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Perennial freshwater systems are valuable natural resources that provide important ecological services globally. However, in highly variable climates, such as Australia, water availability in rivers and streams can vary greatly from year to year and from decade to decade. Further, across Australia and many other regions, perennial river systems are projected to decrease because of anthropogenic climate change, placing the ecosystems they support under additional pressure. Quantifying the potential impacts of climate change on perennial freshwater systems requires robust databases of existing water features with accurate classifications. This is a challenge for rivers that display a high degree of interannual variability since the river classification can be dependent on the period of available data. In this study, we carry out a regional scale comparison of three different spatial databases commonly used in environmental and ecological assessments of perennial systems of Australia, namely Geodata, Geofabric and Water Observations from Space (WOfS). Focusing on the southern Murray Darling Basin (MDB), due to its national and international significance and its highly variable flow regimes, we show that no single spatial database is reliable by itself in terms of perennial water classification, with notable differences likely arising from variations in the periods analysed and methods used to classify the systems. Further, an analysis of high-quality gauged streamflow data (with approximately 40-year daily records) for four sub-catchments, and long-term simulation data (>100 years) for two sub-catchments in the lower MDB, confirm that flow persistence can be non-stationary through time, with some 'perennial' systems exhibiting sustained periods of cease to flow (i.e. becoming non-perennial) during prolonged droughts. This study demonstrates that due consideration is required in developing baseline classification of perennial freshwater systems for assessing future changes and measuring adaptive capacity., Competing Interests: Declaration of competing interest The authors declare no conflicts of interest., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)

This study investigated the effect of five post-weaning supplementation strategies and two weaning weight groups on long-term growth, puberty and pregnancy percentage of Brahman crossbred heifers. Early-weaned (118 ± 6 kg liveweight) and normally-weaned (183 ± 6 kg liveweight) heifers were allocated to group pens (n = 4 and n = 5/pen for early- and normally-weaned respectively) and offered one of five levels of post-weaning protein supplementation: 0, 1, 2.5, 5 and 10 g of supplement/kg liveweight.day with ad libitum access to a low quality sabi grass (Urochloa mosambicensis) hay during the first dry season (169 days) after weaning. After the post-weaning supplementation period, all heifers grazed the same pastures as a single mob until the end of the experiment and were exposed to fertile bulls from January to May 2016. During the first dry season, supplement intake had a positive linear effect on liveweight gain and hip width gain with no difference in the response between weaning groups. Overall, heifers with higher supplement intakes (i.e. 5 and 10 g/kg) had higher hip height gain (P < 0.005), hip width gain (P < 0.001), body condition score (P < 0.001), and concentration of insulin-like growth factor-1 (P = 0.001), triiodothyronine (P = 0.04) and insulin (P = 0.05) in plasma compared to unsupplemented heifers. These changes resulted in thicker proliferative and hypertrophic zones (both P = 0.03) of the tuber coxae growth plate, larger diameter of terminal hypertrophic chondrocytes (both P = 0.004) at the end of the post-weaning supplementation period when comparing the highest level of supplementation with unsupplemented group. Unsupplemented heifers from both weaning weight groups demonstrated compensatory liveweight gain over the first wet season while evidence of catch-up growth in skeletal dimensions was observed in the second wet season. The main determining factor for pregnancy status of two-year-old Brahman crossbred heifers was pre-mating liveweight (P < 0.001), the pre-mating liveweight was in turn affected by post-weaning supplementation (P = 0.02) or weaning weight group (P < 0.001). This study further demonstrated the positive relationship between premating weight and the occurrence of pregnancy, with an approximate 300 kg pre-mating liveweight required to achieve approximately 80% (67.1-90.3% for a 95% confidence interval) probability of pregnancy in two-year-old Brahman crossbred heifers mated for 4 months., Competing Interests: The authors have declared that no competing interests exist.

The objective of this study was to investigate the effect of diet crude protein (CP) content and metabolisable energy (ME) intake on skeletal growth and associated parameters of growing steers prior to and during compensatory growth in weight and catch-up growth in skeletal elongation. The experiment was a factorial design with two cattle genotypes [Brahman crossbred (BX, 178 ± 6 kg) and Holstein-Friesian (HF, 230 ± 34 kg)] and three nutritional treatments; high CP content and high ME intake (HCP-HME), high CP content and low ME intake (HCP-LME) and low CP content and low ME intake (LCP-LME) with the ME intake of HCP-LME matched to that of LCP-LME. Nutritional treatments were imposed over a 103 d period (Phase 1), and after this, all steers were offered ad libitum access to the HCP-HME nutritional treatment for 100 d (Phase 2). Steers fed the high CP content treatment with a low ME intake, showed higher hip height gain (P = 0.04), larger terminal hypertrophic chondrocytes (P = 0.02) and a higher concentration of total triiodothyronine in plasma (P = 0.01) than steers with the same ME intake of the low CP content treatment. In addition, the low CP treatment resulted in significant decreases in bone volume (P = 0.03), bone surface area (P = 0.03) and the concentration of bone-specific alkaline phosphatase in plasma (P < 0.001) compared to steers fed the HCP-HME treatment. A significant interaction between genotype and nutritional treatment existed for the concentration of thyroxine (T4) in plasma where HF steers fed LCP-LME had a lower T4 concentration in plasma (P = 0.05) than BX steers. All steers with a restricted ME intake during Phase 1 demonstrated compensatory growth during Phase 2. However, HF steers fed the LCP treatment during Phase 1 showed a tendency (P = 0.07) for a greater LWG during Phase 2 without any increase in dry matter intake. Results observed at the growth plate and hip height growth suggest that catch-up growth in cattle may also be explained by the growth plate senescence hypothesis. Contrary to our initial hypothesis, the results demonstrate that greater CP intake during ME restriction does not increase compensatory gain in cattle during re-alimentation., Competing Interests: The authors have declared that no competing interests exist.

Musculoskeletal injuries (MSI) continue to affect Thoroughbred racehorses internationally. There is a strong interest in developing training and management strategies to reduce their impact, however, studies of risk factors report inconsistent findings. Furthermore, many injuries and fatalities occur during training rather than during racing, yet most studies report racing data only. By combining racing and training data a larger exposure to risk factors and a larger number of musculoskeletal injuries are captured and the true effect of risk factors may be more accurately represented. Furthermore, modifications to reduce the impact of MSI are more readily implemented at the training level. Our study aimed to: (1) determine the risk factors for musculoskeletal injuries and whether these are different for two-year-old and older horses and (2) determine whether risk factors vary with type of injury. This was performed by repeating analyses by age category and injury type. Data from 202 cases and 202 matched controls were collected through weekly interviews with trainers and analysed using conditional logistic regression. Increasing dam parity significantly reduced the odds of injury in horses of all age groups because of the effect in two-year-old horses (odds ratio (OR) 0.08; 95% confidence interval (CI) 0.02, 0.36; p < 0.001). Increasing total preparation length is associated with higher odds of injury in horses of all ages (OR 5.56; 95% CI 1.59, 19.46; p = 0.01), but particularly in two-year-old horses (OR 8.05; 95% CI 1.92, 33.76; p = 0.004). Increasing number of days exercised at a slow pace decreased the odds of injury in horses of all ages (OR 0.09; 95% CI 0.03, 0.28; p < 0.001). The distance travelled at three-quarter pace and above (faster than 13 m/s; 15 s/furlong; 800 m/min; 48 km/h) and the total distance travelled at a gallop (faster than 15 m/s; 13 s/furlong; 900 m/min; 55 km/h) in the past four weeks significantly affected the odds of injury. There was a non-linear association between high-speed exercise and injury whereby the odds of injury initially increased and subsequently decreased as accumulated high-speed exercise distance increased. None of the racing career and performance indices affected the odds of injury. We identified horses in this population that have particularly high odds of injury. Two-year-old horses from primiparous mares are at increased odds of injury, particularly dorsal metacarpal disease. Two-year-old horses that have had a total preparation length of between 10 and 14 weeks also have increased odds of injury. Horses of all ages that travelled a total distance of 2.4-3.8 km (12-19 furlongs) at a gallop in the last four weeks and horses three years and older that travelled 3.0-4.8 km (15-24 furlongs) at three-quarter pace and above also have increased odds of injury. We recommend that these horses should be monitored closely for impending signs of injury. Increasing the number of days worked at a slow pace may be more effective for preventing injury, if horses are perceived at a higher risk, than resting the horse altogether. Early identification of horses at increased risk and appropriate intervention could substantially reduce the impact of musculoskeletal injuries in Thoroughbred racehorses.

Breeding mares typically foal yearly. Little is known about the dynamics of maternal bone stores during gestation and lactation, the timing of any maternal bone mobilisation, re-accretion post-foaling, or the dynamics of bone metabolism in foals. We measured serum osteocalcin (OC) and serum pyridinoline (PYD) concentrations in 18 mares monthly from 6months gestation to foaling, and in both mares and foals for 4months after birth. From 6 to 11months of gestation, there was no change in mean monthly OC. Serum PYD increased between 7months gestation and foaling. After foaling, mean serum OC was low up to 14days, rising to peak at 1month. Serum PYD rose concomitantly during this period, but subsequently declined. The mare OC:PYD ratio fell to a nadir within 14days of birth, before rising to a peak at 2months. In foals, OC rose within the first 24h of birth to peak at 3months. PYD fell from birth levels by 1month of age. Maternal bone mobilisation occurs progressively from 8months of gestation until term, before increasing markedly in very early lactation. Net mobilisation switches to accretion by one to two months of foaling, suggesting that this is a period during which mares replenish their own bone stores. Changes in the ratio of OC to PYD indicate adaptation to the prevailing biological milieu. In foals, the increase in the OC:PYD ratio in early life reflects the dominance of bone accretion., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Stress fractures (SFxs) are debilitating injuries and exact mechanisms that initiate their repair incompletely understood. We hypothesised that osteocyte apoptosis and expression of cytokines and proteins such as sclerostin, VEGF, TGF-β, COX-2 and IL-6 were early signalling events to facilitate the formation of periosteal woven bone and recruitment of osteoclast precursors to the site of remodelling. A SFx was created in the right ulna of mature female wistar rats using cyclic end loading. Rats were killed 1, 4 and 7 days after loading (n=5 per group). Standard histological staining was used to examine SFx morphology and immunohistochemistry to detect the localisation of these proteins and in situ hybridisation to detect mRNA along the SFx line or gene expression to quantify the target genes. Unloaded ulnae served as controls. The labelling index of caspase-3, COX-2 and IL-6 was significantly elevated in the region of SFxs at all time points compared with controls (P<0.001). In addition, the labelling index of sclerostin protein was significantly reduced in osteocytes adjacent to the SFx region when compared with controls at all three time points (P<0.001). Both VEGF and TGF-β expressions were only localised in the woven bone. These data reinforce the involvement of osteocyte apoptosis in the healing of fatigue damage in bone, and demonstrate that local regulation of sclerostin, COX-2 and IL-6 are important signalling events associated with new bone formation and SFx remodelling.

Anti-inflammatory drugs are widely used to manage pain associated with stress fractures (SFxs), but little is known about their effects on healing of those injuries. We hypothesized that selective and non-selective anti-inflammatory treatments would retard the healing of SFx in the rat ulna. SFxs were created by cyclic loading of the ulna in Wistar rats. Ulnae were harvested 2, 4 or 6 weeks following loading. Rats were treated with non-selective NSAID, ibuprofen (30 mg/kg/day); selective COX-2 inhibition, [5,5-dimethyl-3-3 (3 fluorophenyl)-4-(4 methylsulfonal) phenyl-2 (5H)-furanone] (DFU) (2.0 mg/kg/day); or the novel c5a anatagonist PMX53 (10 mg/kg/day, 4 and 6 weeks only); with appropriate vehicle as control. Quantitative histomorphometric measurements of SFx healing were undertaken. Treatment with the selective COX-2 inhibitor, DFU, reduced the area of resorption along the fracture line at 2 weeks, without affecting bone formation at later stages. Treatment with the non-selective, NSAID, ibuprofen decreased both bone resorption and bone formation so that there was significantly reduced length and area of remodeling and lamellar bone formation within the remodeling unit at 6 weeks after fracture. The C5a receptor antagonist PMX53 had no effect on SFx healing at 4 or 6 weeks after loading, suggesting that PMX53 would not delay SFx healing. Both selective COX-2 inhibitors and non-selective NSAIDs have the potential to compromise SFx healing, and should be used with caution when SFx is diagnosed or suspected., (Copyright © 2012 Orthopaedic Research Society.)

Because bisphosphonates (BPs) are potent inhibitors of bone resorption, we hypothesized that they would retard direct remodeling of stress fractures. The aim of this study was to determine the effect of risedronate on direct remodeling and woven bone callus formation following stress fracture formation in the rat ulna. In 135 adult female Wistar rats, cyclic loading of the ulna created stress fractures. Rats were treated daily with oral saline, or risedronate at 0.1 or 1.0 mg/kg. From each bone, histomorphometry was performed on sections stained with toluidine blue at a standard level along the fracture. The high dose of risedronate caused a significant decrease in the percentage of repaired stress fracture and bone resorption along the stress fracture line at 6 and 10 weeks after loading (p < 0.05). At this dose, intracortical resorption was significantly reduced at 10 weeks after loading and intracortical new bone area was significantly reduced at 6 and 10 weeks. Woven bone formation and consolidation phases of stress fracture repair were not affected by low or high doses of risedronate. In conclusion, high dose bisphosphonate treatment impaired healing of a large stress fracture line by reducing the volume of bone resorbed and replaced during remodeling. We also confirmed that periosteal callus formation was not adversely affected by risedronate treatment., (Copyright © 2011 Orthopaedic Research Society.)

Loading of the rat ulna is an ideal model to examine stress fracture healing. The aim of this study was to undertake a detailed examination of the histology, histomorphometry and gene expression of the healing and remodelling process initiated by fatigue loading of the rat ulna. Ulnae were harvested 1, 2, 4, 6, 8, and 10 weeks following creation of a stress fracture. Stress fracture healing involved direct remodelling that progressed along the fracture line as well as woven bone proliferation at the site of the fracture. Histomorphometry demonstrated rapid progression of basic multicellular units from 1 to 4 weeks with significant slowing down of healing by 10 weeks after loading. Quantitative PCR was performed at 4 hours, 24 hours, 4 days, 7 days, and 14 days after loading. Gene expression was compared to an unloaded control group. At 4 hours after fracture, there was a marked 220-fold increase (P<0.0001) in expression of IL-6. There were also prominent peak increases in mRNA expression for OPG, COX-2, and VEGF (all P<0.0001). At 24 hours, there was a peak increase in mRNA expression for IL-11 (73-fold increase, P<0.0001). At 4 days, there was a significant increase in mRNA expression for Bcl-2, COX-1, IGF-1, OPN, and SDF-1. At 7 days, there was significantly increased mRNA expression of RANKL and OPN. Prominent, upregulation of COX-2, VEGF, OPG, SDF-1, BMP-2, and SOST prior to peak expression of RANKL indicates the importance of these factors in mediating directed remodelling of the fracture line. Dramatic, early upregulation of IL-6 and IL-11 demonstrate their central role in initiating signalling events for remodelling and stress fracture healing., ((c) 2009 Elsevier Inc. All rights reserved.)

This paper describes the clinical and radiological features, surgical techniques used and results obtained in 6 horses with fractures of the tibial tuberosity. The horses were presented between 24 h and 8 weeks following injury. In all 6 cases, the fragments were displaced proximocranially and in 2 of these, there was comminution. Four were treated by open reduction and internal fixation using an AO/ASIF narrow dynamic compression plate and in 2 cases the fragments were removed. All horses returned to full athletic function and remained sound in follow-up times of 17-36 months. Implant removal was necessary in 1 case.