) which had been purified by centrifugation from EDL-enriched populations were compared using human and horse erythrocytes . The enzymic activity of OMV, LC- and EDL-enriched populations, as detected by the API ZYM kit, was compared for strains NCTC 9343 and BE3 . Purified OMV from the strains examined exhibited both haemagglutinating and enzymatic activity . Haemagglutination by the EDL-enriched population was sensitive to treatment with sodium periodate . The LC-enriched population haemagglutinated only after ultrasonic removal of the capsule . This indicates that the LC masks a haemagglutinin . The results suggest a potential role for OMV in the virulence of B . fragilis. J Periodontol, 1996 Apr, 67(4), 422 - 7 Microbial flora in the acute phase of periodontitis and the effect of local administration of minocycline; Umeda M et al.; Periodontitis, similar to other infectious diseases, is known to progress as chronic inflammation with recurrent acute phases . The purpose of this study was to clarify the microbiological composition of the acute phase and to compare the bacterial flora with that of comparable chronic periodontal pockets . We also evaluated the effect of application of minocycline gel locally on the change in the microflora in the acute pockets . Microbial flora from the subgingival pockets of 28 patients in the acute phase of periodontitis and of 12 patients in a comparable chronic phase as the control were investigated by various bacterial culture methods including TS blood agar and TSBV plates . Minocycline gel was applied to the acute periodontal pockets . Changes in the microbiological proportion and clinical parameters at one week after baseline examination were followed by dark-field analysis, culture method, and indirect immunofluorescence technique . Characteristic features of bacterial proportions in the acute site were observed as an increase in Bacteroides forsythus . The number of Porphyromonas gingivalis and black pigmented anaerobic rods also increased . Application of minocycline gel in the acute pocket without any debridement produced improvement in clinical symptoms at one week . Black-pigmented anaerobic rods, P . gingivalis, and B . forsythus decreased significantly at one week after the application . Results indicate that periodontopathic bacteria including B . forsythus and P . gingivalis were predominant in the acute phase of periodontitis and a locally delivered antibiotic may be effective as an alternative modality of treating the acute inflammation. Immunology, 1996 Apr, 87(4), 660 - 7 Cleavage of human complement component C5 by cysteine proteinases from Porphyromonas (Bacteroides) gingivalis . Prior oxidation of C5 augments proteinase digestion of C5; Discipio RG et al.; Since severe periodontitis is characterized by an acute inflammatory response with cellular infiltration and microbial overgrowth, plasma proteins could be exposed to both proteinases and oxidants released from the granulocytes, as well as to proteinases from the microorganisms . When human complement component C5 was digested by cysteine proteinases (i.e . gingipain-R and gingipain-K) from Porphyromonas gingivalis, limited cleavage of the C5 molecule was observed . If C5 was first oxidized by hydroxyl radicals, these gingipains converted modified C5 to fragments that exhibited significantly greater pro-inflammatory activity than did digests of unmodified C5 . After cleavage of oxidized C5 by gingipain-R, the digest exhibited measurably greater neutrophil enzyme release and chemotaxis of human polymorphonuclear leukocytes (PMNs) compared with the activities of unoxidized C5 digests . Gingipain-K generates virtually no polarization or chemotactic activity of human PMNs from C5, nor is enzyme release stimulated by these C5 digests . However, when oxidized C5 was digested by gingipain-K, human PMNs were stimulated for polarization, chemotaxis and enzyme release indicating that an active fragment had been generated . Proteolysis of oxidized C5 evokes greater neutrophil activation than does proteolysis of unoxidized protein, a fact which supports the hypothesis that oxidation and proteolysis may be coupled to enhance the destructive effects of the inflammatory process . These results, in which digests of both oxidized and unmodified complement component C5 were evaluated, support the general concept that oxidation and proteolysis may participate cooperatively in amplifying both the severity and duration of the inflammatory reaction. J Nutr, 1996 Apr, 126(4 Suppl), 1181S - 6S Chemistry, nutritional sources, tissue distribution and metabolism of vitamin K with special reference to bone health; Shearer MJ et al.; Vitamin K occurs in nature as a series of compounds with a common 2-methyl- 1,4 naphthoquinone nucleus and differing isoprenoid side chains at the 3 position . They comprise a single major plant form, phylloquinone with a phytyl side chain and a family of bacterially synthesized menaquinones (MKs) with multiprenyl side chains . The major dietary source to humans is phylloquinone for which the chief food contributors are green, leafy vegetables followed by certain vegetable oils (soybean, rapeseed and olive oils) . Recent analyses by high pressure liquid chromatography are now providing a wide-ranging database of phylloquinone in foods . Menaquinones are found in moderate concentrations in only a few foods such as cheeses (MK-8 and MK-9) . A wider spectrum of MKs is synthesized by the gut microflora, and their intestinal absorption probably accounts for most of the hepatic stores, particularly those with very long side chains (MKs-10--13) synthesized by members of the genus Bacteroides . The site of absorption of floral MKs is not known, but reasonable concentrations are found in the terminal ileum where bile salt-mediated absorption is possible . Both phylloquinone and menaquinones are bioactive in hepatic gamma-carboxylation but long-chain MKs are less well absorbed . Liver stores of vitamin K are relatively small and predominantly MKs-7--13 . The hepatic reserves of phylloquinone (approximately 10% of the total) are labile and turn over at a faster rate than menaquinones . Trabecular and cortical bone appear to contain substantial concentrations of both phylloquinone and menaquinones . A majority (approximately 60-70%) of the daily dietary intake of phylloquinone is lost to the body by excretion, which emphasizes the need for a continuous dietary supply to maintain tissue reserves. J Bacteriol, 1996 Apr, 178(7), 1914 - 8 A recent fixation of cfiA genes in a monophyletic cluster of Bacteroides fragilis is correlated with the presence of multiple insertion elements; Ruimy R et al.; Small-subunit ribosomal DNA sequences of 16 strains of Bacteroides fragilis were determined and compared with previously published sequences . Three phylogenetic methods (the neighbor-joining, maximum-likelihood, and maximum-parsimony methods) as well as a bootstrap analysis were used to assess the robustness of each topology . All phylogenetic analyses demonstrated that the B . fragilis strains were clearly divided into two robust monophyletic units which corresponded to the cfiA-negative and cfiA-positive groups . Strains of two previously identified DNA homology groups separated similarly into the two monophyletic units . According to the intensity of the hybridization signal with a cfiA probe, the cfiA-positive cluster could be further divided into two groups . This difference might reflect the existence of two, probably closely related cfiA-type genes . In the strongly hybridizing cfiA-positive strains, the gene is capable of conferring high-level resistance to the carbapenems and to most beta-lactamase inhibitors as well, while in the weakly hybridizing cfiA-positive strains, only the latter type of resistance is known to occur . The presence of the cfiA-type genes within a monophyletic cluster of B . fragilis that apparently represents only a minority of the species B . fragilis is suggestive of a recent acquisition . The fact that this cluster is also the predominant pool of all known B . fragilis insertion elements, which have been found to play an important role in the expression of carbapenem resistance, raises the possibility that both genetic determinants, i.e., the resistance gene(s) and insertion elements, may have coevolved. Infect Immun, 1996 Apr, 64(4), 1342 - 50 Functional chemotactic factor CP-10 and MRP-14 are abundant in murine abscesses; Kocher M et al.; Murine abscesses induced by intraperitoneal injection of a mixture of Escherichia coli, Bacteroides fragilis, and bran are established models for the study of localized infectious and inflammatory lesions . Chemotactic factors are though to mediate the directed migration of large numbers of leukocytes into the abscess . Microorganisms located within the encapsulated lesion are not readily eliminated by the leukocytes, but their numbers are controlled over many weeks . We report the presence of large amounts of two murine S100 proteins, CP-10 and migration inhibition factor-related protein 14 (MRP-14), in abscesses as demonstrated by immunohistochemistry and measured by enzyme-linked immunosorbent assay and Western blotting (immunoblotting) . High levels of CP-10 (7.7 +/- 1 mg/ml) and MRP-14 (5.5 +/- 1 mg/ml) were found throughout the time course of abscess development from early acute-phase lesions, which are predominantly neutrophilic, to late chronic-phase lesions, which contained more mononuclear cells . Approximately one-third of these amounts occurred as monomers (2.0 mg/ml for MRP 14 and 2.2 mg/ml for CP-10) . Abscess fluid was strongly chemotactic, and a portion of the activity was due to CP-10, indicating its important role in leukocyte recruitment . CP-10-MRP-14 complexes were present in abscess fluid, and the proteins were immunoabsorbed together . In analogy with the related human MRP-8-MRP-14 complex, these proteins could be involved in the inhibition of microbial growth . No growth inhibition occurred with 20 microgram of CP-10 or MRP-14 per ml or with mixtures of both, but these concentrations may have been insufficient and were not representative of the high concentrations found within abscesses . CP-10 may contribute indirectly to the antimicrobial response in abscesses by virtue of its strong chemotactic properties and its capacity to modulate the activation state of recruited leukocytes. J Clin Periodontol, 1996 Mar, 23(3 Pt 1), 212 - 9 Porphyromonas gingivalis, Actinobacillus actinomycetemcomitans and Treponema denticola detection in oral plaque samples using the polymerase chain reaction; Watanabe K et al.; Detection of putative pathogens is critical for delineating the etiology and progression of periodontitis . In the present study, we have used a polymerase chain reaction (PCR) assay utilizing primers specific for the lkt A gene of Actinobacillus actinomycetemcomitans, the fimbrial gene of Porphyromonas gingivalis, and tdp A gene of Treponema denticola in order to determine the presence of these pathogens in subgingival plaque samples from periodontitis sites . These gene specific primers were also used to assess the detection of different strains of bacteria in the PCR assays . Primers for P . gingivalis detected P . gingivalis strain 33277, but no product was detected when primers were used with extracts from 4 species of Capnocytophaga, 3 species of Prevotella, 2 species of Porphyromonas other than P . gingivalis, Bacteroides levii, Escherichia coli, 3 strains of A . actinomycetemcomitans and 3 strains of T . denticola . PCR analysis using primers for the lkt A gene of A . actinomycetemcomitans also did not result in a product with any of these bacteria with the exception of a positive result with 3 different strains of A . actinomycetemcomitans . Primers selected from the tdp A gene of T . denticola did not identify any of the bacteria strains tested except T . denticola serovars a, b, and c . Thus, these primers were shown to amplify gene segments that are specific to either P . gingivalis (33277), A . actinomycetemcomitans (33384, 43717 and 43718) or T . denticola (35405, 33521 and 35404) . The PCR assay may be used to rapidly detect the presence of periodontal pathogens in the future. J Clin Pathol, 1996 Mar, 49(3), 255 - 7 Microbiological and serological investigations of oral lesions in Papillon-Lefèvre syndrome; Clerehugh V et al.; Microbiological and serological (enzyme linked immunosorbent assay) investigations were carried out, including karyotyping, on two Asian children with Papillon-Lefevre syndrome . In case 1, a girl aged four years, the most prevalent putative periodontopathogens were Eikenella corrodens, Fusobacterium nucleatum, Porphyromonas gingivalis, Prevotella intermedia (deciduous dentition) and Bacteroides gracilis, E corrodens and F nucleatum (permanent dentition) . In case 2, a boy aged nine years, they were F nucleatum, P intermedia and P loeschii and E corrodens . Serum from case 2 showed a raised specific IgG antibody response to Actinomyces actino-mycetemcomitans serotype b . Thus, a wider range of species than hitherto reported may be associated with Papillon-Lefevre syndrome, including A actino-mycetemcomitans and F nucleatum. Infect Immun, 1996 Mar, 64(3), 1065 - 9 The capsular polysaccharide complex of Bacteroides fragilis induces cytokine production from human and murine phagocytic cells; Gibson FC 3rd et al.; To stimulate early-phase immunologic events following Bacteroides fragilis infection in the peritoneal cavity, we examined the cytokine response of several cell types to purified capsular polysaccharide complex (CPC) and lipopolysaccharide (LPS) of this organism . Cytokines were produced from murine resident peritoneal (MRP) cells as well as human peripheral blood leukocytes . MRP cells cocultured with either B . fragilis CPC of LPS in vitro produced tumor necrosis factor alpha and interleukin-1alpha (IL-1alpha) . In addition, MRP cells challenged with CPC produced IL-10 . Human peripheral blood monocytes and polymorphonuclear leukocytes secreted IL-8 when cultured in the presence of CPC. J Clin Periodontol, 1996 Feb, 23(2), 133 - 9 Prevalence of 6 putative periodontal pathogens in subgingival plaque samples from Romanian adult periodontitis patients; Ali RW et al.; The aim of the present study was to determine by standard cultivation procedures the detection frequencies of Porphyromonas gingivalis, Prevotella intermedia, Fusobacterium nucleatum, Actinobacillus actinomycetemcomitans, Bacteroides forsythus, and Capnocytophaga species as well as various enteric rods in subgingival plaque samples form Romanian adult periodontitis patients . DNA probe analysis (Affirm DP Microbial Identification Test) was also used, parallel to cultivation, to identify P . gingivalis, A . actinomycetemcomitans, and B . forsythus, in deep (> or = 6 mm) and intermediate (4-5 mm) pockets in some of the subjects investigated . Paper points were used to sample 86 deep pockets in 36 patients and 27 intermediate pockets in 9 of the 36 patients . The chi 2 test was used to test for significance of differences between results obtained by cultivation and DNA analysis in both intermediate and deep pockets . P . gingivalis was recovered in a high percentage of the patients (75.8%) and sites (63.6%) examined, followed by P . intermedia, F . nucleatum, and A . actinomycetemcomitans, respectively . Capnocytophaga species were present in almost all subjects . Enteric rods were recovered in 61.1% of the patients and 55.8% of the sites . Except for this high prevalence of enteric rods, the present group of patients had the periodontal species monitored in %s similar to those commonly perceived in the West . The Affirm DP Test and cultivation showed poor correlation in detecting P . gingivalis, A . actinomycetemcomitans, and B . forsythus . The cultivation prevalence of P . gingivalis and P . intermedia in deep pockets was similar to their prevalence in intermediate ones . Overall, the prevalence of the periodontal pathogens investigated in the present Romanian periodontitis patients is similar to what has been revealed in matching Norwegian and other Western periodontitis patient populations . The high prevalence of enteric rods in the Romanian patients may have been an artifact resulting from prolonged transport of the samples in VMGA III. J Antimicrob Chemother, 1996 Feb, 37(2), 345 - 50 Production of metal dependent beta-lactamases by clinical strains of Bacteroides fragilis isolated before 1987; Khushi T et al.; Five imipenem resistant clinical isolates of Bacteroides fragilis isolated before 1987, were examined to determine if they produced metallo-beta-lactamases . The beta-lactamases produced by the clinical isolates all focused as doublet bands at pl 4.8/4.9, characteristic of the B . fragilis CfiA type metallo-beta-lactamase . Each enzyme had a similar substrate profile and were inhibited by EDTA and activated with zinc sulphate . The sequence of the metallo-beta-lactamase gene from B . fragilis ED262 was determined and confirmed to be a CfiA type beta-lactamase . Consequently, the five isolates examined probably produce a CfiA type beta-lactamase, suggesting that metallo-beta-lactamases were present before the widespread use of carbapenems. Appl Environ Microbiol, 1996 Feb, 62(2), 461 - 8 Phylogenetic diversity of the intestinal bacterial community in the termite Reticulitermes speratus; Ohkuma M et al.; The phylogenetic diversity of the intestinal microflora of a lower termite, Reticulitermes speratus, was examined by a strategy which does not rely on cultivation of the resident microorganisms . Small-subunit rRNA genes (16S rDNAs) were directly amplified from the mixed-population DNA of the termite gut by the PCR and were clonally isolated . Analysis of partial 16S rDNA sequences showed the existence of well-characterized genera as well as the presence of bacterial species for which no 16S rDNA sequence data are available . Of 55 clones sequenced, 45 were phylogenetically affiliated with four of the major groups of the domain Bacteria: the Proteobacteria, the spirochete group, the Bacteroides group, and the low-G+C-content gram-positive bacteria . Within the Proteobacteria, the 16S rDNA clones showed a close relationship to those of cultivated species of enteric bacteria and sulfate-reducing bacteria, while the 16S rDNA clones in the remaining three groups showed only distant relationships to those of known organisms in these groups . Of the remaining 10 clones, among which 8 clones formed a cluster, there was only very low sequence similarity to known 16S rRNA sequences . None of these clones were affiliated with any of the major groups within the domain Bacteria . The 16S rDNA gene sequence data show that the majority of the intestinal microflora of R . speratus consists of new, uncultured species previously unknown to microbiologists. J Bacteriol, 1996 Feb, 178(3), 583 - 90 Development of techniques for the genetic manipulation of the gliding bacterium Cytophaga johnsonae; McBride MJ et al.; Cytophaga johnsonae displays many features that make it an excellent model of bacterial gliding motility . Unfortunately, genetic analyses of C . johnsonae, or any related gliding bacteria, were not possible because of a complete lack of selectable markers, cloning vectors, transposons, and convenient methods of gene transfer . As a first step toward a molecular analysis of gliding motility of C . johnsonae, we developed these genetic techniques and tools . Common broad-host-range plasmids and transposons did not function in C . johnsonae . We identified one Bacteroides transposon, Tn4351, that could be introduced into C . johnsonae on plasmid R751 by conjugation from Escherichia coli . Tn4351 inserted in the C . johnsonae genome and conferred erythromycin resistance . Tn-4351 insertions resulted in auxotrophic mutations and motility mutations . We constructed novel plasmids and cosmids for genetic analyses of C . johnsonae . These cloning vectors are derived from a small cryptic plasmid (pCP1) that we identified in the fish pathogen Cytophaga psychrophila D12 . These plasmids contain the ermF (erythromycin resistance) gene from Tn4351 and a variety of features that facilitate propagation and selection in E . coli and conjugative transfer from E . coli to C . johnsonae. Acta Microbiol Pol, 1996, 45(2), 187 - 92 Detection of enterotoxigenic Bacteroides fragilis (ETBF), among strains isolated between 1976 and 1995 in Poland; Meisel-Mikolajczyk F et al.; Thirty five Bacteroides fragilis strains isolated from different, clinical specimens in 1976-1985 and in 1995, preserved in strains collection of Department of Clinical Bacteriology were studied . The cytotoxicity of 35 bacterial culture supernatants, were tested on the human carcinoma cell line HT29/C1 . Two strains proved to be enterotoxigenic . One of those strains was isolated in 1985 from the human, before the existence of enterotoxin producing strain was reported . All 35 strains were metronidazol sensitive showing lack of correlation between enterotoxigenicity and metronidazol resistance. Acta Vet Scand, 1996, 37(3), 251 - 63 Intrauterine bacterial findings and hormonal profiles in post-partum cows with normal puerperium; Bekana M et al.; The post-partum intrauterine bacterial flora, prostaglandin release, uterine involution and resumption of ovarian activity were studied in 9 Swedish dairy cows during the first 8-week period . Uterine involution was monitored by transrectal examinations of the reproductive tract 3 times weekly . Bacteriological examination was performed from twice weekly uterine biopsies . The main PGF2 alpha metabolite (15-ketodihydro-PGF2 alpha) was monitored from twice daily blood plasma samples, while morning samples were used for progesterone determinations . The cows were assigned to 2 groups: Group I (n = 7) with an uncomplicated puerperal period and Group II (n = 2) with signs of intrauterine infections . A total of 143 biopsies were collected, of which 129 (90.2%) were found to be bacteriologically negative . Thirteen (9.1%) of the remaining 14 biopsies were bacteriologically positive, while one (0.7%) was probably a contamination on a single occasion . The 13 bacteriologically positive biopsies belonged to the Group II cows from which 31 isolates contained 6 different genera of facultative and obligate anaerobic bacteria . Actinomyces pyogenes along with Bacteroides sp . and Fusobacterium necrophorum were found to predominate in a mixed flora . The bacteria were rapidly eliminated and disappeared completely from the uteri towards the end of the third week post-partum . The average number of days required for completion of uterine involution was 21.8 +/- 3.0 for all animals . The plasma levels of the PGF2 alpha metabolite were significantly elevated for the first 12-18, and 18 and 27 days in Group I and Group II, respectively . There was no significant relationship between the duration of PGF2 alpha release and the time required for completion of uterine involution (p > 0.05) . Progesterone analysis showed resumption of ovarian activity and subsequent ovulation in 4 of the 9 cows 44-55 days post-partum . Thus, intrauterine infections are not commonly seen in cows with normal calving and comparison between the duration of PGF2 alpha release and the time required for completion of uterine involution showed insignificant correlation . However, the longer duration of PGF2 alpha release recorded in the 2 cows with intrauterine infections are related to the increased frequency of infections. Vet Res Commun, 1996, 20(5), 473 - 9 The effect of induced fever on the biokinetics of norfloxacin and its interaction with probenecid in goats; Jha K et al.; The kinetic profiles of norfloxacin were evaluated in afebrile, febrile and probenecid pre-treated (70 mg/kg orally) febrile goats after a single intravenous (i.v.) dose (5 mg/kg) . Fever was induced and maintained for 12 h by injecting Escherichia coli endotoxin (0.2 microgram/kg, i.v.) and repeating it in half the dose (0.1 microgram/kg) 5 h later . The plasma pharmacokinetic values for norfloxacin were best represented using a two-compartment open model . The peak norfloxacin plasma level of 90.52 +/- 3.18 micrograms/ml attained in the probenecid pre-treated febrile goats was higher than that in the febrile (75.46 +/- 0.72 micrograms/ml) or afebrile goats (62.25 +/- 1.23 micrograms/ml) . ClB and Kel values were significantly (p < 0.01) decreased in febrile compared with afebrile goats . These values were further reduced in febrile goats after probenecid pre-treatment . However, t1/2 beta was not affected by the fever-probenecid interaction . Norfloxacin may be used as an infusion with probenecid in caprine diseases where very high plasma levels are required to combat resistant organisms such as Bacteroides. Enferm Infecc Microbiol Clin, 1996 Jan, 14(1), 16 - 20 {Necrotizing infections of the soft tissues}; de Polavieja MG et al.; OBJECT: In necrotizing soft tissue infections (NSTI) may be implicated skin, subcutaneous tissue, fascia and skeletal muscle, and include different clinical entities associated with high morbidity and mortality rates . Diagnosis must be early and its initial management must be common . METHODS: A retrospective study of 24 patients diagnosed of NSTI is made . Associated factors with the development of these infections, and its treatment are analysed . It is effectuated an statistical analysis of mortality and related factors . RESULTS: Five cases were Necrotizing Cellulitis, fourteen Necrotizing Fasciitis, three Myonecrosis, one gangrenous pyoderma and one case was a disseminated gluteus abscess . Surgical treatment was resection and debridament--in every case reaching to healthy tissue . Broad-spectrum antibiotics were always associated to surgery . E . coli and Bacteroides have been the most frequent microorganism isolated . 23 patients were operated once at least; 66 surgical interventions were made (mean: 2.86; range: 1-7) . Overall mortality of this series was 29% . CONCLUSIONS: Early and aggressive treatment are the corner stone in the management of these patients . With such therapeutic premise, in our series we had a mortality of 29% . Factors associated with a worse outcome have been advanced age, the presence of an underlying disease in the moment of developing the infection and high levels of urea at admittance. J Periodontal Res, 1996 Jan, 31(1), 27 - 35 Comparison of randomly cloned and whole genomic DNA probes for the detection of Porphyromonas gingivalis and Bacteroides forsythus; Wong M et al.; Whole genomic and randomly-cloned DNA probes for two fastidious periodontal pathogens, Porphyromonas gingivalis and Bacteroides forsythus were labeled with digoxigenin and detected by a colorimetric method . The specificity and sensitivity of the whole genomic and cloned probes were compared . The cloned probes were highly specific compared to the whole genomic probes . A significant degree of cross-reactivity with Bacteroides species, Capnocytophaga sp . and Prevotella sp . was observed with the whole genomic probes . The cloned probes were less sensitive than the whole genomic probes and required at least 10(6) target cells or a minimum of 10 ng of target DNA to be detected during hybridization . Although a ten-fold increase in sensitivity was obtained with the whole genomic probes, cross-hybridization to closely related species limits their reliability in identifying target bacteria in subgingival plaque samples. Appl Environ Microbiol, 1996 Jan, 62(1), 196 - 202 Use of a modified Bacteroides-Prevotella shuttle vector to transfer a reconstructed beta-1,4-D-endoglucanase gene into Bacteroides uniformis and Prevotella ruminicola B(1)4; Gardner RG et al.; A carboxymethyl cellulase (CMCase) gene from Prevotella ruminicola B(1)4 was reconstructed by adding a cellulose binding domain from a Thermomonospora fusca cellulase and was conjugally transferred from Escherichia coli to Bacteroides uniformis 0061 by using a chloramphenicol and tetracycline resistance shuttle vector (pTC-COW) . pTC-COW was specifically constructed to facilitate conjugal transfer of vectors from B . uniformis donors to P . ruminicola recipients . B . uniformis transconjugants containing CMCase constructs cloned into pTC-COW expressed Cmr, but they did not produce the reconstructed CMCase until a xylanase promoter from P . ruminicola 23 was added upstream of the CMCase (pTC-XRCMC) . The xylanase promoter allowed the B . uniformis transconjugants to produce large amounts of the reconstructed CMCase, which was present on the outside surface of the cells . Although the reconstructed CMCase alone did not allow B . uniformis to grow on acid-swollen cellulose, rapid growth was observed when two exocellulases were added to the culture supernatant . Under these conditions, the reconstructed CMCase permitted faster growth than the wild-type CMCase . The frequency of transfer of pTC-XRCMC from B . uniformis to P . ruminicola B(1)4 was increased 100-fold when strictly anaerobic conditions, nitrocelluose filters (cell immobilization), and more stringent selections were employed . Although the P . ruminicola B(1)4 (pTC-XRCMC) transconjugates expressed Tcr and had DNA that hybridized with a probe to the shuttle vector, these transconjugants did not produce detectable levels of the reconstructed CMCase even when xylan was the carbon source . On the basis of these results, it appears that not all of the promoters recognized by B . uniformis and P . ruminicola 23 are functional in P . ruminicola B(1)4 . However, the results with B . uniformis suggest that the introduction of a P . ruminicola B(1)4 promoter should allow expression of the reconstructed CMCase in P . ruminicola B(1)4. Infect Immun, 1996 Jan, 64(1), 113 - 9 Bacteroides fragilis enterotoxin induces cytoskeletal changes and surface blebbing in HT-29 cells; Donelli G et al.; Certain strains of the anaerobic bacterium Bacteroides fragilis are known to produce an enterotoxin of about 20 kDa which is able to induce a fluid response in ligated intestinal loops and a cytotoxic response in HT-29 cells . It presents protease activity, belonging to a family of metalloproteases termed metzincins . In order to investigate the mode of action of the enterotoxin in cultured cells, we performed a study with HT-29 cells, using both fluoresence and electron microscopy . Treated cells underwent morphological changes, mainly consisting of the retraction of the cell body and the formation of numerous blebs on the cell surface . The microfilament system was reorganized, the F-actin being condensed as a ring at the cell periphery, whereas other cell organelles appeared to be unaffected . All these changes, clearly visible after 3 h of exposure to the toxin, were reversed within 24 h of treatment . By inhibiting the protease activity of the toxin with specific metal chelators, the cytoskeletal effects were also prevented . Thus, B . fragilis enterotoxin appears to act on cells by reversibly modifying the actin cytoskeleton, an effect probably dependent on its proteolytic activity. N Engl J Med, 1995 Dec 28, 333(26), 1737 - 42 Association between bacterial vaginosis and preterm delivery of a low-birth-weight infant . The Vaginal Infections and Prematurity Study Group; Hillier SL et al.; BACKGROUND . Bacterial vaginosis is believed to be a risk factor for preterm delivery . We undertook a study of the association between bacterial vaginosis and the preterm delivery of infants with low birth weight after accounting for other known risk factors . METHODS . In this cohort study, we enrolled 10,397 pregnant women from seven medical centers who had no known medical risk factors for preterm delivery . At 23 to 26 weeks' gestation, bacterial vaginosis was determined to be present or absent on the basis of the vaginal pH and the results of Gram's staining . The principal outcome variable was the delivery at less than 37 weeks' gestation of an infant with a birth weight below 2500 g . RESULTS . Bacterial vaginosis was detected in 16 percent of the 10,397 women . The women with bacterial vaginosis were more likely to be unmarried, to be black, to have low incomes, and to have previously delivered low-birth-weight infants . In a multivariate analysis, the presence of bacterial vaginosis was related to preterm delivery of a low-birth-weight infant (odds ratio, 1.4; 95 percent confidence interval, 1.1 to 1.8) . Other risk factors that were significantly associated with such a delivery in this population were the previous delivery of a low-birth-weight infant (odds ratio, 6.2; 95 percent confidence interval, 4.6 to 8.4), the loss of an earlier pregnancy (odds ratio, 1.7; 1.3 to 2.2), primigravidity (odds ratio, 1.6; 1.1 to 1.9), smoking (odds ratio, 1.4; 1.1 to 1.7); and black race (odds ratio, 1.4; 1.1 to 1.7) . Among women with bacterial vaginosis, the highest risk of preterm delivery of a low-birth-weight infant was found among those with both vaginal bacteroides and Mycoplasma hominis (odds ratio, 2.1; 95 percent confidence interval, 1.5 to 3.0) . CONCLUSIONS . Bacterial vaginosis was associated with the preterm delivery of low-birth-weight infants independently of other recognized risk factors. Microbiol Rev, 1995 Dec, 59(4), 579 - 90 Conjugative transposons: an unusual and diverse set of integrated gene transfer elements; Salyers AA et al.; Conjugative transposons are integrated DNA elements that excise themselves to form a covalently closed circular intermediate . This circular intermediate can either reintegrate in the same cell (intracellular transposition) or transfer by conjugation to a recipient and integrate into the recipient's genome (intercellular transposition) . Conjugative transposons were first found in gram-positive cocci but are now known to be present in a variety of gram-positive and gram-negative bacteria also . Conjugative transposons have a surprisingly broad host range, and they probably contribute as much as plasmids to the spread of antibiotic resistance genes in some genera of disease-causing bacteria . Resistance genes need not be carried on the conjugative transposon to be transferred . Many conjugative transposons can mobilize coresident plasmids, and the Bacteroides conjugative transposons can even excise and mobilize unlinked integrated elements . The Bacteroides conjugative transposons are also unusual in that their transfer activities are regulated by tetracycline via a complex regulatory network. Environ Health Perspect, 1995 Nov, 103 Suppl 8, 263 - 8 Bacterial infection as a cause of cancer; Parsonnet J; Bacterial infections traditionally have not been considered major causes of cancer . Recently, however, bacteria have been linked to cancer by two mechanisms: induction of chronic inflammation and production of carcinogenic bacterial metabolites . The most specific example of the inflammatory mechanism of carcinogenesis is Helicobacter pylori infection . H . pylori has been epidemiologically linked to adenocarcinoma of the distal stomach by its propensity to cause lifelong inflammation . This inflammation is in turn thought to cause cancer by inducing cell proliferation and production of mutagenic free radicals and N-nitroso compounds . H . pylori is the first bacterium to be termed a definite cause of cancer in humans by the International Agency for Research on Cancer . Mutagenic bacterial metabolites are also suspected to increase risk for cancer . This model is best exemplified in colon cancer . Bile salt metabolites increase colonic cell proliferation . Exogenous compounds such as rutin may be metabolized into mutagens by resident colonic flora . Moreover, Bacteroides species can produce fecapentaenes, potent in vitro mutagens, in relatively high concentrations . In vivo data on human carcinogenesis by bacterial metabolites, however, are inconsistent . Local bacterial infections may also predispose to nonnodal lymphomas, although the mechanisms for this are unknown . Gastric lymphomas and immunoproliferative small intestinal disease have been most strongly linked to underlying bacterial infection . Because bacterial infections can be cured with antibiotics, identification of bacterial causes of malignancy could have important implications for cancer prevention. Clin Infect Dis, 1995 Nov, 21(5), 1114 - 20 Septic thrombophlebitis of the portal vein (pylephlebitis): diagnosis and management in the modern era; Plemmons RM et al.; Pylephlebitis usually occurs secondary to infection in the region drained by the portal venous system . We describe a case of pylephlebitis at our institution and examine 18 other cases culled from the literature since 1979, reviewing diagnostic and management issues . A precipitating focus of infection (most commonly diverticulitis) was identified in 13 (68%) of the cases . Bacteremia (often polymicrobial) was present in 88% of the patients . The most common blood isolate was Bacteroides fragilis . Overall mortality was 32%, but most of the patients who died had severe sepsis prior to the initiation of antibiotic therapy . In no case was improvement in a patient's clinical status clearly attributable to the use of heparin, but some beneficial effect of anticoagulation could not be ruled out . This report is the first to examine the published experience with pylephlebitis during the era of antibiotics and modern imaging and is also the first to review critically the role of anticoagulation in the management of this disease. J Gastroenterol, 1995 Nov, 30 Suppl 8, 45 - 7 The pathogenic role of Bacteroides vulgatus in patients with ulcerative colitis; Bamba T et al.; To clarify the role played by Bacteroides vulgatus in the pathogenesis of ulcerative colitis, the serum antibody titer to the outer membrane protein of B . vulgatus, obtained from the inflamed rectal mucosa, and measured by enzyme-linked immunosorbent assay, was investigated in patients with ulcerative colitis . High serum antibody titers to the outer membrane protein of B . vulgatus were found in these patients . The specific outer membrane protein of B . vulgatus reactive to serum antibody was recognized as a 26-kDa protein . These findings indicate that B . vulgatus containing a 26-kDa protein in the outer membrane may be implicated in the pathogenesis of ulcerative colitis. Br J Nutr, 1995 Nov, 74(5), 733 - 41 Impact of polyunsaturated fatty acids on human colonic bacterial metabolism: an in vitro and in vivo study; Thompson L et al.; Dietary polyunsaturated fatty acids (PUFA) reduce colonic proliferation and exert a mild laxative effect . We have studied the effect of the highly unsaturated eicosapentaenoic acid ethyl ester (EPA-EE) on the growth and metabolism of colonic bacteria in vitro, and in vivo . For the in vitro study, growth was assessed by viable counts . Bacteroides thetaiotaomicron was significantly inhibited in anaerobic media containing EPA-EE at concentrations > 7 milligrams . Escherichia coli was apparently resistant even at 100 milligrams . For the in vivo study, ten healthy volunteers ingested 18 g EPA-EE/d for 7 d . Stool frequency, 24 h stool weight and whole-gut transit time were assessed together with breath H2 and 14CO2 excretion following oral ingestion of 15 g lactitol labelled with 0.18 MBq {14C}lactitol . The area under the breath-H2-time curve was significantly reduced by EPA-EE, from a control value of 690.3 (SE 94.2) ppm.h to 449.5 (SE 91.7) ppm.h . Percentage dose of 14CO2 excreted, total stool weight and whole-gut transit time were unaltered, being respectively 24 (SE 2)%, 281 (SE 66) g and 45 (SE 4) h with EPA-EE v . control values of 27 (SE 1)%, 300 (SE 89) g and 42 (SE 5) h . It is concluded that dietary supplementation with EPA-EE reduces breath H2 excretion without apparently impairing overall colonic carbohydrate fermentation . The observed reduction may reflect utilization of H2 to hydrogenate the five double bonds of EPA-EE. Dig Dis Sci, 1995 Nov, 40(11), 2456 - 9 Effect of Bacteroides melaninogenicus culture supernatant and deconjugated bile salt on lipid absorption; Healy MJ et al.; Lipid malabsorption is a common clinical manifestation of small bowel bacterial overgrowth . Its pathogenesis, however, remains controversial . Bacteroides melaninogenicus ssp . intermedius, an anaerobic bacterium, is commonly isolated from the upper bowel of patients with small intestinal bacterial overgrowth . The effects of a culture supernate of this organism and deoxycholate, an unconjugated bile salt, on intestinal oleic acid absorption were examined using a rat closed-loop model . The supernatant reduced the in vitro uptake of oleic acid by 19% (P< 0.001) . Deoxycholate did not significantly reduce the lipid absorption . Combined supernate and deoxycholate did not have an additive effect on absorption of oleic acid . We conclude that anaerobic bacterial products may contribute to the malabsorption of lipid in the setting of bacterial overgrowth of the small bowel. Carbohydr Res, 1995 Oct 2, 275(2), 333 - 41 Structural elucidation of the capsular polysaccharide of Bacteroides fragilis strain 23745M1; Pavliak V et al.; The capsule of Bacteroides fragilis (ATCC23745) consists of two distinct polysaccharides, the separation of which could not be accomplished . The mouse-passaged strain (23745M1), however, yielded a preponderant polysaccharide which was isolated and purified . Using mainly high resolution NMR spectroscopy, the structure of the polysaccharide was elucidated and it is composed of the following repeating unit: {formula see text} Oral Microbiol Immunol, 1995 Oct, 10(5), 284 - 7 Molecular genetic detection of Bacteroides heparinolyticus in adult periodontitis; Ashimoto A et al.; Bacteroides heparinolyticus in subgingival plaque was identified using a digoxigenin-labeled whole genomic DNA probe and a polymerase chain reaction (PCR) assay based on 16S rRNA species-specific primers (5'-ATG GTG ATT CCG CAT GGT TTC TCC-3' (base position, 188-212) and 5'-CAA ACT TTC ACA GCT GAC TTA AGC-3' (592-615)) . Subgingival specimens obtained by paper points from 3 deep periodontal pockets in each of 113 adults were examined . The DNA probe reacted with all pure isolates tested of B . heparinolyticus and did not react with other oral species tested; the probe showed positive reactions in 74.3% of the patient samples examined . The PCR primers produced the 428 bp species specific amplification product in all B . heparinolyticus test strains and did not reveal detectable amplicons with strains of other subgingival species . The PCR method detected 50 B . heparinolyticus cells dispersed in subgingival plaque . PCR only revealed B . heparinolyticus in 6.2% of the patient samples studied . The higher level of positive specimens with the DNA probe was probably due to false-positive reactions from cross-hybridization with unknown subgingival species . This study suggests that the PCR method amplifying specific 16S rRNA sequences represents an easy and valuable means to detect B . heparinolyticus in subgingival plaque . The low prevalence of subgingival B . heparinolyticus does not incriminate the organism in the etiology of adult periodontitis. J Appl Bacteriol, 1995 Oct, 79(4), 417 - 24 Expression of a cloned cellulase/xylanase gene from Prevotella ruminicola in Bacteroides vulgatus, Bacteroides uniformis and Prevotella ruminicola; Daniel AS et al.; A new shuttle vector, pRH3 (8.7 kb), was constructed for use in Prevotella/Bacteroides host strains . This vector combines the pRRI2 replicon from P . ruminicola, pBluescript sequences and a tetQ marker gene for selection in Prevotella/Bacteroides hosts . Following insertion of a fragment carrying an endoglucanase/xylanase gene from P . ruminicola 23 into the multiple cloning site, the resulting construct, pRH3X, was introduced into B . vulgatus 1447, B . uniformis 1100 and P . ruminicola 2202 . This resulted in increases of between 4 and 50-fold in CM-cellulase and xylanase activities in cells grown with glucose . In contrast activities were barely detectable for the same construct in E . coli DH5 alpha . Most of the total xylanase activity produced was found within the cell in P . ruminicola 2202 and B . vulgatus 1447 transformed with pRH3X, and in P . ruminicola 23 . An osmotic shock experiment indicated that a significant proportion of the xylanase activity in B . vulgatus 1447 cells carrying pRH3X was periplasmic. Infect Immun, 1995 Oct, 63(10), 3820 - 6 Proteolytic activity of the Bacteroides fragilis enterotoxin causes fluid secretion and intestinal damage in vivo; Obiso RJ Jr et al.; Strains of Bacteroides fragilis that produce an enterotoxin have been implicated in diarrheal disease in farm animals and humans during the past decade . Our laboratory has purified and characterized this enterotoxin as a single polypeptide (M(r), approximately 20,000) . Recently, we used PCR to clone and sequence the enterotoxin gene from B . fragilis and showed that it exhibits significant homology with extracellular metalloproteases . Further studies showed that the purified enterotoxin has protease activity . To further characterize the role of this enterotoxin in diarrheal disease, we studied the histological and pathological effects of highly purified B . fragilis enterotoxin in lamb, rabbit, and rat ligated intestinal loops . When the enterotoxin was injected into ligated ileal and colonic loops, there was significant tissue damage and subsequent fluid accumulation . The fluid response in the ileum was greater in lambs than in rabbits and rats, whereas the fluid response in the colon was greater in rabbits than in lambs and rats . Analysis of the intestinal fluid elicited by the enterotoxin revealed an accumulation of chloride and sodium as well as albumin and total protein . Histological examination revealed mild necrosis of epithelial cells, crypt elongation, villus attenuation, and hyperplasia . There was extensive detachment and rounding of surface epithelial cells and an infiltration of neutrophils . Enterotoxic activity was inhibited by the metal chelators EDTA and 1,10-phenanthroline; to some degree, the enterotoxic activity could be reconstituted by the addition of zinc to the chelated enterotoxin . Our results indicate that the enterotoxin elicits a significant fluid response subsequent to tissue damage in the small and large intestine . These data further support the idea that this enterotoxin is an important virulence factor in B . fragilis-associated diarrhea. Clin Immunol Immunopathol, 1995 Oct, 77(1), 82 - 8 CD4+ T cells mediate preexposure-induced increases in murine intraabdominal abscess formation; Sawyer RG et al.; We have previously shown that an increased number of Escherichia coli/Bacteroides fragilis intraabdominal abscesses are produced in mice after preexposure to small numbers of live E . coli or B . fragilis . Splenic lymphocyte subset changes and the importance of different elements of the immune response in this system were studied . Preexposure to bacteria induced a significant increase in the percentage of splenic T cells without altering the CD4/CD8 ratio . The passive transfer of either 10(7) mixed splenic lymphocytes, 5 x 10(6) mixed T cells, or 2.5 x 10(6) CD4+ T cells from preexposed animals to naive siblings 24 hr prior to abscess induction resulted in increased abscess formation . Transfer of serum, B cells, < 10(7) lymphocytes, CD8+ T cells, or any cell type from naive animals did not change abscess number . The bacterial composition of abscesses changed only in animals receiving either serum or B cells from donors preexposed to B . fragilis, where an increased number of B . fragilis per abscess was found . The CD4+ T cell response can be altered by transient infections and is critical to subsequent abscess formation, and a concurrent humoral response may play a role in determining an abscess' ultimate bacterial composition. Curr Microbiol, 1995 Oct, 31(4), 215 - 9 Detection of Bacteroides fragilis in clinical specimens by polymerase chain reaction amplification of the neuraminidase gene; Jotwani R et al.; The polymerase chain reaction (PCR) was used in an attempt to detect Bacteroides fragilis by amplifying a segment of the gene encoding B . fragilis neuraminidase . Forty-five reference strains representing 45 species and 113 clinical isolates were tested . Only B . fragilis was PCR positive, except for Bacteroides merdae ATCC 43184, which gave a band by ethidium bromide staining that showed no signal by Southern hybridization . Using a protocol that employed DNA extraction by Sepa Gene kit and a highly sensitive digoxigenin-chemiluminescence detection system, detection of B . fragilis by PCR was in complete agreement with culture results for 44 clinical specimens from which a wide range of aerobic and anaerobic organisms and fungi were recovered. Zentralbl Veterinarmed B, 1995 Sep, 42(7), 415 - 20 Isolation of Bacteroides ureolyticus from vaginal discharge of mares; Fodor L et al.; A total of seven Bacteroides ureolyticus strains were isolated from the cervix and the clitoral fossa of mares with vaginal discharge . No other bacteria capable of causing metritis or vaginitis were isolated from the samples . The isolated strains resembled Taylorella equigenitalis . Both species are catalase, oxidase and alkaline phosphatase positive, but, in addition to these characteristics, B . ureolyticus strains produced urease and they could not tolerate 10% O2 . They also failed to be agglutinated in a hyperimmune serum raised against T . equigenitalis; however, B . ureolyticus and T . equigenitalis were agglutinated in the slide agglutination test in a serum produced against one of the B . ureolyticus isolates . Further investigations are needed to clarify the pathologic role of B . ureolyticus in genital infections of mares and other animals. J Bacteriol, 1995 Sep, 177(17), 4992 - 9 Location and characteristics of the transfer region of a Bacteroides conjugative transposon and regulation of transfer genes; Li LY et al.; Many Bacteroides clinical isolates contain large conjugative transposons, which excise from the genome of a donor and transfer themselves to a recipient by a process that requires cell-to-cell contact . It has been suggested that the transfer intermediate of the conjugative transposons is a covalently closed circle, which is transferred by the same type of rolling circle mechanism used by conjugative plasmids, but the transfer origin of a conjugative transposon has not previously been localized and characterized . We have now identified the transfer origin (oriT) region of one of the Bacteroides conjugative transposons, TcrEmr DOT, and have shown that it is located near the middle of the conjugative transposon . We have also identified a 16-kbp region of the conjugal transposon which is necessary and sufficient for conjugal transfer of the element and which is located near the oriT . This same region proved to be sufficient for mobilization of coresident plasmids and unlinked integrated elements as well as for self-transfer, indicating that all of these activities are mediated by the same transfer system . Previously, we had reported that disruption of a gene, rteC, abolished self-transfer of the element . rteC is one of a set of rte genes that appears to mediate tetracycline induction of transfer activities of the conjugative transposons . On the basis of these and other data, we had proposed that RteC activated expression of transfer genes . We have now found, however, that when the transfer region of TcrEmr DOT was cloned as a plasmid that did not contain rteC and the plasmid (pLYL72) was tested for transfer out of a Bacteroides strain that did not have a copy of rteC in the chromosome, the plasmid was self-transmissible without tetracycline induction . This and other findings suggest that RteC is not an activator transfer genes but is stimulating transfer in some other way. Am J Gastroenterol, 1995 Sep, 90(9), 1518 - 20 An unusual presentation of colon cancer: purulent pericarditis and cardiac tamponade due to Bacteroides fragilis; Lam S et al.; Extra-abdominal infections caused by colonic anaerobic organisms have been shown to be associated with colorectal cancers . We report a very unusual case of a patient who suffered from cardiac tamponade secondary to anaerobic bacterial pericarditis caused by Bacteroides fragilis as an initial presentation of colon cancer . The medical literature is reviewed. J Bacteriol, 1995 Sep, 177(18), 5270 - 5 Genotypic identification of two groups within the species Bacteroides fragilis by ribotyping and by analysis of PCR-generated fragment patterns and insertion sequence content; Podglajen I et al.; Molecular typing allowed the separation of the species Bacteroides fragilis into two genotypically distinct groups . A unique set of 50 strains of B . fragilis carrying the chromosomal metallo-beta-lactamase gene cfiA was subjected to a comparative analysis with respect to sets of up to 250 randomly collected strains devoid of this gene . The two groups were found to be distinct on the basis of the following results: (i) ribotyping, after DNA digestion with AvaI, revealed a practically homogeneous DNA fragment pattern for the cfiA-positive strains and distinct multiple patterns for the cfiA-negative strains; (ii) PCR, arbitrarily primed with an experimentally selected decamer, generated fragment patterns typical for the strains of each group; (iii) the three insertion sequences described to date in the species B . fragilis, i.e., IS4351, IS942, and IS1186, were all but confined to the cfiA-positive group, in which they were capable of providing promoter sequences for the transcription of cfiA; and (iv) the cepA gene, encoding the so-called endogenous cephalosporinase of B . fragilis, was found exclusively in the cfiA-negative group, in which it was present in ca . 70% of the strains . The cfiA-, cepA-negative fraction was not characterized further . In a natural population of 500 randomly selected strains of B . fragilis, the cfiA-positive and cfiA-negative groups represented ca . 3 and 97% of the strains, respectively . Analysis of 82 metabolic traits revealed no difference between the two groups. Microb Pathog, 1995 Aug, 19(2), 117 - 28 The pentameric structure of IgM is necessary to enhance opsonization of Bacteroides thetaiotaomicron and Bacteroides fragilis via the alternative complement pathway; Bjornson AB et al.; Studies were conducted to investigate the mechanisms by which natural IgM antibodies act together with the alternative complement pathway to promote opsonization and adherence of encapsulated Bacteroides thetaiotaomicron and Bacteroides fragilis to polymorphonuclear leukocytes (PMN) . A model system consisting of the six isolated proteins of the alternative pathway was used . A comparison of the opsonic effects of pentameric and monomeric forms of isolated normal IgM demonstrated that, although the monomeric form bound to Bacteroides as effectively as the pentameric form and promoted complement deposition to the same extent, it was unable to enhance alternative pathway-dependent opsonization and adherence of Bacteroides to PMN . When opsonization was performed in two steps with pentameric IgM added either before or after alternative pathway components, a marked enhancement of adherence to PMN was observed only in the former case, suggesting IgM must act prior to complement to be effective . Electron microscopic studies demonstrated that, when added with complement, pentameric IgM, but not monomeric IgM, stabilized the bacterial capsule to the dehydration in dimethylformamide used for embedding in Lowicryl K4M . A strong correlation was observed between capsular stability and ability to be bound by PMN . The results suggest that pentameric IgM alters the structure of capsular components, perhaps through crosslinking, and this is in turn facilitates interaction of C3bi and C3b with CR3 and CR1, their respective receptors on PMN. J Virol Methods, 1995 Aug, 54(2-3), 121 - 30 Simple concentration method for bacteriophages of Bacteroides fragilis in drinking water; Lucena F et al.; A membrane of inorganic material with a honeycomb pore structure was used to concentrate phages infecting Bacteroides fragilis from drinking water . Phages were removed from the membrane with 0.25 M glycine buffer pH 9.5 . Phages were not inactivated by storage in this buffer neutralized to pH 7.0 for at least 9 days at 4 degrees C . The method allows recovery of around 50% in drinking water . When the turbidity of the water increased, the efficiency of the concentration decreased markedly . The efficiency of concentration was evaluated versus a presence/absence test in 317 water samples with turbidity level below the threshold of drinking water . Results obtained by concentration of 11 provided data which were significantly more informative than the presence/absence tests carried out on 100 ml . A number of additional tests carried out with both somatic and F-specific coliphages indicated that these conclusions can be extended to these groups of bacteriophages. J Bacteriol, 1995 Aug, 177(15), 4466 - 73 Characterization and DNA sequence of the mobilization region of pLV22a from Bacteroides fragilis; Novicki TJ et al.; A 4.2-kb plasmid (pLV22a) native to Bacteroides fragilis LV22 became fused to a transfer-deficient Bacteroides spp.-Escherichia coli shuttle vector by an inverse transposition event, resulting in a transferrable phenotype . The transfer phenotype was attributable to pLV22a, which was also capable of mobilization within E . coli when coresident with the IncP beta R751 plasmid . Transposon mutagenesis with Tn1000 localized the mobilization region to a 1.5-kb DNA segment in pLV22a . The mobilization region has been sequenced, and five open reading frames have been identified . Mutants carrying disruptions in any of the three genes designated mbpA, mbpB, and mbpC and coding for deduced products of 11.3, 30.4, and 17.1 kDa, respectively, cannot be mobilized when coresident with R751 . Mutations in all three genes can be complemented in the presence of the respective wild-type genes, indicating that the products of mbpA, mbpB, and mbpC have roles in the mobilization process and function in trans . The deduced 30.4-kDa MbpB protein contains a 14-amino-acid conserved motif that is also found in the DNA relaxases of a variety of conjugal and mobilizable plasmids and the conjugative transposon Tn4399 . Deletion analysis and complementation experiments have localized a cis-acting region of pLV22a within mbpA. J Med Microbiol, 1995 Aug, 43(2), 99 - 109 Immunological detection of Bacteroides fragilis in clinical samples; Patrick S et al.; A monospecific polyclonal antiserum, prepared against Bacteroides fragilis common polysaccharide antigen purified by polyacrylamide gel immunoblot detected B . fragilis, B . thetaiotaomicron, B . ovatus and Prevotella melaninogenica in pus samples from various anatomical sites by immunofluorescence microscopy of the pus . With standard clinical laboratory culture methods, 36% of 147 samples were positive for one or more of the above bacteria . Of these, B . fragilis accounted for 33% . By immunofluorescent labelling of pus with the common antigen antiserum the detection of these bacteria in the samples increased to 50% . All nine of the blood cultures in which B . fragilis was detected by culture contained bacteria positive for the common antigen . Immunofluorescent labelling of pus samples with a selection of monoclonal antibodies specific for surface polysaccharides which are known to be antigenically variable in culture in vitro and in an animal model of infection showed that these polysaccharides are also variable in natural infection . The results indicate that the common polysaccharide antigen, in contrast to the variable surface polysaccharides, is a suitable target for the immunodetection of B . fragilis in clinical samples from a range of anatomical sites. Kansenshogaku Zasshi, 1995 Aug, 69(8), 903 - 7 {Detection of Bacteroides fragilis enterotoxin from extraintestinal clinical isolates of B . fragilis group organisms}; Kato N et al.; A total of 300 extraintestinal clinical isolates of Bacteroides fragilis group organisms were assayed for enterotoxin using HT29/C1 cells . Of the 179 strains of B . fragilis, 36 (20.0%) were positive for cell culture assay, a positivity which was neutralized by anti-B . fragilis enterotoxin serum; all 36 strains were confirmed to be enterotoxigenic B . fragilis (ETBF) . Among B . fragilis isolated from blood, 18 (31%) of 59 strains were ETBF while 18 (15.0%) of 120 strains isolated from non-blood specimens were ETBF (p < 0.05) . None of the 121 strains of the B . fragilis group other than B . fragilis were positive for cell culture assay . These results demonstrated that isolation of ETBF from extraintestinal clinical specimens is not uncommon and that B . fragilis enterotoxin, a metalloprotease, may play a role as a pathogenic factor in blood stream infection and sepsis. Microbiology, 1995 Aug, 141 ( Pt 8), 1969 - 76 Immune reactions to Bacteroides fragilis populations with three different types of capsule in a model of infection; Patrick S et al.; The survival and growth of populations of the obligately anaerobic pathogenic bacterium Bacteroides fragilis enriched for large capsules (LCs), small capsules (SCs) or an electron-dense layer (EDL; non-capsulate by light microscopy) were examined in a mouse model of infection over a minimum period of 20 d . Chambers which allowed the influx of leukocytes, but not the efflux of bacteria, were implanted in the mouse peritoneal cavity . The LC and EDL populations consistently attained viable cell densities of the order of 10(8)-10(9) c.f.u . ml-1 within 24 h, whereas the SC population did not . However, after 3 d, all three bacterial populations maintained total viable numbers of 10(8)-10(9) c.f.u . ml-1 within the chambers . LC expression was selected against within 24 h in the model, the populations becoming non-capsulate by light microscopy, whereas in the SC population expression of the SC was retained by approximately 90% of the population . The EDL population remained non-capsulate by light microscopy throughout . Lymphocytes infiltrated the chambers to an equal extent for all three B . fragilis populations and at approximately 1000 times higher concentration than chambers which contained only quarter-strength Ringer's solution . The presence of neutrophils within the chambers did not cause a decrease in the total viable bacterial count . Each population elicited antibodies specific for outer-membrane proteins and polysaccharide, as detected by immunoblotting, which cross-reacted with the other populations . Differences were observed in the immunogenicity of the outer-membrane proteins within the three populations . Neutrophils were initially the predominant cell type in the chambers, but as the total leukocyte count increased with incubation time, neutrophils were outnumbered by other leukocytes.(ABSTRACT TRUNCATED AT 250 WORDS) Infect Immun, 1995 Jul, 63(7), 2459 - 66 Characterization of a Tn4351-generated hemin uptake mutant of Porphyromonas gingivalis: evidence for the coordinate regulation of virulence factors by hemin; Genco CA et al.; The ability of Porphyromonas gingivalis to acquire iron in the iron-limited environment of the host is crucial to the colonization of this organism . We report here on the isolation and characterization of a transpositional insertion mutant of P . gingivalis A7436 (designated MSM-3) which is defective in the utilization and transport of hemin . P . gingivalis MSM-3 was selected on the basis of its nonpigmented phenotype on anaerobic blood agar following mutagenesis with the Bacteroides fragilis transposon Tn4351 . P . gingivalis MSM-3 grew poorly when supplied with hemin as a sole source of iron; however, growth was observed with hemoglobin or inorganic iron . P . gingivalis MSM-3 grown in either hemin-replete or hemin-depleted conditions bound and transported less {14C}hemin or {59Fe}hemin than did the parent strain . At 4 h, P . gingivalis MSM-3 grown in hemin-replete conditions transported only 10,000 pmol of hemin per mg of protein, or 14% of the amount transported by P . gingivalis A7436 . Unlike P . gingivalis A7436, hemin binding and transport by P . gingivalis MSM-3 were not tightly regulated by hemin or iron . Examination of P . gingivalis MSM-3 cultures by electron microscopy revealed an overproduction of membrane vesicles, and determination of the dry weight of purified vesicles indicated that P . gingivalis MSM-3 produced twice as much membrane vesicles as did strain A7436 . Extracellular vesicles isolated from P . gingivalis MSM-3 also were found to express increased hemolytic and trypsin-like protease activities compared with the parent strain . When inoculated into subcutaneous chambers implanted in mice, P . gingivalis MSM-3 was highly infectious and more invasive than the parent strain, as indicated by secondary lesion formation and death . Taken together, these results indicate that the decreased transport of hemin by P . gingivalis MSM-3 results in the increased expression of several virulence factors which may be coordinately regulated by hemin. J Bacteriol, 1995 Jul, 177(14), 3940 - 5 The mobilization regions of two integrated Bacteroides elements, NBU1 and NBU2, have only a single mobilization protein and may be on a cassette; Li LY et al.; Bacteroides conjugative transposons can act in trans to excise, circularize, and transfer unlinked integrated elements called NBUs (for nonreplicating Bacteroides units) . Previously, we localized and sequenced the mobilization region of one NBU, NBU1, and showed that this mobilization region was recognized by the IncP plasmids RP4 and R751, as well as by the Bacteroides conjugative transposons . We report here that the single mobilization protein carried by NBU1 appears to be a bifunctional protein that binds to the oriT region and catalyzes the nicking reaction that initiates the transfer process . We have also localized and sequenced the mobilization region of a second NBU, NBU2 . The NBU2 mobilization region was 86 to 90% identical at the DNA sequence to the oriT-mob region of NBU1 . The high sequence similarity between NBU1 and NBU2 ended abruptly after the stop codon of the mob gene and about 1 kbp upstream of the oriT region, indicating that the oriT-mob regions of NBU1 and NBU2 may be on some sort of cassette . A region on NBU1 and NBU2 which lies immediately upstream of the oriT region had 66% sequence identity to a region upstream of the oriT region on a mobilizable transposon, Tn4399, an element that had previously appeared to be completely unrelated to the NBUs. Antimicrob Agents Chemother, 1995 Jul, 39(7), 1458 - 61 Purification and characterization of a new beta-lactamase from Bacteroides uniformis; Hedberg M et al.; A beta-lactam-resistant Bacteroides uniformis strain was isolated from a clinical specimen . The strain produced large amounts of beta-lactamase and was resistant to penicillins and cephalosporins . The specific activity of the unpurified beta-lactamase was 4.8 U/mg of protein with nitrocefin as the substrate . The enzyme was purified 188-fold by Q-Sepharose, Sephacryl S-300, and Mono Q column passages . Kinetic parameters of the enzyme were determined by a micromethod performed in microtiter plates . beta-Lactamase was inhibited by cefoxitin and imipenem and hydrolyzed cephalosporins more rapidly than penicillins . The molecular weight was determined by sodium dodecyl sulfate-gradient gel electrophoresis to be 32,500, and the isoelectric point was 4.5. Pharmacotherapy, 1995 Jul-Aug, 15(4), 479 - 86 Comparison of the pharmacodynamic activity of cefotaxime plus metronidazole with cefoxitin and ampicillin plus sulbactam; Sullivan MC et al.; STUDY OBJECTIVE . To compare the pharmacokinetic and pharmacodynamic activity of three drug regimens: cefotaxime plus metronidazole, cefoxitin, and ampicillin-sulbactam against two organisms frequently isolated in intraabdominal infection, Escherichia coli and Bacteroides fragilis . DESIGN . Open-label, three-way crossover study . SETTING . Hartford Hospital Clinical Research Center . PARTICIPANTS . Nine healthy volunteers . INTERVENTIONS . Subjects received the following regimens: (1) a single 1-g intravenous dose of cefotaxime plus a single 500-mg oral dose of metronidazole; (2) two intravenous doses of cefoxitin, 2 g each dose given every 6 hours; and (3) two intravenous doses of ampicillin-sulbactam, 3 g each dose given every 6 hours . MEASUREMENTS AND MAIN RESULTS . Serum bactericidal titers and drug concentrations were measured over a 12-hour period . The cefotaxime-metronidazole regimen showed superior activity against E . coli compared with ampicillin-sulbactam and cefoxitin . The mean areas under the bactericidal activity curve (AUBC) for the three regimens were 550.2, 68.7, and 48.9, respectively (p = 0.0001) . There was no significant difference in AUBC among the three regimens for B . fragilis . Serum concentrations of cefotaxime remained above the minimum inhibitory concentration (MIC) for E . coli significantly longer than did concentrations of ampicillin-sulbactam and cefoxitin (p = 0.0002 and p = 0.0023, respectively) . Serum concentrations of metronidazole were still at 9 times the MIC for B . fragilis at the end of the 12-hour dosing interval; for ampicillin-sulbactam and cefoxitin concentrations remained above the MIC for one-half and less than one-fourth, respectively, of the dosing interval (p < 0.0001) . The ratio of AUC:MIC was also favorable for metronidazole (212.2) compared with 63.4 for ampicillin-sulbactam and 9.2 for cefoxitin . CONCLUSIONS . The combination of cefotaxime-metronidazole, even at the relatively low doses used in this study, provides coverage against gram-negative and anaerobic pathogens that is at least as effective as that of cefoxitin and ampicillin-sulbactam . In addition, its cost is considerably less expensive than that of the other regimens. Ginekol Pol, 1995 Jun, 66(6), 324 - 9 {Occurrence of Bacteroides fragilis strains in full term and post term pregnancies}; Leszczynski P et al.; Bacteroides fragilis (B.f.) is an aetiological agent of gynecological and obstetrical infections . The aim of this study was to determine occurrence of B.f . in women genital tract . A total number of 118 women in labor between 38 and 43 gestation age and not demonstrating clinical symptoms of infection was examined . B.f . strains were isolated from 9 women (15.9%) . More often B.f . strains were isolated from women delivering post-term and staying previously in Pregnancy Pathology Ward . It was observed that infant mean weight on the 3rd day of life was higher in the vagina . The following features of isolates were compared: presence of the capsule and agglutination of sheet erythrocytes . Differences in haemagglutination and susceptibility to antibiotics were observed. J Infect Dis, 1995 Jun, 171(6), 1475 - 80 The genital flora of women with intraamniotic infection . Vaginal Infection and Prematurity Study Group; Krohn MA et al.; The relationship of genital flora assessed at the end of the second trimester of pregnancy and intraamniotic infection diagnosed by clinical signs and symptoms during labor was evaluated . Women were enrolled at 23-26 weeks of gestation and followed through delivery in the multi-center Vaginal Infections and Prematurity Study (1984-1989) . Among the cohort of 11,989 followed through delivery, 286 (2.4%) developed intraamniotic infection . The recovery of Gardnerella vaginalis (relative risk {RR} = 1.8; 95% confidence interval {CI} = 1.4-2.4), heavy growth of Bacteroides species (RR = 1.5; 95% CI = 1.1-2.1), and isolation of Mycoplasma hominis (RR = 1.7; 95% CI = 1.3-2.1) from the vagina at the end of the second trimester of pregnancy were associated with an increased risk of intraamniotic infection . Bacterial vaginosis was also associated with intraamniotic infection (RR = 1.5; 95% CI = 1.1-2.2) . These findings extend prior studies by showing that prenatal cultures for microorganisms associated with bacterial vaginosis predicted an increased risk of intraamniotic infection. J Bacteriol, 1995 Jun, 177(11), 3158 - 65 Requirements for strand- and site-specific cleavage within the oriT region of Tn4399, a mobilizing transposon from Bacteroides fragilis; Murphy CG et al.; Replicons that contain Tn4399, a conjugal mobilizing transposon isolated from Bacteroides fragilis, can be mobilized in the presence of broad-host-range IncP plasmids RP4 and R751 in Escherichia coli to B . fragilis or E . coli recipients (C . G . Murphy and M . H . Malamy, J . Bacteriol . 175:5814-5823, 1993) . To identify the initial DNA processing events involved in Tn4399-mediated mobilization in E . coli, plasmid DNA from pCGM328 (a pUC7 vector that contains the mobilization region of Tn4399) was isolated from donor cells following the release of plasmid DNA from the relaxation complex . Site- and strand-specific cleavage within the oriT region of Tn4399 was detected by denaturing gel electrophoresis and Southern hybridization analysis of this DNA in the presence or absence of IncP plasmids . Mutations in either mocA or mocB, two genes which are encoded by Tn4399 and are required for mobilization, significantly decrease the amount of specifically nicked DNA detected . These results suggest roles for the MocA and MocB gene products in specific processing of Tn4399-containing plasmid DNA prior to mobilization . By isolation of the nicked strand and primer extension of this template, we mapped the precise 5' end of the single-stranded cleavage reaction . The nucleotide position of nicTn4399 is adjacent to two sets of inverted repeats, a genetic arrangement similar to those of previously characterized oriT regions . Two site-directed mutations which remove nicTn4399 (oriT delta 1 and oriT delta 2) cannot be mobilized to recipients when they are present in trans along with functional MocA and MocB proteins and an IncP mobilizing plasmid; they are cis-dominant loss-of-function mutations. Am Surg, 1995 Jun, 61(6), 521 - 5 Antibiotic penetration of experimental intra-abdominal abscesses; Galandiuk S et al.; Intra-abdominal abscess is seldom adequately treated by systemic antibiotics alone and often requires surgical or computed tomography-guided drainage for resolution . Abscess penetration of six currently used antibiotics was examined in a murine intra-abdominal abscess model . Ampicillin/sulbactam, cefmetazole, clindamycin, and trospectomycin penetrated intra-abdominal abscesses to a greater degree than cefoxitin and ceftriaxone . Abscess pus antibiotic levels were not significantly higher after multiple doses than after a single dose . Pus antibiotic levels below the MIC90 for Bacteroides and E . coli within intra-abdominal abscess were observed for most antibiotics with the doses used in this study . Selection of antibiotics with a greater ability to penetrate abscess may be important in optimally treating patients with abdominal infection. J Antibiot (Tokyo), 1995 Jun, 48(6), 462 - 6 Macquarimicins, microbial metabolites from Micromonospora . I . Discovery, taxonomy, fermentation and biological properties; Jackson M et al.; A novel series of microbial metabolites were discovered in fermentation broths of two soil isolates . Both cultures were identified as strains of Micromonospora chalcea . Production of the metabolites, named macquarimicins, was monitored by an HPLC assay . A seven-day fermentation yielded 27 mg/liter of macquarimicin A . With MICs of 50 to 100 micrograms/ml, macquarimicin A has only very low activity against strains of Bacteroides and other anaerobes . Macquarimicin B has inhibitory activity against the leukemia cell line P-388. Jpn J Clin Oncol, 1995 Jun, 25(3), 86 - 90 Bacteremia caused by the Bacteroides fragilis group in patients with hematologic diseases; Funada H et al.; During a 22-year period, 13 patients with hematologic diseases developed bacteremia caused by the Bacteroides fragilis group, with a frequency which remained almost unchanged . Nine patients (69%) had polymicrobial infections . Acute leukemia was the most common underlying disease . The lower intestinal tract (necrotizing enterocolitis and anorectal abscesses) was the most common source of infection . Prior antibiotic therapy was the most frequent host condition before bacteremia, followed by cancer chemotherapy, neutropenia, thrombocytopenia and hypoproteinemia . Septic shock occurred only in seven patients with polymicrobial infections . Six patients, including five with shock, died within a week of onset, while the other seven survived for at least three weeks . Despite its clinical similarity to aerobic gram-negative infection, bacteremia due to the B . fragilis group may well, therefore, be suspected particularly when neutropenic patients who present with lower intestinal symptomatology develop a persistent fever unresponsive to the initial empiric antibiotic therapy. Oral Microbiol Immunol, 1995 Jun, 10(3), 129 - 37 Patterns of antibody response in subjects with periodontitis; Haffajee AD et al.; Periodontal diseases comprise a heterogeneous group of infections that are difficult to distinguish on a clinical basis alone . The purpose of the present investigation was to group periodontitis subjects according to their elevated serum antibody levels to specific subgingival species . A total of 119 subjects (19-70 years) with evidence of prior periodontal destruction were monitored at 2-month intervals (maximum 8 visits), prior to therapy, using clinical parameters measured at 6 sites per tooth . The probing attachment level was measured twice at each visit, and an increase of > 2.5mm at a site was used to define subjects with progressing disease . Serum samples were obtained from each subject at each visit and the level of antibody determined by enzyme-linked immunosorbent assay to 12 subgingival species . Subgingival plaque samples were taken from the mesial aspect of all teeth in each subject at each visit, and the levels of 14 different subgingival species were determined using a colony-lift method and DNA probes . Subjects were grouped by cluster analysis of their elevated antibody levels using a simple matching coefficient . Ninety-two subjects fell into 9 clusters with 100% similarity; 29 subjects in one cluster group exhibited elevated antibody to none of the test species . Seven subjects in a second cluster group showed elevated antibody to Bacteroides forsythus . Subjects in the other 7 clusters showed elevated antibody to Actinobacillus actinomycetemcomitans serotype a only or in combination with B . forsythus, A . actinomycetemcomitans serotype b, Prevotella intermedia or Porphyromonas gingivalis.(ABSTRACT TRUNCATED AT 250 WORDS) Clin Infect Dis, 1995 Jun, 20 Suppl 2, S356 - 60 Activity of ampicillin/sulbactam, ticarcillin/clavulanate, clarithromycin, and eleven other antimicrobial agents against anaerobic bacteria isolated from infections in children; Citron DM et al.; The activity of 14 antimicrobial agents against 253 clinical isolates of anaerobic bacteria from pediatric infections was assessed by the agar dilution method . Fifty-eight percent of the isolates were from intraabdominal sites . The drugs tested were ampicillin/sulbactam, ticarcillin/clavulanate, ampicillin, sulbactam, piperacillin, cefoxitin, cefotaxime, cefoperazone/sulbactam, clarithromycin, azithromycin, erythromycin, clindamycin, metronidazole, and chloramphenicol . Ticarcillin/clavulanate was active against all isolates . Clarithromycin was the most active macrolide; combination of this agent with its 14-hydroxy metabolite did not result in synergy . Sixty-two percent of Bacteroides fragilis group isolates, 13% of B . fragilis isolates, and 22% of peptostreptococcal isolates were resistant to clindamycin at a concentration of 4 micrograms/mL . The distribution of these strains in clinical specimens and the patterns of antimicrobial susceptibility documented were different from the findings for isolates from adults in the Los Angeles area. Clin Infect Dis, 1995 Jun, 20 Suppl 2, S352 - 5 Antibiotic susceptibility profiles of Bacteroides fragilis and Bacteroides thetaiotaomicron in Japan from 1990 to 1992; Tanaka-Bandoh K et al.; The antimicrobial susceptibility of clinical isolates of Bacteroides fragilis and Bacteroides thetaiotaomicron collected from December 1990 through November 1992 was determined by the agar dilution technique . Metronidazole, imipenem, and cefoperazone/sulbactam remained highly active against both organisms . Rates of resistance to those agents were 0, 2%, and 0.9% in B . fragilis and 0, 0.9%, and 3% in B . thetaiotaomicron, respectively . Cefoxitin and other cephamycins were active against B . fragilis; rates of resistance to these agents did not tend to increase . With the inclusion of these data, the variation of rates of resistance to several agents was summarized for each year from 1987 to 1992 . Rates of resistance to imipenem decreased in 1991 and 1992 among isolates of B . fragilis (2.3% in 1991, 1.8% in 1992) and B . thetaiotaomicron (2.4% in 1991, 0 in 1992) . Rates of resistance to cefoxitin in B . thetaiotaomicron varied from 10% to 38% during these 6 years, though the distributional peak of MIC values did not change . The rate of resistance to ofloxacin in B . fragilis increased from 42% in 1989 to 81% in 1992 . The rate of resistance to ampicillin in B . thetaiotaomicron was 68% in 1992--approximately 30% lower than before . Mostly, however, the rates of resistance to the antimicrobial agents examined did not change significantly. Clin Infect Dis, 1995 Jun, 20 Suppl 2, S346 - 9 Effect of choice of medium on the results of in vitro susceptibility testing of eight antibiotics against the Bacteroides fragilis group; Hecht DW et al.; The susceptibility of isolates of the Bacteroides fragilis group to eight antibiotics was determined by the agar dilution method on three media . At intermediate breakpoints established by the National Committee for Clinical Laboratory Standards (NCCLS), only minimal differences in the susceptibility of each species to any of the antibiotics were documented among the three media . At NCCLS susceptible breakpoints, greater differences among media were found for cefoxitin and cefotetan than for the other drugs tested . Only minimal differences among the three media were evident in comparisons of MIC50, MIC90, and geometric mean MIC values for all antibiotics tested . Thus, use of any one of these three media does not significantly affect agar-dilution MIC results for the B . fragilis group. Clin Infect Dis, 1995 Jun, 20 Suppl 2, S342 - 5 Susceptibility results for the Bacteroides fragilis group: comparison of the broth microdilution and agar dilution methods; Hecht DW et al.; The antimicrobial susceptibilities of members of the Bacteroides fragilis group were compared using the agar dilution and broth microdilution methods . A total of 455 B . fragilis group isolates were tested against 10 antibiotics . Significant disparity in susceptibility results for most antibiotics was observed between the two methods . Broth microdilution susceptibility results were most similar to agar dilution results when a twofold lower breakpoint was used . In addition, broth microdilution failed to detect resistance to some antibiotics when the recommended agar dilution breakpoint was used . MIC50, MIC90, and geometric mean MIC values for broth microdilution were consistently twofold to fourfold less than those for agar dilution . Susceptibilities of Bacteroides that are determined with use of broth microdilution may more accurately correspond to those determined with use of agar dilution if lower breakpoints are used for interpretation. Clin Infect Dis, 1995 Jun, 20 Suppl 2, S283 - 8 Foot infections in diabetic patients: the role of anaerobes; Gerding DN; The prevalence of anaerobic bacteria in cultures of specimens from foot infections in diabetic patients is dependent upon the method of obtaining the specimen, the care with which it is transported anaerobically, and the sophistication of the laboratory methods . The rate at which anaerobes are isolated with use of the best methods ranges from 74% to 95% of patients, but it is only 41% to 53% in clinical studies of patients with limb-threatening infection . The mean number of bacterial isolates from an infected foot ranges from 4.1 to 5.8, of which 1.2 to 2.6 isolates are anaerobic . Most anaerobic isolates are gram-positive, and Peptostreptococcus species are most common . Bacteroides species are the most common anaerobic gram-negative isolates . Treatment covering anaerobic bacteria is included in most empirical regimens, but the use of agents that are modestly active against anaerobic organisms (i.e., fluoroquinolones or trimethoprim-sulfamethoxazole) has also been successful . Surgical debridement and drainage are essential adjuncts to antimicrobial therapy and may assist in the control of anaerobic infection . Questions regarding management of such infections parallel those regarding management of polymicrobial intraabdominal infections, but to date, studies of the former have been much less sophisticated than those of the latter. Clin Infect Dis, 1995 Jun, 20 Suppl 2, S251 - 6 Clinical correlates of anaerobic bacteriology in peritonitis; Wilson SE et al.; Recent surgical reports indicate that for patients with secondary bacterial peritonitis, surgeons do not routinely use the identification of the bacterial pathogens and determination of their antimicrobial susceptibilities in choosing antimicrobial therapy . Further, some surgeons advocate abandoning the routine practice of obtaining culture specimens from patients with complicated appendicitis, because the data from the clinical laboratory have not been found to have an impact on postoperative care . We review the rationale for continued surveillance of and implementation of bacteriological data in treatment of secondary peritonitis . We also describe in detail the anaerobic flora of secondary peritonitis, the unique susceptibility patterns of these organisms, and the specific virulence factors of anaerobes, particularly Bacteroides fragilis . The fact that clinical investigations sometimes result in treatment failure when gram-negative anaerobes are resistant to the antimicrobials used or when complete antimicrobial susceptibility data are not available emphasizes the need for accurate and early knowledge of the bacteriologic characteristic of the flora of the operative site . We emphasize the relationship of in vitro susceptibility of intraoperative isolates with clinical outcome . We propose a cooperative trial that would demonstrate that successful antimicrobial therapy should be based on the susceptibility of the flora of the operative site, which correlates with clinical outcome. Clin Infect Dis, 1995 Jun, 20 Suppl 2, S187 - 91 Update on the taxonomy and the clinical and laboratory characteristics of pigmented anaerobic gram-negative rods; Jousimies-Somer HR; Pigmented anaerobic gram-negative rods are currently categorized as 17 species distributed in three genera: Prevotella, Porphyromonas, and Bacteroides . These organisms are often encountered in clinical specimens but are also found as part of the indigenous flora on various mucosal surfaces . Several studies are presently assessing the association of individual species with health and disease . For example, Porphyromonas gingivalis and Porphyromonas endodontalis are key putative pathogens in adult periodontitis and root canal infections, respectively . Porphyromonas asaccharolytica is prevalent in extraoral infections . The Porphyromonas species of animal origin have been isolated from infected bite wounds in humans . Isolates closely resembling Bacteroides levii have been recovered from various types of human infections . According to preliminary reports, Prevotella intermedia tends to be associated more often with periodontal disease than with a healthy oral cavity . In the laboratory, enzyme profiling facilitates the identification of these pigmented rods . Beta-Lactamase production is more common among prevotella species (30%-50%) than among Porphyromonas species (< 10%). Clin Infect Dis, 1995 Jun, 20 Suppl 2, S149 - 53 Biological activity of Bacteroides lipopolysaccharide--reappraisal; Poxton IR et al.; Lipopolysaccharide (LPS) was extracted from six species of Bacteroides by the phenol/water, petroleum/chloroform/ phenol, and Triton/magnesium methods . Yields and chemical analysis demonstrated that the products were different . Biological activity (endotoxicity) was assessed by the limulus amebocyte lysate (LAL) assay, induction of tumor n