Water pH and urinary excretion in silver catfish Rhamdia quelen

The effect of acute exposure to different water pH levels on urinary excretion and plasma ion levels in silver catfish Rhamdia quelen was analysed. Fish were exposed to pH 4·0, 5·0, 7·5, 8·0, and 9·0 for 4 days and urine was collected. Other specimens were also exposed to the experimental pH for 24 h and blood was sampled. Urine flow rate, urine and plasma pH showed a significant trend to increase with the increase of water pH. Urinary Na⁺ excretion rate also increased and ammonia urinary excretion rate decreased with the increase of water pH. There was a significant trend to decrease volume, ammonia, Cl⁻ and Na⁺ urinary excretion rate with increasing mass in fish exposed to all pH levels studied. Plasma ammonia levels showed a slight decrease in fish exposed to water pH from 4·0 to 8·0, but those exposed to water pH 9·0 presented the highest ammonia levels. Most plasma ions and urinary excretion changes observed in silver catfish exposed to acidic or alkaline water were similar to those already detected in rainbow trout Oncorhynchus mykiss . In addition, the kidney and urinary bladder might participate on acid–base balance in silver catfish, since urine pH changed according to plasma pH.     

Survival and reproductive performance of the desert pupfish, Cyprinodon n. nevadensis (Eigenmann and Eigenmann), in acid waters

Desert pupfish, Cyprinodon n. nevadensis , were exposed to pH levels of 8·3 (control), 7·0, 6·5, 6·0, 5·5 and 5·0 to determine the effects of acidity on reproduction. Egg production was significantly reduced at every pH level tested below the control. Egg-laying virtually ceased at pH 5·0, while egg viability was reduced to less than 50% of the control value at pH 6·5, when eggs were tested at the same pH at which they were laid. The 96 h LC₅₀ for this species was pH 4·56, considerably below that for successful reproduction. Larvae were less tolerant to acid stress than were adults.
Acclimation of reproductive performance to increased acid levels did not occur. Reproductive performance did not fully recover to control levels when the fish were placed in more favourable conditions after prolonged exposure to low pH.
Reduction in the number and development of oocytes was observed in the ovaries of acid stressed fish resulting in the decreased reproductive potential.     

Salinity tolerance of the mosquito fish, Gambusia affinis (Baird and Girard)

Experiments were conducted to determine the adaptability of mosquito fish to various salt concentrations. Mosquito fish, Gambusia affinis , (total length (T.L.) 27–37 mm) were subjected to abrupt and gradual changes from fresh water to various salinities (S). Low mortality (10%) occurred when the fish were transferred from fresh water (S = 0·5‰) to 50% sea water (S = 19·5‰ Through gradual adaptation 65·0% of the fish were able to tolerate 100% sea water for 7 days. After a 7 -day stay in sea water, fish were readapted to fresh water during a 3-h period. The results indicate that they were well able to tolerate the abrupt change. Through a gradual adaptation fish were also able to tolerate salinities ranging from 39·0‰ (100% sea water) to 58·8‰     

The effect of acid water on the hatching of salmonid eggs

The survival of salmon Salmo salar , sea trout and brown trout Salmo trutta eggs in acid water in the range pH 3·5±7·0 was investigated over a period of three winters. The effects of fertilising the eggs in acid water at pH 4·0, 4·5 and 5·0, and dilution of the test media were also studied. There was considerable yearly variation in the numbers of eggs which hatched, but there were no marked differences between the three species in the tolerances of the eggs to acid water; pH 3·5 was lethal within 10 days to all the eggs, but at pH 4·5 and higher pH levels there was no obvious difference in hatching attributable to acidity.     

Survival of Listeria monocytogenes in sea water and effect of exposure on thermal resistance

Survival, recoverability and sublethal injury of two strains of Listeria monocytogenes , Scott A and an environmental strain KM, on exposure to sea water at 12·8 or 20·8 °C was determined using in situ diffusion chambers. Plate counts were used to assess recoverability and injury while 5-cyano-2,3-ditolyl tetrazolium chloride (CTC) reduction was used to determine respiratory activity. T₉₀ values (times for 10-fold decreases in numbers of recoverable cells) on non-selective medium (trypticase soya agar with 0·6% yeast extract) at 12·8 and 20·8 °C were 61·7 and 69·2 h for L. monocytogenes Scott A, and 103·0 and 67·0 h for L. monocytogenes KM, respectively. On selective medium (Oxford agar), T₉₀ values at 12·8 and 20·8 °C were 60·6 and 56·9 h for L. monocytogenes Scott A, and 83·0 and 65·9 h for L. monocytogenes KM, respectively. With Scott A, the percentage of sublethally injured cells at 12·8 and 20·8 °C was 1·7 and 17·7%, respectively, while for KM the values were 19·0 and 1·6%, respectively. The fraction of cells reducing CTC but which were not recoverable on plating progressively increased on exposure to sea water. Listeria monocytogenes KM challenged at 58 °C showed an apparent increase in heat resistance after exposure to sea water at 20·8 °C for 7 d ( D ₅₈= 2·64 min) compared with before exposure ( D ₅₈= 1·24). This increase in thermal resistance was not apparent at temperatures greater than 63 °C, and analysis of the best-fit regression lines fitted to the thermal data obtained from the two cell populations indicated that their thermal resistance was not significantly different ( P > 0·05) over the temperature range tested (58–62 °C).     

Glucose-induced acid tolerance appearing at neutral pH in log-phase Escherichia coli and its reversal by cyclic AMP

Escherichia coli shifted from broth at external pH (pH₀) 7·0 to pH₀ 7·0 broth plus glucose rapidly induced marked acid tolerance which also appeared, albeit to a lesser extent, plus maltose, sucrose or lactose. Tolerance appeared without the medium pH becoming acidic. Tolerance was most substantial when glucose was added at pH₀ 7·0 but was also appreciable at pH₀ 7·5, 8·0 and 8·5. Induction of tolerance by glucose was markedly reduced by cyclic AMP and essentially abolished plus NaCl or sucrose ; the induction process was also reduced but not fully inhibited by chloramphenicol, tetracycline and nalidixic acid. Glucose-induced organisms showed less acid damage to DNA and β-galactosidase and it is likely that this is because glucose induces a new pH homeostatic mechanism which keeps internal pH close to neutrality at acidic pH₀. In conclusion, it is clear that glucose induces a novel acid tolerance response in log-phase E. coli at pH₀ 7·0 ; it is now known that induction of this response involves the functioning of extracellular induction components including an extracellular induction protein.     

Habituation to acid in Escherichia coli: conditions for habituation and its effects on plasmid transfer

Induction of acid resistance (habituation) in Escherichia coli at pH 5·0 took ca 5 min in broth at 37°C and 30–60 min in minimal medium. Induction occurred at a range of pH values from 4·0 to 6·0; it was dependent on continuing protein and RNA synthesis but substantial acid resistance appeared in the presence of nalidixic acid. Acid resistance was long-lasting; organisms grown at pH 5·0 retained most of their resistance after 2 h growth at pH 7·0. Organisms grown at pH 5·0 showed increased synthesis of a number of cytoplasmic proteins compared with the level in cells grown at pH 7·0. DNA repair-deficient strains carrying recA, uvrA or polAl mutations were more acid-sensitive than the repair-proficient parents but were able to habituate at pH 5·0. Organisms grown at pH 5·0 transferred the ColV plasmid much more effectively at acid pH than did those grown at pH 7·0 and habituated recipients appeared better able to repair incoming acid-damaged plasmid DNA than did those that were non-habituated. Induction of acid resistance at pH 5·0 may be significant for the survival of organisms exposed to periodic discharges of acid effluent in the aquatic environment and habituation may also allow plasmid transfer and repair of acid-damaged plasmid DNA during or after such exposure.     

Survival and sublethal responses of early life stages of pike exposed to low pH in artificial post-mining lake water

The threshold pH for survival of early life stages of pike Esox lucius in post-mining lakes was determined in a laboratory experiment using artificial water characterized by high concentrations of dissolved Ca, Al, Fe, Mn and SO2/4⁻. At pH 3·50–4·00, high mortality was observed before hatching. At pH 4·00 and 4·25, hatching rates were reduced compared with the controls (pH 7·40), and many embryos died in a partly hatched state. Hatching success of embryos exposed to pH levels of 4·50 or higher was not affected. However, at pH 5·00 many newly hatched embryos were deformed. Furthermore, pike exposed to pH 4·00–5·00 did not start feeding. At pH 4·75 and below, mortality increased to 100%, and at pH 5·00, only few eleutheroembryos which were in extremely poor condition survived to the end of the experiment. At pH 5·50, survival was in the same range as in the control group, but growth was reduced. Therefore, early life stages of pike are expected to survive in Lustian post-mining lakes when a pH of 5·50 and above is reached and maintained.     

The preparation of ampicillin dextrin agar for the enumeration of Aeromonas in water

Introduction of ampicillin dextrin agar (ADA) has revealed problems in details of the preparation. The final pH of the medium varied substantially between different laboratories. Measuring temperature has a pronounced effect on the pH (0·7 units lower at 50°C than at 6°C). Addition of agar during medium preparation resulted in a fall in pH of 0·5 units. If poured plates were stored in the refrigerator, the pH was reduced by 0·1–0·4 units, in particular during the first day. Recovery of Aeromonas from pure cultures and naturally polluted samples was unaffected by variation in pH between 7·1 and 8·3 but colony differentiation was optimal at a higher pH. The use of ADA at a final pH of 7·8 ± 0·2 (at 25°C) is recommended. Different types of dextrin differed in respect of solubility, fermentability and colony differentiation. Optimal results were obtained with Difco 161 and Merck 3006.     

THE LOSS OF VIABILITY OF BACTERIA IN SUSPENSION DUE TO CHANGING THE IONIC ENVIRONMENT

SUMMARY: Following the accidental discovery and subsequent confirmation of an 80% loss of viability in a suspension of Escherichia coli when the ionic environment was changed from phosphate buffer to Ringer's solution, and of similar losses after changes in the reverse direction, the behaviour of other organisms was studied. Clostridium butyricum, Streptococcus lactis, Escherichia coli , and Micrococcus candidus gave positive results, Mycobacterium smegrmatis and a brewer's yeast did not. The survey included variations in the conditions, mostly with E. coli. Minor constituents of the solutions or alteration of their pH modified the effect, e.g. that of 1% NaCl in place of Ringer's solution or of pH 8·0 instead of the usual pH 7·2, was quite different. Also, the addition of glucose, sodium thioglycollate, or methylene blue diminished it, with one exception in the case of glucose. Mic. candidus responded if the solutions were chilled, but not at room temperature. With electrolyte concentrations below about 0·02 M there was no effect. Various other changes of anions were effective, as also was change from a univalent to a bivalent cation. A second treatment, following the necessary intermediate reverse change which had no effect, caused another smaller mortality. This reduced sensitivity gave a diminished loss which persisted through many subsequent subculturings.     

Effects of environmental factors on survival, growth, and production of American shad larvae

Episodic increases in temperature of 5°C above 20° C, over 48 h or declines in pH of 1·0 unit from pH 7·0 reduced survival of yolk-sac and feeding-stage larvae of American shad Alosa sapidissima . Over 16 days all measures of survival, growth, and production were more favourable at each higher temperature in the 15–25° C range. More favourable responses were also obtained at the higher prey level (500 v . 50 Artemia nauplii l^-1) and at the higher pH (7·5 v . 6·5). Combinations of high temperature and high prey levels, at pH 7·5, led to highest larval production. Little growth or production occurred at 15° C, regardless of pH or prey level. The effect of pH was strong with respect to survival, but weak with respect to growth. In attempts to restore American shad populations by larval stocking, release times and sites can be critical to optimize survival and eventual returns. Releases of larvae potentially will be most effective when made at temperatures >20° C, pH>7·0, and prey levels >50 1^-1. These conditions are most likely to occur in Maryland tributaries of Chesapeake Bay between mid-May and early June.     

Unusual motility characteristics of sperm of the spotted wolffish

Unlike the sperm of most teleosts, that of the spotted wolffish Anarhichas minor is motile on stripping, remains motile for at least 2 days and loses motility when exposed to sea water. Computer assisted sperm analysis (CASA) was used to quantitatively examine the motility characteristics of spotted wolffish sperm. Straight line velocity (VSL), beat cross frequency (BCF) and percentage motility were the most sensitive indicators of movement. Sperm trajectories were very different to those of other teleosts examined, showing large side-to-side movements of the sperm head and a more 'wiggly' behaviour which may be an adaptation to swimming in the viscous gelatinous egg mass. VSL was not altered by pH from 5·0 to 9·0, but was lower at pH 4·5. It was highest at 200 to 500 mOsm and decreased rapidly at <200 mOsm and more slowly at >500 mOsm. It is suggested that the unusual characteristics of spotted wolffish sperm in its trajectory and duration of motility, its release in a fully activated state and its greatly decreased motility in both fresh and sea water are related to a spawning strategy involving mixing of sperm with eggs contained in a gelatinous mass rather than release directly into water in proximity to the ova.     

Induction of acid tolerance at neutral pH in log-phase Escherichia coli by medium filtrates from organisms grown at acidic pH

Escherichia coli grown at pH 5·0 became acid-tolerant (acid-habituated) but, in addition, neutralized medium filtrates from cultures of E. coli grown to log-phase or stationary-phase at pH 5·0 (pH 5·0 filtrates) induced acid tolerance when added to log-phase E. coli growing at pH 7·0. In contrast, filtrates from pH 7·0-grown cultures were ineffective. The pH 5·0 filtrates were inactivated by heating in a boiling water-bath but there was less activity loss at 75 °C. Protease also inactivated such filtrates, which suggested that a heat-resistant protein (or proteins) in the filtrates was essential for the induction of acid tolerance. Filtrates from cells grown at pH 5·0 plus phosphate or adenosine 3':5'-cyclic monophosphate (cAMP) were much less effective in inducing acid tolerance, while the conversion of pH 7·0-grown log-phase cells to acid tolerance by pH 5·0 filtrates was inhibited by cAMP and bicarbonate. It seems likely that the acid tolerance response (acid habituation) involved the functioning of the extracellular protein(s) as protease reduces tolerance induction if added during acid habituation. Most inducible responses are believed to involve the functioning of only intracellular reactions and components ; the present results suggest that this is not the case for acid habituation, as an extracellular protein (or proteins) is needed for induction.     

Survival and growth of Cronobacter species (Enterobacter sakazakii) in wheat-based infant follow-on formulas

Aims:  To determine the survival and growth characteristics of Cronobacter species ( Enterobacter sakazakii ) in infant wheat-based formulas reconstituted with water, milk, grape juice or apple juice during storage.
Methods and Results:  Infant wheat-based formulas were reconstituted with water, ultra high temperature milk, pasteurized grape or apple juices. The reconstituted formulas were inoculated with Cronobacter sakazakii and Cronobacter muytjensii and stored at 4, 25 or 37°C for up to 24 h. At 25 and 37°C, Cronobacter grew more (>5 log₁₀) in formulas reconstituted with water or milk than those prepared with grape or apple juices ( c. 2–3 log₁₀). The organism persisted, but did not grow in any formulas stored at 4°C. Formulas reconstituted with water and milk decreased from pH 6·0 to 4·8–5·0 after 24 h, whereas the pH of the formulas reconstituted with fruit juices remained at their initial pH values, c. pH 4·8–5·0.
Conclusions:  Cronobacter sakazakii and C. muytjensii can grow in reconstituted wheat-based formulas. If not immediately consumed, these formulas should be stored at refrigeration temperatures to reduce the risk of infant infection.
Significance and Impact of the Study:  The results of this study will be of use to regulatory agencies and infant formula producers to recommend storage conditions that reduce the growth of Cronobacter in infant wheat-based formulas.     

Anaemia and salmonid swimming performance: the potential effects of sub‐lethal sea lice infection

The effect of two known rates of repeated blood loss on rainbow trout Oncorhynchus mykiss swimming performance was measured and blood‐feeding rates of sea lice Lepeophtheirus salmonis were calculated to predict the point at which blood ingestion causes anaemia in infected fish. Known quantities of blood were sampled from rainbow trout over a 5 day period followed by critical swimming performance ( U _crit) testing. A predictive equation was developed using masses of blood‐feeding sea lice and host blood loss calculated for increasing levels of sea lice infection. Blood loss of 8% total blood volume caused a decrease in U _crit for rainbow trout. Total blood volume losses of 3·2% reduced erythrocyte stores, but did not affect fish swimming performance. The predictive feeding rate model suggests that 15–25% of the tissue consumed by sea lice is blood. This consumption of blood at higher sub‐lethal infection levels (≥0·5 sea lice g⁻¹) may cause anaemia and a further decrease in swimming performance. Anaemia would compound the osmotic balance problems due to infection and potentially precipitate the morbidity seen at lethal sea lice levels (0·75–1·0 lice g⁻¹).     

Tilapia are able to withstand long-term exposure to low environmental pH, judged by their energy status, ionic balance and plasma cortisol

Tilapia Oreochromis mossambicus were exposed to water at pH 4·0 for 37 days. The water was acidified slowly over 6 h enabling the animals to acclimate and preventing damage of the gill epithelium. Additional stressors, e.g. aluminium ions and handling stress, were avoided. No mortality or decreased food consumption was observed during the exposure period. No significant changes were observed between the control and acid exposed groups for the energy rich compounds and related parameters, i.e. the adenylate energy charge, the pool of total adenine nucleotides, and the IMP load of white muscle and liver, indicating maintenance of homeostasis. Moreover, there were no significant differences between control groups and acidified groups at 3, 17 and 37 days for plasma sodium, chloride, cortisol and glucose, implying that ionic balance was maintained and that there was no activation of the pituitary-interrenal axis. It is concluded that tilapia can acclimate to water at pH 4·0 when the acidification rate is slow and additional stressors are avoided.     

Habituation to normally lethal acidity by prior growth of Escherichia coli at a sub-lethal acid pH value

Both Col^- and ColV, I-K94⁺ strains of Escherichia coli , grown at pH 7–0, failed to grow after relatively short periods of exposure to pH 3·0 or 3·5. After growth in exposure medium initially at pH 5·0, both strains were almost unaffected by exposure to such acid pH values. Addition of catalase to nutrient agar only slightly increased plating efficiency after acid treatment and very slightly reduced the difference in survival, after acid treatment, between organisms grown from pH 5·0 and those grown from pH 7·0. Accordingly, acid resistance of organisms grown from pH 5·0 is not chiefly due to greater resistance to hydrogen peroxide already present in nutrient media.     

Effects of sporulation pH on the heat resistance and the sporulation of Bacillus cereus

Spores of Bacillus cereus ATCC 7004, 4342 and 9818 were obtained in nutrient agar at several pH from 5·9 to 8·3. The optimum pH for sporulation was around 7, but good production of spores was obtained in the range 6·5–8·3. With all three strains, D -values clearly dropped with sporulation pH, decreasing by about 65% per pH unit. z -Values were not significantly modified ( P > 0·05) by this factor. Mean z -values of 7·13 °C ± 0·16 for strain 7004, 7·67 °C ± 0·04 for 4342 and 8·80 °C ± 0·64 for 9818 were obtained.     

The trout (Salmo trutta) population of the Afon Cwm, a small tributary of the Afon Dyfi, mid-Wales

Data are presented from a 10-year (1984 to 1993) study of a Salmo trutta population in the Afon Cwm, a small tributary of the Afon Dyfi, mid-Wales. The stream is a spawning and nursery area for sea trout. Growth of trout within the stream can be summarized by a von Bertalanffy growth coefficient ( K ) of 0·310, with asymptotic length (1∞) 21·6 cm and with length at age 1 of 7·6 cm. Mean population density in the whole stream varied from year to year between 0·05 and 0·60 0-group trout m⁻² and between 0·05 and 0·70 older trout m⁻². Mean biomass varied, between years, from 0·1 to 3·5 g m⁻² for 0-group and from 1·3 to 10·4g m⁻² for older trout. Loss between 3 and 5 months of age appeared to be proportionate at about 50 to 60% and instantaneous loss rate from 5 to 53 months of age varied from 0·04 to 0·10 month⁻¹ and was positively correlated with cohort number at 3 months of age. Production between 3 and 53 months of age varied between cohorts from 3 to 8 g m ⁻² live weight.     

Factors affecting production and activity of phospholipase C by Yersinia enterocotttica

The production of phospholipase C by Yersinia enterocolitica strain SG was optimum at 37†C at pH 6·5. No enzyme activity could be detected when the organism was grown at extreme pH values (pH > 8·5 or <5·0). The enzyme production was maximum when the organism was grown under static conditions in TSB medium. All solvents and salts inhibited the enzyme activity, whereas loss of activity was 95% in presence of methanol (20%) and 99% in presence of sodium azide (0·2 mol/l). The enzyme activity was increased twofold in the presence of cyste-ine and decreased by 98% in the presence of sodium perchlorate (0·2 mol/1).