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Biomedical subjects

L L Blackall

Publications and source records attributed to L L Blackall.

At least 19 recordsLinked to original sources
PubMed2006

Changes in equine hindgut bacterial populations during oligofructose-induced laminitis.

In the horse, carbohydrate overload is thought to play an integral role in the onset of laminitis by drastically altering the profile of bacterial populations in the hindgut. The objectives of this study were to develop and validate microbial ecology methods to monitor changes in bacterial populations throughout the course of experimentally induced laminitis and to identify the predominant oligofructose-utilizing organisms. Laminitis was induced in five horses by administration of oligofructose. Faecal specimens were collected at 8 h intervals from 72 h before to 72 h after the administration of oligofructose. Hindgut microbiota able to utilize oligofructose were enumerated throughout the course of the experiment using habitat-simulating medium. Isolates were collected and representatives identified by 16S rRNA gene sequencing. The majority of these isolates collected belonged to the genus Streptococcus, 91% of which were identified as being most closely related to Streptococcus infantarius ssp. coli. Furthermore, S. infantarius ssp. coli was the predominant oligofructose-utilizing organism isolated before the onset of lameness. Fluorescence in situ hybridization probes developed to specifically target the isolated Streptococcus spp. demonstrated marked population increases between 8 and 16 h post oligofructose administration. This was followed by a rapid population decline which corresponded with a sharp decline in faecal pH and subsequently lameness at 24-32 h post oligofructose administration. This research suggests that streptococci within the Streptococcus bovis/equinus complex may be involved in the series of events which precede the onset of laminitis in the horse.

Animal Feed↗
PubMed2006

Fluorescence in situ hybridization and spectral imaging of coral-associated bacterial communities.

Microbial communities play important roles in the functioning of coral reef communities. However, extensive autofluorescence of coral tissues and endosymbionts limits the application of standard fluorescence in situ hybridization (FISH) techniques for the identification of the coral-associated bacterial communities. This study overcomes these limitations by combining FISH and spectral imaging.

Animals↗
PubMed2006

Applicability of experience from laboratory reactors with biological phosphorus removal in full-scale plants.

Enhanced biological phosphorus removal (EBPR) has been used at many wastewater treatment plants all over the world for many years. In this study a full-scale sludge with good EBPR was tested with P-release batch tests and combined FISH/MAR (fluorescence in situ hybridisation and microautoradiography). Proposed models of PAOs and GAOs (polyphosphate- and glycogen-accumulating organisms) and microbial methods suggested from studies of laboratory reactors were found to be applicable also on sludge from full-scale plants. Dependency of pH and the uptake of both acetate and propionate were studied and used for calculations for verifying the models and results from microbial methods. All rates found from the batch tests with acetate were higher than in the batch tests with propionate, which was explained by the finding that only those parts of the bacterial community that were able to take up acetate anaerobically were able to take up propionate anaerobically.

Acetates↗
PubMed2006

Construction and analysis of a metagenomic library from an enhanced biological phosphorus removal biomass.

The enhanced biological phosphorus removal (EBPR) process is regularly used for the treatment of wastewater, but suffers from erratic performance. Successful EBPR relies on the growth of bacteria called polyphosphate-accumulating organisms (PAOs), which store phosphorus intracellularly as polyphosphate, thus removing it from wastewater. Metabolic models have been proposed which describe the measured chemical transformations, however genetic evidence is lacking to confirm these hypotheses. The aim of this research was to generate a metagenomic library from biomass enriched in PAOs as determined by phenotypic data and fluorescence in situ hybridisation (FISH) using probes specific for the only described PAO to date, "Candidatus Accumulibacter phosphatis". DNA extraction methods were optimised and two fosmid libraries were constructed which contained 93 million base pairs of metagenomic data. Initial screening of the library for 16S rRNA genes revealed fosmids originating from a range of nonpure-cultured wastewater bacteria. The metagenomic libraries constructed will provide the ability to link phylogenetic and metabolic data for bacteria involved in nutrient removal from wastewater.

Base Sequence↗
PubMed2004

Enhanced biological phosphorus removal in a sequencing batch reactor using propionate as the sole carbon source.

An enhanced biological phosphorus removal (EBPR) system was developed in a sequencing batch reactor (SBR) using propionate as the sole carbon source. The microbial community was followed using fluorescence in situ hybridization (FISH) techniques and Candidatus 'Accumulibacter phosphatis' were quantified from the start up of the reactor until steady state. A series of SBR cycle studies was performed when 55% of the SBR biomass was Accumulibacter, a confirmed polyphosphate accumulating organism (PAO) and when Candidatus 'Competibacter phosphatis', a confirmed glycogen-accumulating organism (GAO), was essentially undetectable. These experiments evaluated two different carbon sources (propionate and acetate), and in every case, two different P-release rates were detected. The highest rate took place while there was volatile fatty acid (VFA) in the mixed liquor, and after the VFA was depleted a second P-release rate was observed. This second rate was very similar to the one detected in experiments performed without added VFA.A kinetic and stoichiometric model developed as a modification of Activated Sludge Model 2 (ASM2) including glycogen economy, was fitted to the experimental profiles. The validation and calibration of this model was carried out with the cycle study experiments performed using both VFAs. The effect of pH from 6.5 to 8.0 on anaerobic P-release and VFA-uptake and aerobic P-uptake was also studied using propionate. The optimal overall working pH was around 7.5. This is the first study of the microbial community involved in EBPR developed with propionate as a sole carbon source along with detailed process performance investigations of the propionate-utilizing PAOs.

Acetate-CoA Ligase↗
PubMed2004

The influence of substrate kinetics on the microbial community structure in granular anaerobic biomass.

The development of a strong, active granular sludge bed is necessary for optimal operation of upflow anaerobic sludge blanket reactors. The microbial and mechanical structure of the granules may have a strong influence on desirable properties such as growth rate, settling velocity and shear strength. Theories have been proposed for granule microbial structure based on the relative kinetics of substrate degradation, but contradict some observations from both modelling and microscopic studies. In this paper, the structures of four granule types were examined from full-scale UASB reactors, treating wastewater from a cannery, a slaughterhouse, and two breweries. Microbial structure was determined using fluorescence in situ hybridisation probing with 16S rRNA-directed oligonucleotide probes, and superficial structure and microbial density (volume occupied by cells and microbial debris) assessed using scanning electron microscopy (SEM), and transmission electron microscopy (TEM). The granules were also modelled using a distributed parameter biofilm model, with a previously published biochemical model structure, biofilm modelling approach, and model parameters. The model results reflected the trophic structures observed, indicating that the structures were possibly determined by kinetics. Of particular interest were results from simulations of the protein grown granules, which were predicted to have slow growth rates, low microbial density, and no trophic layers, the last two of which were reflected by microscopic observations. The primary cause of this structure, as assessed by modelling, was the particulate nature of the wastewater, and the slow rate of particulate hydrolysis, rather than the presence of proteins in the wastewater. Because solids hydrolysis was rate limiting, soluble substrate concentrations were very low (below Monod half saturation concentration), which caused low growth rates.

Abattoirs↗
PubMed2004

Identification, detection, and spatial resolution of Clostridium populations responsible for cellulose degradation in a methanogenic landfill leachate bioreactor.

An anaerobic landfill leachate bioreactor was operated with crystalline cellulose and sterile landfill leachate until a steady state was reached. Cellulose hydrolysis, acidogenesis, and methanogenesis were measured. Microorganisms attached to the cellulose surfaces were hypothesized to be the cellulose hydrolyzers. 16S rRNA gene clone libraries were prepared from this attached fraction and also from the mixed fraction (biomass associated with cellulose particles and in the planktonic phase). Both clone libraries were dominated by Firmicutes phylum sequences (100% of the attached library and 90% of the mixed library), and the majority fell into one of five lineages of the clostridia. Clone group 1 (most closely related to Clostridium stercorarium), clone group 2 (most closely related to Clostridium thermocellum), and clone group 5 (most closely related to Bacteroides cellulosolvens) comprised sequences in Clostridium group III. Clone group 3 sequences were in Clostridium group XIVa (most closely related to Clostridium sp. strain XB90). Clone group 4 sequences were affiliated with a deeply branching clostridial lineage peripherally associated with Clostridium group VI. This monophyletic group comprises a new Clostridium cluster, designated cluster VIa. Specific fluorescence in situ hybridization (FISH) probes for the five groups were designed and synthesized, and it was demonstrated in FISH experiments that bacteria targeted by the probes for clone groups 1, 2, 4, and 5 were very abundant on the surfaces of the cellulose particles and likely the key cellulolytic microorganisms in the landfill bioreactor. The FISH probe for clone group 3 targeted cells in the planktonic phase, and these organisms were hypothesized to be glucose fermenters.

Base Sequence↗
PubMed2004

Short-term effects of carbon source on the competition of polyphosphate accumulating organisms and glycogen accumulating organisms.

The effectiveness of enhanced biological phosphorus removal (EBPR) systems is directly affected by the competition of polyphosphate accumulating organisms (PAOs) and glycogen accumulating organisms (GAOs). This study investigated the short-term effects of carbon source on PAO and GAO performance. The tests were designed to clearly determine the impact of volatile fatty acid (VFA) composition on the performance of two types of biomass, one enriched for PAOs and the other for GAOs. The two populations were enriched in separate reactors using identical operating conditions and very similar influent compositions with acetate as the sole carbon source. The only difference was that a very low level of phosphorus was present in the influent to the GAO reactor. The abundance of PAOs and GAOs was quantified using fluorescence in-situ hybridisation. The results clearly show that there are some very distinctive differences between PAOs and GAOs in their ability to utilise different carbon substrates. While both are able to take up acetate rapidly and completely, the GAOs are far slower at consuming propionate than the PAOs during short-term substrate changes. This provides a potentially highly valuable avenue to influence the competition between PAOs and GAOs. Other VFAs studied seem to be less usable in the short term by both PAOs and GAOs, as indicated by their much lower uptake rates.

Bacteria, Anaerobic↗
PubMed2004

Structure and microbial composition of nitrifying microbial aggregates and their relation to internal mass transfer effects.

This paper presents an analysis of the structure and microbial composition of nitrifying aggregates, formed as either flocs or granules, in sequencing batch reactors (SBR) operated with a high ammonium load. The structure and microbial community of the aggregates was determined by fluorescence in situ hybridisation (FISH). The aggregate structure and size was related to mass transfer limitations observed by measurements of OURs measured by either a titrimetric and off-gas analysis sensor (TOGA) or by microsensors. The FISH analysis showed that the spatial arrangement of the microbial consortia correlated well with the oxygen gradients inside the aggregates. In the larger aggregates, the ammonium- and nitrite-oxidising bacteria were mainly concentrated to the outer 100-200 microm, whereas in the floc system, the bacteria were distributed throughout the entire aggregate. This indicates that the internal mass transfer resistance is considerably larger when the aggregate size increases which is directly supported by TOGA measurements.

Bacteria↗
PubMed2003

PCR detection of Clostridium chauvoei in pure cultures and in formalin-fixed, paraffin-embedded tissues.

The polymerase chain reaction (PCR) was used to amplify specific segments of the 16S ribosomal RNA gene of Clostridium chauvoei, a major pathogen of ruminants. Three sets of primers were used to produce amplicons of 159, 836 and 959 base pairs (bp), respectively. The PCR was evaluated by testing clinically important strains of Clostridium, including 21 strains of C. chauvoei, five strains each of Clostridium septicum and Clostridium perfringens and two strains each of Clostridium novyi, Clostridium histolyticum and Clostridium sordellii. Both purified DNA and biomass from pure cultures of each of these microorganisms were evaluated as templates in the PCR. In addition, extracts of formalin-fixed, paraffin-embedded tissues of eight sheep experimentally inoculated with C. chauvoei or C. septicum (four animals each) were also tested by the PCR using the three sets of primers. Purified DNA template of all C. chauvoei strains produced PCR amplicons of the expected size for all three primer pairs. However, when biomass from pure cultures of C. chauvoei or tissue extracts were used as templates, only the primer pair designed to produce the 159bp amplicon gave consistently positive results. No positive results were obtained with any primer pair when purified DNA or biomass from pure cultures of non-target clostridial species were used as templates. Therefore, the PCR primer sets appear to be very specific for identifying C. chauvoei in both cultures and tissues.

Animals↗
PubMed2003

The effect of GAOs (glycogen accumulating organisms) on anaerobic carbon requirements in full-scale Australian EBPR (enhanced biological phosphorus removal) plants.

Glycogen-accumulating organisms (GAOs) were present in six full-scale plants investigated and in all but one made a significant contribution to the amount of volatile fatty acid (VFA) taken up anaerobically. While most plants surveyed contain GAOs, it was demonstrated that it is possible for a full-scale plant to operate with an insignificant GAO population. "Candidatus Accumulibacter phosphatis" were the significant polyphosphate-accumulating organisms (PAOs) in all plants surveyed. "Candidatus Competibacter phosphatis" were found in all plants along with other possible GAOs that were observed but not identified. A significant GAO population will increase the carbon requirements by removing VFA that could otherwise have been used by PAOs. Process optimization minimizing GAOs in full-scale plants would lead to a more efficient use of VFA.

Bioreactors↗
PubMed2003

Microbial quantification in activated sludge: the hits and misses.

Since the implementation of the activated sludge process for treating wastewater, there has been a reliance on chemical and physical parameters to monitor the system. However, in biological nutrient removal (BNR) processes, the microorganisms responsible for some of the transformations should be used to monitor the processes with the overall goal to achieve better treatment performance. The development of in situ identification and rapid quantification techniques for key microorganisms involved in BNR are required to achieve this goal. This study explored the quantification of Nitrospira, a key organism in the oxidation of nitrite to nitrate in BNR. Two molecular genetic microbial quantification techniques were evaluated: real-time polymerase chain reaction (PCR) and fluorescence in situ hybridisation (FISH) followed by digital image analysis. A correlation between the Nitrospira quantitative data and the nitrate production rate, determined in batch tests, was attempted. The disadvantages and advantages of both methods will be discussed.

Bioreactors↗
PubMed2003

Enrichment of nitrifying microbial communities from shrimp farms and commercial inocula.

Nitrifying bacteria were selected from shrimp farm water and sediment ("natural" seed) in Thailand and from commercial seed cultures. The microbial consortia from each source giving the best ammonia removal during batch culture pre-enrichments were used as inocula for two sequencing batch reactors (SBRs). Nitrifiers were cultivated in the SBRs with 100 mg NH4-N/l and artificial wastewater containing 25 ppt salinity. The two SBRs were operated at a 7 d hydraulic retention time (HRT) for 77 d after which the HRT was reduced to 3.5 d. The amounts of ammonia removed from the influent by microorganisms sourced from the natural seed were 85% and 92% for the 7 d HRT and the 3.5 d HRT, respectively. The ammonia removals of microbial consortia from the commercial seed were 71% and 83% for these HRTs respectively. The quantity of ammonia-oxidizing bacteria (AOB) and nitrite-oxidizing bacteria (NOB) was determined in the SBRs using the most probable number (MPN) technique. Both AOB and NOB increased in number over the long-term operation of both SBRs. According to quantitative fluorescence in situ hybridisation (FISH) probing, AOB from the natural seed and commercial seed comprised 21 +/- 2% and 30 +/- 2%, respectively of all bacteria. NOB could not be detected with currently-reported FISH probes, suggesting that novel NOB were enriched from both sources. Taken collectively, the results from this study provide an indication that the nitrifiers from shrimp farm sources are more effective at ammonia removal than those from commercial seed cultures.

Ammonia↗
PubMed2002

The influence of calcium on granular sludge in a full-scale UASB treating paper mill wastewater.

Calcium precipitation can have a number of effects on the performance of high-rate anaerobic performance including cementing of the sludge bed, limiting diffusion, and diluting the active biomass. The aim of this study was to observe the influence of precipitation in a stable full-scale system fed with high-calcium paper factory wastewater. Granules were examined from an upflow anaerobic sludge blanket reactor (volume 1,805 m3) at a recycled paper mill with a loading rate of 5.7-6.6 kgCOD.m(-3).d(-1) and influent calcium concentration of 400-700 gCa m(-3). The granules were relatively small (1 mm), with a 200-400 microm core of calcium precipitate as observed with energy dispersive X-ray spectroscopy. Compared to other granules, Methanomicrobiales not Methanobacteriales were the dominant hydrogen or formate utilisers, and putative acidogens were filamentous. The strength of the paper mill fed granules was very high when compared to granules from other full-scale reactors, and a partial linear correlation between granule strength and calcium concentration was identified.

Biomass↗
PubMed2002

The "Nostocoida limicola" story: resolving the phylogeny of this morphotype responsible for bulking in activated sludge.

On the basis of 16S rRNA sequence analyses of several isolates of "Nostocoida limicola" from activated sludge plants in Australia and other countries, it is clear that "N. limicola" I, II and III are not three morphological variants of a single bacterium but at least three phylogenetically different bacteria. Data show that "N. limicola" I are members of at least two genera in the low mol% G+C gram-positive bacteria, while some isolates of "N. limicola" II belong to the high mol% G+C gram positive bacteria, and "N. limicola" III is a member of the Planctomycetales. Design and application of 16S rRNA targeted probes for each to biomass samples suggests that their phylogeny is more diverse than pure culture studies would suggest.

Biomass↗
PubMed2002

The development and use of real-time PCR for the quantification of nitrifiers in activated sludge.

Chemical analytical data has long been used to monitor the performance of activated sludge plants even though the process relies on the performance of microorganisms. It is now evident that a rapid and reliable quantitative method is required, to be able to monitor the organisms responsible for nutrient transformation and their activities, allowing avenues for more efficient nutrient removal. The development of real-time or quantitative polymerase chain reaction (PCR) also known as TaqMan or 5'-nuclease assay has allowed the rapid, quantitative analysis of DNA templates, eliminating some of the variability traditionally associated with other quantitative techniques. In this study analysis of Nitrospira spp., one of the key organisms in nitrite oxidation in wastewater treatment, was used to validate real-time PCR for the their quantification in activated sludge. A probe and primer set, targeting the 16S rRNA gene of Nitrospira spp. was designed according to the constraints of the TaqMan specifications. Samples used to evaluate the method included DNA from the sludge from full-scale wastewater treatment plants and laboratory scale systems. The reproducibility, quantitative efficiency and specificity were assessed in the evaluation. It was concluded that the method is sensitive and reproducible but has some constraints on the quantitative efficiency. A survey of full-scale systems for Nitrospira spp. was carried out and the results are presented here.

Automation↗