Unidentified Bacteria Research Articles

Activated carbon decreases invasive plant growth by mediating plant-microbe interactions.

There is growing appreciation for the idea that plant-soil interactions (e.g. allelopathy and plant-microbe feedbacks) may explain the success of some non-native plants. Where this is the case, native plant restoration may require management tools that change plant-soil interactions. Activated carbon (AC) is one such potential tool. Previous research has shown the potential for high concentrations of AC to restore native plant growth to areas dominated by non-natives on a small scale (1 m × 1 m plots). Here we (i) test the efficacy of different AC concentrations at a larger scale (15 m × 15 m plots), (ii) measure microbial responses to AC treatment and (iii) use a greenhouse experiment to identify the primary mechanism, allelopathy versus microbial changes, through which AC impacts native and non-native plant growth. Three years after large-scale applications, AC treatments decreased non-native plant cover and increased the ratio of native to non-native species cover, particularly at concentrations >400 g m(-2). Activated carbon similarly decreased non-native plant growth in the greenhouse. This effect, however, was only observed in live soils, suggesting that AC effects were microbially mediated and not caused by direct allelopathy. Bacterial community analysis of field soils indicated that AC increased the relative abundance of an unidentified bacterium and an Actinomycetales and decreased the relative abundance of a Flavobacterium, suggesting that these organisms may play a role in AC effects on plant growth. Results support the idea that manipulations of plant-microbe interactions may provide novel and effective ways of directing plant growth and community development (e.g. native plant restoration).

Direct identification of Gram-positive bacteria and resistance determinants from blood cultures using a microarray-based nucleic acid assay: in-depth analysis of microarray data for undetermined results.

The Verigene Gram-Positive Blood Culture (BC-GP) nucleic acid assay (Nanosphere, Inc., Northbrook, IL, USA) is a newly developed microarray-based test with which 12 Gram-positive bacterial genes and three resistance determinants can be detected using blood culture broths. We evaluated the performance of this assay and investigated the signal characteristics of the microarray images. At the evaluation stage, we tested 80 blood cultures that were positive for various bacteria (68 bacteria covered and 12 not covered by the BC-GP panel) collected from the blood of 36 patients and 44 spiked samples. In instances where the automated system failed and errors were called, we manually inspected microarray images, measured the signal intensities of target spots, and reclassified the results. With the manual analysis of the microarray images of 14 samples for which error calls were reported, we could obtain correct identification results for 12 samples without the need for retesting, because strong signals in the target spots were clearly discriminable from background noise. With our interpretation strategy, we could obtain 97.1% sensitivity and 100% specificity for bacterial identification by using the BC-GP assay. The two unidentified bacteria were viridans group streptococci, which produced weaker target signals. During the application stage, among 25 consecutive samples positive for Gram-positive bacteria, we identified two specimens with error calls as Streptococcus spp. by using manual analysis. With help of the manual review of the microarray images, the BC-GP assay could successfully identify species and resistance markers for many clinically important Gram-positive bacteria.

Antibiotic Therapy of H. Pylori Negative Gastric MALT Lymphoma

Abstract Introduction The causal association between H. pylori and gastric MALT lymphoma has been well demonstrated and H. pylori eradication with antibiotics has emerged as the standard therapy for Stage I H. pylori positive (HPP) gastric MALT lymphoma. As per the NCCN Guidelines, radiation therapy is the preferred treatment option for early stage H. pylori negative (HPN) gastric MALT lymphoma and antibiotic therapy is not indicated. However, successful treatment of HPN early stage gastric MALT lymphoma with antibiotics was reported in small series of patients by Raderer et al, Gut, 2006 [(5/6 patients achieved complete remission (CR)] and Asano et al, Tohoku J Exp Med, 2012 (5/17 patients achieved CR). Here, we report the outcome of Stage I HPP and HPN gastric MALT lymphoma patients treated with antibiotics at MD Anderson Cancer Center, Houston, Texas, USA over a period of 20 years. Methods We reviewed medical records of all pathologically proven gastric MALT lymphoma patients (n=128) referred to MD Anderson Cancer Center at initial diagnosis between 1991 and 2011. Only patients with Stage IAE disease were considered for this analysis. Patients with large cell transformation were excluded. H. pylori status was determined by histopathology and serum antibody assay. Clinical staging was determined by upper GI endoscopy with biopsy, bone marrow biopsy, and CT scans of neck, chest, abdomen, and pelvis and/or PET-CT scan. Response was assessed by upper GI endoscopy every 3-6 months until complete resolution of lymphoma. Complete remission was defined as the absence of histopathologic evidence of lymphoma on endoscopic biopsies. Results Of the 128 patients reviewed, 81 patients had Stage IAE disease without histologic evidence of large cell transformation. The 81 Stage IAE patients were assigned to HPP (39/81, 48%) or HPN group (42/81, 52%) based on histopathologic evidence of H. pylori. The higher-than-expected proportion of HPN patients might be due to referral bias to a tertiary care cancer center. The results of H. pylori antibody serology are shown in Table 1. There was no significant difference in age, gender, and race between HPP and HPN groups. First-line antibiotic therapy was administered for all 39 (100%) HPP patients and 28/42 (67%) HPN patients. The CR rate after antibiotic therapy was significantly higher in HPP compared with HPN patients (22/39, 56% vs 7/28, 25%; p=0.019). The median time to achieve a CR was similar for the two groups (HPP: 7.8 mo, range 3-40 mo; HPN: 9.7 mo, range 3-35 mo; p=0.385). After a median follow-up of 110 mo for the HPP group and 91.5 mo for the HPN group, 3/22 (14%) and 2/7 (28%) responders relapsed, respectively (p=0.362). Patients who failed to achieve a CR with antibiotic therapy were mostly treated with radiation: 14/17 in the HPP group and 19/21 in the HPN group. All patients that received radiation achieved a CR. Of the patients in both groups who received upfront antibiotic therapy, there were no differences in time to progression (HPP, 90% vs HPN, 92% at 8 years; p=0.543) and overall survival (HPP, 93% vs HPN, 100% at 8 years; p=0.068). Conclusions Our results demonstrate that a substantial proportion of patients with Stage IAE HPN gastric MALT lymphoma achieve durable remission with antibiotic therapy alone. It is possible that such responses may be because of false-negative H. pylori test results or due to association of HPN gastric MALT lymphoma with other unidentified bacteria. We also observed that HPN gastric MALT lymphoma patients failing antibiotic therapy could be effectively salvaged with radiation therapy and their long-term outcome is not affected by delay from initial trial of antibiotic therapy. Thus, the results of this largest series-to-date of HPN patients treated with first-line antibiotic therapy, combined with results of smaller series reported previously, suggest that 1) antibiotic therapy should be considered as first-line therapy for Stage IAE HPN gastric MALT lymphoma patients, and 2) radiation therapy could be avoided in a subset of these patients. Table 1. H. pylori status in Stage IAE gastric MALT lymphoma as determined by histopathology and H. pylori antibody. H. pylori positive by histology (n=39) H. pylori negative by histology (n=28) CR (n=22) Non-CR(n=17) CR (n=7) Non-CR(n=21) H. pylori Ab Positive 4 4 0 3 H. pylori Ab Negative 8 2 1 7 Not available 10 11 6 11 Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

The effect of omeprazole on the development of experimental autoimmune encephalomyelitis in C57BL/6J and SJL/J mice.

BackgroundGastric disturbances such as dyspepsia are routinely encountered by multiple sclerosis (MS) patients, and these conditions are often treated with gastric acid suppressors such as proton pump inhibitors, histamine H2 receptor antagonists, or antacids. The proton pump inhibitor omeprazole can alter the gut flora and immune responses, both of which can influence the course of experimental autoimmune encephalomyelitis (EAE), an animal model of MS. The objective of the current study was to examine the effect of omeprazole treatment on the development of EAE. Bacterial microbiome analysis of mouse fecal pellets was determined in C57BL/6J EAE mice chronically treated with omeprazole, and spleen immune cell content, clinical scores, weight, rotarod latency, and histopathology were used as outcome measures in C57BL/6J and SJL/J mice with EAE.ResultsOmeprazole treatment resulted in decreases in Akkermansia muciniphila and Coprococcus sp. and an increase in unidentified bacteria in the family S24-7 (order Bacteroidales) in C57BL/6J mice with EAE. Omeprazole did not alter spleen immune cell content compared to vehicle in EAE mice, but differences independent of treatment were observed in subsets of T cells between early and advanced disease in C57BL/6J mice as well as between the two strains of mice at an advanced disease stage. Omeprazole caused no difference in clinical scores in either strain, but significantly lowered weight gain compared to vehicle in the C57BL/6J mice with EAE. Omeprazole also did not alter rotarod behavior or hindbrain inflammatory cell infiltration compared to vehicle in both strains of mice with EAE. Rotarod latency did reveal a negative correlation with clinical scores during active disease in both mouse strains, but not during clinical remission in SJL/J mice, suggesting that rotarod can detect disability not reflected in the clinical scores.ConclusionsDespite alterations in the gut microbiota and weight gain in the C57BL/6J EAE model, omeprazole had no effect on multiple measures of disease activity in C57BL/6J and SJL/J mice with EAE, supporting the notion that omeprazole does not substantially influence disease activity in MS patients.Electronic supplementary materialThe online version of this article (doi:10.1186/1756-0500-7-605) contains supplementary material, which is available to authorized users.

Microbial diversity in formation water and enrichment cultures from the Gangxi bed of the Dagang terrigenous oilfield (PRC)

Microbial diversity and biogeochemical processes of the Gangxi bed with low-mineral water and a temperature gradient from 35 to 54°C were studied. The 16S rRNA gene clone libraries (over 800 clones) were obtained from microbial DNA isolated from formation water and from the primary enrichment cultures for fermenting, sulfate-reducing, methanogenic, and aerobic organotrophic prokaryotes. While both sulfate reduction and methanogenesis were registered in formation water by radioisotope techniques, the genes of sulfate-reducing prokaryotes were not revealed in the 16S rRNA gene clone library from formation water. The 16S rRNA genes of Methanobacterium congolense and Methanococcus vannielii predominated among archaeal sequences retrieved from formation water, while the genes of Methanothermobacter thermoautotrophicus, Methanomethylovorans thermophila, and Methanoculleus sp. predominated in the combined library from enrichment cultures. In the library of Bacteria 16S rRNA genes from formation water, the genes of thermophilic fermentative bacteria of the family Thermoanaerobacteriaceae predominated; the remaining sequences belonged to mesophiles (genera Brevundimonas, Sphingomonas, Oxalicibacterium, and Stenotrophomonas), the phylum Chloroflexi, and unidentified bacteria. The combined library from enrichment cultures, contained, apart from the sequences of the family Thermoanaerobacteriaceae, the genes of fermentative bacteria (genera Anaerobaculum, Coprothermobacter, Thermanaerovibrio, Soehngenia, Bacteroides, and Aminobacterium and the order Thermotogales), of aerobic hydrocarbon-oxidizing bacteria (genera Pannonibacter and Pseudomonas), and of sulfate reducers (genera Desulfomicrobium, Thermodesulfovibrio, and Desulfotomaculum). High coverage was shown for bacterial (97.6%) and archaeal (100%) clone libraries, indicating that a significant portion of the microbial diversity in the studied communities was revealed.

Aerosolized Microbes from Organic Rich Materials: Case Study of Bat Guano from Caves in Romania

Cave air, bat guano, and swabs of bat fur from caves with bat guano in Romania were analyzed by using RIDAHCOUNT cultivation plates and standard selective media for Staphylococcus and Streptococcus. The samples of guano varied in concentration of cultivable chemoheterotrophic bacteria (max. 1.9310 10 CFU/g), coliforms (max. 2.2310 8 CFU/g), Escherichia coli (max. 1.0310 8 CFU/g), and yeasts and molds (max. 1.7310 7 CFU/g). The gravity-settling principle was applied to sample airborne microorganisms, and a new method was developed for evaluation of aerosolization potential. In cave air, the concentration of total bacteria was higher than yeast and molds. In addition to coliforms, enterobacteria, E. coli, and unidentified cultivable bacteria in the air samples, we also identified Chryseomonas luteola, Klebsiella pneumoniae, Micrococcus, Salmonella, Staphylococcus, and Streptococcus. In the experiment that provoked microbial aerosolization from guano, 3.35% of total cultivable fungi were aerosolized, up to 0.10% of bacteria, and 0.00% of E. coli. The concentration of Staphylococcus in the air exceeded counts of Streptococcus. The highest concentrations of airborne microorganisms were on the ground level. Using cultivation plates as a robust method we demonstrated that the relative proportion of microbial subgroups in the air remained constant in different seasons, with lower concentrations of airborne microbiota in the autumn. Caves as simplified natural systems demonstrated complex relationships between atmospheric parameters and microorganisms. Bats introduce into caves varying, but not negligible, concentrations of microbes on their fur. Caves with guano had relative high concentration of airborne microbes that may represent a biohazard for animals and humans.

Biogenicity and Characterization of Moonmilk in the Grotta Nera (Majella National Park, Abruzzi, Central Italy)

Observations and hypotheses on the possible influence of unidentified calcifying bacteria on moonmilk speleothem formation in the Grotta Nera are reported for the first time. The Majella Massif hosts a complex karst system of several caves; the accessible Grotta Nera is the most interesting one. Despite its name, the cave is characterized by particularly abundant ivory-white deposits of moonmilk. Two samples of moonmilk were analyzed to determine the geochemistry, fabric, depositional setting, and extent of biogenicity. For this, we combined geochemical, scanning electron microscopic, microbiological, and in vitro precipitation studies. X-ray diffraction of the moonmilk deposits gave clear evidence for the presence of calcite. Scanning electron microscopy showed that moonmilk in the Grotta Nera consists of a network of calcite fibers oriented in all directions, resembling a felted mat. The cultivation on specific medium of moonmilk and drip-water samples showed the presence of fungi, actinomycetes, and other bacteria, but the dominant cultivable microorganisms were bacteria, which produced significant crystallization. Examination of Gram-stained smears taken from the fifteen different colony types showed that the majority (66.7%) of the bacterial isolates were Gram-negative. Single small rods and rod chains were the most common bacteria isolated from the Grotta Nera. None of the molds isolated from the Grotta Nera samples were able to precipitate CaCO3 crystals, suggesting a major bacterial contribution to moonmilk deposition in the cave. Bacteria were capable of precipitating CaCO3 on B-4 solid medium at 15 (cave temperature), 22, and 32 uC. The calcifying bacteria isolated from the Grotta Nera showed a greater capability to solubilize CaCO3 than those associated with consolidated stalactites sampled from previously studied caves. The electron microscopy and microbiological evidences, together with the geochemistry and environmental data, allowed us to postulate the biogenic nature of the moonmilk in the Grotta Nera Cave.

Arsenite oxidation by the phyllosphere bacterial community associated with Wolffia australiana.

Speciation is a key determinant in the toxicity, behavior, and fate of arsenic (As) in the environment. However, little is known about the transformation of As species mediated by floating macrophytes and the phyllosphere bacteria in aquatic and wetland environment. In this study, Wolffia australiana, a rootless floating duckweed, was cultured with (W+B) or without (W-B) phyllosphere bacteria to investigate its ability in arsenite (As(III)) oxidation. Results showed that sterile W. australiana did not oxidize As(III) in the growth medium or in plant tissue, whereas W. australiana with phyllpsphere bacteria displayed substantial As(III) oxidation in the medium. Quantitative PCR of As redox-related functional genes revealed the dominance of the arsenite oxidase (aioA) gene in the phyllosphere bacterial community. These results demonstrate that the phyllosphere bacteria were responsible for the As(III) oxidation in the W+B system. The rapid oxidation of As(III) by the phyllosphere bacterial community may suppress As accumulation in plant tissues under phosphate rich conditions. The aioA gene library showed that the majority of the phyllosphere arsenite-oxidizing bacteria related either closely to unidentified bacteria found in paddy environments or distantly to known arsenite-oxidizing bacteria. Our research suggests a previously overlooked diversity of arsenite-oxidizing bacteria in the phyllosphere of aquatic macrophytes which may have a substantial impact on As biogeochemistry in water environments, warranting further exploration.

High Abundance of Escherichia During the Establishment of Fecal Microbiota in Brazilian Children

The sequence of bacterial events that occurs during the colonization of the gastrointestinal tract may affect the future health of the host. A clear understanding of the colonization process of the human neonatal gut in developing countries is lacking because the few available studies were mostly performed using culture techniques. Using molecular approaches, this study analyzed the fecal microbiota of children of low socioeconomic status in São Paulo, Brazil, during their first year of life. We collected fecal samples of healthy children at 3, 6, and 12 months of life. Total DNA was extracted directly from feces, and the bacteria-specific primers 27F-1492R were used to construct 16S rRNA libraries. Clones were randomly selected and partially sequenced. The main phylogenetic groups identified at 3 months were Streptococcus, unidentified bacteria, and Escherichia. At 6 months, Escherichia remained predominant, while the unidentified bacterial population increased significantly. At 12 months, a more complex composition of fecal microbiota was observed, represented by unidentified bacteria and microorganisms found at low rates at earlier ages. The genus Escherichia remained the most abundant microorganism (34% relative abundance and 75% prevalence). Principal component analysis (PCA) revealed changes in the composition of the microbiota at 6 months and an increase of diversity at 12 months of life. Bifidobacterium was identified by quantitative PCR (qPCR) and showed a high incidence in the microbiota at 3 months. The present results corroborate the global observation of inter-individual variability with an early establishment of microbial complexity at the end of the first year of life and highlight the presence of the Escherichia as abundant in microbiota composition of this group of children.

Bacteriological Quality of Reclaimed Wastewater Used for Irrigation of Public Parks in the United Arab Emirates

Bacteriological quality of reclaimed wastewater used for irrigation of thirty public parks located in Dubai and Sharjah was evaluated. Bacterial contaminants (Escherichia coli, Salmonella, Shigella and Enterobacter aerogenes) were tested by membrane filtration technique using Bismuth Sulphite Agar (BSA) and Violet Red Bile Agar (VRBA) media. The results showed the presence of Salmonella, Shigella, E. coli, Entereobacter aerogenes and other unidentified bacterial species on various selective media with different occurrence rates. The most common type of bacteria observed on the BSA media were Shigella (63%), E. coli (30%), Salmonella (10%) and a large percentage of unidentified bacteria (97%). Whereas on VRBA media the most common bacteria detected were E. coli (83%), unidentified bacteria (76%) followed by Enterobacter aerogenes (70%). The overall results obtained in the present study demonstrated that only 40% of the reclaimed wastewater samples examined complied with the local and widely accepted international standards of less than 1000 fecal coliforms/100mL for unrestricted irrigation. Further studies are recommended to determine the survival rate of Salmonella and other coliform group bacteria in the soil irrigated by reclaimed wastewater. In addition, detailed research should be conducted to establish the identity of Salmonella, Shigella (up to the species and sub-species level) and other unidentified bacteria which were detected in most of the reclaimed water samples.

Outbreak Investigation of Cholera in a Slum of Northern India

Cholera is one of the oldest and best understood endemic diseases. An actual bacterial enteric disease, it is characterized in its severe form by sudden onset, profuse painless watery stools (rice-water stool), nausea and profuse vomiting early in the course of illness. Endemic and pandemics are strongly linked to the consumption of unsafe water, poor hygiene, poor sanitation and crowded living conditions. A rapid survey was conducted for the outbreak investigation on August 4, 2008. The objectives were to investigate the outbreak, risk factors for cholera and recommend control measures immediately. Questionnaire based random convenient sample based investigative study. 60 families were contacted by the public health students. Approximate 300 individuals were screened for cholera cases. Simultaneously six water samples were also collected from the contacted family. Microbiological test for Vibrio cholera and E. coli was conducted. Randomly six water samples were collected from the Rajive colony. Microbiological test for Vibrio cholera and E. coli was conducted from the department of Microbiology, PU. Vibrio cholera and E. coli were found absent, in all the samples. One sample was found positive for unidentified bacteria.

Bacterial structure and spatiotemporal distribution in a horizontal subsurface flow constructed wetland

In this study, bacterial community structure in a horizontal subsurface flow constructed wetland (HSF-CW) planted with Phragmites australis was investigated using the 16S rRNA cloning-sequencing technique. Two layer depths were considered: the rhizosphere zone (RH) and the deep-layer zone (DL) in different sampling periods. Bacteria-specific primers 008F and 1492R were used to amplify the 16S rRNA genes and construct six clone libraries. A total of 1,284 individual sequences were used to assess the HSF-CW diversity. Phylogenetic analysis of RH and DL clone libraries shows that 41.57 and 42.17 % of the 16S rRNA sequences are affiliated with the Proteobacteria in the RH and the DL, respectively. The remaining major phylogenetic groups are Bacteroidetes, Planctomycetes, and Chloroflexi with 11.78, 9.36, and 7.6 %, respectively, in the RH and 11.38, 6.48, and 7.65 % in the DL, respectively. Minor divisions such as Verrucomicrobia, TM7, Nitrospira, and Gemmatimonadetes represented <6 % of the total sequences, while 14.2 % were unidentified Bacteria. Among the Proteobacteria, the Alphaproteobacteria subclass is represented in both locations, while the Deltaproteobacteria and Gammaproteobacteria subclasses were predominant in the RH and the DL, respectively. Results suggest that Archaea and Bacteria in the HSF-CW are the essential actors in the nitrogen cycle and that the established microbial community is efficient in nitrogen removal from wastewater.

Enrichment of a microbial consortium capable of reductive deiodination of 2,4,6-triiodophenol

An anaerobic microbial consortium capable of reductively dehalogenating 2,4,6-triiodophenol (2,4,6-TIP) to 4-iodophenol was enriched from sediments collected from an iodine-producing industry, Chiba, Japan. In the presence of lactate, the enrichment reductively deiodinated 2,4,6-TIP, 2,4-diiodophenol and 2-iodophenol, suggesting preferential removal of ortho-substituted iodines. PCR-denaturing gradient gel electrophoresis and 16S rRNA gene sequence analysis showed that at least 4 bacteria, including Pseudomonas stutzeri, Clostridium sp., Sedimentibacter sp., and unidentified Chloroflexi bacterium, were predominant in the enrichment. Interestingly, the DGGE band corresponding to the Chloroflexi bacterium disappeared when the enrichment was grown in the absence of 2,4,6-triiodophenol. In addition, the DGGE band with a nearly identical gene sequence was detected in another enrichment that consumed 3-amino-2,4,6-triiodobenzoic acid (ATBA). Phylogenetic analysis of 1416 bp of 16S rRNA gene sequence for this putative deiodinating bacterium revealed that it was closely related (93% sequence similarity) with an anaerobic bacterium MO-CFX2 belonging to the class Anaerolineae, which was recently isolated from subseafloor sediments. The sequence similarities with other known reductive dehalogenating bacteria such as Dehalococcoides mccartyi and Dehalobium chlorocoercia were relatively low (78%-80%). Quantitative PCR analysis targeting specific 16S rRNA gene region of the putative deiodinating bacterium showed that the enrichments consuming 2,4,6-TIP or ATBA contained 18 to 1070 times much higher amounts of 16S rRNA gene copies than those in the enrichments that do not consume these iodoaromatic compounds. These results suggest that a novel anaerobic bacterium in the class Anaerolineae is capable of reductively deiodinating 2,4,6-iodobenzene derivatives.

Preliminary Analysis of Life within a Former Subglacial Lake Sediment in Antarctica

Since the first descriptions of Antarctic subglacial lakes, there has been a growing interest and awareness of the possibility that life will exist and potentially thrive in these unique and little known environments. The unusual combination of selection pressures, and isolation from the rest of the biosphere, might have led to novel adaptations and physiology not seen before, or indeed to the potential discovery of relic populations that may have become extinct elsewhere. Here we report the first microbiological analysis of a sample taken from a former subglacial lake sediment in Antarctica (Lake Hodgson, on the Antarctic Peninsula). This is one of a number of subglacial lakes just emerging at the margins of the Antarctic ice sheet due to the renewed onset of deglaciation. Microbial diversity was divided into 23.8% Actinobacteria, 21.6% Proteobacteria, 20.2% Planctomycetes and 11.6% Chloroflexi, characteristic of a range of habitat types ( Overall, common sequences were neither distinctly polar, low temperature, freshwater nor marine). Twenty three percent of this diversity could only be identified to “unidentified bacterium”. Clearly these are diverse ecosystems with enormous potential.

Effects of a dietary β-(1,3)(1,6)-D-glucan supplementation on intestinal microbial communities and intestinal ultrastructure of mirror carp (Cyprinus carpio L.)

To assess the effects of dietary Saccharomyces cerevisiae β-(1,3)(1,6)-D-glucan supplementation (MacroGard(®)) on mirror carp (Cyprinus carpio L.) intestinal microbiota and ultrastructure of the enterocyte apical brush border. Carp were fed either a control diet or diets supplemented with 0.1, 1 or 2% w/w MacroGard(®). Culture-dependent microbiology revealed that aerobic heterotrophic bacterial levels were unaffected by dietary MacroGard(®) after 2 and 4 weeks. No effects were observed on the allochthonous lactic acid bacteria (LAB) populations at either time point; however, reduced autochthonous LAB populations were observed at week 4. PCR-DGGE confirmed these findings through a reduction in the abundance of autochthonous Lactococcus sp. and Vagococcus sp. in MacroGard(®)--fed fish compared with the control-fed fish. Overall, sequence analysis detected microbiota belonging to the phyla Proteobacteria, Firmicutes, Fusobacteria and unidentified uncultured bacteria. DGGE analyses also revealed that dietary MacroGard(®) reduced the number of observed taxonomical units (OTUs) and the species richness of the allochthonous microbiota after 2 weeks, but not after 4 weeks. In contrast, dietary MacroGard(®) reduced the number of OTUs, the species richness and diversity of the autochthonous microbiota after 2 weeks, and those parameters remained reduced after 4 weeks. Transmission electron microscopy revealed that intestinal microvilli length and density were significantly increased after 4 weeks in fish fed diets supplemented with 1% MacroGard(®). This study indicates that dietary MacroGard(®) supplementation modulates intestinal microbial communities of mirror carp and influences the morphology of the apical brush border. To the authors' knowledge, this is the first study to investigate the effects of β-(1,3)(1,6)-D-glucans on fish gut microbial communities, using culture-independent methods, and the ultrastructure of the apical brush border of the enterocytes in fish. This prebiotic-type effect may help to explain the mechanisms in which β-glucans provide benefits when fed to fish.

Isolation and Enrichment of Pathogens with a Surface‐Modified Aluminium Chip for Raman Spectroscopic Applications

We developed a Raman-compatible chip for isolating microorganisms from complex media. The isolation of bacteria is achieved by using antibodies as capture molecules. Due to the very specific interaction with the targets, this approach is promising for isolation of bacteria even from complex matrices such as body fluids. Our choice of capture molecules also enabled the investigation of samples containing yet unidentified bacteria, as the antibodies can capture a large variety of bacteria based on their analogue cell wall surface structures. The capability of our system is demonstrated for a broad range of different Gram-positive and Gram-negative germs. Subsequent identification is done by recording Raman spectra of the bacteria. Further, it is shown that classification with chemometric methods is possible.

한국산 꽃소금과 천일염의 이화학적 특성 및 미생물 분석

한국산 꽃소금의 특성을 알아보기 위하여 국내에서 생산된 꽃소금 3종과 천일염 1종의 이화학적 및 미생물 분석을 실시하였다. 수분함량은 10.54±0.10∼13.82±0.12%, 나머지 조단백, 조지방, 조섬유는 거의 존재하지 않았다. NaCl은 신의도산 꽃소금이 78.81±0.28%, 비금산 꽃소금이 81.67±0.34%, 도초산 꽃소금 84.61±0.21%, 도초산 천일염이 80.82±0.17%로 나타났다. 불용분, 사분은 각각 0.01∼0.05%, 0.01∼0.03%로 낮게 검출되었다. 미네랄 분석은 도초산 꽃소금에서 K과 Mg 함량이 2,975.23 mg/kg, 9,886.72 mg/kg으로 비교적 낮게 나타났다. Ca은 도초산 소금 2종이 945.53 mg/kg, 942.43 mg/kg으로 낮은 함량을 보였다. 중금속 As, Cd, Pb, Hg은 모두 규격 이하로 검출되었다. 소금결정을 관찰한 결과로 천일염과 꽃소금 모두 핵이 중복되어 겹으로 적층된 것을 확인하였으며 크기는 비금산 꽃소금이 0.067×0.067 mm로 가장 작고 도초산 천일염이 0.112×0.124 mm로 꽃소금에 비해 크게 나타났다. 소금의 색도는 꽃소금의 L값이 천일염에 비해 높아 밝게 확인되었다. 관능검사 결과로 짠맛은 NaCl 함량과 유사하며, 쓴맛은 K과 Mg 함량이 적은 도초산 꽃소금이 낮게 나타났다. 천일염에 비해 꽃소금의 단맛과 기호도가 더 높은 것으로 확인되었다. 호염균 동정 결과 19종 모두 Firmicutes로 꽃소금에서 Marinibacillus, Paenibacillus, Bacillus 속이 12종으로 확인되며, 천일염은 Planococcus, Staphylococcus, Bacillus 속 3종으로 검출되었다. DGGE 실험 결과로 도초산 꽃소금에서 16개의 band와 도초산 천일염에서 15개의 band를 확인하였다. 동정 결과 도초산 꽃소금에서 Cupriavidus sp. ATHA3(14.43%), Cupriavidus sp. TSA5(10.41%), Maritimibacter sp. YCSD61-2(8.13%), uncultured bacteria(68%)로 확인되었고, 도초산 천일염에서는 Dunaliella salina(4.76%), Cupriavidus sp. ATHA3(15.80%), uncultured Mycoplasmataceae bacteria(51.72%), uncultured bacteria(27%)로 확인되었다.

Novel bacterial phylotypes associated with the healthy feline oral cavity and feline chronic gingivostomatitis

Feline chronic gingivostomatitis (FCGS) is a painful inflammatory disease of the oral cavity. Treatment options for FCGS are very limited and little is known regarding its aetiology. The aim of this study was to investigate the presence of putative novel species in the oral cavity of cats with and without FCGS. Bacterial DNA was extracted from oral swabs and identified by 16S rRNA gene sequencing. The 16S rRNA genes of 54 clones representing distinct potentially novel species were sequenced (1202–1325 base pairs). Obtained sequences were compared to the BLAST database, aligned using the ClustalW2 alignment tool and a phylogenetic tree created. Twenty-two clones (18 from control and four from FCGS samples) had a similarity of less than 97% and were considered novel. The proportion of novel phylotypes in each group was 19.6% (control) and 2.3% (FCGS). In the derived phylogenetic tree, 15 novel phylotypes clustered together and branched away from known species and phyla. This suggests the presence of a group of novel, previously unidentified bacteria that are associated with the feline oral cavity in both health and disease.

The intestinal microbial diversity in mud crab (Scylla paramamosain ) as determined by PCR-DGGE and clone library analysis

To identify the intestinal microbial diversity in mud crab and to investigate the bacterial difference in the intestinal microbiology between wild crabs (WC), pond-raised healthy and diseased crabs (DC). The intestinal microbial community of mud crab Scylla paramamosain from WC, pond-raised healthy crabs (HC) and DC were examined by Denaturing Gradient Gel Electrophoresis (DGGE) and clone library analysis of 16S rDNA gene. Eight of 21 representative DGGE bands were affiliated with unidentified or unclassified bacteria. Vibrio, Pseudoalteromonas and Shewanella were also found from the DGGE gel. Analysis of clone libraries revealed that all sequenced clones were grouped into either of the following phyla: Proteobacteria, Firmicutes, Tenericutes, Bacteroidetes, Fusobacteria, Cyanobacteria and unidentified or unclassified bacteria. The phylotypes affiliated with Firmicutes were not found in DC library, yet DC had a little portion of Cyanobacteria which did not exist in both WC and HC library. Real-time PCR showed that the abundance of the total bacterial load in WC were significantly three times higher than that in healthy and DC, the abundance of Bacteriodes in healthy and WC were as much four times, three times as that in DC, respectively. Statistical analysis showed that the bacterial communities in intestine of the mud crab from these three populations were significantly different. The phylotypes of the Bacteriodes and Tenericutes were the dominant population in the gut of the mud crab. This study demonstrated significant differences in the intestinal bacterial composition of three crab populations. This knowledge will increase our understanding of the effect of growth conditions on bacterial community composition in the crab gut and provide necessary data for further development of probiotic products for diseases prevention in crab farming.

Isolation and identification of mycobacteria from captive reptiles

The occurrence of Mycobacterium species in clinically healthy pet reptiles was studied in Italy during the period 2004–2006. The feces samples of 223 animals were examined bacteriologically. Thirty-seven strains were isolated, in particular from 13/18 (72.2%) ophidians, 13/134 (9.7%) saurians and 11/71 (15.5%) chelonians. The isolates were classified, after HPLC analysis of bromophenacyl esters of cell wall mycolic acids, as Mycobacterium fortuitum (14 strains, 37.8%), Mycobacterium fortuitum-like (17, 45.9%), Mycobacterium peregrinum (4, 10.8%), and Mycobacterium chelonae (1, 2.7%). M. fortuitum was isolated from seven pythons, five saurians and two turtles; M. fortuitum-like from six saurians, six pythons and five turtles; M. peregrinum from four turtles; M. chelonae from one lizard. One isolate from an Iguana iguana could not be identified by HPLC analysis showing a previously unreported profile. Comparative 16S rDNA sequencing showed a low similarity with Mycobacterium triviale (97.2%) and Mycobacterium confluentis (97.1%). On the basis of such data the unidentified bacterium turned out to belong to a not yet described Mycobacterium species.