Comparison of Gelman and Millipore Membrane Filters for Enumerating Fecal Coliform Bacteria

Tests of two leading brands of membrane filters used for enumerating fecal coliform bacteria showed that Gelman GN-6 filters recovered statistically more colonies of bacteria than did Millipore HAWG 047SO filters from pure cultures incubated at either 35 C (the optimal growth temperature) or 44.5 C (the standard temperature for the fecal coliform test). Standard membrane filter procedures with M-FC broth base were used to enumerate the organisms. Densities of colonies incubated on Gelman filters at 44.5 C averaged 2.3 times greater than those on Millipore filters. Plate counts of the bacteria at both temperatures indicated that incubation at 44.5 C did not inhibit propagation of fecal coliform bacteria. For the pour plates, M-FC broth base plus 1.5% agar was used. This modified medium compared favorably to plate count agar for enumerating Escherichia coli. At 35 and 44.5 C, colony counts on Gelman filters agreed closely with plate counts prepared concurrently, but Millipore counts were consistently lower than plate counts, especially at 44.5 C. Comparative analyses of river water for fecal coliform bacteria by the membrane filter technique gave results comparable to those for the pure cultures.     

Species Diversity of Periphyton Communities in the Littoral of a Temperate Lake

Species diversity of littoral periphyton sampled from Elk Lake (B. C., Canada) was dependent on station location and period of substrate exposure. Site differences were not statistically related to physico-chemical variables. However, diversity was negatively correlated with age of the communities and standing crop, and appeared to be associated with biological interaction or competition for available substrate surface. Further investigation revealed that the Shannon-Weaver index, as commonly applied, was inappropriate for data summarization since it reflected only the changes in evenness of the most abundant species. Components of the diversity index, as illustrated by the periphyton, were discussed.     

Microbial Communities and Fecal Indicator Bacteria Associated with Cladophora Mats on Beach Sites along Lake Michigan Shores

A high biomasses of Cladophora, a filamentous green alga, is found mainly during the summer along the shores of Lake Michigan. In this study, the abundance and persistence of the fecal indicator bacterium Escherichia coli and sulfate-reducing bacteria (SRB) on Cladophora mats collected at Lake Michigan beaches were evaluated using both culture-based and molecular analyses. Additionally, 16S rRNA gene cloning and sequencing were used to examine the bacterial community composition. Overall, E. coli was detected in all 63 samples obtained from 11 sites, and the average levels at most beaches ranged from 2,700 CFU/100 g (wet weight) of Cladophora to 7,500 CFU/100 g of Cladophora. However, three beaches were found to have site average E. coli densities of 12,800, 21,130, and 27,950 CFU/100 g of Cladophora. The E. coli levels in the lake water collected at the same time from these three sites were less than the recommended U.S. Environmental Protection Agency limit, 235 CFU/100 ml. E. coli also persisted on Cladophora mats in microcosms at room temperature for more than 7 days, and in some experiments it persisted for as long as 28 days. The SRB densities on Cladophora mats were relatively high, ranging from 4.4 × 10⁶ cells/g (6.64 log CFU/g) to 5.73 × 10⁶ cells/g (6.76 log CFU/g) and accounting for between 20% and 27% of the total bacterial counts. Partial sequences of the 16S rRNA gene clones revealed a phylogenetically diverse community, in which the Cytophaga-Flavobacterium-Bacteroides cluster and the low-G+C-content gram-positive bacteria were the dominant organisms, accounting for 40% and 12.8%, respectively, of the total clone library. These results further reveal the potential public health and ecological significance of Cladophora mats that are commonly found along the shoreline of Lake Michigan, especially with regard to the potential to harbor microorganisms associated with fecal pollution and odor-causing bacteria.     

Comparison of Fecal Coliform Agar and Violet Red Bile Lactose Agar for Fecal Coliform Enumeration in Foods

A 24-h direct plating method for fecal coliform enumeration with a resuscitation step (preincubation for 2 h at 37 ± 1°C and transfer to 44 ± 1°C for 22 h) using fecal coliform agar (FCA) was compared with the 24-h standardized violet red bile lactose agar (VRBL) method. FCA and VRBL have equivalent specificities and sensitivities, except for lactose-positive non-fecal coliforms such as Hafnia alvei, which could form typical colonies on FCA and VRBL. Recovery of cold-stressed Escherichia coli in mashed potatoes on FCA was about 1 log unit lower than that with VRBL. When the FCA method was compared with standard VRBL for enumeration of fecal coliforms, based on counting carried out on 170 different food samples, results were not significantly different (P > 0.05). Based on 203 typical identified colonies selected as found on VRBL and FCA, the latter medium appears to allow the enumeration of more true fecal coliforms and has higher performance in certain ways (specificity, sensitivity, and negative and positive predictive values) than VRBL. Most colonies clearly identified on both media were E. coli and H. alvei, a non-fecal coliform. Therefore, the replacement of fecal coliform enumeration by E. coli enumeration to estimate food sanitary quality should be recommended.     

Optimum Membrane Structures for Growth of Coliform and Fecal Coliform Organisms

The purpose of this study was to determine the optimum membrane filter structure and characteristics for recovery of coliform organisms. Additionally, other factors such as sterilization method and membrane composition were examined. Fecal coliform growth tests with varied samples indicated that the most critical factor in recovery was surface pore morphology and not other factors previously suspected. Fecal coliform counts showed a dramatic increase, with increasing surface opening sizes. Membrane structures with surface openings large enough to surround the entrapped bacteria are required for optimum growth of fecal coliform organisms. Maximum fecal coliform recoveries are obtained using membranes composed of mixed esters of cellulose exhibiting a surface opening diameter of 2.4 μm and a retention pore size of 0.7 μm.     

Evaluating Spatial and Temporal Variability of Fecal Coliform Bacteria Loads at the Pelahatchie Watershed in Mississippi

Bacterial contaminations of surface waters are an increasing concern for scientists and public health agencies because pathogenic bacteria can cause adverse effects on human health. This research was performed to investigate spatial and seasonal variability of fecal coliform bacteria (FCB) concentrations in the Pelahatchie watershed (527 km²) in Mississippi, USA. Livestock manure, poultry litter, and effluent from failing septic systems were identified as major sources of FCB in the Pelahatchie watershed. The Soil and Water Assessment Tool (SWAT)/microbial sub-model was applied, and model-simulated FCB concentrations were compared with the monthly measured FCB concentrations (years 2001–2008) at the outlet of the watershed. New methodologies were introduced to incorporate bacteria loads into the bacteria model. Results showed coefficients of determination (R ²) of 0.71 to 0.75, and Nash-Sutcliffe efficiency index (NSE) of 0.67 to 0.75 during the bacteria model's calibration and validation periods, respectively. Seasonal analysis of the model-simulated results determined the highest bacteria concentrations in January, whereas the lowest concentrations were simulated in June. Furthermore, the FCB contributions to the watershed outlet from the sources of contamination varied with time of year. This study will help watershed managers to implement best management practices for improvement of water quality.     

Isolation of Fecal Coliform Bacteria from the Diamondback Terrapin (Malaclemys terrapin centrata)

Total and fecal coliform bacteria were isolated from the cloaca and feces of the estuarine diamondback terrapin. The majority of samples contained fecal coliforms. Escherichia coli was the predominant fecal coliform species isolated, and members of the genus Salmonella were isolated from 2 of 39 terrapins. Fecal coliform numbers are used to regulate shellfish harvests, and diamondback terrapins inhabit the brackish-water habitats where oyster beds are found; therefore, these findings have implications for the efficacy of current regulatory parameters in shellfishing waters.     

Diversity and Physiological and Biochemical Properties of Heterotrophic Bacteria Isolated from Lake Baikal Epilithic Biofilms

Microbiology - A total of 170 heterotrophic bacterial strains were isolated from Lake Baikal epilithic biofilms. Identification of the isolates was carried out using the 16S rRNA gene sequencing...     

Permanent Presence of Grazing-Resistant Bacteria in a Hypertrophic Lake

The size structure of planktonic bacteria from a hypertrophic lake was investigated at 5- to 15-day intervals by means of a semiautomatic image analysis system during 1 year. Characteristic of this bacterial assemblage was the permanent presence of large filamentous bacteria and small cocci with cell sizes of <0.01 (mu)m(sup3). These filamentous bacteria, sometimes longer than 200 (mu)m and with cell volumes of up to 276 (mu)m(sup3), are larger than nanoflagellates (<20 (mu)m) and, even, metazoans living in the lake. Although they account for only 4 to 16% of bacterial abundance, their contribution to total bacterial biovolume was between 45 and 86%. An analysis of the food web structure indicates that this particular bacterial size structure may be the consequence of a strong bacterivory pressure by nanoflagellates and the absence of other larger bacterivores. The persistence of bacterial forms resistant to grazing has important consequences for the carbon flow within the microbial food web.     

Redox Transformations of Arsenic Oxyanions in Periphyton Communities

Periphyton (Cladophora sp.) samples from a suburban stream lacking detectable dissolved As were able to reduce added As(V) to As(III) when incubated under anoxic conditions and, conversely, oxidized added As(III) to As(V) with aerobic incubation. Both types of activity were abolished in autoclaved controls, thereby demonstrating its biological nature. The reduction of As(V) was inhibited by chloramphenicol, indicating that it required the synthesis of new protein. Nitrate also inhibited As(V) reduction, primarily because it served as a preferred electron acceptor to which the periphyton community was already adapted. However, part of the inhibition was also caused by microbial reoxidation of As(III) linked to nitrate. Addition of [¹⁴C]glucose to anoxic samples resulted in the production of ¹⁴CO₂, suggesting that the observed As(V) reduction was a respiratory process coupled to the oxidation of organic matter. The population density of As(V)-reducing bacteria within the periphyton increased with time and with the amount of As(V) added, reaching values as high as ~10⁶ cells ml⁻¹ at the end of the incubation. This indicated that dissimilatory As(V) reduction in these populations was linked to growth. However, As(V)-respiring bacteria were found to be present, albeit at lower numbers (~10² ml⁻¹), in freshly sampled periphyton. These results demonstrate the presence of a bacterial population within the periphyton communities that is capable of two key arsenic redox transformations that were previously studied in As-contaminated environments, which suggests that these processes are widely distributed in nature. This assumption was reinforced by experiments with estuarine samples of Cladophora sericea in which we detected a similar capacity for anaerobic As(V) reduction and aerobic As(III) oxidation.     

Comparison of the New Millipore HC with Conventional Membrane Filters for the Enumeration of Fecal Coliform Bacteria

Fecal coliform recoveries were determined for six types of membrane filters using 65 nonchlorinated water samples. Results showed that the membranes could be ranked in order of decreasing recovery as follows: Millipore HC > Gelman > Johns-Manville ~ Sartorius > Millipore HA > Schleicher & Schuell.     

Genetic Characterization of Epilithic Cyanobacteria and Their Associated Bacteria

Epilithic phototrophic biofilms develop inside Roman Necropolis and Catacombs on rock surfaces exposed to artificial light sources and are composed by a microbial consortium dominated by cyanobacteria. In this work, six non-axenic cultures of Leptolyngbya sp. strains isolated from biofilms from different Roman hypogea and maintained in cultures from 11 to 20 years were analysed along with their associated bacteria isolated in culture. The employment of PCR-fingerprinting techniques, using HIP1 and ERIC derived primers, allowed the clustering in three groups of the six Leptolyngbya strains and the typing of their isolated bacteria. The bacterial fingerprinting patterns were in agreement with the 16S rRNA gene sequencing and showed the presence in Leptolyngbya isolates of Pseudomonas, Stenotrophomonas, Agrobacterium and Bacillus representatives that were detected also in biofilms sampled from catacombs.     

Presence and sources of fecal coliform bacteria in epilithic periphyton communities of Lake Superior

Epilithic periphyton communities were sampled at three sites on the Minnesota shoreline of Lake Superior from June 2004 to August 2005 to determine if fecal coliforms and Escherichia coli were present throughout the ice-free season. Fecal coliform densities increased up to 4 orders of magnitude in early summer, reached peaks of up to 1.4x10(5) CFU cm-2 by late July, and decreased during autumn. Horizontal, fluorophore-enhanced repetitive-PCR DNA fingerprint analyses indicated that the source for 2% to 44% of the E. coli bacteria isolated from these periphyton communities could be identified when compared with a library of E. coli fingerprints from animal hosts and sewage. Waterfowl were the major source (68 to 99%) of periphyton E. coli strains that could be identified. Several periphyton E. coli isolates were genotypically identical (>or=92% similarity), repeatedly isolated over time, and unidentified when compared to the source library, suggesting that these strains were naturalized members of periphyton communities. If the unidentified E. coli strains from periphyton were added to the known source library, then 57% to 81% of E. coli strains from overlying waters could be identified, with waterfowl (15 to 67%), periphyton (6 to 28%), and sewage effluent (8 to 28%) being the major potential sources. Inoculated E. coli rapidly colonized natural periphyton in laboratory microcosms and persisted for several weeks, and some cells were released to the overlying water. Our results indicate that E. coli from periphyton released into waterways confounds the use of this bacterium as a reliable indicator of recent fecal pollution.     

Presence and growth of naturalized Escherichia coli in temperate soils from Lake Superior watersheds

The presence of Escherichia coli in water is used as an indicator of fecal contamination, but recent reports indicate that soil populations can also be detected in tropical, subtropical, and some temperate environments. In this study, we report that viable E. coli populations were repeatedly isolated from northern temperate soils in three Lake Superior watersheds from October 2003 to October 2004. Seasonal variation in the population density of soilborne E. coli was observed; the greatest cell densities, up to 3 x 10(3) CFU/g soil, were found in the summer to fall (June to October), and the lowest numbers, < or =1 CFU/g soil, occurred during the winter to spring months (February to May). Horizontal, fluorophore-enhanced repetitive extragenic palindromic PCR (HFERP) DNA fingerprint analyses indicated that identical soilborne E. coli genotypes, those with > or =92% similarity values, overwintered in frozen soil and were present over time. Soilborne E. coli strains had HFERP DNA fingerprints that were unique to specific soils and locations, suggesting that these E. coli strains became naturalized, autochthonous members of the soil microbial community. In laboratory studies, naturalized E. coli strains had the ability to grow and replicate to high cell densities, up to 4.2 x 10(5) CFU/g soil, in nonsterile soils when incubated at 30 or 37 degrees C and survived longer than 1 month when soil temperatures were < or =25 degrees C. To our knowledge, this is the first report of the growth of naturalized E. coli in nonsterile, nonamended soils. The presence of significant populations of naturalized populations of E. coli in temperate soils may confound the use of this bacterium as an indicator of fecal contamination.     

Presence and Growth of Naturalized Escherichia coli in Temperate Soils from Lake Superior Watersheds

The presence of Escherichia coli in water is used as an indicator of fecal contamination, but recent reports indicate that soil populations can also be detected in tropical, subtropical, and some temperate environments. In this study, we report that viable E. coli populations were repeatedly isolated from northern temperate soils in three Lake Superior watersheds from October 2003 to October 2004. Seasonal variation in the population density of soilborne E. coli was observed; the greatest cell densities, up to 3 × 10³ CFU/g soil, were found in the summer to fall (June to October), and the lowest numbers, ≤1 CFU/g soil, occurred during the winter to spring months (February to May). Horizontal, fluorophore-enhanced repetitive extragenic palindromic PCR (HFERP) DNA fingerprint analyses indicated that identical soilborne E. coli genotypes, those with ≥92% similarity values, overwintered in frozen soil and were present over time. Soilborne E. coli strains had HFERP DNA fingerprints that were unique to specific soils and locations, suggesting that these E. coli strains became naturalized, autochthonous members of the soil microbial community. In laboratory studies, naturalized E. coli strains had the ability to grow and replicate to high cell densities, up to 4.2 × 10⁵ CFU/g soil, in nonsterile soils when incubated at 30 or 37°C and survived longer than 1 month when soil temperatures were ≤25°C. To our knowledge, this is the first report of the growth of naturalized E. coli in nonsterile, nonamended soils. The presence of significant populations of naturalized populations of E. coli in temperate soils may confound the use of this bacterium as an indicator of fecal contamination.