Roseovarius pelagicus sp. nov., a facultatively anaerobic bacterium with potential for degrading polypropylene, isolated from Arctic seawater.

A Gram-stain-negative, facultatively anaerobic, non-motile, rod-shaped bacterial strain, designated HL-MP18T, was isolated from Arctic seawater after a prolonged incubation employing polypropylene as the sole carbon source. Phylogenetic analyses of the 16S rRNA gene sequence revealed that strain HL-MP18T was affiliated to the genus Roseovarius with close relatives Roseovarius carneus LXJ103T (96.8 %) and Roseovarius litorisediminis KCTC 32327T (96.5 %). The complete genome sequence of strain HL-MP18T comprised a circular chromosome of 3.86 Mbp and two circular plasmids of 0.17 and 0.24 Mbp. Genomic comparisons based on average nucleotide identity and digital DNA-DNA hybridization showed that strain HL-MP18T was consistently discriminated from its closely related taxa in the genus Roseovarius. Strain HL-MP18T showed optimal growth at 25 °C, pH 7.0 and 2.5 % (w/v) sea salts. The major cellular fatty acids were C18 : 1 ω6c and/or C18 : 1 ω7c (49.6 %), C19 : 0 cyclo ω8c (13.5 %), and C16 : 0 (12.8 %). The major respiratory quinone was ubiquinone-10. The polar lipids consisted of phosphatidylcholine, phosphatidylglycerol, an unidentified aminolipid and three unidentified lipids. The genomic DNA G+C content of the strain was 59.2 mol%. The phylogenetic, genomic, phenotypic and chemotaxonomic results indicate that strain HL-MP18T is distinguishable from the recognized species of the genus Roseovarius. Therefore, we propose that strain HL-MP18T represents a novel species belonging to the genus Roseovarius, for which the name Roseovarius pelagicus sp. nov. is proposed. The type strain is HL-MP18T (=KCCM 90405T=JCM 35639T).

Antimicrobial susceptibility pattern of oral gram negative anaerobes from Indian subjects.

OBJECTIVES: There is paucity of information on the antimicrobial susceptibility pattern of oral anaerobic bacteria. In this study, an attempt has been made to evaluate the antimicrobial susceptibility/resistance trend of oral Gram negative bacteria from Indian subjects. METHODS: Minimum inhibitory concentrations (MIC) of 304 isolates against twelve different antibiotics were determined using gradient diffusion MIC strips. The organisms were isolated and identified based on phenotypic characteristics and included Porphyromonas gingivalis, Prevotella species, Tannerella forsythia, Fusobacterium nucleatum, Aggregatibacter actinomycetemcoitans, Eickenella corrodens and Capnocytophaga species. For each antimicrobial agent, MIC50 and MIC90 were calculated and expressed. RESULTS: Resistance to azithromycin, clindamycin, and amoxicillin was observed in most of the anaerobic bacterial species studied. High degree of susceptibility was observed to amoxillin-clavulanic acid, doxycycline and moxifloxacin. A single strain of P. melaninogenica was resistant to moxifloxacin. The susceptibility pattern varied with cephalosporins among species. Ceftriaxone showed highest and cefazolin least efficacy among cephalosporins. All anaerobic bacteria tested were susceptible to metronidazole. Strains of T. forsythia were more resistant to several antibiotics than other anaerobic bacteria. All three species of capnophilic bacteria displayed high degree of resistance to metronidazole and significant resistance to amoxicillin, azithromycin, clindamycin, cefazolin and cefuroxime. CONCLUSIONS: Amoxicillin-clavulanic acid, doxycycline, moxifloxacin and metronidazole appeared to be the most effective drugs against gram negative anaerobic bacteria. However, the MIC50 and MIC90 values against metronidazole were on the higher side of the normal indicating a potential for developing resistance.

Development and application of aerobic, chemically defined media for Dysgonomonas.

Members of Dysgonomonas are Gram-stain-negative, non-motile, facultatively anaerobic coccobacilli originally described in relation to their isolation from stool and wounds of human patients (CDC group DF-3). More recently, Dysgonomonas have been found to be widely distributed in terrestrial environments and are particularly enriched in insect systems. Their prevalence in xylophagous insects such as termites and wood-feeding cockroaches, as well as in soil-fed microbial fuel cells, elicit interest in lignocellulose degradation and biofuel production, respectively. Their occurrence in mosquito and fruit fly have implications relating to symbiosis, host immunology and developmental biology. Additionally, their presence in termite, mosquito and nematode present novel opportunities for pest and vector control. Currently, the absolute growth requirements of Dysgonomonas are unknown, and they are commonly cultured under anaerobic conditions on complex media containing blood, peptones, tryptones, and yeast, plant or meat extracts. Restrictive and undefined culturing conditions preclude physiological and genetic studies, and thus further understanding of their metabolic potential. Here we describe the requirements for growth of termite-derived Dysgonomonas isolates and create parallel complex, defined and minimal media that permit vigorous and reliable aerobic growth. Furthermore, we show that these media can be used to easily enrich for Dysgonomonas isolates from densely-colonized and microbially-diverse environmental samples.

Surveillance of antimicrobial resistance in recent clinical isolates of Gram-negative anaerobic bacteria in a Greek University Hospital.

The aim of our study was to determine the antimicrobial susceptibility profiles of 267 Gram-negative clinically significant anaerobes, isolated between October 2016 and October 2019, in a Greek university hospital. The species identification was performed by conventional methods and using the Vitek 2 automated system. Antimicrobial susceptibility testing to determine the MICs was performed by the E-test method. The antimicrobial agents tested were penicillin, ampicillin, amoxicillin-clavulanic acid, piperacillin-tazobactam, cefoxitin, imipenem, meropenem, clindamycin, metronidazole, moxifloxacin, chloramphenicol and tigecycline. The results were interpreted using the CLSI and FDA breakpoints. The majority of the isolates belonged to Bacteroides fragilis group (58.8%), followed by Prevotella spp. (23.2%), Fusobacterium spp. (11.2%) and Veillonella spp. (6.4%). The most prevalent types of infection were skin and soft tissue infections (34.8%), and inta-abdomonal infections (29.6%). Among all isolates tested, the lowest rates of resistance (<5%) were detected to carbapenems, metronidazole, chloramphenicol and tigecycline. Resistance to piperacillin-tazobactam was observed in 5.4%, 24.6%, 3.3% and 17.6%, of B. fragilis, B. fragilis group, Fusobacterium spp. and Veillonella spp. isolates, respectively. Although a high prevalence of resistance to clindamycin, cefoxitin, and moxifloxacin, was detected particularly among members of the B. fragilis group, cefoxitin resistance was low for Prevotella spp. (3.2%), Fusobacterium spp. (3.3%) and Veillonella spp. (0%). Our findings underscore the need for periodic monitoring of antimicrobial resistance in order to guide empirical therapy.

Abditibacterium utsteinense sp. nov., the first cultivated member of candidate phylum FBP, isolated from ice-free Antarctic soil samples.

Most bacterial lineages are known only by molecular sequence data from environmental surveys and represent the uncultivated majority. One of these lineages, candidate phylum FBP, is widespread in extreme environments on Earth, ranging from polar and desert ecosystems to wastewater and contaminated mine sites. Here we report on the characterization of strain LMG 29911T, the first cultivated representative of the FBP lineage. The strain was isolated from a terrestrial surface sample from Utsteinen, Sør Rondane Mountains, East Antarctica and is a Gram-negative, aerobic, oligotrophic chemoheterotrophic bacterium. It displays growth in a very narrow pH range, use of only a limited number of carbon sources, but also a metabolism optimized for survival in low-nutrient habitats. Remarkably, phenotypic and genome analysis indicated an extreme resistance against antibiotics and toxic compounds. We propose the names Abditibacterium utsteinense for this bacterium and Abditibacteriota for the former candidate phylum FBP. Furthermore, inter- and intra-phylum relationships indicate Armatimonadetes, a neighboring lineage to the Abditibacteriota, to be a superphylum.

Aerobic bacteriological profile and antimicrobial susceptibility pattern of pus isolates from tertiary care hospital in India.

INTRODUCTION: Pyogenic infections are an important cause of sepsis. These infections are difficult to treat because of the pathogens with increasing antibiotic resistance. It is important to know the pathogens causing the infections and its antibiotic susceptibility for proper management of the patients. METHODOLOGY: A retrospective analysis of 1428 culture positive pus and tissue samples received in the department of microbiology from various departments in the hospital between January 2012 to 2017 was performed. Data regarding the pathogen isolated and its antimicrobial susceptibility were collected and analyzed. The specimens were primarily processed, as per standard methods. Identification and susceptibility testing was done using the Vitek-2C system. RESULTS: Among the samples males outnumbered females (M: F-2.5:1) and the median age was 47 years. The total number of patients were 1428 with total number of isolates being 1525 as in our study monomicrobial infections were seen in 93.2% (1331/1428) patients whereas combined infections with growth of two pathogens in 6.8% (97/1428). Gram-negative bacilli were isolated in 68.3% (1042/1525). Among the Gram-negative bacilli Escherichia coli was the major pathogen isolated (38.6%, 403/1042). Gram positive organisms were isolated in 31.6% (483/1525) of cases and Staphylococcus aureus was the predominant organism isolated (91.7%, 443/483). Rare pathogens like Burkholderia pseudomallei in 3 patients and Nocardia in one patient were also isolated. CONCLUSION: This study emphasizes to understand the common organisms isolated from wound infections and it helps in empirical treatment of patients based on antibiotic susceptibility patterns.

Bacterial translocation and mortality on rat model of intestinal ischemia and obstruction

Abstract Purpose: To develop an experimental model of intestinal ischemia and obstruction followed by surgical resection of the damaged segment and reestablishment of intestinal transit, looking at bacterial translocation and survival. Methods: After anesthesia, Wistar rats was subject to laparotomy, intestinal ischemia and obstruction through an ileal ligature 1.5cm of ileum cecal valve; and the mesenteric vessels that irrigate upstream of the obstruction site to approximately 7 to 10 cm were ligated. Abdominal wall was closed. Three, six or twenty-four hours after, rats were subject to enterectomy followed by an end to end anastomosis. After 24h, mesenteric lymph nodes, liver, spleen and lung tissues were surgically removed. It was studied survival rate and bacterial translocation. GraphPadPrism statistical program was used. Results: Animals with intestinal ischemia and obstruction for 3 hours survived 24 hours after enterectomy; 6hx24h: survival was 70% at 24 hours; 24hx24h: survival was 70% and 40%, before and after enterectomy, respectively. Culture of tissues showed positivity on the 6hx24h and negativity on the 3hx24h. Conclusion: The model that best approached the clinic was the one of 6x24h of ischemia and intestinal obstruction, in which it was observed bacterial translocation and low mortality rate.

The Deep-Sea Polyextremophile Halobacteroides lacunaris TB21 Rough-Type LPS: Structure and Inhibitory Activity towards Toxic LPS.

The structural characterization of the lipopolysaccharide (LPS) from extremophiles has important implications in several biomedical and therapeutic applications. The polyextremophile Gram-negative bacterium Halobacteroideslacunaris TB21, isolated from one of the most extreme habitats on our planet, the deep-sea hypersaline anoxic basin Thetis, represents a fascinating microorganism to investigate in terms of its LPS component. Here we report the elucidation of the full structure of the R-type LPS isolated from H. lacunaris TB21 that was attained through a multi-technique approach comprising chemical analyses, NMR spectroscopy, and Matrix-Assisted Laser Desorption Ionization (MALDI) mass spectrometry. Furthermore, cellular immunology studies were executed on the pure R-LPS revealing a very interesting effect on human innate immunity as an inhibitor of the toxic Escherichia coli LPS.

Use of MALDI-TOF mass spectrometry after liquid enrichment (BD Bactec™) for rapid diagnosis of bone and joint infections.

Advantages of MALDI-TOF MS (MS) were evaluated for diagnosis of bone and joint infections after enrichment of synovial fluid (SF) or crushed osteoarticular samples (CSs). MS was performed after enrichment of SF or crushed osteoarticular samples CS (n = 108) in both aerobic and anaerobic vials. Extraction was performed on 113 vials (SF: n = 47; CS: n = 66), using the Sepsityper® kit prior identification by MS. The performances of MS, score and reproducibility results on bacterial colonies from blood agar and on pellets after enrichment in vials, were compared. MS analysis of the vial resulted in correct identification of bacteria at a species and genus level (80.5% and 92% of cases, respectively). The reproducibility was superior for aerobic Gram-positive bacteria (Staphylococci and Gram-positive bacilli: 100% colonies), as compared to aerobic Gram-negative bacilli (89.7%), anaerobes (83.3%) and Streptococcus/Enterococcus (58.8%). MS performance was significantly better for staphylococci than for streptococci on all identification parameters. For polymicrobial cultures, identification (score>1.5) of two species by MS was acceptable in 92.8% of cases. Use of MS on enrichment pellets of bone samples is an accurate, rapid and robust method for bacterial identification of clinical isolates from osteoarticular infections, except for streptococci, whose identification to species level remains difficult.

Bacterial diversity of symptomatic primary endodontic infection by clonal analysis.

The aim of this study was to explore the bacterial diversity of 10 root canals with acute apical abscess using clonal analysis. Samples were collected from 10 patients and submitted to bacterial DNA isolation, 16S rRNA gene amplification, cloning, and sequencing. A bacterial genomic library was constructed and bacterial diversity was estimated. The mean number of taxa per canal was 15, ranging from 11 to 21. A total of 689 clones were analyzed and 76 phylotypes identified, of which 47 (61.84%) were different species and 29 (38.15%) were taxa reported as yet-uncultivable or as yet-uncharacterized species. Prevotella spp., Fusobacterium nucleatum, Filifactor alocis, and Peptostreptococcus stomatis were the most frequently detected species, followed by Dialister invisus, Phocaeicola abscessus, the uncharacterized Lachnospiraceae oral clone, Porphyromonas spp., and Parvimonas micra. Eight phyla were detected and the most frequently identified taxa belonged to the phylum Firmicutes (43.5%), followed by Bacteroidetes (22.5%) and Proteobacteria (13.2%). No species was detected in all studied samples and some species were identified in only one case. It was concluded that acute primary endodontic infection is characterized by wide bacterial diversity and a high intersubject variability was observed. Anaerobic Gram-negative bacteria belonging to the phylum Firmicutes, followed by Bacteroidetes, were the most frequently detected microorganisms.

Distributions of Synergistetes in clinically-healthy and diseased periodontal and peri-implant niches.

Bacterial taxa belonging to the phylum Synergistetes are commonly detected within diseased periodontal niches, but are rarely found within healthy oral sites. However, as they typically constitute a minor fraction of the oral microbiota, their precise distributions and disease-associations remain to be fully established. Here, we surveyed the Synergistetes taxa present within individual periodontal/subgingival and peri-implant/submucosal sites, within Chinese subjects (n = 18) affected by both peri-implantitis and periodontitis. Four individual, clinically-distinct sites were analyzed in each patient: healthy sulcus; periodontitis lesion; healthy peri-implant space; peri-implantitis lesion. We employed a clone library-based approach, using PCR-primers that specifically amplified ca. 650bp regions of the 16S rRNA gene from oral cluster A and B Synergistetes taxa. Twenty-one of the 72 sites (from 12/18 subjects) yielded Synergistetes 16S rRNA PCR products. Sequencing of cloned amplicon libraries yielded 1338 quality-filtered 16S rRNA sequences, which were assigned to 26 Synergistetes operational taxonomic units (OTUs; oral taxon SH01-SH26) using a 98.5% identity cut-off. We identified 25 Synergistetes oral cluster A OTUs (genus Fretibacterium; corresponding to Human Oral Taxon (HOT) numbers 358, 359, 360, 361, 362, 363, 452, and 453), and one oral cluster B OTU (Pyramidobacter piscolens oral taxon SH04, HOT-357). Three OTUs predominated: Fretibacterium oral taxon SH01 (HOT-360), Fretibacterium oral taxon SH02 (HOT-452), and Fretibacterium fastidiosum oral taxon SH03 (HOT-363). The Synergistetes community compositions within the respective periodontal and peri-implant sites were variable and complex, and no statistically-significant correlations could be established. However, the detection frequency of F. fastidiosum SH03 and Fretibacterium oral taxon SH01 were both positively associated with plaque index at healthy subgingival sites. Taken together, our results show that diverse Synergistetes populations inhabit both diseased and healthy periodontal and peri-implant niches, with considerable site-to-site variations in composition occurring within the same oral cavity.

Clinical association of Spirochaetes and Synergistetes with peri-implantitis.

OBJECTIVES: The microbial composition of peri-implantitis-associated biofilms may resemble that of periodontitis, with some distinctive differences, as identified by various conventional or molecular detection methods. Yet, the complete microbiome of peri-implantitis awaits further characterization. The present clinical study was undertaken with the aim to investigate the association of Spirochaetes, and the more recently identified phylum Synergistetes, with peri-implantitis. MATERIALS AND METHODS: Submucosal biofilms were obtained from single sites of patients with peri-implantitis (n = 43) or individuals with peri-implant health (n = 41). The samples were analysed by fluorescence in situ hybridization (FISH) and epifluorescence microscopy, using 16S rRNA-based oligonucleotide probes for Synergistetes cluster A, subclusters A1 and A2, and Treponema groups I-III and IV. RESULTS: Treponema group IV was barely detectable, whereas Treponema groups I-III were detected at low prevalence in health, but their prevalence and numbers were significantly increased in peri-implantitis by 48% and 2.4-log, respectively. Synergistetes cluster A was detected in half of the healthy sites, and its prevalence and numbers were significantly increased in peri-implantitis by 30% and 2.5-log, respectively. No quantitative differences were found between Synergistetes subclusters A1 and A2 numbers, as both increased by 2.8-log. Synergistetes cluster A displayed strong correlations with several clinical peri-implant parameters, but Treponema groups I-III only with probing pocket depth. CONCLUSION: The present clinical cross-sectional study demonstrates that Spriochaetes of the Treponema groups I-III, but not group IV, and Synergistetes of the cluster A are highly associated with peri-implantitis. Synergistetes cluster A appears to display a stronger association with peri-implantitis than Spirochaetes.

The Microbiologic Profile Associated with Peri-Implantitis in Humans: A Systematic Review.

PURPOSE: To qualitatively investigate the microbiologic profile in peri-implantitis by systematically reviewing the published literature on peri-implant infection. MATERIALS AND METHODS: Searches of the US National Institutes of Health free digital archives of the biomedical and life sciences journal literature (PubMed) and The Cochrane Library of the Cochrane Collaboration (CENTRAL), as well as a hand search of other literature, were conducted to identify articles potentially relevant for the review. Randomized clinical trials, prospective cohort studies, longitudinal studies, case-control studies, and cross-sectional studies in humans reporting microbiologic findings in patients with diagnosed peri-implantitis were considered eligible for this review. Screening, data extraction, and quality assessment were conducted independently and in duplicate. RESULTS: Twenty-one articles were eligible for inclusion in this review. Early studies focused on the identification of target periopathogens, whereas more recent studies used advanced molecular techniques for comprehensive overview of the peri-implantitis-associated microbiome. In summary, the microbiologic profile in peri-implantitis (1) is complex and variable, (2) consists of gram-negative anaerobic periopathogens and opportunistic microorganisms in almost the same ratio, (3) is frequently associated with the Epstein-Barr virus and nonsaccharolytic anaerobic gram-positive rods, (4) is not so strictly associated with Staphylococcus aureus, and (5) is different from that of periodontitis. A meta-analysis could not be performed because of the heterogeneity of the reviewed studies. CONCLUSION: Although a comparison of the published results was limited because of the inhomogeneity of the studies, it is clear that the microbiologic profile of peri-implantitis consists of aggressive and resistant microorganisms and is distinct from that of periodontitis. It seems that the quantitative characteristics of the microflora cohabitants represent the key determinant of disease, rather than the qualitative composition, which is very similar in healthy and peri-implantitis states.

Bacterial diversity of symptomatic primary endodontic infection by clonal analysis

Abstract The aim of this study was to explore the bacterial diversity of 10 root canals with acute apical abscess using clonal analysis. Samples were collected from 10 patients and submitted to bacterial DNA isolation, 16S rRNA gene amplification, cloning, and sequencing. A bacterial genomic library was constructed and bacterial diversity was estimated. The mean number of taxa per canal was 15, ranging from 11 to 21. A total of 689 clones were analyzed and 76 phylotypes identified, of which 47 (61.84%) were different species and 29 (38.15%) were taxa reported as yet-uncultivable or as yet-uncharacterized species. Prevotella spp., Fusobacterium nucleatum, Filifactor alocis, and Peptostreptococcus stomatis were the most frequently detected species, followed by Dialister invisus, Phocaeicola abscessus, the uncharacterized Lachnospiraceae oral clone, Porphyromonas spp., and Parvimonas micra. Eight phyla were detected and the most frequently identified taxa belonged to the phylum Firmicutes (43.5%), followed by Bacteroidetes (22.5%) and Proteobacteria (13.2%). No species was detected in all studied samples and some species were identified in only one case. It was concluded that acute primary endodontic infection is characterized by wide bacterial diversity and a high intersubject variability was observed. Anaerobic Gram-negative bacteria belonging to the phylum Firmicutes, followed by Bacteroidetes, were the most frequently detected microorganisms.

Description of a novel pectin-degrading bacterial species Prevotella pectinovora sp. nov., based on its phenotypic and genomic traits.

Five strictly anaerobic Gram-negative bacterial strains, P4-65, P4-76(T), P5-60, P5-119, and P5-125, presumably belonging to the genus Prevotella were isolated from pig fecal samples. Strains were tested for various phenotypic traits and nearcomplete genome sequences were obtained and analyzed. Phylogenetic analysis based on 16S rRNA gene sequences and multilocus sequence analysis based on five conserved genes confirmed that the strains belong to the genus Prevotella, revealing that they represent a novel and discrete lineage distinct from other known species of this genus. The size of the genome of the isolated strains is 3-3.3 Mbp, and the DNA G+C content is 47.5-48.1 mol%. The isolates are strictly anaerobic, rod-shaped with rounded ends, non-motile and non-spore-forming. The main fermentation products are succinate and acetate, with minor concentrations of isovalerate, propionate and isobutyrate. Hydrogen is also produced. Major cellular fatty acids consist of anteiso-C(15:0) and iso-C(15:0), and a number of additional acids are present in lower concentrations. A substantial portion of genes involved in carbohydrate utilization is devoted to pectin degradation and utilization, while those supporting growth on xylan in ruminal Prevotella could not have been revealed. On the basis of the presented results, a novel species, Prevotella pectinovora sp. nov. is proposed. The type strain is P4-76(T) (=DSM 29996(T) =ZIM B1020(T)).

Characterization of Sporohalobacter salinus sp. nov., an anaerobic, halophilic, fermentative bacterium isolated from a hypersaline lake.

Halophilic, obligately anaerobic, Gram-stain-negative bacterial strains were isolated from a sediment sample taken from under the salt crust of El-Jerid hypersaline lake in southern Tunisia by using tryptone or glucose as the substrate. One strain, CEJFT1B(T), was characterized phenotypically and phylogenetically. Cells were non-motile, non-spore-forming, short rods. Strain CEJFT1B(T) was able to grow in the presence of 5-30 % (w/v) NaCl (optimum 20 %) and at 30-60 °C (optimum 45 °C). It grew at pH 5.5-7.8 and the optimum pH for growth was 6.8. The isolate required yeast extract for growth. Substrates utilized by strain CEJFT1B(T) as the sole carbon source included glucose, fructose, sucrose, pyruvate, Casamino acids and starch. Individual amino acids such as glutamate, lysine, methionine, serine, tyrosine, and amino acid mixtures formed by the Stickland reaction such as alanine-glycine, valine-proline, leucine-proline, isoleucine-proline were also utilized. Products of glucose fermentation were acetate (major product), butyrate, H2 and CO2. The genomic DNA G+C content of strain CEJFT1B(T) was 32.3 mol%. Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain CEJFT1B(T) should be assigned to the genus Sporohalobacter. The sequence similarity between strain CEJFT1B(T) and Sporohalobacter lortetii was 98.5 %, but DNA-DNA hybridization between the two strains revealed a relatedness value of 56.4 %, indicating that they are not related at the species level. The combination of phylogenetic analysis, DNA-DNA hybridization data, and differences in substrate utilization support the view that strain CEJFT1B(T) represents a novel species of the genus Sporohalobacter, for which the name Sporohalobacter salinus sp. nov. is proposed. The type strain is CEJFT1B(T) ( = DSM 26781(T) = JCM 19279(T)).

Centipeda periodontii in human periodontitis.

This study assessed the subgingival occurrence of the flagellated, Gram-negative, anaerobic rod Centipeda periodontii in chronic periodontitis and periodontal health/gingivitis with species-specific nucleic acid probes, and evaluated the in vitro resistance of subgingival isolates to therapeutic levels of amoxicillin, metronidazole, and doxycycline. Subgingival plaque biofilm specimens from 307 adults with chronic periodontitis, and 48 adults with periodontal health/localized gingivitis, were evaluated with digoxigenin-labeled, whole-chromosomal, DNA probes to C. periodontii ATCC 35019 possessing a 10(4) cell detection threshold. Fifty-two C. periodontii subgingival culture isolates were assessed on antibiotic-supplemented enriched Brucella blood agar for in vitro resistance to either amoxicillin at 2 µg/ml, metronidazole at 4 µg/ml, or doxycycline at 2 µg/ml. A significantly greater subgingival occurrence of C. periodontii was found in chronic periodontitis subjects as compared to individuals with periodontal health/gingivitis (13.4 vs. 0 %, P < 0.003), although high subgingival counts of the organism (≥ 10(6) cells) were rarely detected (1.3 % of chronic periodontitis subjects). In vitro resistance was not found to amoxicillin or metronidazole, and to doxycycline in only 2 (3.9 %) of the 52 C. periodontii clinical isolates studied. These findings indicate that C. periodontii is not a major constituent of the subgingival microbiome in chronic periodontitis or periodontal health/gingivitis. The potential contribution of C. periodontii to periodontal breakdown in the few chronic periodontitis subjects who yielded high subgingival levels of the organism remains to be delineated. C. periodontii clinical isolates were susceptible in vitro to therapeutic concentrations of three antibiotics frequently used in treatment of human periodontitis.

The subgingival microbiome of clinically healthy current and never smokers.

Dysbiotic oral bacterial communities have a critical role in the etiology and progression of periodontal diseases. The goal of this study was to investigate the extent to which smoking increases risk for disease by influencing the composition of the subgingival microbiome in states of clinical health. Subgingival plaque samples were collected from 200 systemically and periodontally healthy smokers and nonsmokers. 16S pyrotag sequencing was preformed generating 1,623,713 classifiable sequences, which were compared with a curated version of the Greengenes database using the quantitative insights into microbial ecology pipeline. The subgingival microbial profiles of smokers and never-smokers were different at all taxonomic levels, and principal coordinate analysis revealed distinct clustering of the microbial communities based on smoking status. Smokers demonstrated a highly diverse, pathogen-rich, commensal-poor, anaerobic microbiome that is more closely aligned with a disease-associated community in clinically healthy individuals, suggesting that it creates an at-risk-for-harm environment that is primed for a future ecological catastrophe.