scholarly journals Population Dynamics of Phenol-Degrading Bacteria in Activated Sludge Determined by gyrB-Targeted Quantitative PCR

1998 ◽  
Vol 64 (4) ◽  
pp. 1203-1209 ◽  
Author(s):  
Kazuya Watanabe ◽  
Satoshi Yamamoto ◽  
Sanae Hino ◽  
Shigeaki Harayama
Keyword(s):  
Population Dynamics ◽  
Activated Sludge ◽  
Quantitative Pcr ◽  
Pcr Amplification ◽  
Plate Count ◽  
Bacterial Populations ◽  
Pcr Product ◽  
Degrading Bacteria ◽  
Pcr Method ◽  
Kinetics Of

ABSTRACT A method for quantifying bacterial populations introduced into an activated-sludge microbial community is described. The method involves extraction of DNA from activated sludge, appropriate dilution of the extracted DNA with DNA extracted from nonintroduced activated sludge, PCR amplification of a gyrB gene fragment from the introduced strain with a set of strain-specific primers, and quantification of the electrophoresed PCR product by densitometry. The adequacy of the method was examined by analyzing the population dynamics of two phenol-degrading bacteria, Pseudomonas putida BH and Comamonas sp. strain E6, that had been introduced into phenol-digesting activated sludge. The density of each of the two populations determined by the PCR method immediately after the introduction was consistent with the density estimated from a plate count of the inoculum. This quantitative PCR method revealed different population dynamics for the two strains in the activated sludge under different phenol-loading conditions. The behavior of both of these strains in the activated sludge reflected the growth kinetics of the strains determined in laboratory axenic cultures.

scholarly journals Molecular Detection, Isolation, and Physiological Characterization of Functionally Dominant Phenol-Degrading Bacteria in Activated Sludge

1998 ◽  
Vol 64 (11) ◽  
pp. 4396-4402 ◽  
Author(s):  
Kazuya Watanabe ◽  
Maki Teramoto ◽  
Hiroyuki Futamata ◽  
Shigeaki Harayama
Keyword(s):  
Activated Sludge ◽  
Chemostat Culture ◽  
Nucleotide Sequences ◽  
Pcr Analysis ◽  
Bacterial Populations ◽  
Direct Plating ◽  
Physiological Characterization ◽  
Degrading Bacteria ◽  
Culture Enrichment ◽  
Gradient Gel Electrophoresis

ABSTRACT DNA was isolated from phenol-digesting activated sludge, and partial fragments of the 16S ribosomal DNA (rDNA) and the gene encoding the largest subunit of multicomponent phenol hydroxylase (LmPH) were amplified by PCR. An analysis of the amplified fragments by temperature gradient gel electrophoresis (TGGE) demonstrated that two major 16S rDNA bands (bands R2 and R3) and two major LmPH gene bands (bands P2 and P3) appeared after the activated sludge became acclimated to phenol. The nucleotide sequences of these major bands were determined. In parallel, bacteria were isolated from the activated sludge by direct plating or by plating after enrichment either in batch cultures or in a chemostat culture. The bacteria isolated were classified into 27 distinct groups by a repetitive extragenic palindromic sequence PCR analysis. The partial nucleotide sequences of 16S rDNAs and LmPH genes of members of these 27 groups were then determined. A comparison of these nucleotide sequences with the sequences of the major TGGE bands indicated that the major bacterial populations, R2 and R3, possessed major LmPH genes P2 and P3, respectively. The dominant populations could be isolated either by direct plating or by chemostat culture enrichment but not by batch culture enrichment. One of the dominant strains (R3) which contained a novel type of LmPH (P3), was closely related toValivorax paradoxus, and the result of a kinetic analysis of its phenol-oxygenating activity suggested that this strain was the principal phenol digester in the activated sludge.

scholarly journals Detection, Survival, and Sources of Inoculum for Bacterial Diseases of Leafy Crucifers in Oklahoma

Plant Disease ◽  
2002 ◽  
Vol 86 (8) ◽  
pp. 883-888 ◽  
Author(s):  
Youfu Zhao ◽  
John P. Damicone ◽  
Carol L. Bender
Keyword(s):  
Pseudomonas Syringae ◽  
Xanthomonas Campestris ◽  
Bacterial Pathogens ◽  
Pcr Amplification ◽  
Soil Surface ◽  
In Planta ◽  
Bacterial Diseases ◽  
Pcr Product ◽  
Pathogenicity Tests ◽  
Pcr Method

Xanthomonas campestris pv. campestris, X. campestris pv. armoraciae, and Pseudomonas syringae pv. maculicola are bacterial pathogens that cause leaf spot diseases on leafy crucifers in Oklahoma. Polymerase chain reaction (PCR) amplification of the cfl gene from the gene cluster encoding the phytotoxin coronatine was used to identify coronatine-producing strains of P. syringae, and the expected 0.65-kb PCR product was detected in 19 strains of P. syringae pv. maculicola originating from diseased crucifers in Oklahoma. A simple, rapid PCR method based on primers from the cfl gene was developed to detect coronatine-producing strains of P. syringae in planta. Pathogenicity tests confirmed the cfl-positive strains to be P. syringae pv. maculicola. To monitor the survival of X. campestris pv. armoraciae and P. syringae pv. maculicola in the field, turnip and collards were inoculated with rifampicin-resistant strains and were buried beneath the soil or left on the soil surface. Both pathogens were recovered from turnip and collard debris up to 2 months following burial, but neither pathogen was recovered from soil after the debris had decomposed. However, both pathogens were recovered from plant debris left on the soil surface for up to 5 months. Four production fields were surveyed for sources of inoculum of the bacterial pathogens from October 1999 to May 2000. X. campestris pv. campestris was isolated from the weed shepherd's purse (Capsella bursa-pastoris) in all fields, and from volunteer turnip and kale in three fields. X. campestris pv. campestris and P. syringae pv. maculicola were isolated from surface debris and regrowth from crop stubble left in one field after harvest in the fall. X. campestris pv. campestris was detected in 6 of 51 lots of crucifer seed assayed. X. campestris pv. armoraciae and P. syringae pv. maculicola were not recovered from weeds, volunteer plants, or seed lots.

scholarly journals Development of the Quantitative PCR Method for Candidatus ‘Accumulibacter phosphatis’ and Its Application to Activated Sludge

2007 ◽  
Vol 5 (1) ◽  
pp. 37-43 ◽  
Author(s):  
Toshikazu Fukushima ◽  
Naoki Uda ◽  
Motoharu Onuki ◽  
Hiroyasu Satoh ◽  
Takashi Mino
Keyword(s):  
Activated Sludge ◽  
Quantitative Pcr ◽  
Pcr Method ◽  
Accumulibacter Phosphatis

Quantitative Selective PCR of 16S Ribosomal DNA Correlates Well with Selective Agar Plating in Describing Population Dynamics of Indigenous Pseudomonas spp. in Soil Hot Spots

1999 ◽  
Vol 65 (4) ◽  
pp. 1786-1788 ◽  
Author(s):  
Kaare Johnsen ◽  
Øivind Enger ◽  
Carsten S. Jacobsen ◽  
Laila Thirup ◽  
Vigdis Torsvik
Keyword(s):  
Population Dynamics ◽  
Quantitative Pcr ◽  
Ribosomal Dna ◽  
Hot Spots ◽  
Competitive Pcr ◽  
Specific Detection ◽  
Indigenous Bacteria ◽  
16S Ribosomal Dna ◽  
Selective Agar ◽  
Pcr Method

ABSTRACT We used a quantitative PCR method targeting 16S ribosomal DNA using competitive PCR for specific detection of indigenousPseudomonas DNA in soil hot spots. The amount ofPseudomonas DNA corresponded to the number of culturablePseudomonas bacteria on Gould’s S1 agar. This represents the first use of PCR for quantification of indigenous bacteria in more than one sample of soil.

sequences allows for the design of PCR primer sets that specifically amplify the genes encoding the enzyme from DNA samples extracted from the natural environment (Henckel et al. 1999, Meyer et al. 1999, Mesarch et al. 2000). Although the PCR method is both specific and sensitive, such standard reactions are not quantitative. To obtain quantitative data from PCR-based analyses, statistical methods based on most probable number (MPN) estimations have been used (Wand et al. 1997). In MPN-PCR, DNA extracts are diluted before PCR amplification and limits are set on the number of genes in the sample by reference to known control dilutions. Another way to quantify PCR-amplified products for comparison is to include an internal control in the PCR reaction (Leser et al. 1995, Mesarch et al. 2000). Here, a known amount of target DNA is added to a PCR reaction containing DNA from the mixed microbial population. The known target DNA is complementary to the same primers and thus competes with the target sequences in the extracted DNA sample. By preparing a dilution series of the known and unknown DNA species, it is possible to quantify the amount of product produced from die complementary gene in the extracted DNA. The known DNA target can be generated by cloning the gene of interest or purifying the PCR-amplified product after which a deletion is introduced to give a differently sized PCR product. There exists many variations of the standard PCR technique. The sensitivity and specificity of the PCR may be improved by adopting a nested approach. The initial amplification is carried out with a pair of primers that are not organism-specific, whereafter a second round of amplification is conducted on the product using primers specific for an organism with target sites internal to the first primer pair (el Fantroussi et 1997, Levesque et al. 1997, Rheims and Stackebrandt 1999).

2003 ◽  
pp. 223-223
Keyword(s):  
Internal Control ◽  
Pcr Amplification ◽  
Most Probable Number ◽  
Probable Number ◽  
Pcr Product ◽  
Target Dna ◽  
Genes Encoding ◽  
Pcr Method ◽  
Primer Sets ◽  
Nested Approach

COUNTING TOTAL BACTERIA IN LAND ORGANIC WASTE HOUSEHOLD AND LAND INORGANIC WITH TOTAL PLATE COUNT METHOD (TPC)

2017 ◽  
Vol 4 (2) ◽  
pp. 87-91
Author(s):  
Ekamaida Ekamaida
Keyword(s):  
Soil Fertility ◽  
Bacterial Population ◽  
Organic Soil ◽  
Biological Properties ◽  
Plate Count ◽  
Bacterial Populations ◽  
Fertility Data ◽  
Plate Count Method ◽  
The Mean ◽  
Total Bacteria

The soil fertility aspect is characterized by the good biological properties of the soil. One important element of the soil biological properties is the bacterial population present in it. This research was conducted in the laboratory of Microbiology University of Malikussaleh in the May until June 2016. This study aims to determine the number of bacterial populations in soil organic and inorganic so that can be used as an indicator to know the level of soil fertility. Data analysis was done by T-Test that is by comparing the mean of observation parameter to each soil sample. The sampling method used is a composite method, which combines 9 of soil samples taken from 9 sample points on the same plot diagonally both on organic soil and inorganic soil. The results showed the highest bacterial population was found in total organic soil cfu 180500000 and total inorganic soil cfu 62.500.000

The Impact of a Chemostat Discharge Containing Oil Degrading Bacteria on the Biological Kinetics of a Refinery Activated Sludge Process

1988 ◽  
Vol 20 (11-12) ◽  
pp. 131-136 ◽  
Author(s):  
A. D. Wong ◽  
C. D. Goldsmith
Keyword(s):  
Activated Sludge ◽  
Retention Time ◽  
Waste Treatment ◽  
Treatment Plant ◽  
Utilization Rate ◽  
Activated Sludge Process ◽  
Kinetic Evaluation ◽  
Degrading Bacteria ◽  
Waste Treatment Plant ◽  
The Impact

The effect of discharging specific oil degrading bacteria from a chemostat to a refinery activated sludge process was determined biokinetically. Plant data for the kinetic evaluation of the waste treatment plant was collected before and during treatment. During treatment, the 500 gallon chemostatic growth chamber was operated on an eight hour hydraulic retention time, at a neutral pH, and was fed a mixture of refinery wastewater and simple sugars. The biokinetic constants k (days−1), Ks (mg/L), and K (L/mg-day) were determined before and after treatment by Monod and Lineweaver-Burk plots. Solids discharged and effluent organic concentrations were also evaluated against the mean cell retention time (MCRT). The maximum utilization rate, k, was found to increase from 0.47 to 0.95 days−1 during the operation of the chemostat. Subsequently, Ks increased from 141 to 556 mg/L. Effluent solids were shown to increase slightly with treatment. However, this was acceptable due to the polishing pond and the benefit of increased ability to accept shock loads of oily wastewater. The reason for the increased suspended solids in the effluent was most likely due to the continual addition of bacteria in exponential growth that were capable of responding to excess substrate. The effect of the chemostatic addition of specific microbial inocula to the refinery waste treatment plant has been to improve the overall organic removal capacity along with subsequent gains in plant stability.

Relationships between Population Dynamics and Operating Parameters in an Activated Sludge-Plant by Statistical Analysis

1992 ◽  
Vol 25 (4-5) ◽  
pp. 399-400 ◽  
Author(s):  
L. Cingolani ◽  
M. Cossignani ◽  
R. Miliani
Keyword(s):  
Population Dynamics ◽  
Statistical Analysis ◽  
Activated Sludge ◽  
Treatment Plant ◽  
Operating Conditions ◽  
Statistical Analyses ◽  
Operating Parameters ◽  
Aerobic Treatment

Statistical analyses were applied to data from a series of 38 samples collected in an aerobic treatment plant from November 1989 to December 1990. Relationships between microfauna structure and plant operating conditions were found. Amount and quality of microfauna groups and species found in activated sludge proved useful to suggest the possible causes of disfunctions.

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