Experimental considerations of acute heat stress assays to quantify coral thermal tolerance.

Understanding the distribution and abundance of heat tolerant corals across seascapes is imperative for predicting responses to climate change and to support novel management actions. Thermal tolerance is variable in corals and intrinsic and extrinsic drivers of tolerance are not well understood. Traditional experimental evaluations of coral heat and bleaching tolerance typically involve ramp-and-hold experiments run across days to weeks within aquarium facilities with limits to colony replication. Field-based acute heat stress assays have emerged as an alternative experimental approach to rapidly quantify heat tolerance in many samples yet the role of key methodological considerations on the stress response measured remains unresolved. Here, we quantify the effects of coral fragment size, sampling time point, and physiological measures on the acute heat stress response in adult corals. The effect of fragment size differed between species (Acropora tenuis and Pocillopora damicornis). Most physiological parameters measured here declined over time (tissue colour, chlorophyll-a and protein content) from the onset of heating, with the exception of maximum photosynthetic efficiency (Fv/Fm) which was surprisingly stable over this time scale. Based on our experiments, we identified photosynthetic efficiency, tissue colour change, and host-specific assays such as catalase activity as key physiological measures for rapid quantification of thermal tolerance. We recommend that future applications of acute heat stress assays include larger fragments (> 9 cm2) where possible and sample between 10 and 24 h after the end of heat stress. A validated high-throughput experimental approach combined with cost-effective genomic and physiological measurements underpins the development of markers and maps of heat tolerance across seascapes and ocean warming scenarios.

Cardiovascular, muscular, and skeletal adaptations to recreational team handball training: a randomized controlled trial with young adult untrained men.

PURPOSE: The prevalence of lifestyle diseases has escalated, and effective exercise training programmes are warranted. This study tested the hypothesis that regular participation in small-sided team handball training could provide beneficial health effects on cardiovascular, skeletal, and muscular parameters in young adult untrained men. METHOD: Twenty-six untrained 20-30-year-old men were randomly allocated to either a team handball training group (HG; n = 14), which completed 1.9 ± 0.3 training sessions per week over 12 weeks, or an inactive control group (CG; n = 12). Physiological training adaptations were assessed pre- and post interventions by DXA scans, blood samples, muscle biopsies, and physical tests. RESULTS: The average heart rate during training was equivalent to 84 ± 4% of maximal heart rate. Compared to CG, HG displayed significant increases in VO2max (11 ± 6%), proximal femur bone mineral density (2 ± 1%), whole-body bone mineral content (2 ± 1%), intermittent endurance performance (32 ± 16%), incremental treadmill test performance (16 ± 7%) and muscle citrate synthase activity (22 ± 28%) as well as decreases in total fat mass (7 ± 7%) and total fat percentage (6 ± 7%) (all p < 0.05). There were no significant changes in muscle mass, blood pressure, resting heart rate, muscle hydroxyl-acyl-dehydrogenase activity, or blood lipids (all p > 0.05). CONCLUSION: Participation in regular recreational team handball training was associated with positive cardiovascular, skeletal, and muscular adaptations, including increased maximal oxygen uptake, increased muscle enzymatic activity, and improved bone mineralization as well as lower fat percentage. These findings suggest that recreational team handball training may be an effective health-promoting activity for young adult men.

Correction to: Cardiovascular, muscular, and skeletal adaptations to recreational team handball training: a randomized controlled trial with young adult untrained men.

The author would like to correct the errors in the publication of the original article. The corrected details are given below for your reading.

Effects of long-term football training on the expression profile of genes involved in muscle oxidative metabolism.

We investigated whether long-term recreational football training affects the expression of health-related biochemical and molecular markers in healthy untrained subjects. Five untrained healthy men trained for 1 h 2.4 times/week for 12 weeks and 1.3 times/week for another 52 weeks. Blood samples and a muscle biopsy from the vastus lateralis were collected at T0 (pre intervention) and at T1 (post intervention). Gene expression was measured by RTqPCR on RNA extracted from muscle biopsies. The expression levels of the genes principally involved in energy metabolism (PPARγ, adiponectin, AMPKα1/α2, TFAM, NAMPT, PGC1α and SIRT1) were measured at T0 and T1. Up-regulation of PPARγ (p < 0.0005), AMPKα1 (p < 0.01), AMPKα2 (p < 0.0005) and adiponectin was observed at T1 vs T0. Increases were also found in the expression of TFAM (p < 0.001), NAMPT (p < 0.01), PGC1α (p < 0.01) and SIRT1 (p < 0.01), which are directly or indirectly involved in the glucose and lipid oxidative metabolism. Multiple linear regression analysis revealed that fat percentage was independently associated with NAMPT, PPARγ and adiponectin expression. In conclusion, long-term recreational football training could be a useful tool to improve the expression of muscle molecular biomarkers that are correlated to oxidative metabolism in healthy males.

Effect of football or strength training on functional ability and physical performance in untrained old men.

The effects of 16 weeks of football or strength training on performance and functional ability were investigated in 26 (68.2 ± 3.2 years) untrained men randomized into a football (FG; n = 9), a strength training (ST; n = 9), or a control group (CO; n = 8). FG and ST trained 1.6 ± 0.1 and 1.5 ± 0.1 times per week, respectively, with higher (P < 0.05) average heart rate (HR) (∼140 vs 100 bpm) and time >90%HRmax (17 vs 0%) in FG than ST, and lower (P < 0.05) peak blood lactate in FG than ST (7.2 ± 0.9 vs 10.5 ± 0.6 mmol/L). After the intervention period (IP), VO2 max (15%; P < 0.001), cycle time to exhaustion (7%; P < 0.05), and Yo-Yo Intermittent Endurance Level 1 performance (43%; P < 0.01) were improved in FG, but unchanged in ST and CO. HR during walking was 12% and 10% lower (P < 0.05) in FG and ST, respectively, after IP. After IP, HR and blood lactate during jogging were 7% (P < 0.05) and 30% lower (P < 0.001) in FG, but unchanged in ST and CO. Sit-to-stand performance was improved (P < 0.01) by 29% in FG and 26% in ST, but not in CO. In conclusion, football and strength training for old men improves functional ability and physiological response to submaximal exercise, while football additionally elevates maximal aerobic fitness and exhaustive exercise performance.

Street football is a feasible health-enhancing activity for homeless men: biochemical bone marker profile and balance improved.

This case-control study investigated the feasibility of street football as a health-enhancing activity for homeless men, specifically the musculoskeletal effects of 12 weeks of training. Twenty-two homeless men participated in the football group (FG) and 10 served as controls (C). Plasma osteocalcin, TRACP5b, leptin, and postural balance were measured, and whole-body DXA scanning was performed. The attendance rate was 75% (2.2 ± 0.7 sessions per week). During 60 min of training, the total distance covered was 5534 ± 610 m, with 1040 ± 353, 2744 ± 671, and 864 ± 224 m covered by high-intensity, low-intensity, and backwards/sideways running, respectively. In FG, osteocalcin increased by 27% from 20.1 ± 11.1 to 25.6 ± 11.8 ng/mL (P = 0.007). Postural balance increased by 39% (P = 0.004) and 46% (P = 0.006) in right and left leg. Trunk bone mineral density increased by 1.0% from 0.959 ± 0.095 to 0.969 ± 0.090 g/cm(2) (P = 0.02). No effects were observed in C. In conclusion, street football appears to be a feasible training activity with musculoskeletal health benefits for homeless men. The attendance rate and the training intensity were high, and 12 weeks of training resulted in a substantial anabolic response in bone metabolism. Postural balance improved markedly, and the overall risk of falling, and hospitalization due to sudden trauma, could be reduced by street football for homeless men.

Physiological response and activity profile in recreational small-sided football: no effect of the number of players.

We examined the effect of the number of players on the activity profile and physiological response to small-sided recreational football games with fixed relative pitch size. Twelve untrained men (age: 33.0 ± 6.4 (± standard deviation) years, fat%: 22.4 ± 6.1%, VO2 max: 43.3 ± 5.2 mL/min/kg) completed three football sessions of 4 times 12 min with 3v3, 5v5, or 7v7 in a randomized order. Pitch sizes were 80 m(2) per player. Activity profile (10 Hz global positioning system), heart rate (HR), and rating of perceived exertion (RPE) were measured, and blood samples were collected before and during games. Average HR was 84.1 ± 3.9, 84.5 ± 5.0, and 82.8 ± 5.1 %HRmax for 3v3, 5v5, and 7v7, respectively, with no difference between game formats. High blood lactate (5.9 ± 2.9, 5.9 ± 2.4, and 5.5 ± 2.9 mmol/L) and plasma NH3 concentrations (124 ± 48, 112 ± 38, and 126 ± 55 µmol/L, respectively) were observed during 3v3, 5v5, and 7v7, respectively, with no difference between formats. Similar total distance (3676 ± 478, 3524 ± 467, and 3577 ± 500 m), high-intensity distance (349 ± 145, 406 ± 134, and 409 ± 165 m), and RPE (4.7 ± 1.6, 4.9 ± 2.1, and 4.6 ± 1.8) were also observed. The number of intense accelerations (500 ± 139 vs 459 ± 143 and 396 ± 144) were higher (P < 0.05) during 3v3 than 5v5 and 7v7. In conclusion, the intensity is high during small-sided recreational football games, with similar physiological responses for 6-14 players when pitch size is adapted, providing further evidence that effective recreational football training is easy to organize.

Positive performance and health effects of a football training program over 12 weeks can be maintained over a 1-year period with reduced training frequency.

We examined whether improvements in the performance and health profile of an intensive 12-week football intervention could be maintained with a reduced training frequency. Seventeen healthy untrained males completed the study. Ten subjects trained 2.4 times/week for 12 weeks and another 52 weeks with 1.3 sessions/week [football group (FG)] and seven subjects acted as controls [control group (CG)]. For FG, fat mass (3.2 kg) and systolic blood pressure (8 mmHg) were lower (P<0.05) after 64 than 0 weeks, and VO(2max) (8%) and Yo-Yo intermittent endurance level 2 test performance (49%) were higher (P<0.05), with no difference between 64 and 12 weeks. After 64 weeks, quadriceps muscle mass (11%), mean fiber area (10%) and citrate synthase activity (18%) were higher (P<0.05) than those at 0 weeks. Leg bone mass (3.5%) and density (2.0%) were higher (P<0.05) after 64 than 0 weeks, but not different between 12 and 0 weeks. Plantar jump force (17-18%), 30-m sprinting velocity (1.3-3.0%) and muscle glycogen concentration (19-21%) were higher (P<0.05) and blood lactate during submaximal exercise was lower (27-72%, P<0.05) after 64 than after 12 and 0 weeks. The above-mentioned variables were unaltered for CG. In conclusion, positive adaptations in cardiovascular fitness obtained over 12 weeks of regular recreational football training can be maintained over a 1-year period with a reduced training frequency, with further development in musculo-skeletal fitness.

Activity profile and physiological response to football training for untrained males and females, elderly and youngsters: influence of the number of players.

The present study examined the activity profile, heart rate and metabolic response of small-sided football games for untrained males (UM, n=26) and females (UF, n=21) and investigated the influence of the number of players (UM: 1v1, 3v3, 7v7; UF: 2v2, 4v4 and 7v7). Moreover, heart rate response to small-sided games was studied for children aged 9 and 12 years (C9+C12, n=75), as well as homeless (HM, n=15), middle-aged (MM, n=9) and elderly (EM, n=11) men. During 7v7, muscle glycogen decreased more for UM than UF (28 +/- 6 vs 11 +/- 5%; P<0.05) and lactate increased more (18.4 +/- 3.6 vs 10.8 +/- 2.1 mmol kg(-1) d.w.; P<0.05). For UM, glycogen decreased in all fiber types and blood lactate, glucose and plasma FFA was elevated (P<0.05). The mean heart rate (HR(mean)) and time >90% of HR(max) ranged from 147 +/- 4 (EM) to 162 +/- 2 (UM) b.p.m. and 10.8 +/- 1.5 (UF) to 47.8 +/- 5.8% (EM). Time >90% of HR(max) (UM: 16-17%; UF: 8-13%) and time spent with high speed running (4.1-5.1%) was similar for training with 2-14 players, but more high-intensity runs were performed with few players (UM 1v1: 140 +/- 17; UM 7v7: 97 +/- 5; P<0.05): Small-sided games were shown to elucidate high heart rates for all player groups, independently of age, sex, social background and number of players, and a high number of intense actions both for men and women. Thus, small-sided football games appear to have the potential to create physiological adaptations and improve performance with regular training for a variety of study groups.

Performance enhancements and muscular adaptations of a 16-week recreational football intervention for untrained women.

The present study investigated the performance effects and physiological adaptations over 16 weeks of recreational football training and continuous running for healthy untrained premenopausal women in comparison with an inactive control group [Football group (FG): n=21; running group (RG): n=18; CO: n=14]. Two weekly 1-h training sessions were performed in FG and RG. After 4 and 16 weeks of training VO(2max) was elevated (P<0.05) by 7% and 15%, respectively, in FG, and by 6% and 10%, respectively, in RG. After 16 weeks, Yo-Yo intermittent endurance level 2 performance was 33% and 19% better (P<0.05) for FG and 29% and 21% better (P<0.05) for RG than after 4 and 0 weeks, respectively. Peak sprinting speed was 12% higher (21.0 +/- 0.6 vs 18.8 +/- 0.7 km/h; P<0.05) for FG after the training period, whereas no difference was observed for RG. After 4 weeks citrate synthase (CS) and 3-hydroxyacyl-CoA dehydrogenase (HAD) activity was 9% and 8%, respectively, higher (P<0.05) than before training in FG with no further changes during the last 12 weeks. In RG, CS increased (P<0.05) by 12% after 4 weeks and no significant increase was observed for HAD. In FG, the number of capillaries per fiber was 18% higher (P<0.05) after 16 weeks (2.44 +/- 0.15 vs 2.07 +/- 0.05 cap/fiber), with no significant difference for RG. No differences were observed between 0 and 16 weeks for CO. In conclusion, recreational women's football leads to significant increases in VO(2max), performance and muscular adaptations throughout a 16-week training period. Thus, football can be used as an activity to elevate the physical capacity of untrained women.

Recreational soccer is an effective health-promoting activity for untrained men.

To examine the effects of regular participation in recreational soccer on health profile, 36 healthy untrained Danish men aged 20-43 years were randomised into a soccer group (SO; n = 13), a running group (RU; n = 12) and a control group (CO; n = 11). Training was performed for 1 h two or three times per week for 12 weeks; at an average heart rate of 82% (SEM 2%) and 82% (1%) of HR(max) for SO and RU, respectively. During the 12 week period, maximal oxygen uptake increased (p<0.05) by 13% (3%) and 8% (3%) in SO and RU, respectively. In SO, systolic and diastolic blood pressure were reduced (p<0.05) from 130 (2) to 122 (2) mm Hg and from 77 (2) to 72 (2) mm Hg, respectively, after 12 weeks, with similar decreases observed for RU. After the 12 weeks of training, fat mass was 3.0% (2.7 (0.6) kg) and 1.8% (1.8 (0.4) kg) lower (p<0.05) for SO and RU, respectively. Only SO had an increase in lean body mass (1.7 (0.4) kg, p<0.05), an increase in lower extremity bone mass (41 (8) g, p<0.05), a decrease in LDL-cholesterol (2.7 (0.2) to 2.3 (0.2) mmol/l; p<0.05) and an increase (p<0.05) in fat oxidation during running at 9.5 km/h. The number of capillaries per muscle fibre was 23% (4%) and 16% (7%) higher (p<0.05) in SO and RU, respectively, after 12 weeks. No changes in any of the measured variables were observed for CO. In conclusion, participation in regular recreational soccer training, organised as small-sided drills, has significant beneficial effects on health profile and physical capacity for untrained men, and in some aspects it is superior to frequent moderate-intensity running.

Muscle temperature and sprint performance during soccer matches--beneficial effect of re-warm-up at half-time.

The relationship between quadriceps muscle temperature (T(m)) and sprint performance was evaluated during soccer matches in 25 competitive players. In one game, T(m) was determined frequently (n=9). In another game, eight players performed low-intensity activities at half-time (re-warm-up, (RW), whereas another eight players recovered passively (CON). T(m) was 36.0+/-0.2 degrees C at rest and increased (P<0.05) to 39.4+/-0.2 degrees C before the game and remained unaltered during the first half. At half-time, T(m) decreased (P<0.05) to 37.4+/-0.2 degrees C, but increased (P<0.05) to 39.2+/- degrees C during the second half. In CON and RW, T(m) and core temperature (T(c)) were similar before and after the first half, but 2.1+/-0.1 and 0.9+/-0.1 degrees C higher (P<0.05), respectively, in RW prior to the second half. At the onset of the second half, the sprint performance was reduced (P<0.05) by 2.4% in CON, but unchanged in RW. The decrease in T(m) was correlated to the decrease in performance (r=0.60, P<0.05, n=16). This study demonstrates that in soccer, the decline in T(m) and T(c) during half-time is associated with a lowered sprint capacity at the onset of the second half, whereas sprint performance is maintained when low-intensity activities preserve muscle temperature.

Direct detection and identification of African trypanosomes by fluorescence in situ hybridization with peptide nucleic acid probes.

We have developed a rapid and easy to perform fluorescence in situ hybridization test that allows specific identification of trypanosomes from the subgenus Trypanozoon, using peptide nucleic acid probes. Probes were designed to target subgenus-specific sequences on the multiple-copy 18S rRNA, greatly facilitating the detection of a single trypanosome.

Identification of indicator microorganisms using a standardized PNA FISH method.

A standardized fluorescent in situ hybridization (FISH) method using Peptide Nucleic Acid (PNA) probes for analysis of gram-negative and gram-positive bacteria, as well as yeast, has been developed. Fluorescently labeled PNA probes targeting specific rRNA sequences of Escherichia coli, Pseudomonas aeruginosa, Staphyloccocus aureus, Salmonella were designed, as well as PNA probes targeting eubacteria and eucarya. These PNA probes were evaluated by PNA FISH using 27 bacterial and 1 yeast species, representing both phylogenetically closely related species, as well as species important to both clinical and industrial settings. The S. aureus and P. aeruginosa PNA probes did not cross react with any of the organisms tested, whereas the E. coli PNA probe, as expected from sequence data, also detected Shigella species. The Salmonella PNA probe reacted with all of the 13 Salmonella strains, representing the 7 subspecies of Salmonella, however, it is also complementary to a few other bacterial species. The eubacteria- and eucarya-specific PNA probes detected all bacterial species and one yeast species, respectively. The general applicability of the PNA FISH method made simultaneous identification of multiple species, both gram-negative and gram-positive, in a mixed population an attractive possibility never accomplished using DNA probes. Four color images using differently labeled PNA probes showed simultaneous identification of E. coli, P. aeruginosa, S. aureus and Salmonella, thereby demonstrating the potential of multiplex FISH for various diagnostic applications within both clinical and industrial microbiology.

Differentiation of Candida albicans and Candida dubliniensis by fluorescent in situ hybridization with peptide nucleic acid probes.

The recent discovery of Candida dubliniensis as a separate species that traditionally has been identified as Candida albicans has led to the development of a variety of biochemical and molecular methods for the differentiation of these two pathogenic yeasts. rRNA sequences are well-established phylogenetic markers, and probes targeting species-specific rRNA sequences have been used in diagnostic assays for the detection and identification of microorganisms. Peptide nucleic acid (PNA) is a DNA mimic with improved hybridization characteristics, and the neutral backbone of PNA probes offers significant advantages in whole-cell in situ hybridization assays. In this study, we developed PNA probes targeting the rRNAs of C. albicans and C. dubliniensis and applied them to a fluorescence in situ hybridization method (PNA FISH) for differentiation between C. albicans and C. dubliniensis. Liquid cultures were smeared onto microscope slides, heat fixed, and then hybridized for 30 min. Unhybridized PNA probe was removed by washing, and smears were examined by fluorescence microscopy. Evaluation of the PNA FISH method using smears of 79 C. dubliniensis and 70 C. albicans strains showed 100% sensitivity and 100% specificity for both PNA probes. We concluded that PNA FISH is a powerful tool for the differentiation of C. albicans and C. dubliniensis.

Self-reporting PNA/DNA primers for PCR analysis.

We report a new fluorogenic method for sealed-tube PCR analysis using a quencher-labeled peptide nucleic acid (Q-PNA) probe. The Q-PNA hybridizes to a complementary tag sequence located at the 5' end of a 5' fluorophore-labeled oligonucleotide primer, quenching the primer's fluorescence. Incorporation of the primer into a doublestranded amplicon causes displacement of the Q-PNA such that the fluorescence of the sample is a direct indication of the amplicon concentration. The Q-PNA is able to quench multiple primers bearing distinct 5' fluorophores in a single reaction. We show realtime quantitative detection of a single-copy gene, K-ras, from human genomic DNA, as well as an endpoint multiplex assay for Chlamydia trachomatis and Neisseria gonorrhoeae targets. Because the Q-PNA may be used to quench any primer that contains the 5' tag sequence, it is possible to inexpensively adapt an existing primer set for use in a self-reporting fluorescent assay by including the tag sequence in one of the primers.

Identification of Dekkera bruxellensis (Brettanomyces) from wine by fluorescence in situ hybridization using peptide nucleic acid probes.

A new fluorescence in situ hybridization method using peptide nucleic acid (PNA) probes for identification of Brettanomyces is described. The test is based on fluorescein-labeled PNA probes targeting a species-specific sequence of the rRNA of Dekkera bruxellensis. The PNA probes were applied to smears of colonies, and results were interpreted by fluorescence microscopy. The results obtained from testing 127 different yeast strains, including 78 Brettanomyces isolates from wine, show that the spoilage organism Brettanomyces belongs to the species D. bruxellensis and that the new method is able to identify Brettanomyces (D. bruxellensis) with 100% sensitivity and 100% specificity.

Filter-based PNA in situ hybridization for rapid detection, identification and enumeration of specific micro-organisms.

AIMS: A method for rapid and simultaneous detection, identification and enumeration of specific micro-organisms using Peptide Nucleic Acid (PNA) probes is presented. METHODS AND RESULTS: The method is based on a membrane filtration technique. The membrane filter was incubated for a short period of time. The microcolonies were analysed by in situ hybridization, using peroxidase-labelled PNA probes targeting a species-specific rRNA sequence, and visualized by a chemiluminescent reaction. Microcolonies were observed as small spots of light on film, thereby providing simultaneous detection, identification and enumeration. The method showed 95-100% correlation to standard plate counts along with definitive identification due to the specificity of the probe. CONCLUSION: Using the same protocol, results were generated approximately three times faster than culture methods for Gram-positive and -negative bacterial species and yeast species. SIGNIFICANCE AND IMPACT OF THE STUDY: The method is an improvement on the current membrane filtration technique, providing rapid determination of the level of specific pathogens, spoilage or indicator micro-organisms.

Rapid detection, identification, and enumeration of Escherichia coli cells in municipal water by chemiluminescent in situ hybridization.

A new chemiluminescent in situ hybridization (CISH) method provides simultaneous detection, identification, and enumeration of culturable Escherichia coli cells in 100 ml of municipal water within one working day. Following filtration and 5 h of growth on tryptic soy agar at 35 degrees C, individual microcolonies of E. coli were detected directly on a 47-mm-diameter membrane filter using soybean peroxidase-labeled peptide nucleic acid (PNA) probes targeting a species-specific sequence in E. coli 16S rRNA. Within each microcolony, hybridized, peroxidase-labeled PNA probe and chemiluminescent substrate generated light which was subsequently captured on film. Thus, each spot of light represented one microcolony of E. coli. Following probe selection based on 16S ribosomal DNA (rDNA) sequence alignments and sample matrix interference, the sensitivity and specificity of the probe Eco16S07C were determined by dot hybridization to RNA of eight bacterial species. Only the rRNA of E. coli and Pseudomonas aeruginosa were detected by Eco16S07C with the latter mismatch hybridization being eliminated by a PNA blocker probe targeting P. aeruginosa 16S rRNA. The sensitivity and specificity for the detection of E. coli by PNA CISH were then determined using 8 E. coli strains and 17 other bacterial species, including closely related species. No bacterial strains other than E. coli and Shigella spp. were detected, which is in accordance with 16S rDNA sequence information. Furthermore, the enumeration of microcolonies of E. coli represented by spots of light correlated 92 to 95% with visible colonies following overnight incubation. PNA CISH employs traditional membrane filtration and culturing techniques while providing the added sensitivity and specificity of PNA probes in order to yield faster and more definitive results.