Factors affecting the survival and growth of bacteria introduced into lake water

The populations of Pseudomonas sp. B4, Escherichia coli, Klebsiella pneumoniae, Micrococcus flavus, and Rhizobium leguminosarum biovar phaseoli declined rapidly in lake water. The initially rapid decline of the two pseudomonads and R. phaseoli was followed by a period of slow loss of viability, but viable cells of the other species were not found after 10 days. The rapid initial phase of decline was not a result of Bdellovibrio spp., bacteriophages, or toxins in the water since Bdellovibrio spp. were not present and passage of the lake water through filters that should not have removed bacteriophages or soluble toxins led to the elimination of the rapid phase of decline. The addition of 250 g of cycloheximide and 30 g of nystatin per ml eliminated viable protozoa form the lake water, and the population of Pseudomonas sp. B4 did not fall and the decline of E. coli and K. pneumoniae was delayed or slowed under these conditions. Pseudomonas sp. L2 proliferated rapidly in lake water amended with glucose, phosphate, and NH₄NO₃, but its numbers subsequently fell abruptly; however, in water amended with cycloheximide and nystatin, which killed indigenous protozoa, the population density was higher and the fall in numbers was delayed. Of the nutrients, the chief response was to carbon, but when glucose was added, phosphorus and nitrogen stimulated growth further. Removing other bacteria by filtering the lake water before inoculation with Pseudomonas sp. L2 suggested that competition reduced the extent of response of the pseudomonad to added nutrients. We suggest that the decline in lake water of bacteria that are resistant to starvation may be a result of protozoan grazing and that the extent of growth of introduced species may be limited by the supply of available carbon and sometimes of nitrogen and phosphorus, and by predation by indigenous protozoa.     

The effects of nutrients on the survival of Escherichia coli in lake water

Escherichia coli was shown to survive without decline in viable counts for at least 12 d in filtered-autoclaved lake water. In unfiltered lake water there was a rapid decline in the viable count of E. coli. The addition of synthetic sewage to filtered-autoclaved lake water led to an increase in the viable count of E. coli at 15 degrees C and 37 degrees C and to an increase in the survival time of the E. coli in unfiltered water. The addition of phosphate and carbon sources (glucose, glycerol, succinate, acetate and lactose) did not significantly increase the survival time of E. coli in unfiltered water over the controls. The addition of ammonium sulphate and some amino acids (as nitrogen sources) to the unfiltered lake water did lead to an increase in the survival times for E. coli and this increase was proportional to the concentration of the added nitrogen source.     

The effects of nutrients on the survival of Escherichia coli in lake water

Escherichia coli was shown to survive without decline in viable counts for at least 12 d in filtered-autoclaved lake water. In unfiltered lake water there was a rapid decline in the viable count of E. coli. The addition of synthetic sewage to filtered-autoclaved lake water led to an increase in the viable count of E. coli at 15°C and 37°C and to an increase in the survival time of the E. coli in unfiltered water. The addition of phosphate and carbon sources (glucose, glycerol, succinate, acetate and lactose) did not significantly increase the survival time of E. coli in unfiltered water over the controls. The addition of ammonium sulphate and some amino acids (as nitrogen sources) to the unfiltered lake water did lead to an increase in the survival times for E. coli and this increase was proportional to the concentration of the added nitrogen source.     

Effect of organic amendments on bacterial multiplication in lake water

In Cayuga Lake water amended with 30 g of glucose or amino acids per ml, an added strain of Pseudomonas fluorescens and indigenous bacteria grew extensively, Pseudomonas sp. B4 and two rhizobia multiplied at a moderate extent, and introduced Escherichia coli and Klebsiella pneumoniae multiplied but to only a slight degree. The amendments did not enhance growth of Micrococcus flavus and Arthrobacter citreus, and an asporogenous strain of Bacillus subtilis decreased in numbers. The pseudomonads, rhizobia, E. coli and K. pneumoniae multiplied extensively when inoculated into sterile lake water amended with amino acids, A. citreus grew to a slight extent, but the numbers of M. flavus and B. subtilis did not change appreciably. In nonsterile lake water amended with 30 g of Trypticase soy broth per ml, the indigenous bacteria greatly increased in abundance, the pseudomonads, rhizobia, and E. coli developed to a lesser extent, the numbers of K. pneumoniae, A. citreus and M. flavus showed little increase, and B. subtilis decreased in numbers. Tests in pure culture containing 2 to 64 g of Trypticase soy broth per ml demonstrated good growth of P. fluorescens, Pseudomonas sp. B4, and the rhizobia at all concentrations; an initial decline followed by growth of E. coli, K. pneumoniae and R. leguminosarum biovar phaseoli at low concentrations; little or no growth or decline at the low levels but multiplication at the high levels by A. citreus and B. subtilis; and decline of M. flavus. It is proposed that the apparent K_s value, _max value, length of lag phase and resistance to stress can be used to predict behavior of bacteria in lake water receiving low levels of organic nutrients.     

Relationship between respiratory enzymes and survival of Escherichia coli under starvation stress in lake water

Survival, electron transport system (ETS) activity and the activity of NADH and succinate dehydrogenase of Escherichia coli ML30 were studied under starvation stress at different temperatures in a filtered-autoclaved lake water microcosm. ETS activity in E. coli declined rapidly at 30 degrees C but more slowly at 4 degrees and 15 degrees C over a 20 d starvation period. The decrease in ETS activity in E. coli only started after 6 d of incubation at 4 degrees C and 15 degrees C. Viability of E. coli, as determined by plate counts, declined faster at 37 degrees C than at the other temperatures and remained highest at 4 degrees C in filtered-autoclaved lake water. There was also a significant cell size reduction at 37 degrees C in filtered-autoclaved lake water but not at 4 degrees C. ETS activity after up to 16 d of starvation increased after the addition of nutrient broth to the filtered-autoclaved lake water at 15 degrees C and 30 degrees C suggesting that cells were still able to respond to nutrients, even after prolonged starvation. The response to the addition of nutrient broth, however, declined with the length of the starvation period. The activity of both succinate and NADH dehydrogenase declined over a 13 d starvation period. The loss of activity was fastest at 37 degrees C compared to lower incubation temperatures but even at 4 degrees C, a significant proportion of the activity was lost over the 13 d period.     

Production and purification ofEscherichia coli hemolysin

Eight strains of Escherichia coli, isolated from patients with a urinary tract infection were investigated for production of hemolysin. Six of these produced hemolysin and one revealed maximum hemolytic activity. Three urinary and two faecal isolates were positive for mannose-resistant hemagglutination. One isolate positive for hemagglutination and giving maximum hemolytic activity was then used. Hemolysin was present in the supernatant broth and the medium of choice to obtain the optimum yield was the alkaline meat extract broth followed by brain heart infusion broth. The highest yield appeared in the exponential phase of growth. Hemolysin is a heat-labile protein, being produced optimally at pH 8. A three-stage procedure was the best method for its purification.     

Pathogen survival during anaerobic digestion: Fatty acids inhibit anaerobic growth ofEscherichia coli

Summary The fate of pathogens in anaerobic digesters has been studied in a laboratory model system in which glucose-nutrient broth cultures of genetically-defined strains ofEscherichia coli received additions of fatty acids at concentrations similar to those attained during anaerobic treatment of farm wastes. Marked concentration-dependent inhibition of growth was observed for both antibiotic resistant and sensitive strains, and the effects increased with increasing chain lengths up to C₈. Survival of enteric organisms during anaerobic digestion may be limited by fatty acid toxicity.     

Overproduction and purification ofEscherichia coli tRNALeu

Chemically synthesized genes encodingEscherichia coli tRNA ₁ ^Leu and tRNA ₂ ^Leu were ligated into the plasmid pTrc99B. then transformed intoEscherichia coli MT102, respectively. The positive transformants, named MT-Leu1 and MT-Leu2, were confirmed by DNA sequencing, and the conditions of cultivation for the two transformants were optimized. As a result, leucinc accepting activity of their total tRNA reached 810 and 560 pmol/A₂₆₀, respectively: the content of tRNA ₁ ^Leu was 50% of total tRNA from MT-Leu1, while that of tRNA ₂ ^Leu was 30% of total tRNA from MT-Leu2. Both tRNA^Leus from their rotal tRNs were fractionated to 1 600 pmol/A₂₆₀ after DEAE-Sepharose and BD-cellulose column chromatography. The accurate kinetic constants of aminoacylation of the two isoacceptors of tRNA^Leu catalyzed by leucyl-tRNA synthetase were determined.     

Influence of phosphate ions and alkaline phosphatase activity of cells on survival ofEscherichia coli in seawater

When grown in a minimal medium and suspended for 2 hours in distilled water, seawater, phosphate buffer or a polyphosphate solution,E. coli MC4100 cells with high alkaline phosphatase activity survived in seawater for longer periods than cells with low or no activity. However, mutant cells totally deprived of alkaline phosphatase activity held in phosphate-containing media before transfer to seawater showed survival almost as high as the wild type strain, indicating that alkaline phosphatase activity is not the only factor influencing survival. Alkaline phosphatase activity also increased the protection of cells provided by glycine betaine. Survival was enhanced when cells were preincubated in the presence of phosphate or polyphosphate. Thus, the transfer of cells in wastewater could influence their subsequent survival in seawater.     

Factors involved in the distribution pattern of ciliates in the water column of a transparent alpine lake

The recurrent depth preference of three ciliate species (two prostomatids and one haptorid) in a transparent alpine lake indicates the existence of niche partitioning among them involving potential factors such as avoidance of high ultraviolet radiation levels and zooplankton predation, as well as competition for food resources.     

Cell survival and multiplication

Abstract There are clear similarities in the control mechanisms for cell survival and multiplication in the two eukaryotes, the ciliate Tetrahymena thermophila and the yeast, Saccharomyces cerevisiae . Cell multiplication in both organisms is activated by the same compounds (phorbol esters, diacylglycerol, tetrapyrroles, etc.). These compounds also affect cell multiplication and other activities in mammalian cell systems. This homology in control mechanisms in two distinct groups of unicellular eukaryotes on the one hand, and in cells from multicellular animals on the other, leads us to propose that these cytoplasmic control mechanisms for cell survival and multiplication originated in the unicellular eukaryotes.     

Survival ofEscherichia coli andYersinia enterocolitica in stream water: Comparison of field and laboratory exposure

Experiments were done to compare the influence of three aquatic exposure methods on the behavior of pathogenic and nonpathogenic enteric bacteria (Yersinia enterocolitica andEscherichia coli). Bacterial suspensions were exposed to stream water in membrane diffusion chambers in situ as well as in the laboratory using a large vessel of stream water and in enclosed bottles. The persistence of culturability of the bacterial suspensions was dependent upon the method of aquatic exposure. This difference was most apparent during the initial six days of each experiment. A steady decline in colony forming units was seen after a short stationary period in chambers in situ, while there was an abrupt increase in bacteria within chambers exposed in the laboratory. A rapid initial decrease was observed in the experimental variation using bottles, accompanied by higher levels of injury inE. coli and reduced expression of plasmid-borne virulence phenotypes inY. enterocolitica. However, there were no changes in the plasmid profiles of either organism throughout the 21-day duration of the experiments. In addition, the survival and injury of pathogenic and nonpathogenic strains of both test bacteria was very similar with aquatic exposure. These results suggest that the response of enteric bacteria in aquatic environments is influenced by experimental design as well as other factors and that the comparison of survival data should only be attempted when similar methods are used.     

Adenyl cyclase in cell-free extracts ofEscherichia coli

The adenyl cyclase enzyme system was detected in the cells ofEscherichia coli disrupted by sonic treatment. This enzyme activity is located mainly in the cell fraction sedimenting at 2,000×g, i.e. in the cytoplasmic membrane fraction. A prolonged sonication treatment of the cell suspension was followed by the disappearance of activity in the membrane preparation. The pH optimum of the adenyl cyclase inEscherichia coli was on the alkaline side, around pH 9.     

Properties ofEscherichia coli grown in deuterated media

The effects on a number of parameters of transferringEscherichia coli between protonated and deuterated media were studied; these included growth, oxygen consumption and the synthesis of DNA, RNA, total protein, and β-galactosidase. Similar measurements were made on cells fully adapted to growth on deuterated media. The amino acid compositions of deuterated and protonated cellular protein were similar, but in deuterated cells the ratio protein: DNA was doubled. Deutero- and protio-β-galactosidase had similarK _M values and turnover numbers in D₂O and H₂O. The kinetics of β-galactosidase synthesis were not changed by deuteration, but it was found that lower concentrations of inducer were required to achieve particular levels of induction. Brief exposure to inducer in one medium, followed by removal of inducer and expression of enzyme-forming-potential in either D₂O or H₂O, showed that mRNA synthesized by deuterated cells was translated equally well in both media. mRNA synthesized by protonated cells was translated about twice as efficiently in H₂O. Inducible strains (but not a regulator constitutive) lost the capacity to synthesize enzymically active β-galactosidase after more than 100 generations in D₂O-acetate. The defect persisted when such cells were grown in H₂O-acetate, but enzyme activity was restored by growth in H₂O-glycerol. The failure to produce active enzyme was not due to a failure of the induction mechanism; gel electrophoresis revealed the presence of an inactive protein species. The nature of adaptation to deuteration is discussed.     

Factors influencing the survival and multiplication of bacteriophages in calves and in their environment

Seven phages were fairly susceptible in vitro to the lethal effect of acidified whey, more so than the enteropathogenic Escherichia coli strains on which they were active. The low acidity that prevailed in the abomasum contents of calves shortly after a milk feed had little harmful effect on orally administered organisms of these phages; they flooded into the small intestine. The high acidity that prevailed later was lethal to orally administered phage organisms; few entered the small intestine. The lethal effect could be counteracted by giving CaCO3 in the feed. Low concentrations of phage-neutralizing antibodies were found in some serum samples from human beings, cattle and pigs. Antibodies to one of the seven phages were common in the human samples and antibodies to another, phage B44/1, were common in the cattle and pig samples and in bovine colostrum. Phage B44/1 antibodies in a sample of colostral whey were destroyed at pH 3.25 or less. Giving colostrum containing phage B44/1 antibodies with CaCO3 to a calf greatly reduced the numbers of orally administered phage B44/1 organisms in its alimentary tract. Antibodies to another phage were induced in the serum of a calf suffering from E. coli diarrhoea by treating it with that phage. The phages were as susceptible as the E. coli strains to the lethal action of formaldehyde and sodium hypochlorite. In contrast to the E. coli strains, they were almost completely resistant to phenol and chloroxylenol. The in vitro virulence of 21 phages varied according to the temperature at which tests were performed.(ABSTRACT TRUNCATED AT 250 WORDS)     

The hydrogenases and formate dehydrogenases ofEscherichia coli

Escherichia coli has the capacity to synthesise three distinct formate dehydrogenase isoenzymes and three hydrogenase isoenzymes. All six are multisubunit, membrane-associated proteins that are functional in the anaerobic metabolism of the organism. One of the formate dehydrogenase isoenzymes is also synthesised in aerobic cells. Two of the formate dehydrogenase enzymes and two hydrogenases have a respiratory function while the formate dehydrogenase and hydrogenase associated with the formate hydrogenlyase pathway are not involved in energy conservation. The three formate dehydrogenases are molybdo-selenoproteins while the three hydrogenases are nickel enzymes; all six enzymes have an abundance of iron-sulfur clusters. These metal requirements alone invoke the necessity for a profusion of ancillary enzymes which are involved in the preparation and incorporation of these cofactors. The characterisation of a large number of pleiotropic mutants unable to synthesise either functionally active formate dehydrogenases or hydrogenases has led to the identification of a number of these enzymes. However, it is apparent that there are many more accessory proteins involved in the biosynthesis of these isoenzymes than originally anticipated. The biochemical function of the vast majority of these enzymes is not understood. Nevertheless, through the construction and study of defined mutants, together with sequence comparisons with homologous proteins from other organisms, it has been possible at least to categorise them with regard to a general requirement for the biosynthesis of all three isoenzymes or whether they have a specific function in the assembly of a particular enzyme. The identification of the structural genes encoding the formate dehydrogenase and hydrogenase isoenzymes has enabled a detailed dissection of how their expression is coordinated to the metabolic requirement for their products. Slowly, a picture is emerging of the extremely complex and involved path of events leading to the regulated synthesis, processing and assembly of catalytically active formate dehydrogenase and hydrogenase isoenzymes. This article aims to review the current state of knowledge regarding the biochemistry, genetics, molecular biology and physiology of these enzymes.     

Influence of ecosystematic factors on survival of Escherichia coli after large-scale release into lake water mesocosms.

Mass cultures of an Escherichia coli K-12 strain were released into exposed mesocosms in a eutrophic lake. The release was performed with and without additional input of the E. coli culture medium to stimulate the scenario of leakage of a production fermenter on one hand and to compare the influence of the added organic nutrients with that of the added strain on the other hand. The survival of the introduced strain and the influence on ecological processes in the mesocosms were monitored for 10 weeks after release. For comparison, survival of the strain in microcosms with sterile lake water was also monitored. Survival of the strain was determined by means of immunofluorescence and growth on selective agar medium. In lake mesocosms, E. coli showed a rapid and constant dieback during the first week. After 4 days, cells were mostly restricted to particles, which seemed to provide niches for survival. From the second week onward, survival was improved in mesocosms with culture medium added. In microcosms with sterile lake water, plate counts of E. coli showed a strong decrease within 2 weeks, while total cell numbers remained approximately the same. The rapid elimination of E. coli from the free-water phase of the mesocosms was probably due to the combined effect of the inability to grow in lake water and grazing. The better survival of E. coli (mainly on particles) in mesocosms with added medium was attributed to the medium-induced enhancement of primary production, which was the source of a large quantity of particles. These particles, in turn, may have functioned as niches for prolonged survival as well as transport vehicles for sedimentation of the E. coli cells.     

Influence of ecosystematic factors on survival of Escherichia coli after large-scale release into lake water mesocosms.

Mass cultures of an Escherichia coli K-12 strain were released into exposed mesocosms in a eutrophic lake. The release was performed with and without additional input of the E. coli culture medium to stimulate the scenario of leakage of a production fermenter on one hand and to compare the influence of the added organic nutrients with that of the added strain on the other hand. The survival of the introduced strain and the influence on ecological processes in the mesocosms were monitored for 10 weeks after release. For comparison, survival of the strain in microcosms with sterile lake water was also monitored. Survival of the strain was determined by means of immunofluorescence and growth on selective agar medium. In lake mesocosms, E. coli showed a rapid and constant dieback during the first week. After 4 days, cells were mostly restricted to particles, which seemed to provide niches for survival. From the second week onward, survival was improved in mesocosms with culture medium added. In microcosms with sterile lake water, plate counts of E. coli showed a strong decrease within 2 weeks, while total cell numbers remained approximately the same. The rapid elimination of E. coli from the free-water phase of the mesocosms was probably due to the combined effect of the inability to grow in lake water and grazing. The better survival of E. coli (mainly on particles) in mesocosms with added medium was attributed to the medium-induced enhancement of primary production, which was the source of a large quantity of particles. These particles, in turn, may have functioned as niches for prolonged survival as well as transport vehicles for sedimentation of the E. coli cells.