BMC Microbiology

Journal: BMC Microbiology

Abbreviation

BMC microbiol. (Online)

Publisher

BioMed Central

ISSN

1471-2180

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Publications1 - 10 of 30

Mixed infections with Chlamydia and porcine epidemic diarrhea virus - a new in vitro model of chlamydial persistence
Borel, Nicole; Dumrese, Claudia; Ziegler, Urs; et al. (2010)
BMC Microbiology
Background Chlamydiae induce persistent infections, which have been associated with a wide range of chronic diseases in humans and animals. Mixed infections with Chlamydia and porcine epidemic diarrhea virus (PEDV) may result in generation of persistent chlamydial infections. To test this hypothesis, an in vitro model of dual infection with cell culture-adapted PEDV and Chlamydia abortus or Chlamydia pecorum in Vero cells was established. Results Infected cultures were investigated by immunofluorescence (IF), transmission electron microscopy (TEM) and re-infection experiments. By IF, Chlamydia-infected cells showed normal inclusions after 39 hpi. Dual infections with Chlamydia abortus revealed a heterogenous mix of inclusion types including small inclusions consisting of aberrant bodies (ABs), medium-sized inclusions consisting of ABs and reticulate bodies and normal inclusions. Only aberrant inclusions were observable in dual infection experiments with Chlamydia pecorum and PEDV. TEM examinations of mixed infections with Chlamydia abortus and Chlamydia pecorum revealed aberrant chlamydial inclusions containing reticulate-like, pleomorphic ABs, which were up to 2 μm in diameter. No re-differentiation into elementary bodies (EBs) was detected. In re-infection experiments, co-infected cells produced fewer EBs than monoinfected cells. Conclusions In the present study we confirm that PEDV co-infection alters the developmental cycle of member species of the family Chlamydiaceae, in a similar manner to other well-described persistence induction methods. Interestingly, this effect appears to be partially species-specific as Chlamydia pecorum appears more sensitive to PEDV co-infection than Chlamydia abortus, as evidenced by TEM and IF observations of a homogenous population of aberrant inclusions in PEDV - Chlamydia pecorum co-infections.
Influence of growth stage on activities of polyhydroxyalkanoate (PHA) polymerase and PHA depolymerase in Pseudomonas putida U
Ren, Qun; de Roo, Guy; Witholt, Bernard; et al. (2010)
BMC Microbiology
Background Medium chain length (mcl-) polyhydroxyalkanoates (PHA) are synthesized by many bacteria in the cytoplasm as storage compounds for energy and carbon. The key enzymes for PHA metabolism are PHA polymerase (PhaC) and depolymerase (PhaZ). Little is known of how mcl-PHA accumulation and degradation are controlled. It has been suggested that overall PHA metabolism is regulated by the β-oxidation pathway of which the flux is governed by intracellular ratios of [NADH]/[NAD] and [acetyl-CoA]/[CoA]. Another level of control could relate to modulation of the activities of PhaC and PhaZ. In order to investigate the latter, assays for in vitro activity measurements of PhaC and PhaZ in crude cell extracts are necessary. Results Two in vitro assays were developed which allow the measurement of PhaC and PhaZ activities in crude cell extracts of Pseudomonas putida U. Using the assays, it was demonstrated that the activity of PhaC decreased 5-fold upon exponential growth on nitrogen limited medium and octanoate. In contrast, the activity of PhaZ increased only 1.5-fold during growth. One reason for the changes in the enzymatic activity of PhaC and PhaZ could relate to a change in interaction with the phasin surface proteins on the PHA granule. SDS-PAGE analysis of isolated PHA granules demonstrated that during growth, the ratio of [phasins]/[PHA] decreased. In addition, it was found that after eliminating phasins (PhaF and PhaI) from the granules PhaC activity decreased further. Conclusion Using the assays developed in this study, we followed the enzymatic activities of PhaC and PhaZ during growth and correlated them to the amount of phasins on the PHA granules. It was found that in P. putida PhaC and PhaZ are concomitantly active, resulting in parallel synthesis and degradation of PHA. Moreover PhaC activity was found to be decreased, whereas PhaZ activity increased during growth. Availability of phasins on PHA granules affected the activity of PhaC.
The role of nitric-oxide-synthase-derived nitric oxide in multicellular traits of Bacillus subtilis 3610: biofilm formation, swarming, and dispersal
Schreiber, Frank; Beutler, Martin; Enning, Dennis; et al. (2011)
BMC Microbiology
Background Bacillus subtilis 3610 displays multicellular traits as it forms structurally complex biofilms and swarms on solid surfaces. In addition, B. subtilis encodes and expresses nitric oxide synthase (NOS), an enzyme that is known to enable NO-mediated intercellular signalling in multicellular eukaryotes. In this study, we tested the hypothesis that NOS-derived NO is involved in the coordination of multicellularity in B. subtilis 3610. Results We show that B. subtilis 3610 produces intracellular NO via NOS activity by combining Confocal Laser Scanning Microscopy with the NO sensitive dye copper fluorescein (CuFL). We further investigated the influence of NOS-derived NO and exogenously supplied NO on the formation of biofilms, swarming motility and biofilm dispersal. These experiments showed that neither the suppression of NO formation with specific NOS inhibitors, NO scavengers or deletion of the nos gene, nor the exogenous addition of NO with NO donors affected (i) biofilm development, (ii) mature biofilm structure, and (iii) swarming motility in a qualitative and quantitative manner. In contrast, the nos knock-out and wild-type cells with inhibited NOS displayed strongly enhanced biofilm dispersal. Conclusion The results suggest that biofilm formation and swarming motility in B. subtilis represent complex multicellular processes that do not employ NO signalling and are remarkably robust against interference of NO. Rather, the function of NOS-derived NO in B. subtilis might be specific for cytoprotection against oxidative stress as has been proposed earlier. The influence of NOS-derived NO on dispersal of B. subtilis from biofilms might be associated to its well-known function in coordinating the transition from oxic to anoxic conditions. Here, NOS-derived NO might be involved in fine-tuning the cellular decision-making between adaptation of the metabolism to (anoxic) conditions in the biofilm or dispersal from the biofilm.
The effects of chemical interactions and culture history on the colonization of structured habitats by competing bacterial populations
van Vliet, Simon; Hol, Felix J.H.; Weenink, Tim; et al. (2014)
BMC Microbiology
Background Bacterial habitats, such as soil and the gut, are structured at the micrometer scale. Important aspects of microbial life in such spatial ecosystems are migration and colonization. Here we explore the colonization of a structured ecosystem by two neutrally labeled strains of Escherichia coli. Using time-lapse microscopy we studied the colonization of one-dimensional arrays of habitat patches linked by connectors, which were invaded by the two E. coli strains from opposite sides. Results The two strains colonize a habitat from opposite sides by a series of traveling waves followed by an expansion front. When population waves collide, they branch into a continuing traveling wave, a reflected wave and a stationary population. When the two strains invade the landscape from opposite sides, they remain segregated in space and often one population will displace the other from most of the habitat. However, when the strains are co-cultured before entering the habitats, they colonize the habitat together and do not separate spatially. Using physically separated, but diffusionally coupled, habitats we show that colonization waves and expansion fronts interact trough diffusible molecules, and not by direct competition for space. Furthermore, we found that colonization outcome is influenced by a culture’s history, as the culture with the longest doubling time in bulk conditions tends to take over the largest fraction of the habitat. Finally, we observed that population distributions in parallel habitats located on the same device and inoculated with cells from the same overnight culture are significantly more similar to each other than to patterns in identical habitats located on different devices inoculated with cells from different overnight cultures, even tough all cultures were started from the same −80°C frozen stock. Conclusions We found that the colonization of spatially structure habitats by two interacting populations can lead to the formation of complex, but reproducible, spatiotemporal patterns. Furthermore, we showed that chemical interactions between two populations cause them to remain spatially segregated while they compete for habitat space. Finally, we observed that growth properties in bulk conditions correlate with the outcome of habitat colonization. Together, our data show the crucial roles of chemical interactions between populations and a culture’s history in determining the outcome of habitat colonization.
Effects of increasing n3:n6 ratio by replacing extruded soybeans with extruded flaxseed on dry matter intake, rumen fluid bacteria, and liver lipid metabolism in transition cows
Liu, Xiaojing; Zhang, Xinyue; He, Qiongyu; et al. (2025)
BMC Microbiology
Background The drop of dry matter intake (DMI) and rise of milk production in transitional dairy cows would mobilize reserved fat and disrupt lipid metabolism, eventually attributed to negative energy balance (NEB) and immune injury. The positive effect of n-3 polyunsaturated fatty acids (PUFA) on regulating energy metabolism and inflammation has been elucidated, however, the lack of regulatory mechanism of dairy cows deserves further investigation. In this study, 30 Holstein transition cows were divided into the control (CON) and HN3 groups based on the n-3: n-6 PUFA ratio in the diet. Results The results showed that compared to the CON group, high n-3: n-6 PUFA ratio-supplemented cows in the prepartum phase reduced the relative abundance of gram-negative bacteria in the rumen, the concentration of lipopolysaccharide in the plasma and liver also significantly decreased (P < 0.05). Transcriptomic analysis of the liver showed that the NF-κB signaling pathway significantly down-regulated and the taste transduction pathway up-regulated (P < 0.05) in the HN3 group. In the postpartum phase, a high n-3/n-6 PUFA ratio in the diet increased the relative abundance of Prevotella, Succinimonas and Treponema in the rumen, at the same time, orexins in plasma were also changed (P < 0.05). Further, the insulin resistance pathway significantly down-regulated and the taste transduction pathway up-regulated (P < 0.05) in the liver. Conclusions Overall, these results showed that a high n-3: n-6 PUFA ratio in the diet attenuates inflammatory responses in the prepartum phase and increases milk protein in the postpartum phase of transitional dairy cows. Appropriate increase in the proportion of n-3: n-6 PUFA ratio in the diet may be an effective measure to alleviate postpartum metabolic disease in dairy cows.
Phylogenetic, epidemiological and functional analyses of the Streptococcus bovis/Streptococcus equinus complex through an overarching MLST scheme
Jans, Christoph; Wouters, Tomas de; Bonfoh, Bassirou; et al. (2016)
BMC Microbiology
Background The Streptococcus bovis/Streptococcus equinus complex (SBSEC) comprises seven (sub)species classified as human and animal commensals, emerging opportunistic pathogens and food fermentative organisms. Changing taxonomy, shared habitats, natural competence and evidence for horizontal gene transfer pose difficulties for determining their phylogeny, epidemiology and virulence mechanisms. Thus, novel phylogenetic and functional classifications are required. An SBSEC overarching multi locus sequence type (MLST) scheme targeting 10 housekeeping genes was developed, validated and combined with host-related properties of adhesion to extracellular matrix proteins (ECM), activation of the immune responses via NF-KB and survival in simulated gastric juice (SGJ). Results Commensal and pathogenic SBSEC strains (n = 74) of human, animal and food origin from Europe, Asia, America and Africa were used in the MLST scheme yielding 66 sequence types and 10 clonal complexes differentiated into distinct habitat-associated and mixed lineages. Adhesion to ECMs collagen I and mucin type II was a common characteristic (23 % of strains) followed by adhesion to fibronectin and fibrinogen (19.7 %). High adhesion abilities were found for East African dairy and human blood isolate branches whereas commensal fecal SBSEC displayed low adhesion. NF-KB activation was observed for a limited number of dairy and blood isolates suggesting the potential of some pathogenic strains for reduced immune activation. Strains from dairy MLST clades displayed the highest relative survival to SGJ independently of dairy adaptation markers lacS/lacZ. Conclusion Combining phylogenetic and functional analyses via SBSEC MLST enabled the clear delineation of strain clades to unravel the complexity of this bacterial group. High adhesion values shared between certain dairy and blood strains as well as the behavior of NF-KB activation are concerning for specific lineages. They highlighted the health risk among shared lineages and establish the basis to elucidate (zoonotic-) transmission, host specificity, virulence mechanisms and enhanced risk assessment as pathobionts in an overarching One Health approach.
Differential transcriptome analysis of enterohemorrhagic Escherichia coli strains reveals differences in response to plant-derived compounds
Bufe, Thorsten; Hennig, André; Klumpp, Jochen; et al. (2019)
BMC Microbiology
Background Several serious vegetable-associated outbreaks of enterohemorrhagic Escherichia coli (EHEC) infections have occurred during the last decades. In this context, vegetables have been suggested to function as secondary reservoirs for EHEC strains. Increased knowledge about the interaction of EHEC with plants including gene expression patterns in response to plant-derived compounds is required. In the current study, EHEC O157:H7 strain Sakai, EHEC O157:H− strain 3072/96, and the EHEC/enteroaggregative E. coli (EAEC) hybrid O104:H4 strain C227–11φcu were grown in lamb’s lettuce medium and in M9 minimal medium to study the differential transcriptional response of these strains to plant-derived compounds with RNA-Seq technology. Results Many genes involved in carbohydrate degradation and peptide utilization were similarly upregulated in all three strains, suggesting that the lamb’s lettuce medium provides sufficient nutrients for proliferation. In particular, the genes galET and rbsAC involved in galactose metabolism and D-ribose catabolism, respectively, were uniformly upregulated in the investigated strains. The most prominent differences in shared genome transcript levels were observed for genes involved in the expression of flagella. Transcripts of all three classes of the flagellar hierarchy were highly abundant in strain C227–11φcu. Strain Sakai expressed only genes encoding the basal flagellar structure. In addition, both strains showed increased motility in presence of lamb’s lettuce extract. Moreover, strain 3072/96 showed increased transcription activity for genes encoding the type III secretion system (T3SS) including effectors, and was identified as a powerful biofilm-producer in M9 minimal medium. Conclusion The current study provides clear evidence that EHEC and EHEC/EAEC strains are able to adjust their gene expression patterns towards metabolization of plant-derived compounds, demonstrating that they may proliferate well in a plant-associated environment. Moreover, we propose that flagella and other surface structures play a fundamental role in the interaction of EHEC and EHEC/EAEC with plants.
Protective effect of probiotics on Salmonella infectivity assessed with combined in vitro gut fermentation cellular models
Zihler, Annina; Gagnon, Mélanie; Chassard, Christophe; et al. (2011)
BMC Microbiology
Background Accurate assessment of probiotics with targeted anti-Salmonella activity requires suitable models accounting for both, microbe-microbe and host-microbe interactions in gut environments. Here we report the combination of two original in vitro intestinal models closely mimicking the complex in vivo conditions of the large intestine. Effluents from continuous in vitro three-stage fermentation colonic models of Salmonella Typhimurium infection inoculated with immobilized child microbiota and Salmonella were directly applied to confluent mucus-secreting HT29-MTX cell layers. The effects of Salmonella, addition of two bacteriocinogenic strains, Bifidobacterium thermophilum RBL67 (thermophilicin B67) and Escherichia coli L1000 (microcin B17), and inulin were tested on Salmonella growth and interactions with epithelial cell layers. Salmonella adhesion and invasion were investigated and epithelial integrity assessed by transepithelial electrical resistance (TER) measurements and confocal microscopy observation. Data from complex effluents were compared with pure Salmonella cultures. Results Salmonella in effluents of all reactors of the colonic fermentation model stabilized at mean values of 5.3 ± 0.8 log10 cfu/ml effluent. Invasion of cell-associated Salmonella was up to 50-fold lower in complex reactor samples compared to pure Salmonella cultures. It further depended on environmental factors, with 0.2 ± 0.1% being measured with proximal, 0.6 ± 0.2% with transverse and 1.3 ± 0.7% with distal reactor effluents, accompanied by a similar high decrease of TER across cell monolayers (minus 45%) and disruption of tight junctions. Subsequent addition of E. coli L1000 stimulated Salmonella growth (6.4 ± 0.6 log10 cfu/ml effluent of all 3 reactors) and further decreased TER, but led to 10-fold decreased invasion efficiency when tested with distal reactor samples. In contrast, presence of B. thermophilum RBL67 revealed a protective effect on epithelial integrity compared to previous E. coli L1000 periods, as reflected by a significant mean increase of TER by 58% in all reactors. Inulin addition enhanced Salmonella growth and invasion when tested with distal and proximal reactor samples, respectively, but induced a limited decrease of TER (minus 18%) in all reactors. Conclusions Our results highlight the benefits of combining suitable cellular and colonic fermentation models to assess strain-specific first-level host protection properties of probiotics during Salmonella infection, providing an efficient system biology tool for preclinical development of new antimicrobials.
Characterisation of atypical enteropathogenic E. coli strains of clinical origin
Tennant, Sharon M.; Tauschek, Marija; Azzopardi, Kristy; et al. (2009)
BMC Microbiology
Background Enteropathogenic E. coli (EPEC) is a prominent cause of diarrhoea, and is characterised in part by its carriage of a pathogenicity island: the locus for enterocyte effacement (LEE). EPEC is divided into two subtypes according to the presence of bundle-forming pili (BFP), a fimbrial adhesin that is a virulence determinant of typical EPEC (tEPEC), but is absent from atypical EPEC (aEPEC). Because aEPEC lack BFP, their virulence has been questioned, as they may represent LEE-positive Shiga toxin-producing E. coli (STEC) that have lost the toxin-encoding prophage, or tEPEC that have lost the genes for BFP. To determine if aEPEC isolated from humans in Australia or New Zealand fall into either of these categories, we undertook phylogenetic analysis of 75 aEPEC strains, and compared them with reference strains of EPEC and STEC. We also used PCR and DNA hybridisation to determine if aEPEC carry virulence determinants that could compensate for their lack of BFP. Results The results showed that aEPEC are highly heterogeneous. Multilocus sequence typing revealed that 61 of 75 aEPEC strains did not belong to known tEPEC or STEC clades, and of those that did, none expressed an O:H serotype that is frequent in tEPEC or STEC strains associated with disease. PCR for each of 18 known virulence-associated determinants of E. coli was positive in less than 15% of strains, apart from NleB which was detected in 30%. Type I fimbriae were expressed by all aEPEC strains, and 12 strains hybridised with DNA probes prepared from either bfpA or bfpB despite being negative in the PCR for bfpA. Conclusion Our findings indicate that clinical isolates of aEPEC obtained from patients in Australia or New Zealand are not derived from tEPEC or STEC, and suggest that functional equivalents of BFP and possibly type I fimbriae may contribute to the virulence of some aEPEC strains.
Classification of a moderately oxygen-tolerant isolate from baby faeces as Bifidobacterium thermophilum
von Ah, Ueli; Mozzetti, Valeria; Lacroix, Christophe; et al. (2007)
BMC Microbiology
Background Bifidobacteria are found at varying prevalence in human microbiota and seem to play an important role in the human gastrointestinal tract (GIT). Bifidobacteria are highly adapted to the human GIT which is reflected in the genome sequence of a Bifidobacterim longum isolate. The competitiveness against other bacteria is not fully understood yet but may be related to the production of antimicrobial compounds such as bacteriocins. In a previous study, 34 Bifidobacterium isolates have been isolated from baby faeces among which six showed proteinaceous antilisterial activity against Listeria monocytogenes. In this study, one of these isolates, RBL67, was further identified and characterized. Results Bifidobacterium isolate RBL67 was classified and characterized using a polyphasic approach. RBL67 was classified as Bifidobacterium thermophilum based on phenotypic and DNA-DNA hybridization characteristics, although 16S rDNA analyses and partial gro EL sequences showed higher homology with B. thermacidophilum subsp. porcinum and B. thermacidophilum subsp. thermacidophilum, respectively. RBL67 was moderately oxygen-tolerant and was able to grow at pH 4 and at a temperature of 47°C. Conclusion In order to assign RBL67 to a species, a polyphasic approach was used. This resulted in the classification of RBL67 as a Bifidobacterium thermophilum strain. To our knowledge, this is the first report about B. thermophilum isolated from baby faeces since the B. thermophilum strains were related to ruminants and swine faeces before. B. thermophilum was previously only isolated from animal sources and was therefore suggested to be used as differential species between animal and human contamination. Our findings may disapprove this suggestion and further studies are now conducted to determine whether B. thermophilum is distributed broader in human faeces. Furthermore, the postulated differentiation between human and animal strains by growth above 45°C is no longer valid since B. thermophilum is able to grow at 47°C. In our study, 16S rDNA and partial gro EL sequence analysis were not able to clearly assign RBL67 to a species and were contradictory. Our study suggests that partial gro EL sequences may not be reliable as a single tool for species differentiation.

Publications1 - 10 of 30

Journal: BMC Microbiology

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