The metabolic and biochemical responses of tropical whitespotted bamboo shark Chiloscyllium plagiosum to alterations in environmental temperature

The capacity of tropical whitespotted bamboo sharks Chiloscyllium plagiosum to metabolically compensate, at both the whole‐animal and biochemical levels, to prolonged exposure to temperatures higher (30° C) and lower (20 and 15° C) than their native temperature (24·5° C) was examined. As expected, whitespotted bamboo shark oxygen consumption increased upon exposure to 30° C and decreased at 20 and 15° C. Initial changes in oxygen consumption were maintained even after months at the experimental temperature, indicating that whitespotted bamboo sharks did not compensate metabolically to the experimental temperatures. Maximal activities and thermal sensitivity of citrate synthase and lactate dehydrogenase from whitespotted bamboo shark white locomotor muscle were similar between control animals maintained at 24·5° C and those maintained at 15° C, indicating that cold‐exposed animals did not compensate at the biochemical level. Similarly, lactate dehydrogenase activity did not change following prolonged exposure to 30° C. White muscle from whitespotted bamboo sharks maintained at 30° C had significantly lower citrate synthase activity than did control animals. This result was surprising given the lack of metabolic compensation at the whole‐animal level. Overall, whole‐animal oxygen consumption measurements supported the hypothesis that animals from thermally stable environments lacked the capacity to metabolically compensate to altered temperatures. Enzymatic results, however, suggested that the metabolic potential of muscle could change following temperature acclimation even in the absence of metabolic compensation at the whole‐animal level.     

Stress induced by hooking, net towing, elevated sea water temperature and air in sablefish: lack of concordance between mortality and physiological measures of stress

In a series of laboratory studies designed to simulate bycatch processes, sablefish Anoplopoma fimbria were either hooked for up to 24 h or towed in a net for 4 h and then subjected to an abrupt transfer to elevated sea water temperature and air. Mortality did not result from hooking or net towing followed by exposure to air, but increased for both capture methods as fish were exposed to elevated temperatures, reflecting the magnifying effect of elevated temperature on mortality. Hooking and exposure to air resulted in increased plasma cortisol and lactate concentrations, while the combination of hooking and exposure to elevated temperature and air resulted in increased lactate and potassium oncentrations. In fish that were towed in a net and exposed to air, cortisol, lactate, potassium and sodium concentrations increased, but when subjected to elevated temperature and air, no further increases occurred above the concentrations induced by net towing and air, suggesting a possible maximum of the physiological stress response. The results suggest that caution should be exercised when using physiological measures to quantify stress induced by capture and exposure to elevated temperature and air, that ultimately result in mortality, since the connections between physiological stress and mortality in bycatch processes remain to be fully understood.     

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).     

Contrasting behavioral responses to cold temperatures by two marine fish species during their pelagic juvenile interval

Motor activity of juvenile walleye pollock, Theragra chalcogramma, and sablefish, Anoplopoma fimbria, was monitored in the laboratory during high and low light levels under a changing temperature regime over a 5 d period. Water temperatures were ambient (12 °C) for the first 24 h of observation, rapidly lowered to 3 °C for the next 48 h, then raised back to 12 °C for the final 48 h. We hypothesized that the fishes' behavior would either follow a simple bioenergetic response of lowered activity associated with reduced metabolic rates at the colder temperature, or an avoidance response, with increased activity at decreased temperatures. Results for walleye pollock were consistent with a bioenergetic response, with activity decreasing in the presence of cold water under both high and low light levels, then returning to initial levels when temperatures increased. The response of sablefish, in contrast, indicated avoidance of cold temperatures, depending on light level. During high light, when sablefish were typically highly active, cold water induced a slight but insignificant decrease in activity. At low light, however, the presence of cold water caused a marked increase in sablefish movement through the experimental tanks, with a seven fold increase in the measured index of activity. When water temperatures were raised back to 12 °C, sablefish activity at low light returned to its normal, minimal level. The sharp increase in activity of sablefish in cold water, followed by a decrease in activity when the temperature was raised to pre-test levels, is suggestive of an avoidance response. The contrasting responses of the two species to thermal changes are consistent with their separate life history patterns and natural distributions.     

Modelling the time–temperature relationship in cold injury and effect of high-temperature interruptions on survival in a chill-sensitive collembolan

1. Temperature- and time-dependent mortalities were studied and modelled in insects exposed in regimes with constant and alternating temperatures. In these experiments, freezing was not a cause of death.
2. Survival rates at a range of constant low temperatures (– 5 to + 1 °C) and for different exposure periods (1–14 days) were measured in the summer acclimated springtail Orchesella cincta .
3. Daily interruptions of the cold exposure with short intervals at high temperature reduced mortality or slowed the increase of mortality. This effect was stronger at higher temperature (19 vs 5 and 12 °C) and increased with the duration of the interruption (0·25–2 h).
4. The injury was reversible when the cold exposure was limited to 2 days.
5. Survival in desiccated animals (14% water loss) was reduced.
6. It is suggested that the mortality of summer acclimated springtails is caused by a complex metabolic disorder and membrane changes at low temperatures.     

Behavioural thermoregulation by subyearling fall (autumn) Chinook salmon Oncorhynchus tshawytscha in a reservoir

This study investigated behavioural thermoregulation by subyearling fall (autumn) Chinook salmon Oncorhynchus tshawytscha in a reservoir on the Snake River, Washington, U.S.A. During the summer, temperatures in the reservoir varied from 23° C on the surface to 11° C at 14 m depth. Subyearlings implanted with temperature-sensing radio transmitters were released at the surface at temperatures >20° C during three blocks of time in summer 2004. Vertical profiles were taken to measure temperature and depth use as the fish moved downstream over an average of 5·6–7·2 h and 6·0–13·8 km. The majority of the subyearlings maintained average body temperatures that differed from average vertical profile temperatures during most of the time they were tracked. The mean proportion of the time subyearlings tracked within the 16–20° C temperature range was larger than the proportion of time this range was available, which confirmed temperature selection opposed to random use. The subyearlings selected a depth and temperature combination that allowed them to increase their exposure to temperatures of 16–20° C when temperatures <16 and >20° C were available at lower and higher positions in the water column. A portion of the subyearlings that selected a temperature c. 17·0° C during the day, moved into warmer water at night coincident with an increase in downstream movement rate. Though subyearlings used temperatures outside of the 16–20° C range part of the time, behavioural thermoregulation probably reduced the effects of intermittent exposure to suboptimal temperatures. By doing so, it might enhance growth opportunity and life-history diversity in the population of subyearlings studied.     

Hypoxia tolerance in Atlantic cod

Oxygen saturation levels that killed 50 and 5% of cod Gadus morhua over 96 h averaged 21·2 and 27·7%, respectively. No fish survived at 10% saturation and only a few survived at 16% saturation, whereas no mortality occurred at 34 and 40% oxygen saturation. Since metabolic rate and oxygen consumption increase with increasing temperature, we hypothesized that cod would be less tolerant to hypoxic conditions at 6 than at 2° C. However, temperature (2 and 6° C) had no measurable impact on cod survival. Small (mean & S.D.; 45·2 ± 4·2 cm) and large (57·5 ± 3·8 cm) cod had the same tolerance to hypoxia. At the end of the experiments, hypoxia had a significant effect on blood haematocrit, mean cellular haemoglobin content, liver lactate, plasma glucose and plasma lactate, but accounted for only a small fraction (< 10%) of the variation, except for plasma lactate which exhibited a strong response with concentrations increasing progressively with decreasing levels of oxygen saturation. Temperature had a significant effect on most variates in normoxia and hypoxia. Variates also affected by oxygen level showed significant interactions between oxygen and size or temperature effects. However, these interactions accounted for only a small proportion of the variation. Physiological parameters indicated that extending the duration of our tests beyond 96 h would not have changed our estimates of the lethal thresholds. Hypoxic conditions are a permanent feature of the deep waters of the Gulf of St Lawrence. This study shows that a significant portion of the benthic habitats in the Gulf are uninhabitable for cod which would be expected to avoid waters below 28% oxygen saturation.     

The critical thermal limits for juvenile Arctic charr Salvelinus alpinus

The chief objective was to determine the critical thermal limits for alevins, fry and parr of Arctic charr, Salvelinus alpinus , (L.) from four races living in Windermere (northwest England). The experimental fish were reared in a hatchery but were the progeny of wild parents. As comparisons between tethal temperatures at four acclimation temperatures (5, 10, 15, 20° C) revealed few significant racial differences, the data were pooled to estimate the lethal values for survival over 7 days (incipient lethal temperature) and over only 10 min (ultimate lethal temperature) for each life stage. Upper lethal values increased with acclimation temperatures for alevins but this effect was negligible for fry and parr, Alevins were generally less tolerant than fry and parr at lower, but not higher, acclimation temperatures; e.g. after acclimation at 5° C, mean upper ultimate values were 23·3, 25·1 and 25·7° C and mean upper incipient values were 18·7, 21·5 and 21·5° C for alevins, fry and parr respectively; after acclimation at 20° C, mean upper ultimate and incipient values were 26·2, 26·1 and 26·6° C and 20·8, 20·8 and 21·6° C for alevins, fry and parr respectively. The area of the temperature tolerance polygon (expressed as ° C²) for juvenile Arctic charr is amongst the lowest recorded for salmonids; being 409, 439 and 461° C² for alevins, fry and parr respectively. These low values are due to lower upper tolerance limits, not high lower tolerance limits; the latter being close to 0° C (<1°C for parr and fry, <0·3° C for alevins) at all acclimation temperatures. Arctic charr are therefore amongst the least resistant of salmonids to high temperatures but probably the most resistant to low temperatures.     

The influence of thermal parameters on the acclimation responses of pinfish Lagodon rhomboides exposed to static and decreasing low temperatures

Pinfish Lagodon rhomboides acclimation rates were determined by modelling changes in critical thermal minimum ( T _{crit min}, ° C) estimates at set intervals following a temperature decrease of 3–4° C. The results showed that pinfish gained a total of 3·7° C of cold tolerance over a range of acclimation temperatures ( T _acc, ° C) from (23–12° C), that cold tolerance increased with exposure time to the reduced temperature at all T _acc, but that the rate of cold tolerance accruement (mean 0·14° C day⁻¹) was independent of T _acc. A highly significant ( P < 0·001) multivariate predictive model was generated that described the acclimation rates and thermal tolerance of pinfish exposed to reduction in water temperature: log₁₀ T _{crit min}= 0·41597 − 0·01704 T _acc+ 0·04320 T _plunge− 0·08376[log₁₀ ( t + 1)], where T _plunge is plunge temperature (° C) and t is the time (days). A comparison of the present data, with acclimation rate data for other species, suggests that factors such as latitude or geographic range may play a more important role than ambient temperature in determining cold acclimation rates in fishes.     

The influence of environmental temperature and oxygen concentration on the recovery of largemouth bass from exercise: implications for live–release angling tournaments

The impact of variation in water temperature and dissolved oxygen on recovery of largemouth bass Micropterus salmoides from exercise was examined. For this, largemouth bass were first exercised and recovered for either 1, 2 or 4 h at ambient water temperatures (25° C) in fully oxygenated water. Results showed that exercise forced fish to utilize anaerobic metabolism to meet energy demands, and resulted in reductions in anaerobic energy stores adenosine triphosphate (ATP), Phosphocreatine (PCr) and glycogen. Exercise also resulted in a seven‐fold increase in lactate within white muscle. After 2 h of recovery in oxygenated water at acclimation temperature, physiological recovery from exercise was under way, and by 4 h most variables examined had returned to control levels. Next, largemouth bass were exercised at ambient temperatures and recovered for 2 h in environments with either elevated temperature (32° C), reduced temperature (14 and 20° C), hypoxia or hyperoxia. Both elevated and reduced temperature impaired recovery of tissue lactate and tissue ATP relative to fish recovered in water at acclimation temperature, while hyperoxic water impaired recovery of tissue ATP. Moderately hypoxic waters impaired the recovery of plasma glucose, plasma lactate and tissue PCr relative to fish recovered in fully oxygenated water. Results from this study are discussed in the context of critical oxygen and temperature guidelines for largemouth bass. In addition, several recommendations are made concerning remedial treatments used in livewells (tanks) during angling tournaments when fish are recovering from exercise associated with angling.     

Early development of the skull of Sander lucioperca (L.) (Teleostei: Percidae) relating to growth and mortality

The early osteological development of the skull (chondrocranium and osteocranium) of the pikeperch Sander lucioperca was studied. Specimens were reared at two temperatures, 15·5 and 18·0° C, from hatching until 47 and 43 days after fertilization (DAF), respectively. The skeletal elements characteristic for the different developmental stages were the same at both rearing temperatures, but pikeperch reared at 15·5° C reached the developmental stages later. The formation of the functional complexes, the neurocranium, jaws and suspensorium, branchial basket and hyoid arch, was evaluated chronologically. The focus was on skull development during several functional changes: at hatching, at the shift from endogenous to mixed feeding, the shift to exclusively exogenous feeding and upon reaching the final prey-capture mechanism. Growth in total length differed between fishes reared at the two temperatures, except during a phase of very slow growth from the end of the embryonic stage until the second larval stage. The latter phase, in which most of the bony elements of the viscerocranium started to form, was marked by high mortality. When exogenous feeding began, the growth rates at both temperatures increased distinctly and the first bony elements were formed in the neurocranium. Specimens reared at 18·0° C grew continuously, but those at 15·5° C showed a second period of slow growth and high mortality. Fish reared at 18·0° C reached the successive larval stages distinctly earlier than fish reared at 15·5° C.     

Stress and survival of small pike-perch Sander lucioperca (L.) after trawling and chilling

The mortality and stress responses (plasma cortisol concentration) of undersized (total length, L _T, <370 mm) pike-perch Sander lucioperca caught as by-catch in a trawl fishery were assessed. The effects of three different holding methods on pike-perch caught by trawl were compared: (1) fish that were transferred to ambient-temperature water (15·0–21·4° C) immediately after capture and not exposed to chilling water, (2) fish held in chilling water for 10 min and (3) fish exposed to chilling water for 2 h. The sample fish were held and monitored in individual plastic restrainers at ambient-water temperature for 48 h after each of the handling procedures. Total mortality was similar for fish held in chilling tanks for 10 min (27·2% total mortality) compared to fish that were only held in ambient-temperature water (28·2% mortality). Total mortality reached 91·3% when fish were held in chilled water for 2 h. Mortality and plasma cortisol concentrations correlated inversely with size (96–368 mm L _T) and directly with lake-water temperature (15·0–21·4° C) after treatment. In addition, the effect of chilling was observed in elevated cortisol concentration and delayed start of recovery. The results show that quickly removing (<10 min) undersized pike-perch from chilled water could markedly improve the survival of released fish. Due to delayed recovery, however, fish should be allowed to recover (30–40 min) in a deck tank before release.     

Oxygen consumption, ammonia excretion and protein use in response to thermal changes in juvenile Atlantic salmon Salmo salar

Experiments were designed to examine the effects of various temperature challenges on oxygen consumption and ammonia excretion rates and protein utilization in juvenile Atlantic salmon Salmo salar . Fish acclimated to 15° C were acutely and abruptly exposed to either 20 or 25° C for a period of 3 h. To simulate a more environmentally relevant temperature challenge, a third group of fish was exposed to a gradual increase in temperature from 15 to 20° C over a period of 3 h ( c. 1·7° C h⁻¹). Oxygen consumption and ammonia excretion rates were monitored before, during and after the temperature shift. From the ammonia excretion and oxygen consumption rates, protein utilization rates were calculated. Acute temperature changes (15–20° C or 15–25° C) caused large and immediate increases in the oxygen consumption rates. When the temperature was gradually changed ( i.e. 1·7° C h⁻¹), however, the rates of oxygen consumption and ammonia excretion were only marginally altered. When fish were exposed to warmer temperatures ( i.e. 15–20° C or 15–25° C) protein use generally remained at pre-exposure (15° C) levels. A rapid transfer back to 15° C (20–15° C or 25–15° C) generally increased protein use in S. salar . These results indicate that both the magnitude and the rate of temperature change are important in describing the physiological response in juvenile salmonids.     

The effects of temperature on muscle pH, adenylate and phosphogen concentrations in Oreochromis alcalicus grahami, a fish adapted to an alkaline hot-spring

The concentrations of phosphorylcreatine (PCr), adenosine triphosphate (ATP), adenosine diphosphate (ADP), adenosine monophosphate (AMP), inorganic phosphate (Pi), pyruvate and lactate were determined in freeze-clamped fast muscle samples from Oreochromis alcalicus grahami a fish adapted to extreme alkalinity (∼ pH 10·0) and high temperatures (Lake Magadi, Kenya). Specimens were analysed from both geothermally heated hotsprings (35–37°C) and from isolated cool pools (28°C) and from stocks acclimated to 20°C in the laboratory. The ratios of (ATP)/(ADP) and (ATP)/(ADP) (Pi) decreased with increasing body temperature consistent with an increase in glycolysis and tissue respiration rates, respectively. The apparent equilibrium constant of creatine kinase (K_CK), (creatine) (ATP)/(phosphorylcreatine) (ADP) was found to decrease with increasing temperature: 20·2 (20°C), 13·9 (28°C), 8·0 (37°C). A near constant muscle and blood pH (or slight increase in alkalinity with higher temperatures) was found regardless of body temperature (Blood pH 7·64, 7·74, muscle pH 7·27, 7·51 at 20°C and 35°C, respectively). These results are consistent with an unusual pattern of acid-base regulation in this species.     

The relative importance of temperature and diet to larval development and adult size of the winter stonefly, Soyedina carolinensis (Piecoptera: Nemouridae)

SUMMARY. 1. Soyedina carolinensis Claassen, a leaf shredding stonefly, was reared in a series of three laboratory experiments from early instar to adult on different species of deciduous leaves and at various constant and fluctuating temperature regimes.
2. Experiment 1, which involved rearing larvae on fourteen different leaf diets at ambient stream temperatures, showed that diet significantly affected larval growth and adult size but did not affect overall developmental time.
3. Experiment 2, which involved rearing larvae on five different leaf diets at each of three fluctuating temperature regimes (viz ambient White Clay Creek (WCC), ambient WCC+3°C, and ambient WCC+6°C), showed that: (i) adding 6°C to the normal temperature regime of WCC was lethal to 99% of the larvae regardless of diet; and (ii) warming WCC by 3°C did not affect developmental time but did significantly reduce adult size relative to adults reared at WCC temperatures on certain diets.
4. Experiment 3, which involved rearing larvae on five different leaf diets at each of five constant temperatures (viz 5, 10, 15, 20, 25°C), showed that: (i) temperature significantly affected the mortality, growth, and development time of larvae whereas diet only affected larval growth and mortality; (ii) temperatures at or near 10°C yielded maximum larval growth and survival for most diets; (iii) at 5°C, larval mortality was high and growth was low resulting in a few small adults for most diets; (iv) larval mortality was at or near 100% at 15°C regardless of diet; and (v) no larvae survived at 20 and 25°C.     

Early development and growth of the laboratory reared north-east Atlantic mackerel Scomber scombrus L.

The early development, growth and morphological changes of mackerel Scomber scombrus were investigated at different incubation temperatures (8, 10, 13, 15 and 18° C). Details on the early life history are illustrated with special reference to morphological transformations. Culture techniques to rear larval mackerel stages are described using laboratory cultured foods. Artificially fertilized eggs were hatched after 80·6 h at 18·4° C and 256·8 h at 8·7° C. The standard length ( L _S) of the individuals at first feeding was 4·71 ± 0·18 mm. Four mortality critical periods and cannibalistic behaviour were identified. A maximum average larval size of 37·5 ± 4·41 mm L _S was attained 30 days post-hatch (dph) at 18·4° C. Development and growth were affected significantly by temperature during both endogenous and exogenous feeding periods. Larvae grew more rapidly at high, than at low temperatures. Daily specific growth rate (in mass) ranged from 2·4% at 10·6° C to 16·9% at 18·4° C. Likewise, average growth rate (in length) ranged from 0·05 mm day⁻¹ at 8·4° C to 0·37 mm day⁻¹ at 18·4° C. The allometric relationship of L _S, with several body measurements was not affected by temperature. Comparison with larvae collected in the Bay of Biscay did not show any significant difference in the dry mass and L _S relationship; conversely, the growth rate in length differed significantly between both laboratory and field conditions. The trends observed in the laboratory are described in relation to some aspects of the year-class strength regulation.     

Elevated red myotomal muscle temperatures in the most basal tuna species, Allothunnus fallai

The present study tested the hypothesis that Allothunnus fallai can elevate its slow-oxidative red myotomal muscle (RM) temperature. Measurements on 30 A. fallai (750–850 mm fork length) captured by hook and line off the coast of southern New Zealand revealed that RM temperatures are elevated by mean ± s . d . 8·1 ± 1·3° C (range 6·7–10·0° C) above the mean ± s . d . ambient sea surface temperature 15·3 ± 0·8° C (range 14·3 to 16·4° C). These data provide evidence that the vascular modifications to the central circulation of A. fallai act as a counter-current heat exchanger and that RM heat conservation is a character state present in all extant tuna species.     

Temperature-induced changes of survival, development and yolk partitioning in Chondrostoma nasus

Fertilized Chondrostoma nasus eggs were incubated at 10, 13, 16 and 19° C until full resorption of the yolk sac. High survival was observed at 10–16° C (89–92% at the onset of external feeding), whereas at 19) C survival was depressed (76%). The time at which 5, 50 and 95% of individuals had hatched, filled the swim bladder, ingested the first food and fully resorbed the yolk sac was determined. An increase in temperature accelerated development and made it more synchronous. Within the period from fertilization to hatching embryonic development was theoretically arrested (t_{0 dev}) at 8·8° C, and growth was arrested (t_0gr) at 8·86° C. For the whole endogenous feeding period (from fertilization to full yolk resorption) the amount of matter transformed into tissue was temperature independent between 10° and 19° C. Respiration increased exponentially with age; the respiration increase was faster at higher temperatures, but, in general, metabolic expenditures of C. nasus were low. As a consequence, the efficiency of utilizing yolk energy for growth was high as compared with other fish species (57% during the whole endogenous feeding period); it was temperature independent. However, time was used less efficiently at low temperatures, increasing a risk of predation. Within the endogenous feeding period a shift from lower to higher temperatures for optimal yolk utilization efficiency was observed. The temperatures optimal for survival and energetic performance seem to be 13–16° C for egg incubation and 15–18° C for rearing of yolk-feeding larvae. Chondrostoma nasus is a potential candidate for aquaculture for restocking purposes.     

Moist-heat disinfection of pathogenic Acanthamoeba cysts

The effect of moist-heat in the disinfection of pathogenic Acanthamoeba cysts was investigated. Temperatures of 56°C or 60°C were not effective in killing cysts of A. polyphaga even after a contact time of 60 min, A 4 log reduction in viability was achieved within 15 min at 65°C and 2 min at 70°C giving decimal reduction rates (D-values) of 3·75 min and 30 s respectively. The ability of a commercial moist-heat contact lens disinfection unit to kill 1 times 10⁵ cysts of Acanthamoeba isolated from contact lens storage cases was also shown. Holding temperatures inside the cases of 65°C for 5·5 min and 70°C (the maximum temperature obtained) for 1 min were recorded during the disinfection cycle.