Nasopharyngeal Microbiome Signature in COVID-19 Positive Patients: Can We Definitively Get a Role to Fusobacterium periodonticum?

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) caused the pandemic Coronavirus Disease 2019 (COVID-19). This virus is highly transmissible among individuals through both droplets and aerosol leading to determine severe pneumonia. Among the various factors that can influence both the onset of disease and the severity of its complications, the microbiome composition has also been investigated. Recent evidence showed the possible relationship between gut, lung, nasopharyngeal, or oral microbiome and COVID-19, but very little is known about it. Therefore, we aimed to verify the relationships between nasopharyngeal microbiome and the development of either COVID-19 or the severity of symptoms. To this purpose, we analyzed, by next generation sequencing, the hypervariable V1-V2-V3 regions of the bacterial 16S rRNA in nasopharyngeal swabs from SARS-CoV-2 infected patients (n=18) and control (CO) individuals (n=12) using Microbiota solution A (Arrow Diagnostics). We found a significant lower abundance of Proteobacteria and Fusobacteria in COVID-19 patients in respect to CO (p=0.003 and p<0.0001, respectively) from the phylum up to the genus (p<0.001). The Fusobacterium periodonticum (FP) resulted as the most significantly reduced species in COVID-19 patients respect to CO. FP is reported as being able to perform the surface sialylation. Noteworthy, some sialic acids residues on the cell surface could work as additional S protein of SARS-CoV-2 receptors. Consequently, SARS-CoV-2 could use sialic acids as receptors to bind to the epithelium of the respiratory tract, promoting its clustering and the disease development. We can therefore speculate that the significant reduction of FP in COVID-19 patients could be directly or indirectly linked to the modulation of sialic acid metabolism. Finally, viral or environmental factors capable of interfering with sialic metabolism could determine a fall in the individual protection from SARS-CoV-2. Further studies are necessary to clarify the precise role of FP in COVID-19.

An inexpensive anaerobic chamber for the genetic manipulation of strictly anaerobic bacteria.

Strictly anaerobic bacteria are important to both human health and industrial usage. These bacteria are sensitive to oxygen, therefore, it is preferable to manipulate these microbes in an anaerobic chamber. However, commercial anaerobic chambers (CACs) are expensive, making them less accessible to scientists with a limited budget, especially to those in developing countries. The high price of commercial chambers has hindered, at least partially, the progress of research on anaerobes in developing countries. In the research presented here, we developed an inexpensive and reliable anaerobic chamber and successfully achieved routine maintenance of eleven strictly anaerobic bacterial strains. Furthermore, genetic manipulation examples have been set for both Clostridioidesdifficile 630 and Clostridiumbeijerinckii NCIMB 8052 strains to validate that the chamber could applied to advanced genetic engineering of strictly anaerobes. C. difficile and C. beijerinckii were both genetically manipulated in this chamber, showing it's utility for the genetic engineering of anaerobes. Most importantly, the anaerobic chamber was 76% - 88% less expensive than a CACs and has similar functionality with regards to the cultivation and manipulation of strictly anaerobic bacteria. The anaerobic chamber described in this study will promote the research of anaerobes in developing counties and scientists who have limited research budgets.

Specific increase of Fusobacterium in the faecal microbiota of neonatal calves infected with Cryptosporidium parvum.

The faecal microbiota plays a critical role in host health, with alterations in the human faecal microbial composition associated with various conditions, particularly diarrhoeal diseases. However, little is known about microbial changes during cryptosporidiosis, one of the most important diarrhoeal diseases caused by protozoa in cattle. In this study, alterations in the faecal microbiota of neonatal calves as a result of Cryptosporidium parvum infection were investigated on a C. parvum-positive farm. Comparisons were made among groups of C. parvum-infected, rotavirus-infected, and the pathogen-negative calves. A specific increase in the abundance of Fusobacterium was observed in the faecal microbiota of C. parvum-infected animals. Diarrhoea severity increased in accordance with the abundance of C. parvum and Fusobacterium. Moreover, the specific increase of Fusobacterium appeared to be a universal feature of C. parvum infection, since neonatal calves from geographically separated areas showed the same result. These observations indicated that the growth of Fusobacterium may be an important aggravating factor of cryptosporidiosis.

Effects and efficacy of different sterilization and disinfection methods on electrospun drug delivery systems.

Microbiological quality of a pharmaceutical product is an essential requirement ensuring patient safety, thus effective sterilization/disinfection methods need to be found. The aim of this study was to evaluate the efficacy of different sterilization/disinfection methods on drug-loaded electrospun matrices and the impact of these treatments on the functionality related characteristics of these matrices. The sterilization efficacy of gamma-irradiation, ultraviolet-irradiation, in situ generated chlorine gas and low-pressure argon plasma treatment were evaluated on two different chloramphenicol-loaded electrospun matrices using pristine polycaprolactone (PCL) as a carrier polymer or PCL in combination with polyethylene oxide. Drug stability, solid state properties, morphology, mechanical properties, swelling, biodegradation and drug release kinetics were studied before and after the treatments. It was shown that all tested methods help to reduce bioburden and only plasma treated matrices were not sterile. At the same time drug degradation after the treatment can be considerable and depends not only on the susceptibility of the drug to degradation, but also on matrix properties (e.g. the nature of carrier polymers). Even though no morphological changes were observed, gamma sterilization increased the hardness and elasticity of PCL matrices as a result of increased crystallinity of the polymer. Plasma treatment was able to significantly enhance water absorption to otherwise hydrophobic PCL/CAM matrix and had tremendous impact on its drug release kinetics as the drug was instantly released from otherwise prolonged release formulation.

Effects of Intranasal Pseudorabies Virus AH02LA Infection on Microbial Community and Immune Status in the Ileum and Colon of Piglets.

Pseudorabies virus (PRV) variants broke out in china since 2011, causing high fever, respiratory distress, systemic neurological symptoms, and diarrhea in piglets. This study investigated the effect of intranasal PRV variant (AH02LA) infection on ileal and colonic bacterial communities and immune status in piglets. Ten piglets (free of PRV) were assigned to PRV variant and control groups (uninfected). At day 5 after inoculation, all piglets were euthanized. No PRV was detected in the ileal and colonic mucosa. In the PRV group, we observed up-regulation of specific cytokines gene expression, down-regulation of intestinal barrier-related gene expression, and reduction of secretory immunoglobulin A (sIgA) concentration in the ileum and colon. PRV infection increased the diversity of ileal bacterial community composition. PRV infection reduced the abundance of some beneficial bacteria (Lactobacillus species in the ileum and colon; butyrate-producing bacteria species in the colon) and increased the abundance of potentially pathogenic Fusobacterium nucleatum in the ileum and Sphingomonas paucimobilis in the colon. Moreover, PRV infection decreased concentrations of the beneficial lactate in the ileum and butyrate in the colon. However, this study does not allow to evaluate whether the observed changes are directly due to the PRV infection or rather to indirect effects (fever, clinical signs and changes in diet), and will be our next research content. In summary, our findings provide evidence that intranasal PRV infection directly or indirectly brings gut health risks and implications, although no PRV was detected in the ileum and colon.

Enrichment of gut-derived Fusobacterium is associated with suboptimal immune recovery in HIV-infected individuals.

We explored the gut microbiota profile among HIV-infected individuals with diverse immune recovery profiles following long-term suppressive ART and investigated the relationship between the altered bacteria with markers of immune dysfunction. The microbiota profile of rectal swabs from 26 HIV-infected individuals and 20 HIV-uninfected controls were examined. Patients were classified as suboptimal responders, sIR (n = 10, CD4 T-cell <350 cells/ul) and optimal responders, oIR (n = 16, CD4 T-cell >500 cells/ul) after a minimum of 2 years on suppressive ART. Canonical correlation analysis(CCA) and multiple regression modelling were used to explore the association between fecal bacterial taxa abundance and immunological profiles in optimal and suboptimal responders. We found Fusobacterium was significantly enriched among the HIV-infected and the sIR group. CCA results showed that Fusobacterium abundance was negatively correlated with CD4 T-cell counts, but positively correlated with CD4 T-cell activation and CD4 Tregs. Multiple linear regression analysis adjusted for age, baseline CD4 T-cell count, antibiotic exposure and MSM status indicated that higher Fusobacterium relative abundance was independently associated with poorer CD4 T-cell recovery following ART. Enrichment of Fusobacterium was associated with reduced immune recovery and persistent immune dysfunction following ART. Modulating the abundance of this bacterial taxa in the gut may be a viable intervention to improve immune reconstitution in our setting.

Dysbiosis of oral microbiota and its association with salivary immunological biomarkers in autoimmune liver disease.

The gut microbiota has recently been recognized to play a role in the pathogenesis of autoimmune liver disease (AILD), mainly primary biliary cholangitis (PBC) and autoimmune hepatitis (AIH). This study aimed to analyze and compare the composition of the oral microbiota of 56 patients with AILD and 15 healthy controls (HCs) and to evaluate its association with salivary immunological biomarkers and gut microbiota. The subjects included 39 patients with PBC and 17 patients with AIH diagnosed at our hospital. The control population comprised 15 matched HCs. Salivary and fecal samples were collected for analysis of the microbiome by terminal restriction fragment length polymorphism of 16S rDNA. Correlations between immunological biomarkers measured by Bio-Plex assay (Bio-Rad) and the oral microbiomes of patients with PBC and AIH were assessed. Patients with AIH showed a significant increase in Veillonella with a concurrent decrease in Streptococcus in the oral microbiota compared with the HCs. Patients with PBC showed significant increases in Eubacterium and Veillonella and a significant decrease in Fusobacterium in the oral microbiota compared with the HCs. Immunological biomarker analysis showed elevated levels of inflammatory cytokines (IL-1ß, IFN-γ, TNF-α, IL-8) and immunoglobulin A in the saliva of patients with AILD. The relative abundance of Veillonella was positively correlated with the levels of IL-1ß, IL-8 and immunoglobulin A in saliva and the relative abundance of Lactobacillales in feces. Dysbiosis of the oral microbiota is associated with inflammatory responses and reflects changes in the gut microbiota of patients with AILD. Dysbiosis may play an important role in the pathogenesis of AILD.

In Vitro Activities of Pexiganan and 10 Comparator Antimicrobials against 502 Anaerobic Isolates Recovered from Skin and Skin Structure Infections.

Pexiganan, a cationic peptide, exhibited a broad range of anti-anaerobic antimicrobial activity. The MIC90s of studied isolates were as follows: Bacteroides fragilis, 16 µg/ml; other B. fragilis group spp., 4 µg/ml; Prevotella and Fusobacterium spp., 32 µg/ml; Porphyromonas spp., 64 µg/ml; Propionibacterium acnes, 4 µg/ml; Eggerthella lenta and Peptostreptococcus anaerobius, 32 µg/ml; other Gram-positive rods and cocci, 4 µg/ml; Clostridium perfringens, 128 µg/ml; and other clostridia, 256 µg/ml. Pexiganan cream shows potential as adjunctive therapy for skin and skin structure infections (SSSIs) involving anaerobes.

A reproducible microcosm biofilm model of subgingival microbial communities.

OBJECTIVE: To develop a reproducible subgingival microcosm biofilm model. MATERIAL AND METHODS: Subgingival plaque samples were collected from four deep pockets (probing pocket depth ≥6 mm) in each of seven patients with periodontitis and from shallow pockets (probing pocket depth ≤3 mm) in two periodontally healthy donors. An active attachment model and a peptone medium (Thompson et. al., Appl Environ Microbiol 2015;81:8307-8314) supplemented with 30% serum was used. Biofilms were harvested at 2 and 4 weeks. DNA of dead cells was blocked for amplification by propidium monoazide treatment. Composition was analyzed using 16S rRNA gene amplicon pyrosequencing. Similarities between the biofilm samples were assessed by non-metric multidimensional scaling using the Bray-Curtis similarity index and similarity percentage analysis. Data from duplicate experiments, different biofilm sources and different biofilm age were compared. RESULTS: The non-metric multidimensional scaling revealed a strong clustering by the inoculum source, the donor and their periodontal status. Statistically significant differences were found between the sources of inoculum (P=.0001) and biofilm age (P=.0016). Furthermore, periodontitis biofilms (P) were distinct in composition from health-derived biofilms (H) by genera: Porphyromonas (P=19%; H=0%), Filifactor (P=10%; H=0%), Anaeroglobus (P=3%; H=0%), Phocaeicola (P=1.5%; H=0%), Parvimonas (P=19%; H=14%), Fusobacterium (P=2%; H=26%), Peptostreptococcus (P=20%; H=30%), Veillonella (P=7%; H=8%) and 57 other genera. Similarity distances (Bray-Curtis) (mean 0.73, SD 0.15) and the Shannon diversity index (mean 2, SD 0.2) revealed no differences between duplicate experiments (P=.121). CONCLUSION: This biofilm model allows reproducible production of complex subgingival microbial communities.

Amino Acid Block Copolymers with Broad Antimicrobial Activity and Barrier Properties.

Antimicrobial properties of a long-chain, synthetic, cationic, and hydrophobic amino acid block copolymer are reported. In 5 and 60 min time-kill assays, solutions of K100 L40 block copolymers (poly(l-lysine·hydrochloride)100 -b-poly(l-leucine)40 ) at concentrations of 10-100 µg mL-1 show multi-log reductions in colony forming units of Gram-positive and Gram-negative bacteria, as well as yeast, including multidrug-resistant strains. Driven by association of hydrophobic segments, K100 L40 copolymers form viscous solutions and self-supporting hydrogels in water at concentrations of 1 and 2 wt%, respectively. These K100 L40 preparations provide an effective barrier to microbial contamination of wounds, as measured by multi-log decreases of tissue-associated bacteria with deliberate inoculation of porcine skin explants, porcine open wounds, and rodent closed wounds with foreign body. Based on these findings, amino acid copolymers with the features of K100 L40 can combine potent, direct antimicrobial activity and barrier properties in one biopolymer for a new approach to prevention of wound infections.

Reducing exposure to pathogens in the horse: a preliminary study into the survival of bacteria on a range of equine bedding types.

AIMS: To compare the rate of growth of four microbial strains that cause disease in the horse, on four commonly used types of bedding. The moisture-holding capacity of each bedding type was also tested. METHODS AND RESULTS: Microbial strains included Streptococcus equi, Streptococcus zooepidemicus, Fusobacterium necrophorum, Dichelobacter nodosus and Dermatophilus congolensis. The bedding types tested were Pinus sylvestris (Scots pine shavings), Pinus nigra (Corsican pine shavings), Picea sitchensis (Sitka spruce shavings), Cannabis sativa (hemp) and chopped wheat straw. A suspension of each microbial strain was spread in triplicate on agar media and incubated in its optimal growth conditions. The viable count (colony-forming unit per ml) was determined for each bacterial strain for the five different bedding types. Pinus sylvestris bedding resulted in significantly less (P = 0·001) bacterial growth of all strains tested. CONCLUSIONS: Factors resulting in the inhibition of bacterial growth include the antibacterial effects reported in the Pinacea family and the physical properties of the bedding substrate. Research is currently focussed on the diagnosis and management of disease. Prevention of disease is also important for matters of biosecurity. Strategies should include the provision of a hygienic environment and the use of specific types of bedding. SIGNIFICANCE AND IMPACT OF THE STUDY: Bedding choice has implications for global equine health and disease prevention as well as potential benefits in other animal species.

Anaerobic bacteraemia: a 10-year retrospective epidemiological survey.

In order to identify current trends in anaerobic bacteraemia, a 10-year retrospective study was performed in the University Hospital Brussel, Belgium. All clinically relevant bacteraemia detected from 2004 until 2013 were included. Medical records were reviewed in an attempt to define clinical parameters that might be associated with the occurrence of anaerobic bacteraemia. 437 of the isolated organisms causing anaerobic bacteraemia were thawed, subcultured and reanalyzed using matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF). There were an average of 33 cases of anaerobic bacteraemia per year during 2004-2008 compared to an average of 27 cases per year during 2009-2013 (P = 0.017), corresponding to a decrease by 19% between the first and the latter period. Also, the total number of cases of anaerobic bacteraemia per 100,000 patient days decreased from 17.3 in the period from 2004 to 2008 to 13.7 in the period 2009 to 2013 (P = 0.023). Additionally, the mean incidence of anaerobic bacteraemia decreased during the study period (1.27/1000 patients in 2004 vs. 0.94/1000 patients in 2013; P = 0.008). In contrast, the proportion of isolated anaerobic bacteraemia compared to the number of all bacteraemia remained stable at 5%. Bacteroides spp. and Parabacteroides spp. accounted for 47.1% of the anaerobes, followed by 14.4% Clostridium spp., 12.6% non-spore-forming Gram-positive rods, 10.5% anaerobic cocci, 8.2% Prevotella spp. and other Gram-negative rods and 7.1% Fusobacterium spp. The lower gastrointestinal tract (47%) and wound infections (10%) were the two most frequent sources for bacteraemia, with the origin remaining unknown in 62 cases (21%). The overall mortality rate was 14%. Further studies focusing on the antimicrobial susceptibility and demographic background of patients are needed to further objectify the currently observed trends.

Antimicrobial activity of a quaternary ammonium methacryloxy silicate-containing acrylic resin: a randomised clinical trial.

Quaternary ammonium methacryloxy silicate (QAMS)-containing acrylic resin demonstrated contact-killing antimicrobial ability in vitro after three months of water storage. The objective of the present double-blind randomised clinical trial was to determine the in vivo antimicrobial efficacy of QAMS-containing orthodontic acrylic by using custom-made removable retainers that were worn intraorally by 32 human subjects to create 48-hour multi-species plaque biofilms, using a split-mouth study design. Two control QAMS-free acrylic disks were inserted into the wells on one side of an orthodontic retainer, and two experimental QAMS-containing acrylic disks were inserted into the wells on the other side of the same retainer. After 48 hours, the disks were retrieved and examined for microbial vitality using confocal laser scanning microscopy. No harm to the oral mucosa or systemic health occurred. In the absence of carry-across effect and allocation bias (disks inserted in the left or right side of retainer), significant difference was identified between the percentage kill in the biovolume of QAMS-free control disks (3.73 ± 2.11%) and QAMS-containing experimental disks (33.94 ± 23.88%) retrieved from the subjects (P ≤ 0.001). The results validated that the QAMS-containing acrylic exhibits favourable antimicrobial activity against plaque biofilms in vivo. The QAMS-containing acrylic may also be used for fabricating removable acrylic dentures.

Oxygen Affects Gut Bacterial Colonization and Metabolic Activities in a Gnotobiotic Cockroach Model.

The gut microbiota of termites and cockroaches represents complex metabolic networks of many diverse microbial populations. The distinct microenvironmental conditions within the gut and possible interactions among the microorganisms make it essential to investigate how far the metabolic properties of pure cultures reflect their activities in their natural environment. We established the cockroach Shelfordella lateralis as a gnotobiotic model and inoculated germfree nymphs with two bacterial strains isolated from the guts of conventional cockroaches. Fluorescence microscopy revealed that both strains specifically colonized the germfree hindgut. In diassociated cockroaches, the facultatively anaerobic strain EbSL (a new species of Enterobacteriaceae) always outnumbered the obligately anaerobic strain FuSL (a close relative of Fusobacterium varium), irrespective of the sequence of inoculation, which showed that precolonization by facultatively anaerobic bacteria does not necessarily favor colonization by obligate anaerobes. Comparison of the fermentation products of the cultures formed in vitro with those accumulated in situ indicated that the gut environment strongly affected the metabolic activities of both strains. The pure cultures formed the typical products of mixed-acid or butyrate fermentation, whereas the guts of gnotobiotic cockroaches accumulated mostly lactate and acetate. Similar shifts toward more-oxidized products were observed when the pure cultures were exposed to oxygen, which corroborated the strong effects of oxygen on the metabolic fluxes previously observed in termite guts. Oxygen microsensor profiles of the guts of germfree, gnotobiotic, and conventional cockroaches indicated that both gut tissue and microbiota contribute to oxygen consumption and suggest that the oxygen status influences the colonization success.

Mechanistic analysis of PLGA/HPMC-based in-situ forming implants for periodontitis treatment.

In-situ forming implant formulations based on poly(lactic-co-glycolic acid) (PLGA), acetyltributyl citrate (ATBC), minocycline HCl, N-methyl pyrrolidone (NMP) and optionally hydroxypropyl methylcellulose (HPMC) were prepared and thoroughly characterized in vitro. This includes electron paramagnetic resonance (EPR), nuclear magnetic resonance ((1)H NMR), mass change and drug release measurements under different conditions, optical microscopy, size exclusion chromatography (SEC) as well as antibacterial activity tests using gingival crevicular fluid samples from periodontal pockets of periodontitis patients. Based on these results, deeper insight into the physico-chemical phenomena involved in implant formation and the control of drug release could be gained. For instance, the effects of adding HPMC to the formulations, resulting in improved implant adherence and reduced swelling, could be explained. Importantly, the in-situ formed implants effectively hindered the growth of bacteria present in the patients' periodontal pockets. Interestingly, the systems were more effectively hindering the growth of pathogenic bacterial strains (e.g., Fusobacterium nucleatum) than that of strains with a lower pathogenic potential (e.g., Streptococcus salivarius). In vivo, such a preferential action against the pathogenic bacteria can be expected to give a chance to the healthy flora to re-colonize the periodontal pockets.

Evolution of invasion in a diverse set of Fusobacterium species.

UNLABELLED: The diverse Fusobacterium genus contains species implicated in multiple clinical pathologies, including periodontal disease, preterm birth, and colorectal cancer. The lack of genetic tools for manipulating these organisms leaves us with little understanding of the genes responsible for adherence to and invasion of host cells. Actively invading Fusobacterium species can enter host cells independently, whereas passively invading species need additional factors, such as compromise of mucosal integrity or coinfection with other microbes. We applied whole-genome sequencing and comparative analysis to study the evolution of active and passive invasion strategies and to infer factors associated with active forms of host cell invasion. The evolution of active invasion appears to have followed an adaptive radiation in which two of the three fusobacterial lineages acquired new genes and underwent expansions of ancestral genes that enable active forms of host cell invasion. Compared to passive invaders, active invaders have much larger genomes, encode FadA-related adhesins, and possess twice as many genes encoding membrane-related proteins, including a large expansion of surface-associated proteins containing the MORN2 domain of unknown function. We predict a role for proteins containing MORN2 domains in adhesion and active invasion. In the largest and most comprehensive comparison of sequenced Fusobacterium species to date, we have generated a testable model for the molecular pathogenesis of Fusobacterium infection and illuminate new therapeutic or diagnostic strategies. IMPORTANCE: Fusobacterium species have recently been implicated in a broad spectrum of human pathologies, including Crohn's disease, ulcerative colitis, preterm birth, and colorectal cancer. Largely due to the genetic intractability of member species, the mechanisms by which Fusobacterium causes these pathologies are not well understood, although adherence to and active invasion of host cells appear important. We examined whole-genome sequence data from a diverse set of Fusobacterium species to identify genetic determinants of active forms of host cell invasion. Our analyses revealed that actively invading Fusobacterium species have larger genomes than passively invading species and possess a specific complement of genes-including a class of genes of unknown function that we predict evolved to enable host cell adherence and invasion. This study provides an important framework for future studies on the role of Fusobacterium in pathologies such as colorectal cancer.

Individual growth detection of bacterial species in an in vitro oral polymicrobial biofilm model.

Most in vitro studies on the antibacterial effects of antiseptics have used planktonic bacteria in monocultures. However, this study design does not reflect the in vivo situation in oral cavities harboring different bacterial species that live in symbiotic relationships in biofilms. The aim of this study was to establish a simple in vitro polymicrobial model consisting of only three bacterial strains of different phases of oral biofilm formation to simulate in vivo oral conditions. Therefore, we studied the biofilm formation of Actinomyces naeslundii (An), Fusobacterium nucleatum (Fn), and Enterococcus faecalis (Ef) on 96-well tissue culture plates under static anaerobic conditions using artificial saliva according to the method established by Pratten et al. that was supplemented with 1 g l(-1) sucrose. Growth was separately determined for each bacterial strain after incubation periods of up to 72 h by means of quantitative real-time polymerase chain reaction and live/dead staining. Presence of an extracellular polymeric substance (EPS) was visualized by Concanavalin A staining. Increasing incubation times of up to 72 h showed adhesion and propagation of the bacterial strains with artificial saliva formulation. An and Ef had significantly higher growth rates than Fn. Live/dead staining showed a median of 49.9 % (range 46.0-53.0 %) of living bacteria after 72 h of incubation, and 3D fluorescence microscopy showed a three-dimensional structure containing EPS. An in vitro oral polymicrobial biofilm model was established to better simulate oral conditions and had the advantage of providing the well-controlled experimental conditions of in vitro testing.

Activity of ceftolozane-tazobactam against a broad spectrum of recent clinical anaerobic isolates.

We evaluated in vitro activity of ceftolozane-tazobactam (TOL-TAZ), formerly CXA-201, against recent clinical anaerobic isolates with emphasis on the Bacteroides fragilis group. Ceftolozane-tazobactam showed good activity against B. fragilis species and intermediate to limited activity against other species of Bacteroides. Ceftolozane-tazobactam showed very good activity against Prevotella spp., Fusobacterium spp., and Propionibacterium spp., varying activities against Gram-positive cocci, and limited activity against Clostridium spp.

Establishment of ruminal bacterial community in dairy calves from birth to weaning is sequential.

AIM: Establishment of ruminal bacterial community in dairy calves. METHODS AND RESULTS: Rumen bacterial community was analysed on 6 calves bred according to commercial practices from day one to weaning at day 83 of age, using 454 16S rRNA-based pyrosequencing. Samples taken at day 1 did not produce amplicons. Analysis of data revealed a three-stage implantation process with a progressive but important shift of composition. At day 2, the bacterial community was mainly composed of Proteobacteria (70%) and Bacteroidetes (14%), and Pasteurellaceae was the dominant family (58%). The bacterial community abruptly changed between days 2 and 3, and until day 12, dominant genera were Bacteroides (21%), Prevotella (11%), Fusobacterium (5%) and Streptococcus (4%). From 15 to 83 days, when solid food intake rapidly increased, Prevotella became dominant (42%) and many genera strongly decreased or were no longer detected. A limited number of bacteria genera correlated with feed intake, rumen volatile fatty acids and enzymatic activities. CONCLUSION: The ruminal bacterial community is established before intake of solid food, but solid food arrival in turn shapes this community. SIGNIFICANCE AND IMPACT OF THE STUDY: This study provides insight into the establishment of calves' rumen bacterial community and suggests a strong effect of diet.

Nested culture method improves detection of Fusobacterium from stool in patients with ulcerative colitis.

Fusobacterium varium is an elusive pathogenic factor in ulcerative colitis (UC); conventional methods of fecal culture rarely recover F. varium. We have developed a nested culture method to recover Fusobacterium and we used it to investigate whether F. varium could be isolated from UC patients. We enrolled 50 consecutive patients in this study; 26 received combination antibiotic therapy that included amoxicillin, tetracycline, and metronidazole (ATM) for 2 weeks and were thus assigned to the ATM group, and the remaining 24 were assigned to the non-ATM group and did not receive any antibiotics. Stool samples were added to 10 ml of GAM broth that contained neomycin and crystal violet. The samples were vortexed and incubated under anaerobic conditions. The preincubated broth was streaked onto a Fusobacterium-selective agar plate and then incubated under anaerobic conditions. The species of the colonies isolated were identified using the Vitek Automated system and PCR analysis. We recoverd F. varium from 7 of the 24 non-ATM patients (29.2%) and none from the ATM patients (0%) (P = 0.0035). All of the F. varium isolates were susceptible to ATM. This study suggests that the recovery of F. varium is related to UC, which aligns with results from previous studies that used mucosal culture, immunostaining, real-time PCR, and serological studies.