Heterologous microarray experiments used to identify the early gene response to heat stress in a coral reef fish

Coral reef fishes are expected to experience rising sea surface temperatures due to climate change. How well tropical reef fishes will respond to these increased temperatures and which genes are important in the response to elevated temperatures is not known. Microarray technology provides a powerful tool for gene discovery studies, but the development of microarrays for individual species can be expensive and time-consuming. In this study, we tested the suitability of a Danio rerio oligonucleotide microarray for application in a species with few genomic resources, the coral reef fish Pomacentrus moluccensis. Results from a comparative genomic hybridization experiment and direct sequence comparisons indicate that for most genes there is considerable sequence similarity between the two species, suggesting that the D. rerio array is useful for genomic studies of P. moluccensis. We employed this heterologous microarray approach to characterize the early transcriptional response to heat stress in P. moluccensis. A total of 111 gene loci, many of which are involved in protein processing, transcription, and cell growth, showed significant changes in transcript abundance following exposure to elevated temperatures. Changes in transcript abundance were validated for a selection of candidate genes using quantitative real-time polymerase chain reaction. This study demonstrates that heterologous microarrays can be successfully employed to study species for which specific microarrays have not yet been developed, and so have the potential to greatly enhance the utility of microarray technology to the field of environmental and functional genomics.

No effect of mild heat stress on the regulation of carbohydrate metabolism at the onset of exercise

To investigate the influence of heat stress on the regulation of skeletal muscle carbohydrate metabolism, six active, but not specifically trained, men performed 5 min of cycling at a power output eliciting 70% maximal O(2) uptake in either 20 degrees C (Con) or 40 degrees C (Heat) after 20 min of passive exposure to either environmental condition. Although muscle temperature (T(mu)) was similar at rest when comparing trials, 20 min of passive exposure and 5 min of exercise increased (P < 0.05) T(mu) in Heat compared with Con (37.5 +/- 0.1 vs. 36.9 +/- 0.1 degrees C at 5 min for Heat and Con, respectively). Rectal temperature and plasma epinephrine were not different at rest, preexercise, or 5 min of exercise between trials. Although intramuscular glycogen phosphorylase and pyruvate dehydrogenase activity increased (P < 0.05) at the onset of exercise, there were no differences in the activities of these regulatory enzymes when comparing Heat with Con. Accordingly, glycogen use in the first 5 min of exercise was not different when comparing Heat with Con. Similarly, no differences in intramuscular concentrations of glucose 6-phosphate, lactate, pyruvate, acetyl-CoA, creatine, phosphocreatine, or ATP were observed at any time point when comparing Heat with Con. These results demonstrate that, whereas mild heat stress results in a small difference in contracting T(mu), it does not alter the activities of the key regulatory enzymes for carbohydrate metabolism or glycogen use at the onset of exercise, when plasma epinephrine levels are unaltered.

Evaluation of heat stress indices using physiological comparisons in an alumina refinery in a sub-tropical climate

The production of alumina involves the use of a process known as the Bayer process. This method involves the digestion of raw bauxite in sodium hydroxide at temperatures around 250°C. The resultant pregnant liquor then goes through a number of filtering and precipitation processes to obtain the aluminium oxide crystals which are then calcined to obtain the final product. The plant is situated in a sub tropical climate in Northern Australia and this combined with the hot nature of the process results in a potential for heat related illnesses to develop. When assessing a work environment for heat stress a heat stress index is often employed as a guideline and to date the Wet Bulb Globe Temperature (WBGT) has been the recommended index. There have been concerns over the past that the WBGT is not suited to the Northern Australian climate and in fact studies in other countries have suggested this is the case. This study was undertaken in the alumina plant situated in Gladstone Queensland to assess if WBGT was in fact the most suitable index for use or if another was more applicable. To this end three indices, Wet Bulb Globe Temperature (WBGT), Heat Stress Index (HSI) and Required Sweat Rate (SWreq) were compared and assessed using physiological monitoring of heart rate and surrogate core temperature. A number of different jobs and locations around the plant were investigated utilising personal and environmental monitoring equipment. These results were then collated and analysed using a computer program written as part of the study for the manipulation of the environmental data . Physiological assessment was carried out using methods approved by international bodies such as National Institute for Occupational Safety & Health (NIOSH) and International Standards Organisation (ISO) and incorporated the use of a ‘Physiological Factor’ developed to enable the comparison of predicted allowable exposure times and strain on the individual. Results indicated that of the three indices tested, Required Sweat Rate was found to be the most suitable for the climate and in the environment of interest. The WBGT system was suitable in areas in the moderate temperature range (ie 28 to 32°C) but had some deficiencies above this temperature or where the relative humidity exceeded approximately 80%. It was however suitable as a first estimate or first line indicator. HSI over-estimated the physiological strain in situations of high temperatures, low air flows and exaggerated the benefit of artificial air flows on the worker in certain environments ie. fans.

Effect of heat stress on glucose kinetics during exercise

To identify the mechanism underlying the exaggerated hyperglycemia during exercise in the heat, six trained men were studied during 40 min of cycling exercise at a workload requiring 65% peak pulmonary oxygen uptake (V˙o 2 peak) on two occasions at least 1 wk apart. On one occasion, the ambient temperature was 20°C [control (Con)], whereas on the other, it was 40°C [high temperature (HT)]. Rates of glucose appearance and disappearance were measured by using a primed continuous infusion of [6,6-2H]glucose. No differences in oxygen uptake during exercise were observed between trials. After 40 min of exercise, heart rate, rectal temperature, respiratory exchange ratio, and plasma lactate were all higher in HT compared with Con (P < 0.05). Plasma glucose levels were similar at rest (Con, 4.54 ± 0.19 mmol/l; HT, 4.81 ± 0.19 mmol/l) but increased to a greater extent during exercise in HT (6.96 ± 0.16) compared with Con (5.45 ± 0.18;P < 0.05). This was the result of a higher glucose rate of appearance in HT during the last 30 min of exercise. In contrast, the glucose rate of disappearance and metabolic clearance rate were not different at any time point during exercise. Plasma catecholamines were higher after 10 and 40 min of exercise in HT compared with Con (P < 0.05), whereas plasma glucagon, cortisol, and growth hormone were higher in HT after 40 min. These results indicate that the hyperglycemia observed during exercise in the heat is caused by an increase in liver glucose output without any change in whole body glucose utilization.

Firefighter feedback during active cooling: A useful tool for heat stress management?

Monitoring an individual's thermic state in the workplace requires reliable feedback of their core temperature. However, core temperature measurement technology is expensive, invasive and often impractical in operational environments, warranting investigation of surrogate measures which could be used to predict core temperature. This study examines an alternative measure of an individual's thermic state, thermal sensation, which presents a more manageable and practical solution for Australian firefighters operating on the fireground. Across three environmental conditions (cold, warm, hot & humid), 49 Australian volunteer firefighters performed a 20-min fire suppression activity, immediately followed by 20min of active cooling using hand and forearm immersion techniques. Core temperature (Tc) and thermal sensation (TS) were measured across the rehabilitation period at five minute intervals. Despite the decline in Tc and TS throughout the rehabilitation period, there was little similarity in the magnitude or rate of decline between each measure in any of the ambient conditions. Moderate to strong correlations existed between Tc and TS in the cool (0.41, p<0.05) and hot & humid (0.57, p<0.05) conditions, however this was resultant in strong correlation during the earlier stages of rehabilitation (first five minutes), which were not evident in the latter stages. Linear regression revealed TS to be a poor predictor of Tc in all conditions (SEE=0.45-0.54°C) with a strong trend for TS to over-predict Tc (77-80% of the time). There is minimal evidence to suggest that ratings of thermal sensation, which represent a psychophysical assessment of an individual's thermal comfort, are an accurate reflection of the response of an individual's core temperature. Ratings of thermal sensation can be highly variable amongst individuals, likely moderated by local skin temperature. In account of these findings, fire managers require a more reliable source of information to guide decisions of heat stress management.

Short note on the possible occurrence of heat stress in roosting waders on the Banc d'Arguin, Mauritania

The temperature at the Banc d'Arguin can be very high, and may cause problems of heat regulation in waders originating from the Arctic or Antarctic. A formalin-fixed bar-tailed godwit Limosa lapponica mount exposed to the sun had a temperature of 45°C. The temperature of the mount was higher when placed in a lying position than when it was placed upright. Nevertheless observed bar-tailed godwits, dunlins Calidris alpina and Kentish plovers Charadrius alexandrinus in the high water roost were often lying on the hot sand, and most birds avoided contact with humid (cold) substrates. Only the dunlins stood in the water with their bill tucked under the scapular wing feathers. This behavioral trait is thought to conserve heat, and is also observed in the other bird species in the roosts. Heat stress is apparently not a problem for birds awaiting low tide. -M.Dean

Incubation and rearing temperature effects on Hsp70 levels and heat stress response in broilers

Incubation temperature (IT) was changed to evaluate if 6-wk-old birds become more tolerant to heat stress. After 13 d of incubation, 470 eggs were submitted to low (36.8degreesC), normal (37.8degreesC) and high (38.8degreesC) temperatures. At day 7 post-hatching, 144 birds were allocated to three rearing temperatures (48 birds/treatment): control/thermoneutral (35-24degreesC), high (33-30degreesC) or low (27-18degreesC) according to the age of the birds. Hsp70 levels in tissues of birds (1 d and 42 d), stress response (42 d) and performance were evaluated. High IT decreased brain (P < 0.01) and liver (P < 0.01) Hsp70 levels, whereas low IT decreased brain (P < 0.01) but increased heart (P < 0.01) Hsp70 levels in 1-d-old chicks. Birds incubated at a low temperature had higher (P < 0.05) feed intake (1-42d). High rearing temperature decreased feed intake (P<0.01) and liveweight (P<0.01). Colonic temperature was lower in birds incubated at a low temperature (P < 0.05) and higher in birds reared in a high temperature (P < 0.05) before heat stress. Birds reared in low temperature had higher increase in colonic temperature after heat stress (P < 0.05). Tissue Hsp70 levels were differently affected by rearing temperature, which affected broiler performance more than IT. Lower IT seemed to increase the sensitivity of birds to heat stress at market age.

Effect of heat stress or lipopolysaccharide (E-coli) injection on HSP70 levels in the liver and brain of adrenalectomized rats

Hsp70 content (ng Hsp70 mu g total protein(-1)) in the liver and brain of control and adrenalectomized male rats was investigated by Western Blotting after heat stress (40 degrees C) or endotoxin-induced fever (E. coli lipopolysaccharide injection). The increase in rectal temperature was higher after heat stress than after LPS injection, Heat stress affected Hsp70 content of the liver, but not of the brain; however adrenalectomy did not influence any parameter. These results suggest that, under these circumstances, there is no relationship between the hypothalamic-pituitary-adrenal axis and Hsp70 synthesis in liver and brain. (C) 2000 Elsevier B.V. Ltd. All rights reserved.

Effect of moderate and severe heat stress on avian embryonic Hsp70 gene expression

Stress response is a universal mechanism developed by all organisms to deal with adverse changes in the environment, which lead to the synthesis of heat shock proteins (Hsps). In this study, the effect of moderate (41degreesC) and severe (44degreesC) heat stress on Hsp70 transcript expression pattern was investigated during chicken embryogenesis. Acute exposure to severe heat stress for one hour resulted in a fifteen-fold increase in Hsp70 mRNA levels. The return of stressed embryos to normal incubation temperature resulted in Hsp70 mRNA levels five-fold higher than control after three hours and normal levels after six hours. Moderate heat stress did not induce enhancements on Hsp70 mRNA levels. The spatial expression of Hsp70 transcripts was detected in embryos under normal incubation conditions. Whole-mount in situ hybridization analysis showed that Hsp70 transcripts were constitutively present in somite and in distinct encephalic domains (predominantly in prosencephalon and mesencephalon areas) of the chicken embryo. These results showed that Hsp70 induction is dependent on incubation temperature conditions, suggesting that early chicken embryos may induce a quick emergence response to cope with severe heat stress by increasing Hsp70 mRNA levels.

Performance and carcass characteristics of broiler chickens with different growth potential and submitted to heat stress

In order to evaluate the effects of broiler genotype and of heat exposure on performance, carcass characteristics, and protein and fat accretion, six hundred one-day-old male broilers were randomly assigned in a 2 x 3 factorial arrangement, according to the following factors: genetic group (selected and non-selected broilers) and pair-feeding scheme (Ad(32) - reared under heat stress and fed ad libitum; Ad(23) - reared at thermoneutrality and fed ad libitum; Pf(23) - reared at thermoneutrality and pair fed with Ad(32)), with a total of six treatments with four replicates of 25 birds each. Independent of pair-feeding scheme, selected broilers showed better feed conversion, higher carcass yield, and lower abdominal fat deposition rate. However, as compared to non-selected broilers, they reduced more intensively feed intake when heat exposed, which promoted significant breast-yield decrease, and more pronounced changes on carcass chemical composition. These findings allows concluding that, in both genetic groups, both environmental temperature and feed-intake restriction influence abdominal fat deposition rate and other carcass characteristics; however, the impact of heat exposure on broiler performance is more noticeable on the selected line.

Water intake behavior of broiler chickens exposed to heat stress and drinking from bell or and nipple drinkers

The aspects involved in broiler water intake are not well known, despite the importance of water in animal nutrition and physiology. Water intake behavior should be taken into account when deciding on different types of drinkers. Bell and nipple drinkers are the most commonly used in commercial broiler production. Broilers were housed in cages equipped with two different drinker types and raised at two different environmental temperatures (25 and 34 ºC) to evaluate water intake behavior and volume. Broiler water intake behavior was influenced by drinker type. Birds visited bell drinkers less often, but presented higher total water intake per visit to the drinker as compared to those drinking from nipple drinkers. The results of this study suggest that both broilers drinking behavior and water intake volume should be taken into account when deciding on drinker type to equip broiler houses.

Does low-protein diet improve broiler performance under heat stress conditions?

Nutrition for broilers under high temperatures is extremely important for brazilian broiler chicken industry because the amounts of consumed nutrients and environmental temperature have great effects on bird performance and carcass quality. Among diet nutrients, protein has the highest heat increment; thus, during many years, diets with low protein level were recommended in order to reduce heat production in broiler chickens under heat stress. However, reports have shown that low-protein diets have negative effects on broiler performance when environmental temperature is high, because during heat stress, low food intake associated to a low diet protein induce amino acid deficiencies. Other studies have shown that broilers fed low-protein diets increase their energy requirement for maintenance with higher heat production. Thus, with the growth of broiler industry in tropical areas more challenges need to be faced by the farmers. So, both the ambient and nutritional conditions ought to be well managed to avoid negative effects on poultry production once they can affect the metabolism (body heat production under low temperature and body heat dissipation under high temperature) with consequence on poultry performance (meat and eggs).

Brain and hepatic Hsp70 protein levels in heat-acclimated broiler chickens during heat stress

In the present study we have investigated the effects of heat acclimation on brain and hepatic Hsp70 protein levels and body temperature of broiler chickens in response to gradual heat stress. Two groups of broilers were raised up to 47 days of age under distinct temperature conditions: thermoneutral (TN, according to bird age) or hot environmental (HS, 31-33°C). At 46 days of age, the birds reared at high ambient temperature were transferred to thermoneutrality conditions. After 18 h, these birds and the birds reared at thermoneutral temperature were submitted to gradual heat stress in a climatic chamber so that environment temperature was increased from 28 to 40ºC at a rate of 2ºC/h. Colonic temperature was measured using a thermometer sensor probe at each two hours, and hepatic and brain tissues were collected immediately after slaughter in order to assess Hsp70 protein level by Western blotting analysis. The colonic temperatures of birds reared at high temperature increased steeply during the first 2 h of heat stress (1.06ºC/h) and more slowly thereafter (0.59ºC/h). Broilers reared at thermoneutral temperature showed a small increase in the first 4 h of heat stress (0.18ºC/h) and then colonic temperature increased sharply (0.72ºC/h). Nevertheless, both groups presented similar final colonic temperature by the end of the stress period. Hsp70 levels (ng Hsp70 µg total protein-1) did not change in the liver or brain of the birds reared at high temperature. on the other hand, both liver and brain Hsp70 levels increased significantly during heat stress in the animals reared at thermoneutrality, with a higher expression of this peptide in brain tissue.

Influence of sire and sire breed (Gyr versus Holstein) on establishment of pregnancy and embryonic loss in lactating Holstein cows during summer heat stress

Heat stress has negative effects on pregnancy rates of lactating dairy cattle. There are genetic differences in tolerance to heat stress; Bos taurus indicus (B. t. indicus) cattle and embryos are more thermotolerant than Bos taurus taurus (B. t. taurus). In the present study, the effects of sire and sire breed on conception and embryonic/fetal loss rates of lactating Holstein cows during the Brazilian summer were determined. In Experiment 1, cows (n = 302) were AI after estrus detection or at a fixed-time with semen from one Gyr (B. t. indicus) or one Holstein sire (B. t. taurus). Pregnancy was diagnosed 80 days after AI. In Experiment 2, cows (n = 811) were AI with semen from three Gyr and two Holstein sires. Pregnancy was diagnosed at 30-40 and at 60-80 days after AI. Cows diagnosed pregnant at the first examination but non-pregnant at the second were considered as having lost their embryo or fetus. Data were analyzed by logistic regression. The model considered the effect of sire within breed, sire breed, days postpartum, period of lactation, and AI type (AI after estrus versus fixed-time). There was no effect of the AI type, days postpartum or milk production on conception or embryonic loss rates. The use of Gyr bulls increased pregnancy rate when compared to Holstein bulls [9.1% (60/657) versus 5.0% (23/456), respectively, P = 0.008; data from Experiments 1 and 2 combined]. Additionally, in Experiment 2, cows inseminated using semen from sire #4 (Gyr) had lower embryonic loss (10%) when compared with other B. t. indicus (35.3% and 40%) or B. t. taurus sires (18.2% and 38.5%, P = 0.03). In conclusion, the use of B. t. indicus sires may result in higher conception rates in lactating Holstein cows during summer heat stress. Moreover, sire can affect embryonic loss and selection of bulls according to this criterion may result in higher parturition rates in lactating Holstein cows. (c) 2006 Elsevier B.V. All rights reserved.