GOurmet: A tool for quantitative comparison and visualization of gene expression profiles based on gene ontology (GO) distributions

Background  

The ever-expanding population of gene expression profiles (EPs) from specified cells and tissues under a variety of experimental conditions is an important but difficult resource for investigators to utilize effectively. Software tools have been recently developed to use the distribution of gene ontology (GO) terms associated with the genes in an EP to identify specific biological functions or processes that are over- or under-represented in that EP relative to other EPs. Additionally, it is possible to use the distribution of GO terms inherent to each EP to relate that EP as a whole to other EPs. Because GO term annotation is organized in a tree-like cascade of variable granularity, this approach allows the user to relate (e.g., by hierarchical clustering) EPs of varying length and from different platforms (e.g., GeneChip, SAGE, EST library).     

Critical role of Toll‐like receptor 2 in Bacteroides fragilis‐mediated immune responses in murine peritoneal mesothelial cells

In this study, the role of Toll‐like receptor 2 (TLR2) in immune responses of murine peritoneal mesothelial cells against Bacteroides fragilis was investigated. Enzyme linked immunosorbent assay was used to measure cytokines and chemokines. Activation of nuclear factor κB (NF‐κB‐α) and mitogen‐activated protein kinases (MAP kinases) was investigated by western blot analysis. B. fragilis induced production of interleukin‐6, chemokine (C‐X‐C motif) ligand 1 (CXCL1) and chemokine (C‐C motif) ligand 2 (CCL2) in wild type peritoneal mesothelial cells; this was impaired in TLR2‐deficient cells. In addition, in response to B. fragilis, phosphorylation of inhibitory NF‐κB‐α and c‐Jun N‐terminal kinase mitogen‐activated protein kinase (MAPK) was induced in wild type mesothelial cells, but not in TLR2‐deficient cells,. Inhibitor assay revealed that NF‐κB and MAPKs are essential for B. fragilis‐induced production of CXCL1 and CCL2 in mesothelial cells. These findings suggest that TLR2 mediates immune responses in peritoneal mesothelial cells in response to B. fragilis.     

Quorum Sensing in <Emphasis Type="Italic">Aeromonas</Emphasis> Species Isolated from Patients in Malaysia

Bacterial quorum sensing signal molecules called N-acylhomoserine lactone (AHL) controls the expression of virulence determinants in many Gram-negative bacteria. We determined AHL production in 22 Aeromonas strains isolated from various infected sites from patients (bile, blood, peritoneal fluid, pus, stool and urine). All isolates produced the two principal AHLs, N-butanoylhomoserine lactone (C4-HSL) and N-hexanoyl homoserine lactone (C6-HSL). Ten isolates also produced additional AHLs. This report is the first documentation of Aeromonas sobria producing C6-HSL and two additional AHLs with N-acyl side chain longer than C₆. Our data provides a better understanding of the mechanism(s) of this environmental bacterium emerging as a human pathogen.     

Detection of Bacteroides fragilis in clinical specimens by polymerase chain reaction amplification of the neuraminidase gene

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.     

G × E Analysis of Growth Traits of <i>Betula platyphylla</i> Clones at Three Separated Sites in Northeastern China

To select elite materials, the growth traits of 32 Betula platyphylla clones at three separated northern sites in Northeast China were investigated and analyzed. The results showed that there were significant differences among all variation sources in the different investigated traits (P < 0.01). Except for the carbon contents, all the coefficients of phenotypic variation of the other investigated traits were higher than 10%. The repeatability of different traits ranged from 0.760 to 0.998. Correlation analysis showed that tree height were significantly correlated with diameter at breast height, but neither was significantly correlated with leaf traits nor element contents. Additive main effects and multiplicative interaction analysis showed that genotype, environment and genotype × environment interactions were significantly different in diameter at breast height, which indicated that environment had a significant effect on genotype. Comprehensive assessment results showed that three clones with high and stable diameters at breast height were selected, and the genetic gains of diameter at breast height on sites Maoershan, Qingan, and Yongji were 21.24%, 20.58%, and 38.65%, respectively. The results could provide a theoretical basis for elite clone selection in B. platyphylla and other broad leaved species.     

Plasmid-Related Resistance to Cefoxitin in Species of the Bacteroides fragilis Group Isolated from Intestinal Tracts of Calves

Species of the Bacteroides fragilis group are considered the most common anaerobe in human and animal infections and also harbor plasmids conferring resistance to several antibiotics. In this study, resistance to cefoxitin, plasmid profile and β-lactamase production in species of the B. fragilis group isolated from intestinal tracts of calves were evaluated. One hundred sixty-one B. fragilis group bacteria isolated from calves with and without diarrhea were analyzed. Cefoxitin susceptibility was performed using an agar dilution method, β-lactamase production by using a nitrocefin method, and plasmid extraction by using a commercial kit. Minimal inhibitory concentration values for cefoxitin ranged from 32 to > 512 μg/ml, and 47 bacteria (29.2%) were resistant to cefoxitin (breakpoint 16 μl). Only seven isolates harbored plasmids varying from 6.0 to 5.0 kb, and a 5.5-kb plasmid in B. vulgatus Bd26e and B. fragilis Bc5j might be related to cefoxitin resistance. β-lactamase was detected in 33 (70.2%) isolates. The cepA gene was observed in total DNA and in the 5.5-kb plasmid. The plasmid presence in organisms isolated from cattle may be important in ecologic terms, and it needs further study.     

Anaerobic Bacterial Flora of Intra-abdominal Infections and their Antimicrobial Susceptibility Pattern in Kuwait

Clinical samples obtained from 200 patients with intra-abdominal infections were investigated for the presence of anaerobic bacteria. The majority of samples were from patients with appendicitis (108, 54%) followed by peritoneal abscess/peritonitis (37, 18.5%). A total of 153 anaerobes were isolated from 83 culture positive specimens with an isolation rate of 1.8 per sample. Ninety (59%) yielded Bacteroides fragilis group and B. fragilis stricto sensu accounted for half of them. Other isolates were 36 (23.5%) Prevotella species and 15 (9.8%)Peptostreptococcus micros . The susceptibility of the 153 isolates against eight antibiotics was determined by the E-test. All the isolates were susceptible to metronidazole, MIC₉₀s varying between 1–2 μg/mL. ThePrevotella spp., Peptostreptococcus spp., Fusobacterium spp. and Porphyromonas spp. were all susceptible to clindamycin (MIC₉₀s=0.25–2 μg/mL respectively), imipenem (MIC₉₀s=0.12–0.5μg/mL respectively) and meropenem (MIC₉₀=0.25 μg/mL each). About 25% of the B. fragilis group were resistant to clindamycin with MIC more than 256 μg/mL. Piperacillin-tazobactam also exhibited excellent in vitro activity against all the isolates (MIC₉₀=0.25 μg/mL).     

In vitro Activity of Gatifloxacin Against 238 Strains of Anaerobic Bacteria

The activity of gatifloxacin, a new 8-methoxy-fluoroquinolone, was tested against 208 pulmonary pathogens and against an additional 30 isolates of the Bacteroides fragilis group. Pulmonary isolates were from patients with documented anaerobic pleuropulmonary infections and were obtained by appropriate sampling methods. MICs were determined using the NCCLS-approved Wadsworth brucella laked blood agar method and compared to those of clindamycin, imipenem, metronidazole and trovafloxacin. Breakpoints used to define susceptible and [resistant] categories were (in μg/ml): Clindamycin-2, imipenem-4, metronidazole 8 and trovafloxacin. No breakpoint has been defined for gatifloxacin. Gatifloxacin inhibited 99% of all anaerobes tested at 4 μg/ml and 97% of all strains at 2 μg/ml. One strain of B. fragilis was resistant to gatifloxacin at 4 μg/ml; all strains of other B. fragilis group species were susceptible. One strain of Peptostreptococcus sp. was resistant to both gatifloxacin and trovafloxacin (MIC >4 μg/ml). All other strains were susceptible to all agents at ≤μg/ml. All of the non-sporeforming Gram-positive rods were susceptible to gatifloxacin at ≤μg/ml (three strains had an MIC of 4 μg/ml). Trovafloxacin had MICs of 4 μg/ml for two strains, and an MIC of 8 μg/ml for one strain. Five percent of B. fragilis, 21% of other B. fragilis group species and 20% of Clostridium species (other than C. difficile, C. perfringens or C. ramosum) were resistant to clindamycin. No imipenem resistant isolates were found in this study. Gatifloxacin appears to have excellentin vitro activity against pulmonary isolates of anaerobes and very good activity against strains of the B. fragilis group.     

A model of open-loop control of equilibrium position and stiffness of the human elbow joint

According to the equilibrium point theory, the control of posture and movement involves the setting of equilibrium joint positions (EP) and the independent modulation of stiffness. One model of EP control, the α-model, posits that stable EPs and stiffness are set open-loop, i.e. without the aid of feedback. The purpose of the present study was to explore for the elbow joint the range over which stable EPs can be set open-loop and to investigate the effect of co-contraction on intrinsic low-frequency elbow joint stiffness (K _ilf). For this purpose, a model of the upper and lower arm was constructed, equipped with Hill-type muscles. At a constant neural input, the isometric force of the contractile element of the muscles depended on both the myofilamentary overlap and the effect of sarcomere length on the sensitivity of myofilaments to [Ca²⁺] (LDCS). The musculoskeletal model, for which the parameters were chosen carefully on the basis of physiological literature, captured the salient isometric properties of the muscles spanning the elbow joint. It was found that stable open-loop EPs could be achieved over the whole range of motion of the elbow joint and that K _ilf, which ranged from 18 to 42 N m·rad⁻¹, could be independently controlled. In the model, LDCS contributed substantially to K _ilf (up to 25 N m·rad⁻¹) and caused K _ilf to peak at a sub-maximal level of co-contraction.     

Development,polymorphism, and cross-taxon utility of EST–SSR markers from safflower (<Emphasis Type="Italic">Carthamus tinctorius</Emphasis> L.)

The breeding of japonica varieties with erect-pose panicle (EP) has recently progressed in the northern part of China, because these varieties exhibit a far higher grain yield than the varieties with normal-pose panicle (NP). A genetic analysis using the F₂ population from the cross between Liaojing5, the first japonica EP variety in China, and the Japanese japonica NP variety Toyonishiki revealed that EP is governed by a single dominant gene EP. Based on previous studies, map-based cloning of EP locus was conducted using Liaojing5, Toyonishiki, their F₂ population, and a pair of near-isogenic lines for EP locus (ZF14 and WF14) derived from the cross between the two varieties; consequently, the STS marker H90 was found to completely cosegregate with panicle pose. The H90 is located in the coding sequence AK101247 in the database, and the AK101247 of Liaojing5 has a 12 bp sequence in exon 5 replaced with a 637 bp sequence of its wild type allele. It was therefore considered that the AK101247 encodes the protein of the wild type allele at EP locus, and that the sequence substitution in exon 5 of Liaojing5 is crucial for expression of the EP phenotype. The effects of EP gene on agronomic traits were investigated using two pairs of near-isogenic lines (ZF6 vs. WF6 and ZF14 vs. WF14) derived from the cross between the two varieties. Experimental results showed that EP gene markedly enhanced grain yield, chiefly by increasing number of secondary branches and number of grains on the secondary branch. EP gene also produced a remarkable increase in grain density.     

Effect of osmotic stress on plant growth promoting Pseudomonas spp.

In this study we isolated and screened drought tolerant Pseudomonas isolates from arid and semi arid crop production systems of India. Five isolates could tolerate osmotic stress up to −0.73 MPa and possessed multiple PGP properties such as P-solubilization, production of phytohormones (IAA, GA and cytokinin), siderophores, ammonia and HCN however under osmotic stress expression of PGP traits was low compared to non-stressed conditions. The strains were identified as Pseudomonas entomophila, Pseudomonas stutzeri, Pseudomonas putida, Pseudomonas syringae and Pseudomonas monteilli respectively on the basis of 16S rRNA gene sequence analysis. Osmotic stress affected growth pattern of all the isolates as indicated by increased mean generation time. An increase level of intracellular free amino acids, proline, total soluble sugars and exopolysaccharides was observed under osmotic stress suggesting bacterial response to applied stress. Further, strains GAP-P45 and GRFHYTP52 showing higher levels of EPS and osmolytes (amino acids and proline) accumulation under stress as compared to non-stress conditions, also exhibited higher expression of PGP traits under stress indicating a relationship between stress response and expression of PGP traits. We conclude that isolation and screening of indigenous, stress adaptable strains possessing PGP traits can be a method for selection of efficient stress tolerant PGPR strains.     

Diversity of <Emphasis Type="Italic">Burkholderia cepacia</Emphasis> Complex from the Moso Bamboo (<Emphasis Type="Italic">Phyllostachys edulis</Emphasis>) Rhizhosphere Soil

The purpose of this study was to determine the existence of Burkholderia cepacia complex (Bcc) at species level and the predominant species in the environment of moso bamboo plantations in Hangzhou, China. A total of 423 isolates were recovered from moso bamboo rhizhosphere soil samples of three sites on the selective medium during 2007–2008. Isolates were identified by Bcc-specific PCR assays, followed by recA-restriction fragment length polymorphism assays, species-specific PCR analysis, recA gene sequencing, multilocus sequence typing (MLST) scheme, and BOX-PCR fingerprinting for genomic diversity. Out of 423 isolates, 278 isolates were assigned to the following Bcc species, eight B. stabilis, 26 B. anthina, 193 B. pyrrocinia, and 51 B. arboris, which indicated B. pyrrocinia as the most dominant species followed by B. arboris. Moreover, false positives were observed in certain isolates of B. arboris while performing species-specific PCR test. Furthermore, the results of recA gene sequence similarity and MLST data demonstrated that nine isolates formed a single discrete cluster but were PCR negative to species-specific primers representing novel species may exist within the Bcc. In addition, BOX-PCR fingerprinting for all the Bcc isolates also showed the strain diversity. It is the first report of the existence of B. arboris and predominance of B. pyrrocinia in the moso bamboo environment.     

Close encounters of the type‐six kind: injected bacterial toxins modulate gut microbial composition

Bacteria of the phylum Bacteroidetes constitute a substantial portion of the human gut microbiota, including symbionts and opportunistic pathogens. How these bacteria coexist and provide colonization resistance to pathogenic strains is not well understood. In this issue of EMBO Reports, Hecht and colleagues describe a mechanism by which strains of Bacteroides fragilis compete with each other for an intestinal niche 1 . Prompted by the observation that B. fragilis populations appear to be dominated by either commensal, non‐toxigenic strains, or by enterotoxigenic, potentially pathogenic strains, the authors investigated mechanisms of competition between these two subsets. In agreement with two recent studies 2 3 , Hecht et al 1 found that competition between B. fragilis strains is dependent on a type‐6 secretion system (T6SS) apparatus, secreted effectors, and immunity genes. They identify a T6SS effector–immunity gene pair that enables a non‐toxigenic strain to competitively exclude enterotoxigenic B. fragilis, thus providing a proof of principle for the use of T6SS‐mediated killing as a therapeutic strategy to eradicate pathogenic strains.     

Identification of the Metallo-β-Lactamase Gene from Clinical Isolates of Bacteroides fragilis

Bacteroides fragilis is one of the organisms known to produce carbapenem-hydrolysing metallo-β-lactamase, which can confer resistance to a wide variety of β-lactams. The purpose of this study was to identify carbapenem-hydrolysing metallo-β-lactamase-producing B. fragilis strains by means of PCR assay, nucleotide sequencing and enzyme inhibition studies. Ten β-lactam-resistant B. fragilis isolates were investigated. Four imipenem-resistant strains among the 10 isolates gave positive reactions in the PCR assay. The nucleotide sequences of the PCR products from two imipenem-resistant strains shared >98% similarity with the metallo-β-lactamase gene from B. fragilis TAL 3636, which was used as a control. The amino acid sequence homology between the two imipenem-resistant strains and B. fragilis TAL 3636 was 99.2%. These strains produced high amounts of Zn²⁺-dependent β-lactamases which were inactivated by EDTA.     

Expression of prostaglandin E2 receptor subtypes in the canine lower urinary tract varies according to the gonadal status and gender

Locally-synthesised prostaglandin E₂ (PGE₂) is pivotal for the function of the lower urinary tract (LUT). This study aimed at investigating the expression and distribution pattern of the four PGE₂ receptor (EP) subtypes in the LUT of intact and gonadectomised male and female dogs. Expression for EP1, EP2, EP3, and EP4 and their mRNA (EP2, EP3, and EP4) was investigated. Twenty clinically healthy dogs were allotted into 4 groups based on their gonadal status and gender including 5 intact males, 5 anoestrous females, 4 castrated males, and 6 spayed females. In situ hybridization and immunohistochemistry showed variation in the expression of mRNA and protein for the EP subtypes among tissue layers (epithelium, sub-epithelial stroma, and muscle), regions (body and neck of the bladder as well as proximal and distal urethra) and between gonadal statuses and genders. The expression for the four EPs was intense in the luminal epithelium, intermediate to low in the muscle and the sub-epithelial stroma regardless of gonadal status or gender. Higher expression of all EPs and their mRNAs was observed in the proximal urethra compared to other regions in intact dogs. However, in gonadectomised dogs, the expression did not differ among different regions and was generally lower than in intact dogs particularly in the proximal urethra. Differences in the expression between genders were found and depended on EP subtypes. In conclusion, the results have shown that four subtypes of EP receptors and their mRNAs are present in the canine LUT and their expression was affected by the gonadal status and the gender. The results lead to suggest that an impaired LUT function post-neutering may partly be associated with differences in PGE₂ receptor expression between intact and gonadectomised dogs.     

A Selective Medium for Isolation and Presumptive Identification of the Bacteroides fragilis Group

A new selective medium, Bacteroides fragilis ammonium-sulfate gentamicin (BFAG) agar, for isolation and presumptive identification of the Bacteroides fragilis group is presented in this paper. This semisynthetic medium includes 0.2 g of ammonium sulfate, 0.7 g of lactose, 10 mg of gentamicin, 0.1 mg of aminobenzylpenicillin, 60 units of bacitracin, 20 mg of sodium cholate and 1 mg of sodium azide per 100 ml of medium. Stock cultures of the B. fragilis group grew well on this medium. None of the other 126 gram-positive or negative strains belonging to 40 aerobic or 45 anaerobic species tested grew on this medium. Three of the seven specimens in the clinical trials yielded colonies of only the B. fragilis group on BFAG agar plates. Also BFAG agar plates inoculated with human feces and contents of the alimentary tract (stomach, small intestine, cecum and colon) of mice gave rise to colonies of only the B. fragilis group. The high selectivity and good plating efficiency of BFAG agar enabled us to isolate the B. fragilis group rapidly from various clinical specimens.     

Electrophoretic analysis of the lipopolysaccharides ofBacteroides spp.

Lipopolysaccharide (LPS) extracts of reference strains and isolates ofBacteroides spp. prepared by the proteinase K method were resolved by tricine-sodium-dodecyl-sulphate-polyacrylamide gel electrophoresis and located by silver staining. A considerable diversity of LPS profiles was evident within theBacteroides genus although profiles were essentially species-specific, with some minor interstrain variations apparent among isolates ofB. uniformis, B. ovatus, B. eggerthii andB. thetaiotaomicron. The LPS of most species consisted of a major rough LPS component of 2–5 kDa and a series of higher molecular weight bands which varied with species.B. vulgatus LPS was distinctive in showing an extensive ladder of multiple repeating oligosaccharide units with molecular weights ranging from 4 to >17 kDaB. stercoris LPS included a high molecular weight (>17 kDa) ladder of repeating oligosaccharide units.B. fragilis andB. thetaiotaomicron differed from most other species in producing a short ladder of repeating oligosaccharide units interspersing the rough LPS and a 5.6 kDa (B. fragilis) or 9 kDa (B. thetaiotaomicron) yellow-staining component. The heterogeneity of LPS profiles within theBacteroides genus may reflect the differences in pathogenicity among the species and prove useful for typing.     

Prevalence of the Bacteroides fragilis Group and Enterotoxigenic Bacteroides fragilis in Immunodeficient Children

Members of the Bacteroides fragilis group are indigenous to the human and animal intestinal microbiota and they are responsible for several endogenous infections. Enterotoxigenic B. fragilis (ETBF) has been associated with acute diarrhea in children and farm animals. Immunodeficient patients are more predisposed to different opportunistic infections, including anaerobic infections. In this study, 130 stool samples were analysed from 56 immunodeficient and 74 healthy children. Enterotoxin production was detected by cytotoxicity assay on HT-29 cells and by PCR. B. fragilis sensu strictu was prevalent in both groups and ETBF species was detected from a single stool sample belonged to an immunodeficient child with AIDS.     

Genetic and morphological heterogeneity of Lake Baikal endemic gastropod <Emphasis Type="Italic">Benedictia fragilis</Emphasis> (Dybowski, 1875)

Baikal endemic Benedictia fragilis gastropods distributed in a wide range of depths (from sublittoral to abyssal) of three lake basins are studied. The analysis of the nucleotide sequence of the COI mitochondrial gene fragment and internal transcribed nuclear DNA spacer (ITS1) demonstrates that the studied gastropods are represented in Lake Baikal by three genetic groups. The results of the studies on genetic diversity, phenotypic traits, and distribution allow us to assume that the detected groups are incipient allopatric (geographical) species. On the basis of the data obtained and geological and climatic history of Baikal, possible pathways of the B. fragilis resettlement in the lake and the emergence of three genetic groups are hypothesized.