In Vitro Assessment of Prebiotic Activity.

Bifidogenic effect is a main target for the assessment of prebiotic activity. pH-controlled batch processes of bifidobacteria and fecal microbiota are herein presented. Growth of bifidobacteria, carbohydrate breakdown and consumption, organic acid production, and activity of specific glycosyl hydrolases involved in the hydrolysis of di-, oligo-, or polysaccharides are exploited to study and compare substrate preference of bifidobacteria for candidate prebiotics.

Assessment and characterization of biofilm formation among human isolates of Streptococcus dysgalactiae subsp. equisimilis.

The capacity to form biofilm is considered a protective mechanism that allows the bacteria to survive and proliferate in hostile environments, facilitating the maintenance of the infectious process. Recently, biofilm has become a topic of interest in the study of the human pathogen group A Streptococcus (GAS). Although GAS has not been associated with infection on medical implants, the presence of microcolonies embedded in an extracellular matrix on infected tissues has been reported. Despite the similarity between GAS and Streptococcus dysgalactiae subspecies equisimilis (SDSE), there are no studies in the literature describing the production of biofilm by SDSE. In this work, we assessed and characterized biofilm development among SDSE human isolates of group C. The in vitro data showed that 59.3% of the 118 isolates tested were able to form acid-induced biofilm on glass, and 28% formed it on polystyrene surfaces. More importantly, biofilm was also formed in a foreign body model in mice. The biofilm structure was analyzed by confocal laser scanning microscopy, transmission electron microscopy, and scanning electron microscopy. Long fibrillar-like structures were observed by scanning electron microscopy. Additionally, the expression of a pilus associated gene of SDSE was increased for in vitro sessile cells compared with planktonics, and when sessile cells were collected from biofilms formed in the animal model compared with that of in vitro model. Results obtained from the immunofluorescence microscopy indicated the biofilm was immunogenic. Our data also suggested a role for proteins, exopolysaccharide and extracellular DNA in the formation and accumulation of biofilm by SDSE.

Fouling cake layer in a submerged anaerobic membrane bioreactor treating saline wastewaters: curse or a blessing?

The treatment of inhibitory (saline) wastewaters is known to produce considerable amounts of soluble microbial products (SMPs), and this has been implicated in membrane fouling; the fate of these SMPs was of considerable interest in this work. This study also investigated the contribution of SMPs to membrane fouling of the; (a) cake layer/biofilm layer, (b) the compounds below the biofilm/cake layer and strongly attached to the surface of the membrane, (c) the compounds in the inner pores of the membrane, and (d) the membrane. It was found that the cake/biofilm layer was the main reason for fouling of the membrane. Interestingly, the bacteria attached to the cake/biofilm layer showed higher biodegradation rates compared with the bacteria in suspension. Moreover, the bacteria attached to the cake layer showed higher amounts of attached extracellular polysaccharides (EPS) compared with the bacteria in suspension, possibly due to accumulation of the released EPS from suspended biomass in the cake/biofilm layer. Molecular weight (MW) analysis of the effluent and reactor bulk showed that the cake layer can retain a large fraction of the SMPs in the reactor and prevent them from being released into the effluent. Hence, while cake layers lead to lower fluxes in submerged anaerobic membrane bioreactors (SAMBRS), and hence higher costs, they can improve the quality of the reactor effluent.

Laboratory assessment of factors affecting soil clogging of soil aquifer treatment systems.

In this study the effect of soil type, level of pre-treatment, ponding depth, temperature and sunlight on clogging of soil aquifer treatment (SAT) systems was evaluated over an eight week duration in constant temperature and glasshouse environments. Of the two soil types tested, the more permeable sand media clogged more than the loam, but still retained an order of magnitude higher absolute permeability. A 6- to 8-fold difference in hydraulic loading rates was observed between the four source water types tested (one potable water and three recycled waters), with improved water quality resulting in significantly higher infiltration. Infiltration rates for ponding depths of 30 cm and 50 cm were higher than 10 cm, although for 50 cm clogging rates were higher due to greater compaction of the clogging layer. Overall, physical clogging was more significant than other forms of clogging. Microbial clogging becomes increasingly important when the particulate concentrations in the source waters are reduced through pre-treatment and for finer textured soils due to the higher specific surface area of the media. Clogging by gas binding took place in the glasshouse but not in the lab, and mechanical clogging associated with particle rearrangement was evident in the sand media but not in the loam. These results offer insight into the soil, water quality and operating conditions needed to achieve viable SAT systems.

Assessment of potential probiotic properties of Lactobacillus spp., Lactococcus spp., and Pediococcus spp. strains isolated from kefir.

In this study, the metabolic activities (in terms of quantities of the produced lactic acid, hydrogen peroxide, and exopolysaccharides) of 8 strains of Lactobacillus spp., Lactococcus spp., and Pediococcus spp., were determined. Lactic acid levels produced by strains were 8.1 to 17.4 mg/L. The L. acidophilus Z1L strain produced the maximum amount (3.18 µg/mL) of hydrogen peroxide. The exopolysaccharides (EPS) production by the strains was ranged between 173 and 378 mg/L. The susceptibility of 7 different antibiotics against these strains was also tested. All strains were found to be sensitive to ampicillin. The tolerance of the strains to low pH, their resistance to bile salts of strains, and their abilities to autoaggregate and coaggregate with Escherichia coli ATCC 11229 were also evaluated. High EPS-producing strains showed significant autoaggregation and coaggregation ability with test bacteria (P < 0.01). A correlation also was determined between EPS production and acid-bile tolerance (P < 0.05). EPS production possibly affects or is involved in acid-bile tolerance and aggregation of Lactobacillus spp., Lactococcus spp., and Pediococcus spp. strains and supports the potential of L. acidophilus Z1L strain as new probiotic.

Quantitative analysis of neutral and acidic sugars in whole bacterial cell hydrolysates using high-performance anion-exchange liquid chromatography-electrospray ionization tandem mass spectrometry.

A procedure for analysis of a mixture of neutral and acidic sugars in bacterial whole cell hydrolysates using high-performance anion-exchange liquid chromatography-electrospray ionization tandem mass spectrometry (HPAEC-ESI-MS-MS) is described. Certain bacteria (including bacilli), grown under phosphate-limited conditions, switch from producing a teichoic acid (containing ribitol) to a teichuronic acid (characterized by glucuronic acid content). Bacterial cells were hydrolyzed with sulfuric acid to release sugar monomers. The solution was neutralized by extraction with an organic base. Hydrophobic and cationic contaminants (including amino acids) were removed using C18 and SCX columns, respectively. HPAEC is well established as a high-resolution chromatographic technique, in conjunction with a pulsed amperometric detector. Alternatively, for more selective detection, sugars (as M-H- ions) were monitored using ESI-MS. In HPAEC, the mobile phase contains sodium hydroxide and sodium acetate, which are necessary for chromatographic separation of mixtures of neutral and acidic sugars. Elimination of this high ionic content prior to entry into the ESI ion source is vital to avoid compromising sensitivity. This was accomplished using an on-line suppressor and decreasing post-column flow-rates from 1 ml to 50 microliters/min. In the selected ion monitoring mode, background (from the complex sample matrix as well as the mobile phase) was eliminated, simplifying chromatograms. Sugar identification was achieved by MS-MS using collision-induced dissociation.

Antigen detection in the diagnosis and in the prognostic assessment of bacterial pneumonias.

Sputum cultures are not helpful in the immediate management of patients with bacterial pneumonia. Sputum Gram stains may provide a presumptive identification of an etiologic agent; this procedure, however, is insensitive (approximately 50%). Consequently, during the last decade, other more sensitive and specific methods of providing a rapid etiologic diagnosis have been sought. This article discusses data on antigen detection in various body fluids by counterimmunoelectrophoresis and agglutination tests. Results from our own laboratory as well as those reported in the literature are presented. The best estimates of antigen detection rates, by the most sensitive assays, in pneumococcal pneumonia, are as follows: serum, 45%-80%; urine, 50%-64%; and sputum, 75%-100%. There is less information for Haemophilus, Klebsiella, and Pseudomonas pneumonias, but the diagnostic yield is approximately 50%-100%. Data will also be presented on the association between free and complexed antigens and morbidity and mortality in pneumococcal pneumonia. Indicators of morbidity discussed include disseminated intravascular coagulation, duration and severity of illness, and occurrence of nephritis.