Explore the vast range of titles available.

This study aimed at investigating the fecal microbiota, and the fecal and urinary metabolome of non progressor (NP) and progressor (P) patients with immunoglobulin A nephropathy (IgAN). Three groups of volunteers were included in the study: (i) sixteen IgAN NP patients; (ii) sixteen IgAN P patients; and (iii) sixteen healthy control (HC) subjects, without known diseases. Selective media were used to determine the main cultivable bacterial groups. Bacterial tag-encoded FLX-titanium amplicon pyrosequencing of the 16S rDNA and 16S rRNA was carried out to determine total and metabolically active bacteria, respectively. Biochrom 30 series amino acid analyzer and gas-chromatography mass spectrometry/solid-phase microextraction (GC-MS/SPME) analyses were mainly carried out for metabolomic analyses. As estimated by rarefaction, Chao and Shannon diversity index, the lowest microbial diversity was found in P patients. Firmicutes increased in the fecal samples of NP and, especially, P patients due to the higher percentages of some genera/species of Ruminococcaceae, Lachnospiraceae, Eubacteriaceae and Streptococcaeae. With a few exceptions, species of Clostridium, Enterococcus and Lactobacillus genera were found at the highest levels in HC. Bacteroidaceae, Porphyromonadaceae, Prevotellaceae and Rikenellaceae families differed among NP, P and HC subjects. Sutterellaceae and Enterobacteriaceae species were almost the highest in the fecal samples of NP and/or P patients. Compared to HC subjects, Bifidobacterium species decreased in the fecal samples of NP and P. As shown by multivariate statistical analyses, the levels of metabolites (free amino acids and organic volatile compounds) from fecal and urinary samples markedly differentiated NP and, especially, P patients.

Dairy biofilms as a source of contamination of milk and its products are of great concern in the dairy industry. For a reliable risk assessment, knowledge about the microbial community composition of biofilms in the milking systems of dairy farms must be improved. In this work, swab samples of milking machine biofilms of two dairy farms were investigated by a combination of culture-dependent and -independent methods. Spots in the milking system with enhanced microbial colonization were identified by quantification on selective and non-selective media. In addition, stainless steel coupons were placed into the piping system of a milking machine, removed after several milking intervals, and investigated for colonization by cultivation and culture-independently. Isolates were differentiated and identified by a combination of chemotaxonomical methods and 16S rRNA sequencing. The culture-independent approach involved treatment of the samples with the viability dye propidium monoazide prior to direct DNA-extraction by enzymatic cell lysis and cloning to exclude bias from dead biomass. The milking equipment retainers and the outlet of the milk bulk tank were identified as highly colonized spots on both farms. A high bacterial diversity was detected covering the phyla Actinobacteria, Bacteroidetes, Firmicutes, and Proteobacteria. Presence of biofilms was demonstrated on several materials including stainless steel and plastic, which are frequently used in milking machines, but also in dairy processing plants. Growth of mainly Gram-positive bacteria with high percentages of the phylum Actinobacteria was detected on the stainless steel coupons after exposition in the milking system for two to three days. Knowledge about the heterogenic microbial load on different parts of the milking machines and the stainless steel coupons will help to identify primary colonizers of the milking system, to assess the risk potential of biofilms for raw milk, to improve sanitation processes and to identify parts of the milking machine, which should be improved by hygienic design.

Microbial anti-cancer enzymes have been proven to be effective and economical agents for cancer treatment. Aeromonas veronii has been identified as a microorganism with the potential to produce L-glutaminase, an anticancer agent effective against acute lymphocytic leukaemia. In this study, a selective medium of Aeromonas veronii was used to culture the microorganism. Strain improvement was done by adaptive and induced mutational techniques. A selective minimal agar media was incorporated for the growth of the strain which further supports adaptive mutation. Strains were also UV-irradiated and successively treated with N-methyl-N'-nitro-N-nitrosoguanidine to find a resilient strain capable of producing L-glutaminase efficiently. The Plackett-Burman design and central composite designs were used to screen and optimize additional carbon and nitrogen sources. Adaptive mutation resulted in promising yield improvements compared to native strain (P<0.001). The mean yield of 30 treated colonies from the induced mutation was significantly increased compared to the non-induced strain (P< 0.001). The economically feasible statistical designs were found to reinforce each other in order to maximize the yield of the enzyme. The interactions of nutrient factors were understood from the 3D response surface plots. The model was found to be a perfect fit in terms of maximizing enzyme yield, with the productivity improving at every stage to a fourfold output of enzyme (591.11 ±7.97 IU/mL) compared to the native strain (135±3.51 IU/mL).

The recent literature suggests that chronic wound biofilms often consist of multiple bacterial species. However, without appropriate in vivo, polybacterial biofilm models, our understanding of these complex infections remains limited. We evaluate and compare the effect of single- and mixed-species biofilm infections on host wound healing dynamics using a quantitative, in vivo, rabbit ear model. Six-mm dermal punch wounds in New Zealand rabbit ears were inoculated with Staphylococcus aureus strain UAMS-1, Pseudomonas aeruginosa strain PAO1, or both, totaling 10/6 colony-forming units/wound. Bacterial proliferation and maintenance in vivo were done using procedures from our previously published model. Wounds were harvested for histological measurement of wound healing, viable bacterial counts using selective media, or inflammatory cytokine (IL-1β, TNF-α) expression via quantitative reverse-transcription PCR. Biofilm structure was studied using scanning electron microscopy (SEM). For comparison, biofilm deficient mutant UAMS-929 replaced strain UAMS-1 in some mixed-species infections. Bacterial counts verified the presence of both strains UAMS-1 and PAO1 in polybacterial wounds. Over time, strain PAO1 became predominant (p<0.001). SEM showed colocalization of both species within an extracellular matrix at multiple time-points. Compared to each monospecies infection, polybacterial biofilms impaired all wound healing parameters (p<0.01), and increased expression of IL-1β and TNF-α (p<0.05). In contrast, mixed-species infections using biofilm-deficient mutant UAMS-929 instead of wild-type strain UAMS-1 showed less wound impairment (p<0.01) with decreased host cytokine expression (p<0.01), despite a bacterial burden and distribution comparable to that of mixed-wild-type wounds. This study reveals that mixed-species biofilms have a greater impact on wound healing dynamics than their monospecies counterparts. The increased virulence of polybacterial biofilm appears dependent on the combined pathogenicity of each species, verified using a mutant strain. These data suggest that individual bacterial species can interact synergistically within a single biofilm structure.

Campylobacter, Arcobacter, and Helicobacter species have been isolated from many vertebrate hosts, including birds, mammals, and reptiles. Multiple studies have focused on the prevalence of these Epsilonproteobacteria genera in avian and mammalian species. However, little focus has been given to the presence within reptiles, and their potential zoonotic and pathogenic roles. In this study, occurrence, diversity, and host association of intestinal Epsilonproteobacteria were determined for a large variety of reptiles. From 2011 to 2013, 444 cloacal swabs and fecal samples originating from 417 predominantly captive-held reptiles were screened for Epsilonproteobacteria. Campylobacter, Arcobacter, and Helicobacter genus specific PCRs were performed directly on all samples. All samples were also cultured on selective media and screened for the presence of Epsilonproteobacteria. Using a tiered approach of AFLP, atpA, and 16S rRNA sequencing, 432 Epsilonproteobacteria isolates were characterized at the species level. Based on PCR, Campylobacter, Arcobacter, and Helicobacter were detected in 69.3% of the reptiles; 82.5% of the chelonians, 63.8% of the lizards, and 58.0% of the snakes were positive for one or more of these genera. Epsilonproteobacteria were isolated from 22.1% of the reptiles and were isolated most frequently from chelonians (37.0%), followed by lizards (19.6%) and snakes (3.0%). The most commonly isolated taxa were Arcobacter butzleri, Arcobacter skirrowii, reptile-associated Campylobacter fetus subsp. testudinum, and a putative novel Campylobacter taxon. Furthermore, a clade of seven related putative novel Helicobacter taxa was isolated from lizards and chelonians. This study shows that reptiles carry various intestinal Epsilonproteobacteria taxa, including several putative novel taxa.

Background and aims The avocado-producing area of southern Spain includes conventional orchards and organic orchards that use different organic amendments. To gain insight into the effects of these amendments, physicochemical properties and microbial communities of the soil were analysed in a representative set of commercial and experimental orchards. Methods The population size of several groups of culturable microorganisms was determined by plating on different selective media. Bacterial community structure was studied by denaturing gradient gel electrophoresis (DGGE) Results Commercial composts showed the largest effects, especially the animal compost, enhancing the population sizes of some microbial groups and affecting bacterial community structure in superficial and deep soil layers. Moreover, animal and vegetal compost, manure and blood meal addition are related to high bacterial diversity in the superficial soil layer. Conclusions All of the organic amendments used in this study affect soil properties in one or more of the characteristics that were analysed. Culturable microbial population data revealed the most evident effects of some of the organic treatments. However, molecular analysis of soil bacterial communities by DGGE allowed the detection of the influence of all of the analysed amendments on bacterial community composition. This effect was stronger in the superficial layer of the avocado soil.

Diverse species of lactic acid bacteria (LAB) are involved in the spontaneous fermentation of lait caillé, a traditional milk product from Burkina Faso. However, there is little information on the identity of the non-LAB occurring in the fermentation. The objective of this study was to provide information on the identity of non-LAB originating from traditional lait caillé and capable of growing abundantly on standard media used for dairy LAB isolation. Man Rogosa Sharp (MRS) and M17 agar plates previously inoculated with traditional lait caillé were used to screen for gram-negative (catalase-positive or -negative) and gram-positive catalase-positive bacteria. Pure colonies were selected, then the penta-GTG oligonucleotide (GTG)5-based rep-PCR fingerprinting followed by cluster analysis and sequencing of the 16S rRNA gene was performed to identify the isolates. These data will provide key information on the presence of bacteria associated with emerging public health risks. In addition, they may provide insights for a broader investigation of non-LAB occurring in the fermentation of unpasteurized milk and for selective media development.

Culturing is an indispensable technique in microbiological research, and culturing with selective media has played a crucial role in the detection of pathogenic microorganisms and the isolation of commercially useful microorganisms from environmental samples. Although numerous selective media have been developed in empirical studies, unintended microorganisms often grow on such media probably due to the enormous numbers of microorganisms in the environment. Here, we present a novel strategy for designing highly selective media based on two selective agents, a carbon source and antimicrobials. We named our strategy SMART for highly Selective Medium-design Algorithm Restricted by Two constraints. To test whether the SMART method is applicable to a wide range of microorganisms, we developed selective media for Burkholderia glumae, Acidovorax avenae, Pectobacterium carotovorum, Ralstonia solanacearum, and Xanthomonas campestris. The series of media developed by SMART specifically allowed growth of the targeted bacteria. Because these selective media exhibited high specificity for growth of the target bacteria compared to established selective media, we applied three notable detection technologies: paper-based, flow cytometry-based, and color change-based detection systems for target bacteria species. SMART facilitates not only the development of novel techniques for detecting specific bacteria, but also our understanding of the ecology and epidemiology of the targeted bacteria.

The dynamics related to the loss of stx genes from Shiga toxin-producing Escherichia coli remain unclear. Current diagnostic procedures have shortcomings in the detection and identification of STEC. This is partly owing to the fact that stx genes may be lost during an infection or in the laboratory. The aim of the present study was to provide new insight into in vivo and in vitro stx loss in order to improve diagnostic procedures. Results from the study support the theory that loss of stx is a strain-related phenomenon and not induced by patient factors. It was observed that one strain could lose stx both in vivo and in vitro. Whole genome comparison of stx-positive and stx-negative isolates from the same patient revealed that different genomic rearrangements, such as complete or partial loss of the parent prophage, may be factors in the loss of stx. Of diagnostic interest, it was shown that patients can be co-infected with different E. coli pathotypes. Therefore, identification of eae-positive, but stx-negative isolates should not be interpreted as “Shiga toxin-lost” E. coli without further testing. Growth and recovery of STEC were supported by different selective agar media for different strains, arguing for inclusion of several media in STEC diagnostics.

Background Immunomagnetic separation (IMS) and immunoassays are widely used for pathogen detection. However, novel technology platforms with highly selective antibodies are essential to improve detection sensitivity, specificity and performance. In this study, monoclonal antibodies (MAbs) against Internalin A (InlA) and p30 were generated and used on paramagnetic beads of varying diameters for concentration, as well as on fiber-optic sensor for detection. Results Anti-InlA MAb-2D12 (IgG2a subclass) was specific for Listeria monocytogenes and L. ivanovii , and p30-specific MAb-3F8 (IgM) was specific for the genus Listeria . At all bacterial concentrations (10 3 –10 8  CFU/mL) tested in the IMS assay; the 1-μm diameter MyOne beads had significantly higher capture efficiency ( P  < 0.05) than the 2.8-μm diameter M-280 beads with both antibodies. The highest capture efficiency for MyOne-2D12 (49.2% for 10 5  CFU/mL) was significantly higher ( P  < 0.05) than that of MyOne-3F8 (16.6 %) and Dynabeads anti- Listeria antibody (9  % ). Furthermore, capture efficiency for MyOne-2D12 was highly specific for L. monocytogenes and L. ivanovii . Subsequently, we captured L. monocytogenes by MyOne-2D12 and MyOne-3F8 from hotdogs inoculated with mono- or co-cultures of L. monocytogenes and L. innocua (10–40 CFU/g), enriched for 18 h and detected by fiber-optic sensor and confirmed by plating, light-scattering, and qPCR assays. The detection limit for L. monocytogenes and L. ivanovii by the fiber-optic immunosensor was 3 × 10 2  CFU/mL using MAb-2D12 as capture and reporter antibody. Selective media plating, light-scattering, and qPCR assays confirmed the IMS and fiber-optic results. Conclusions IMS coupled with a fiber-optic sensor using anti-InlA MAb is highly specific for L. monocytogenes and L. ivanovii and enabled detection of these pathogens at low levels from buffer or food.