Skin lesion-associated pathogens from Octopus vulgaris: first detection of Photobacterium swingsii, Lactococcus garvieae and betanodavirus.

The common octopus Octopus vulgaris Cuvier, 1798 is extremely important in fisheries and is a useful protein source in most Mediterranean countries. Here we investigated pathogens associated with skin lesions in 9 naturally deceased specimens that included both cultured and wild common octopus. Within 30 min after death, each octopus was stored at 4°C and microbiologically examined within 24 h. Bacterial colonies, cultured from swabs taken from the lesions, were examined using taxonomical and biochemical analyses. Vibrio alginolyticus and V. parahaemolyticus were only isolated from cultured animals. A conventional PCR targeting the 16S ribosomal RNA (rRNA) gene and sequencing were performed on 2 bacterial isolates that remained unidentified after taxonomical and biochemical analysis. The sequence results indicated that the bacteria had a 99% identity with Lactococcus garvieae and Photobacterium swingsii. L. garvieae was confirmed using a specific PCR based on the 16S-23S rRNA internal transcribed spacer region, while P. swingsii was confirmed by phylogenetic analyses. Although all animals examined were found to be infected by the protozoan species Aggregata octopiana localised in the intestines, it was also present in skin lesions of 2 of the animals. Betanodavirus was detected in both cultured and wild individuals by cell culture, PCR and electron microscopy. These findings are the first report of L. garvieae and betanodavirus from skin lesions of common octopus and the first identification of P. swingsii both in octopus skin lesions and in marine invertebrates in Italy.

The visualization of biofilms in chronic diabetic foot wounds using routine diagnostic microscopy methods.

Diabetic foot wounds are commonly colonised by taxonomically diverse microbial communities and may additionally be infected with specific pathogens. Since biofilms are demonstrably less susceptible to antimicrobial agents than are planktonic bacteria, and may be present in chronic wounds, there is increasing interest in their aetiological role. In the current investigation, the presence of structured microbial assemblages in chronic diabetic foot wounds is demonstrated using several visualization methods. Debridement samples, collected from the foot wounds of diabetic patients, were histologically sectioned and examined using bright-field, fluorescence, and environmental scanning electron microscopy and assessed by quantitative differential viable counting. All samples (n = 26) harboured bioburdens in excess of 5 log10 CFU/g. Microcolonies were identified in 4/4 samples by all three microscopy methods, although bright-field and fluorescence microscopy were more effective at highlighting putative biofilm morphology than ESEM. Results in this pilot study indicate that bacterial microcolonies and putative biofilm matrix can be visualized in chronic wounds using fluorescence microscopy and ESEM, but also using the simple Gram stain.

Fecal protease activity is associated with compositional alterations in the intestinal microbiota.

OBJECTIVE: Intestinal proteases carry out a variety of functions in the gastrointestinal (GI) tract. Studies have reported that elevated enteric proteases in patients with GI disease can alter intestinal physiology, however the origin (human vs. microbial) of elevated proteases in patients with GI disease is unclear. AIM: The aim of this study was to investigate the association between protease activity and the microbiota in human fecal samples. DESIGN: In order to capture a wide range of fecal protease (FP) activity stool samples were collected from 30 IBS patients and 24 healthy controls. The intestinal microbiota was characterized using 454 high throughput pyro-sequencing of the 16S rRNA gene. The composition and diversity of microbial communities were determined and compared using the Quantitative Insights Into Microbial Ecology (QIIME) pipeline. FP activity levels were determined using an ELISA-based method. FP activity was ranked and top and bottom quartiles (n=13 per quartile) were identified as having high and low FP activity, respectively. RESULTS: The overall diversity of the intestinal microbiota displayed significant clustering separation (p = 0.001) between samples with high vs. low FP activity. The Lactobacillales, Lachnospiraceae, and Streptococcaceae groups were positively associated with FP activity across the entire study population, whilst the Ruminococcaceae family and an unclassified Coriobacteriales family were negatively associated with FP activity. CONCLUSIONS: These data demonstrate significant associations between specific intestinal bacterial groups and fecal protease activity and provide a basis for further causative studies investigating the role of enteric microbes and GI diseases.

Levofloxacin versus azithromycin on the oropharyngeal carriage and selection of antibacterial- resistant streptococci in the microflora of healthy adults.

OBJECTIVE: To determine the proportion of subjects with oropharyngeal streptococci resistant to either levofloxacin or azithromycin prior to and during antibacterial exposure, and to follow temporal changes in the proportion of resistant and susceptible isolates through 6 weeks post-exposure. This randomized, open-label, single-center study is registered with ClinicalTrials.gov (identifier: NCT00821782). RESEARCH DESIGN AND METHODS: A total of 143 healthy volunteers (levofloxacin, n = 71; azithromycin, n = 72) without antibacterial exposure in the previous 90 days received either levofloxacin 750 mg once daily for 5 days or azithromycin 500 mg once daily on day 1 and 250 mg once daily on days 2 through 5. Oropharyngeal cultures were obtained pre-exposure, at day 5, and at 2, 4, and 6 weeks post-dosing. Bacterial strains were identified and the minimum inhibitory concentrations for levofloxacin and azithromycin were determined. RESULTS: At study entry 117 streptococci were isolated from 72 subjects randomized to azithromycin and 53 (45.3%) were azithromycin-resistant. None of the 121 streptococci isolated from 71 subjects randomized to.levofloxacin were colonized by a levofloxacin-resistant microorganism prior to dosing. At the end of dosing, the number of subjects with resistant streptococci (S. mitis, S. salivarius, S. sanguis, or alpha streptococcus species [spp.]) increased in azithromycin-exposed subjects and resistant isolates remained through 6 weeks post-dosing. In contrast, a small number of levofloxacin-resistant streptococci were observed at the end of dosing but decreased by week 2 post-dosing and continued to decrease through the 6-week evaluation period (p < 0.001 azithromycin vs. levofloxacin for S. mitis, S. salivarius, S. sanguis and alpha streptococcus spp. at week 6). Limitations of this study included the fact that, since previous antibiotic use was self-reported, genetic typing was not done. The results of this study may not be completely generalizable, because subjects in this study received study drug under directly-observed conditions, thus ensuring compliance. CONCLUSIONS: Both antibacterial agents were well tolerated. Levofloxacin 750 mg administered for 5 days was associated with less microbial resistance than that observed with azithromycin in healthy subjects.

Dexamethasone prevents alteration of tight junction-associated proteins and barrier function in porcine choroid plexus epithelial cells after infection with Streptococcus suis in vitro.

Apart from antibiotic treatment in bacterial meningitis supportive therapy including dexamethasone is widely used. In investigations on the pathogenesis of bacterial meningitis we previously demonstrated that Streptococcus suis (S. suis), a relevant cause of bacterial meningitis in pigs and humans, affects porcine choroid plexus epithelial cell (PCPEC) barrier function. The choroid plexus epithelium constitutes the structural basis of the blood-CSF barrier. Now, we investigated the role of tight junction proteins and the actin cytoskeleton of PCPEC in correlation to barrier function after S. suis infection and analyzed the influence of dexamethasone. S. suis caused massive rearrangement of the tight junction proteins ZO-1, occludin and claudin-1, caused loss of actin at the apical cell pole and induced basolateral stress fiber formation. Moreover, tight junctions were shifted from the Triton X insoluble to the Triton X soluble fraction, and additionally occludin was dephosphorylated and degraded. Infection with S. suis leads to an inflammatory response exemplified by the induction of tumor necrosis factor (TNF) alpha and matrix metalloproteinase (MMP)-3 gene activation, which correlated with phosphorylation of extracellular signal regulated kinases (ERKs). Importantly, dexamethasone significantly prevented S.suis-induced protein and morphological tight junction alterations and attenuated ERK activation and MMP-3 expression. It especially improved the barrier function by preventing tight junction protein reorganization and degradation. In the pathogenesis of bacterial meningitis protection of blood-CSF barrier by dexamethasone may prevent the penetration of bacteria and leukocytes into the CSF.

Characterization of Aerococcus viridans isolates from swine clinical specimens.

We present here the biochemical and genetic characterization and antimicrobial susceptibility of 58 isolates of Aerococcus viridans isolated in pure culture from different clinical specimens of normally sterile body sites of pigs. A. viridans isolates were commonly susceptible to beta-lactam antimicrobials and exhibited a great genetic heterogeneity as determined by pulsed-field gel electrophoresis typing. The results indicate that A. viridans might be included in the list of possible etiological agents causing disease in pigs.

Isolation and characterization of lactic acid bacteria from dochi (fermented black beans), a traditional fermented food in Taiwan.

AIMS: To isolate, characterize and identify lactic acid bacteria (LAB) in dochi (fermented black beans), a traditional fermented food in Taiwan. METHODS AND RESULTS: A total of 30 samples were collected from three different dochi producers and analysed after different periods of storage. Fifty-two cultures of LAB were isolated from dochi samples and the isolates were divided into classes by phenotype and then into groups by restriction fragment length polymorphism analysis and sequencing of 16S ribosomal DNA. Phenotypic and biochemical characteristics identified six different bacterial groups (A-F) and showed that the majority of the isolates were homofermentative LAB. Enterococcus faecium was the most abundant of the dochi-isolated LAB. All isolated LAB were able to grow in MRS broth containing 6% NaCl, but only Enterococcus, Pediococcus and Tetragenococcus species could grow in MRS broth containing 10% NaCl. Furthermore, antibacterial activities of isolates were determined, and four isolates showed inhibitory activities against the indicator strain Lactobacillus sakei JCM 1157(T). CONCLUSIONS: These results suggest that Ent. faecium is the main LAB present during the fermentation of dochi. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first report describing the distribution and varieties of LAB that exist in the dochi fermentation process.

Autophagy vs. Group A Streptococcus.

We have identified several mammalian protein components of the autophagy pathway. By using them as powerful tools to examine the functional significance of this degradation pathway, we recently showed that autophagy efficiently kills a pathogenic bacterium, Group A Streptococcus, after it invades host cells. However, the autophagosomes induced by these bacteria have features distinct from those of the canonical pathway.

Bloodstream and endovascular infections due to Abiotrophia defectiva and Granulicatella species.

BACKGROUND: Abiotrophia and Granulicatella species, previously referred to as nutritionally variant streptococci (NVS), are significant causative agents of endocarditis and bacteraemia. In this study, we reviewed the clinical manifestations of infections due to A. defectiva and Granulicatella species that occurred at our institution between 1998 and 2004. METHODS: The analysis included all strains of NVS that were isolated from blood cultures or vascular graft specimens. All strains were identified by 16S rRNA sequence analysis. Patients' medical charts were reviewed for each case of infection. RESULTS: Eleven strains of NVS were isolated during the 6-year period. Identification of the strains by 16S rRNA showed 2 genogroups: Abiotrophia defectiva (3) and Granulicatella adiacens (6) or "para-adiacens" (2). The three A. defectiva strains were isolated from immunocompetent patients with endovascular infections, whereas 7 of 8 Granulicatella spp. strains were isolated from immunosuppressed patients, mainly febrile neutropenic patients. We report the first case of "G. para-adiacens" bacteraemia in the setting of febrile neutropenia. CONCLUSION: We propose that Granulicatella spp. be considered as a possible agent of bacteraemia in neutropenic patients.

[Interaction of bacteria and red blood cells].

The interaction of staphylococci, streptococci, meningococci, enterobacteria, leptospires and other microorganisms with red blood cells is considered. Three forms of the interaction of bacteria and red blood cells are discussed: adhesion, the influence of secretory factors on red blood cells, the action of pathogenic bacteria on hemoglobin. The applied aspects of the interaction of bacteria and red blood cells in the human body are presented in accordance with the results of clinical and laboratory studies.

Reclassification of Pediococcus urinaeequi (ex Mees 1934) Garvie 1988 as Aerococcus urinaeequi comb. nov.

The taxonomic status of Pediococcus urinaeequi is described, and the transfer of the species to the genus Aerococcus with the name Aerococcus urinaeequi comb. nov. is proposed, on the basis of the analysis of 16S rRNA gene sequence and DNA-DNA hybridization data.

Mytilus galloprovincialis filter feeding on the bacterial community in a Mediterranean coastal area (Northern Ionian Sea, Italy).

This study was carried out seasonally, throughout a year, to evaluate the filtering activity on bacteria of Mytilus galloprovincialis. Six microbiological parameters were researched in the water and mussels samples collected along the coastal area of the Northern Ionian Sea in three stations, S. Vito, Lido Gandoli and Lido Silvana. We detected the densities of culturable heterotrophic bacteria by spread plate on Marine Agar, total culturable bacteria at 37 degrees C on Plate Count Agar and vibrios abundance on thiosulphate-citrate-bile-sucrose-salt (TCBS) agar. Total and fecal coliforms as well as fecal streptococci were determined by the Most Probable Number. Bacterial concentrations at 20 and 37 degrees C as well as vibrios concentrations were higher in the mussel samples compared to the corresponding seawater throughout the year. The results obtained could contribute to improve the information relatively either to the natural processes existing between bacteria and mussels or to the risk of human infections related to the consumption of mussels.

Inhibition of fibronectin binding of some bacterial cells by subtle pH increase within the physiological range.

The fibronectin (Fn)-binding ability of microorganisms is considered to be involved in their pathogenicities. Granulicatella adiacens, a member of the oral flora and a causative agent of culture-negative infective endocarditis, showed nearly maximum binding to immobilized Fn at pH 7.2 but greatly reduced binding at a slightly higher pH 7.4 and almost no binding at pH 7.6 in the presence of physiological concentration of NaCl (0.15 M). A similar pH-sensitive Fn-binding property was noted with Escherichia coli and Abiotrophia defectiva, but not with Streptococcus pyogenes nor Staphylococcus aureus. In contrast, bindings to laminin and fibrinogen observed for some of these strains were unaffected by the same pH changes. This fastidious pH-dependency of Fn-binding abilities of some bacteria warns that the pH condition must be seriously considered in the in vitro assay of bacterial adherence to fibronectin.

Classification of Homarus americanus hemocytes and the use of differential hemocyte counts in lobsters infected with Aerococcus viridans var. homari (Gaffkemia).

Hemocytes of the American lobster (Homarus americanus H. Milne Edwards) were classified after examination of Wright-Giemsa stained cytocentrifuge preparations by brightfield light microscopy. Eleven hemocyte types were identified using morphologic criteria. The classification system was then used to monitor changes in the differential hemocyte count (DHC) of lobsters infected with the Gram positive coccus Aerococcus viridans var. homari, etiologic agent of gaffkemia. The appearance of less mature hemocytes in the DHCs of lobsters in the late stages of infection was similar to the 'left shift' of vertebrate inflammation. Results from this study suggest that DHCs can be used to assess and characterize inflammation in H. americanus and possibly other crustaceans.

The lactic acid bacteria: a literature survey.

The purpose of this review article on the lactic acid bacteria grew from an early curiosity and a desire to convey and impart the broad scope of literary information on their functions as starter cultures, in the manufacture of fermentation products such as dairy products and alcoholic beverages, as well as their contribution to better health. This review article is an attempt to empower the reader and to circumvent the difficult task in acquiring and elucidating a large body of information. The intent is to familiarize the reader with the various lactic species, their habitat or source, associated food, physiological characteristics, colonial morphology, biochemical characteristics, culture media (enrichment, nonselective, and selective), classic description, and taxonomy. This review provides information on Lactobacillus, Lactococcus, Leuconostoc, Pediococcus, Carnobacterium, and Enterococcus. Trends are presented, such as the use of nisin to extend food shelf-life and the current research premise that Probiotic strains may alter the intestinal flora and thus prevent intestinal wall penetration by pathogens.

Granulicatella and Abiotrophia species from human clinical specimens.

One hundred one isolates of nutritionally variant streptococci from 97 patients were phenotypically characterized and compared with the type strains of Granulicatella adiacens (formerly Abiotrophia adiacens) (ATCC 49175(T)) Abiotrophia defectiva (ATCC 49176(T)), and Granulicatella elegans (formerly Abiotrophia elegans) (DSM 11693(T)). Of the isolates, 55 and 43 resembled G. adiacens and A. defectiva, respectively, while 3 strains resembled G. elegans. Phenotypic characteristics useful in differentiating between species within the genera Granulicatella and Abiotrophia (G. adiacens, G. elegans, Granulicatella balaenopterae, and A. defectiva) were production of alpha- and beta-galactosidase; production of beta-glucuronidase; hippurate hydrolysis; arginine dihydrolase activity; and acid production from trehalose, sucrose, pullulan, and tagatose. From the reports submitted with the specimens, the clinical diagnosis was endocarditis in 58% of patients and septicemia or bacteremia in 26% of patients.

Flocculation and coflocculation of bacteria by yeasts.

Biotransformations in natural environments frequently involve interactions between microorganisms. Although there are many reports on the interactions between bacteria, interactions between yeasts and bacteria have not been extensively studied. Previously we reported on the flocculation and coflocculation of Pediococcus damnosus by Saccharomyces cerevisiae. Now we report that several other yeasts, such as Candida utilis, Dekkera bruxellensis, Hanseniaspora guilliermondii, Kloeckera apiculata, and Schizosaccharomyces pombe, induce flocculation with several industrially or medically relevant bacteria, including Bacillus subtilis, Pseudomonas aeruginosa, and Staphylococcus aureus. Candida utilis was one of the best flocculation inducers. The results are discussed with respect to interactions between yeasts and bacteria and their applications in industry and medicine.

[Evaluation of bacterial and polluting loads of effluent from activated sludge wastewater treatment plants in Yaounde, Cameroon]. / Evaluation de la charge polluante et de la charge bactérienne des rejets des stations d'épuration a boues activées Yaoundé (Cameroun).

Activated sludge wastewater treatment plants in Yaounde are plagued by failures in their operation. A microbiological and physico-chemical study was carried out on wastewater effluent of an hospital, university campus, and two residential areas made up of more than 2,000 apartments connected to two collective sewase systems. Wastewater sampling was performed in the form of daily cycles during which composite samples were made every two hours with samples collected every half hour. Variables measured included incoming flow rates, suspended solids, pH, electrical conductivity, ammonia nitrogen, total Kjeldahl nitrogen (combination of ammonia and organic nitrogen) and total reactive phosphorus, BOD5 and COD. Microorganisms were isolated on selective culture media using membrane filter procedures. The results obtained show that effluent discharged in receiving waters are, in general, not treated. The suspended solids contents of raw sewage range from 120 to 2,600 mg/l. In General Hospital a mean value of suspended solids of 31.8 mg/l was obtained for treated wastewater. pH values of both raw and treated sewage range from 6 to 8.5 units. Mean values of the conductivity of treated and raw wastewater range from 449.7 to 1,038.3 microseconds/cm. The TKN contents of untreated wastewater are comprised between 36 and 259.2 mg/l. These values range from 5 to 20 mg/l for treated effluent in the General Hospital plant. Large amounts of total hydrolysable phosphorus varying from 3.8 to 27.9 mg/l are present in wastewater from the plants. Influent BOD5 of Grand Messa, Cite Verte and university campus wastewater range from 110 to 530 mgO2/l. Values recorded indicate an important input of organic matter to receiving waters. In the case of biotic components, test germs densities registered in the influent of Grand Messa and those of Cite Verte are comprised between 9 x 106 and 88 x 107 CFU/100 ml for fecal coliforms, and between 9 x 105 and 74 x 107 CFU/100 ml for fecal streptococci. The densities of Aeromonas hydrophila in Grand Messa and Cite Verte influent fluctuate between 30 x 106 and 65 x 108 CFU/100 ml. Influent of university campus harbour 10 x 105. Densities of Pseudomonas aeruginosa remain below 70 x 105 CFU/100 ml. Treated effluent BOD5 values range from 48 to 150 mgO2/l. Such values indicate an inefficient treatment. The decrease in bacterial densities is limited to 2 logarithmic units. Treated effluent still harbour 10 x 103 to 10 x 105 CFU/100 ml for fecal coliforms, and 20 x 102 to 12 x 104 CFU/100 ml for fecal streptococci. These values range from 10 x 104 to 50 x 105 CFU/100 ml for Aeromonas hydrophila. Pseudomonas aeruginosa densities remain in general below 40 x 104 CFU/100 ml. Discharges from these wastewater treatment plants are responsible for the permanent fecal contamination of the environment and a lack of proper hygiene. Furthermore, they produce overfertilization of receiving waters.

Atopobacter phocae gen. nov., sp. nov., a novel bacterium isolated from common seals.

Two strains of a Gram-positive, catalase-negative, facultatively anaerobic, rod-shaped bacterium isolated from common seals were characterized using phenotypic and molecular taxonomic methods. The two strains closely resembled each other based on their biochemical characteristics, and PAGE analysis of whole-cell protein patterns confirmed their close phenotypic affinity. 16S rRNA gene sequencing showed that the two strains were genetically highly related (99.8% sequence similarity) and that they constitute a new line of descent within the lactic acid group of bacteria. The nearest phylogenetic neighbours of the unknown bacterium were Granulicatella spp., with related taxa such as enterococci, carnobacteria, Desemzia incerta, Lactosphaera pasteurii, Melissococcus plutonius, tetragenococci and vagococci more distantly related. Based on phylogenetic and phenotypic evidence it is proposed that the unknown bacterium from seals be classified in a new genus as Atopobacter phocae gen. nov., sp. nov. The type strain of Atopobacter phocae is CCUG 42358T (= CIP 106392T).

The genus Abiotrophia (Kawamura et al.) is not monophyletic: proposal of Granulicatella gen. nov., Granulicatella adiacens comb. nov., Granulicatella elegans comb. nov. and Granulicatella balaenopterae comb. nov.

The genus Abiotrophia currently includes four species, Abiotrophia defectiva, Abiotrophia adiacens, Abiotrophia balaenopterae and Abiotrophia elegans. Recent 16S rRNA gene sequencing studies have demonstrated that the genus is not monophyletic and is in need of taxonomic revision. Phylogenetically, the genus Abiotrophia consists of two distinct lines, A. defectiva, the type species of the genus, and a robust group consisting of A. adiacens, A. balaenopterae and A. elegans. Therefore, it is formally proposed that the genus Abiotrophia should be restricted to A. defectiva and that A. adiacens, A. balaenopterae and A. elegans should be reclassified in a new genus, Granulicatella, as Granulicatella adiacens comb. nov., Granulicatella balaenopterae comb. nov. and Granulicatella elegans comb. nov.