Aerobic and anaerobic microorganisms in tubercles of the Columbus, Ohio, water distribution system.

Aerobic and anaerobic microorganisms were enumerated in tubercles collected from sections of the water distribution pipeline in the Columbus, Ohio, metropolitan area. Coliform bacteria were not detected in the tubercles examined. Sulfate-reducing bacteria were detected in 80% of the samples. Nitrate-reducing heterotrophs were present in all samples. The results, including plate counts of aerobic heterotrophs, indicated variation in bacterial densities depending on the tubercle sample and fraction examined. The associations among the viable counts obtained by the different culture methods were analyzed statistically, using three methods (Pearson, Spearman, and Kendall).     

Evaluation of a cup scrub technique for quantification of the microbial flora on bovine skin

A cup-scrub technique devised for sampling the human skin surface microflora was evaluated in cattle. Scrub samples from bovine skin contained clumps of squama and bacterial microcolonies which were progressively broken down by shaking. This was accelerated in the presence of ballotini beads but aggregations of bacteria were still present after prolonged agitation. Vigorous shaking, particularly with beads, decreased the viability of the bacteria and optimum viable counts were obtained after manual shaking for half a minute. Immersion in buffered detergent, wash and diluting fluids for up to 2 h promoted release of bacteria from microcolonies but decreased the viability of aerobic and anaerobic pleomorphic rods and a Bacillus strain. There was no significant effect on strains of Micrococcaceae. Prolonged exposure of bacteria from scrub samples to these fluids can thus lead to both quantitative and qualitative alterations in the counts obtained, although these effects may be masked by the continuing release of bacteria from microcolonies. The cup-scrub technique provides a convenient means of quantifying changes in the bovine skin microflora but results obtained from different studies should only be compared if closely similar techniques are used.     

Distribution of bacteria in the sediment of an oxbow tropical lake (Lagoa do Infernão,SP, Brazil)

The temporal distribution of bacteria on the surface sediment of the oxbow lake Lagoa do Infernão was studied over a period of one year using different techniques for bacterial counts. Several bacterial groups were differentiated, such a total bacteria and filamentous, heterotrophic, amylolytic, and lipolytic bacteria. The best defined temporal distribution pattern was obtained by total-AO bacterial counts, with a variation of 1.26 × 10¹⁰ to 8.58 × 10¹⁰ bacteria (g dw)^–1 during the dry and rainy seasons, respectively. The electrode potential (Eh) of the surface sediment ranged from – 148 mV during the rainy season to + 371 mV during the dry season. The variations detected on the surface sediment suggest a greater allochthonous contribution of material and bacteria resulting in high decomposing activity during the rainy season.     

A note on 'plotless' methods for estimating bacterial cell densities

R oser , D., N edwell , D.B. & G ordon , A. 1984. A note on "plotless" methods for estimating bacterial cell densities. Journal of Applied Bacteriology 56 , 343–347.
'Plotless' techniques for determining population densities have been developed for, and applied to, higher plant populations. They can often be carried out more rapidly than techniques involving total counts of individuals in plots, or quadrants, but such plotless techniques have not been generally applied to the estimation of densities of bacterial cells. Direct microscopical counting of cell numbers in a field of view, an example of a plot-related method, has been traditionally used for micro-bial cell counts. In this study 'plot' and 'plotless' methods on a variety of bacterial samples are compared. Estimates of bacterial cell density were obtained by measuring the distance of cells from a fixed point in a field of view. The values, which were more rapidly obtained, were directly correlated with total cell counts. Although there was some apparent deviation from a perfect 1:1 relationship with total counts, as indicated by a correlation coefficient less than 1.0, there were no significant differences between the replicated counts of bacteria on samples of tissue from the surface of Hypholoma basidiocarps ( P < 0.05). This indicated that the methods of enumeration were comparable. The distance-related estimates could readily be obtained from fields of view with cell densities varying over several orders of magnitude. It was more rapidly applied, particularly at high density, and the method was applicable not only to random cell distributions but also to the non-random distributions encountered when microbial cells aggregated into micro-colonies. The method appears to be particularly well-suited for automated, digitized, direct counting procedures, as well as to estimating bacterial numbers on membrane filters and natural substrates.     

Evaluation of a cup scrub technique for quantification of the microbial flora on bovine skin

L loyd , D.H. 1984. Evaluation of a cup scrub technique for quantification of the microbial flora on bovine skin. Journal of Applied Bacteriology 56 , 103–107.
A cup-scrub technique devised for sampling the human skin surface microfiora was evaluated in cattle. Scrub samples from bovine skin contained clumps of squames and bacterial microcolonies which were progressively broken down by shaking. This was accelerated in the presence of ballotini beads but aggregations of bacteria were still present after prolonged agitation. Vigorous shaking, particularly with beads, decreased the viability of the bacteria and optimum viable counts were obtained after manual shaking for half a minute. Immersion in buffered detergent, wash and diluting fluids for up to 2 h promoted release of bacteria from micro-colonies but decreased the viability of aerobic and anaerobic pleomorphic rods and a Bacillus strain. There was no significant effect on strains of Micrococcaceae. Prolonged exposure of bacteria from scrub samples to these fluids can thus lead to both quantitative and qualitative alterations in the counts: obtained, although these effects may be masked by the continuing release of bacteria from microcolonies. The cup-scrub technique provides a convenient means of quantifying changes in the bovine skin microfiora but results obtained from different studies should only be compared if closely similar techniques are used.     

Bacterial Counts and Quality of Iced Fish Retailed at a Lusaka Market, Zambia

S ummary . The counts of total viable, coliform, streptococcal and sulphite reducing anaerobic bacteria and the presence of salmonellae were determined on 134 iced fish obtained from Luburma Market, Lusaka, Zambia, during June-December 1970. The quality of the uncooked fish was also assessed by appearance and odour. The purpose of these determinations was to obtain a picture of the variations of the bacterial counts in relation to season, origin, fish species and market quality. Total viable and coliform counts were of the order of millions and tens of thousands/cm² of skin surface, respectively. Higher counts were obtained in the hot season during September-October but with little change in appearance of the fish. There was a significant correlation ( P < 0·01) of both total viable and coliform bacteria with quality scores. A maximum permissible level of 10⁷ cells/cm² of skin surface was proposed for total viable counts and 10⁵/cm² for coliform bacteria, for iced fish of acceptable quality in Zambia.     

Recovery and diversity of heterotrophic bacteria from chlorinated drinking waters.

Heterotrophic bacteria were enumerated from the Seattle drinking water catchment basins and distribution system. The highest bacterial recoveries were obtained by using a very dilute medium containing 0.01% peptone as the primary carbon source. Other factors favoring high recovery were the use of incubation temperatures close to that of the habitat and an extended incubation (28 days or longer provided the highest counts). Total bacterial counts were determined by using acridine orange staining. With one exception, all acridine orange counts in chlorinated samples were lower than those in prechlorinated reservoir water, indicating that chlorination often reduces the number of acridine orange-detectable bacteria. Source waters had higher diversity index values than did samples examined following chlorination and storage in reservoirs. Shannon index values based upon colony morphology were in excess of 4.0 for prechlorinated source waters, whereas the values for final chlorinated tap waters were lower than 2.9. It is not known whether the reduction in diversity was due solely to chlorination or in part to other factors in the water treatment and distribution system. Based upon the results of this investigation, we provide a list of recommendations for changes in the procedures used for the enumeration of heterotrophic bacteria from drinking waters.     

Recovery and diversity of heterotrophic bacteria from chlorinated drinking waters

Heterotrophic bacteria were enumerated from the Seattle drinking water catchment basins and distribution system. The highest bacterial recoveries were obtained by using a very dilute medium containing 0.01% peptone as the primary carbon source. Other factors favoring high recovery were the use of incubation temperatures close to that of the habitat and an extended incubation (28 days or longer provided the highest counts). Total bacterial counts were determined by using acridine orange staining. With one exception, all acridine orange counts in chlorinated samples were lower than those in prechlorinated reservoir water, indicating that chlorination often reduces the number of acridine orange-detectable bacteria. Source waters had higher diversity index values than did samples examined following chlorination and storage in reservoirs. Shannon index values based upon colony morphology were in excess of 4.0 for prechlorinated source waters, whereas the values for final chlorinated tap waters were lower than 2.9. It is not known whether the reduction in diversity was due solely to chlorination or in part to other factors in the water treatment and distribution system. Based upon the results of this investigation, we provide a list of recommendations for changes in the procedures used for the enumeration of heterotrophic bacteria from drinking waters.     

Comparison of methods for detection and enumeration of airborne microorganisms collected by liquid impingement.

Bacterial agents and cell components can be spread as bioaerosols, producing infections and asthmatic problems. This study compares four methods for the detection and enumeration of aerosolized bacteria collected in an AGI-30 impinger. Changes in the total and viable concentrations of Pseudomonas fluorescens in the collection fluid with respect to time of impingement were determined. Two direct microscopic methods (acridine orange and BacLight) and aerodynamic aerosol-size spectrometry (Aerosizer) were employed to measure the total bacterial cell concentrations in the impinger collection fluid and the air, respectively. These data were compared with plate counts on selective (MacConkey agar) and nonselective (Trypticase soy agar) media, and the percentages of culturable cells in the collection fluid and the bacterial injury response to the impingement process were determined'. The bacterial collection rate was found to be relatively unchanged during 60 min of impingement. The aerosol measurements indicated an increased amount of cell fragments upstream of the impinger due to continuous bacterial nebulization. Some of the bacterial clusters, present in the air upstream of the impinger, deagglomerated during impingement, thus increasing the total bacterial count by both direct microscopic methods. The BacLight staining technique was also used to determine the changes in viable bacterial concentration during the impingement process. The percentage of viable bacteria, determined as a ratio of BacLight live to total counts was only 20% after 60 min of sampling. High counts on Trypticase soy agar indicated that most of the injured cells could recover. On the other hand, the counts from the MacConkey agar were very low, indicating that most of the cells were structurally damaged in the impinger. The comparison of data on the percentage of injured bacteria obtained by the traditional plate count with the data on percentage of nonviable bacteria obtained by the BacLight method showed good agreement.     

Viable airborne microbial counts from air-cooling units with and without complaints of urine and body odors

Viable airborne microbial counts are commonly used in indoor air quality (IAQ) assessment, but studies linking the microbial counts to a specific type of indoor microbial contamination are limited. We hypothesize that the airborne microbial counts can differentiate air-cooling units with and without complaints of urine and body odors. The keratinolytic property of some isolated bacteria prompts to the hypothesis that keratinase is present in the units to break down keratins, structural proteins that form human skin scales, as sources of amino acids and ammonium to produce the odors. Seven bacterial species and four fungal species were identified in the units and room air. Airborne Staphylococcus haemolyticus and Methylobacterium organophilum counts contributed the most to the microbial dissimilarities of units with and without odor complaints. Keratinolytic bacteria and a methylotrophic bacterium were abundant in the units. All the units contained ammonium, and keratinase activity was higher in the units with odor complaints. Extracellular keratinase activity was more effective at 20 °C than at 30 or 4 °C. Keratinolytic bacteria produced high levels of ammonium in the culture with skin cells. Viable airborne microbial counts can help IAQ inspectors to identify potential odor-causing air-cooling units. Keratins may be broken down in the units and associated with the odor complaints.     

The cutaneous microbiology of normal human feet

A survey has been made of the bacterial and fungal populations carried at three different sites on the feet of 60 individuals. The bacteria found at the three sites were quantitatively similar and Micrococcaceae and aerobic coryneform bacteria predominated. The carriage of other bacterial groups was generally low. There was a quantitative variation between sites--mean total counts were 1.04 X 10(7) cfu/cm2 skin in the fourth toe cleft, 4.08 X 10(5) cfu/cm2 skin on the sole and 1.21 X 10(3) cfu/cm2 skin on the dorsal surface. Staphylococci were most often dominant on the sole and dorsal surface whereas aerobic coryneforms predominated in the majority of fourth toe clefts. The higher the total count at a given site the more likely it was that aerobic coryneform bacteria predominated. The skin surface pH was significantly higher on the sole (mean value 6.25) than on the dorsal surface (mean value 5.23). Factors controlling the microbial ecology of the foot are discussed.     

Comparison of bacterial populations of the pig cecum and colon based upon enumeration with specific energy sources.

Concentrations of bacteria in the ceca and colons of pigs were measured by determinations of colony counts on rumen fluid-based media in anaerobic roll tubes. With our most complete medium (medium CCA), the mean colony count of cecal samples from 20 pigs was 2.37 X 10(10) +/- 1.0 X 10(10) (+/- standard deviation)/g (wet weight). The mean number of bacteria attached to or associated with cecal epithelial tissues from three pigs on medium CCA was 2.67 X 10(7) +/- 0.81 X 10(7)/cm2 of tissue. The proportions of gut bacterial populations able to use various energy substrates were estimated on the basis of relative colony counts. The following substrates are listed in descending order of their capacity to support growth of cecal bacteria: glucose, starch, cellobiose, xylose, Trypticase, gastric mucin from swine, mannitol, glycerol, and lactate. The effect of diet upon this distribution was not examined. The relative proportions of bacteria from a given population that were able to grow on various selective media were used as population profiles. Comparisons of populations in this way indicated that differences could be detected between (i) populations from the cecum of littermate pigs, (ii) populations from the cecum and colon of the same pig, and (iii) populations in the lumen of the cecum as compared with populations associated with cecal mucosa.     

Comparison of bacterial populations of the pig cecum and colon based upon enumeration with specific energy sources.

Concentrations of bacteria in the ceca and colons of pigs were measured by determinations of colony counts on rumen fluid-based media in anaerobic roll tubes. With our most complete medium (medium CCA), the mean colony count of cecal samples from 20 pigs was 2.37 X 10(10) +/- 1.0 X 10(10) (+/- standard deviation)/g (wet weight). The mean number of bacteria attached to or associated with cecal epithelial tissues from three pigs on medium CCA was 2.67 X 10(7) +/- 0.81 X 10(7)/cm2 of tissue. The proportions of gut bacterial populations able to use various energy substrates were estimated on the basis of relative colony counts. The following substrates are listed in descending order of their capacity to support growth of cecal bacteria: glucose, starch, cellobiose, xylose, Trypticase, gastric mucin from swine, mannitol, glycerol, and lactate. The effect of diet upon this distribution was not examined. The relative proportions of bacteria from a given population that were able to grow on various selective media were used as population profiles. Comparisons of populations in this way indicated that differences could be detected between (i) populations from the cecum of littermate pigs, (ii) populations from the cecum and colon of the same pig, and (iii) populations in the lumen of the cecum as compared with populations associated with cecal mucosa.     

Survival of Staphylococcus epidermidis on the skin of patients of lepromatous leprosy

The effect of hydration on Staphylococcus epidermidis, the predominant resident bacterial flora, was studied on skin affected by leprosy and known to have impaired sweating. Normal areas served as control. Significantly higher bacterial counts were observed in affected areas compared with normal-looking skin in 16/19 of the patients. Artificial application of Staph. epidermidis on leprosy-affected and unaffected areas, however, showed equivocal results, as in only 50% of the patients were higher counts obtained in affected compared with unaffected sites. The possible responsible factors for the present observation are discussed.     

Comparison of techniques to determine the abundance of predatory bacteria attacking chromatiaceae

Abstract This work quantifies the number of bacterial predators attacking the population of Chromatiaceae in the hypolimnion of Lake Estanya to assess the potential role of these microorganisms in controlling phototrophic bacterial populations. The abundance of predators was estimated from total counts of infected prey cells and by counting plaque-forming units. In spite of the large difference between both determinations, their variations with depth and time followed very similar patterns. During the summer, in the hypolimnion, and during the winter in the entire lake, up to 60% of the prey cells had potential predators attached. In comparison, plaque counts showed that viable predators represented less than 1% of the population of the prey. Our results demonstrated that predatory bacteria were far more abundant than indicated by the low viable counts obtained, suggesting that they play a more important role in controlling phototrophic bacterial populations than is currently assumed.     

Survival of Staphylococcus epidermidis on the skin of patients of lepromatous leprosy

The effect of hydration on Staphylococcus epidermidis , the predominant resident bacterial flora, was studied on skin affected by leprosy and known to have impaired sweating. Normal areas served as control. Significantly higher bacterial counts were observed in affected areas compared with normal-looking skin in 16/19 of the patients. Artificial application of Staph. epidermidis on leprosy-affected and unaffected areas, however, showed equivocal results, as in only 50% of the patients were higher counts obtained in affected compared with unaffected sites. The possible responsible factors for the present observation are discussed.     

Studies of Different Sampling Methods for the Determination of Bacterial Counts from Frozen Broilers

The attachment of bacteria to the skin of broilers was not influenced by freezing or thawing of the carcasses. Only a small proportion of indicator organisms which were attached to the skin during processing were recovered in either the thaw water or rinsing fluid; a close correlation was found between bacterial counts from both types of sample. Numbers of Enterobacteriaceae and total counts were readily obtained by sampling the thaw water and this is, moreover, a non-destructive method. In order to detect the presence or absence of pathogenic micro-organisms the rinsing method was considered to be preferable, provided that the whole of the rinsing fluid was tested in each case.     

Isolation of Klebsielleae from within living wood.

Previous studies from this laboratory have documented the presence of coliform bacteria emanating from wooden reservoirs containing finished drinking water. Coliforms were identified as Klebsiella pneumoniae and Enterobacter spp. In the present report, evidence is presented which suggests that the origin of these coliforms is from the wood used to construct the reservoirs. In liquid expressed from freshly cut redwood, total bacterial counts in the range of 10(5) to 10(6)/ml were commonly observed. When present, coliform counts were over 10(3)/ml of expressed liquid. E. agglomerans was the most prevalent coliform present, but Klebsiella was isolated from freshly cut logs. Citrobacter freundii was also occasionally isolated. No fecal coliform-positive Klebsiella were obtained from any of the samples. Highest total bacteria and coliform counts were observed in sapwood specimens. Coliforms were present throughout sapwood as evidenced by contact plating serial sections of freshly cut wood. Scanning electron micrographs illustrate the presence of bacterial colonies within sapwood tracheids. Other wood species also contained coliform bacteria but in numbers lower than found in redwood.     

Bacterial skin flora variation and in vitro inhibitory activity against Saprolegnia parasitica in brown and rainbow trout

Variations in the number and diversity of bacteria from the skin of brown trout Salmo trutta L. and rainbow trout Oncorhynchus mykiss Walbaum were surveyed from different rivers and fish farms in northern Spain. In addition to determining bacterial populations in skin samples of healthy fish, bacterial populations were determined from skin lesions (of brown trout only) infected with Saprolegnia parasitica, the causal agent of saprolegniosis. Mean bacterial counts from skin lesions of brown trout suffering from saprolegniosis were nearly 1000 times greater than from the skin of uninfected brown and rainbow trout. More than 20 different genera of bacteria were identified, with isolates of Aeromonas and Iodobacter being the predominant genera associated with saprolegniosis lesions. The in vitro inhibitory activity of 72 of these skin isolates was tested against S. parasitica using 3 different assays. These included (1) assessing the inhibition by bacteria of colony growth on agar media, (2) the inhibition of colony growth from colonized hemp seeds in liquid media and (3) the inhibition of cyst germination in liquid media. Finally, the fungicidal effect of the 24 most inhibitory bacterial species, and the inhibitory activity of their culture supernatants, was tested in the same way. Isolates identified as Aeromonas piscicola, A. sobria, Pantoea agglomerans and Pseudomonas fluorescens achieved the highest inhibition against S. parasitica. Many of these inhibitory isolates were obtained primarily from skin lesions of fish with saprolegniosis. It is suggested that some of these isolates might be useful in the biological control of saprolegniosis.