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At least 55 records · Page 3Linked to original sources
USGS2007

Peptidomic analysis of skin secretions supports separate species status for the tailed frogs, Ascaphus truei and Ascaphus montanus

The tailed frog Ascaphus truei Stejneger, 1899 is the most primitive extant anuran and the sister taxon to the clade of all other living frogs. The species occupies two disjunct ranges in the Northwest region of North America: the Cascade Mountains and coastal area from British Columbia to Northern California, and an inland range in the northern Rocky Mountains and the Blue and Wallowa mountains. A previous study led to the isolation of eight peptides with antimicrobial activity (termed the ascaphins) from skin secretions of A. truei from the coastal range. The present study has used peptidomic analysis to identify the products of orthologous ascaphin genes in electrically-stimulated skin secretions from inland range specimens. Structural characterization of the peptides demonstrated that ascaphins from the inland range contained the following amino acid substitutions compared with orthologs from the coastal range frogs: ascaphin-1 (Ala 12 → Glu), ascaphin-3 (Asp 4 → Glu), ascaphin-4 (Ala 19 → Ser), ascaphin-5 (Lys 12 → Thr), and ascaphin-7 (Gly 8 → Ser and Ser 20 → Asn). Orthologs of ascaphins-2, -6, and -8 were not identified but a paralog of ascaphin-5, identical to ascaphin-5 from coastal range frogs, was found. The data support the claims, derived from analysis of the nucleotide sequences of mitochondrial genes, that the inland populations of the tailed frog should be recognized as a distinct species, the Rocky Mountain tailed frog Ascaphus montanus and that the divergence of the species from A. truei probably occurred in the late Miocene (approximately 10 Mya).

Comparative Biochemistry and Physiology, Part D: G↗
USGS2007

Gene expression in the liver of rainbow trout, Oncorhynchus mykiss , during the stress response

To better appreciate the mechanisms underlying the physiology of the stress response, an oligonucleotide microarray and real-time RT-PCR (QRT-PCR) were used to study gene expression in the livers of rainbow trout ( Oncorhynchus mykiss ). For increased confidence in the discovery of candidate genes responding to stress, we conducted two separate experiments using fish from different year classes. In both experiments, fish exposed to a 3 h stressor were compared to control (unstressed) fish. In the second experiment some additional fish were exposed to only 0.5 h of stress and others were sampled 21 h after experiencing a 3 h stressor. This 21 h post-stress treatment was a means to study gene expression during recovery from stress. The genes we report as differentially expressed are those that responded similarly in both experiments, suggesting that they are robust indicators of stress. Those genes are a major histocompatibility complex class 1 molecule ( MHC1 ), JunB , glucose 6-phosphatase ( G6Pase ), and nuclear protein 1 ( Nupr1 ). Interestingly, Nupr1 gene expression was still elevated 21 h after stress, which indicates that recovery was incomplete at that time.

Comparative Biochemistry and Physiology, Part D: G↗
USGS2012

Transcriptional profiling of the parr–smolt transformation in Atlantic salmon

The parr–smolt transformation in Atlantic salmon ( Salmo salar ) is a complex developmental process that culminates in the ability to migrate to and live in seawater. We used GRASP 16K cDNA microarrays to identify genes that are differentially expressed in the liver, gill, hypothalamus, pituitary, and olfactory rosettes of smolts compared to parr. Smolts had higher levels of gill Na + /K + -ATPase activity, plasma cortisol and plasma thyroid hormones relative to parr. Across all five tissues, stringent microarray analyses identified 48 features that were differentially expressed in smolts compared to parr. Using a less stringent method we found 477 features that were differentially expressed at least 1.2-fold in smolts, including 172 features in the gill. Smolts had higher mRNA levels of genes involved in transcription, protein biosynthesis and folding, electron transport, oxygen transport, and sensory perception and lower mRNA levels for genes involved in proteolysis. Quantitative RT-PCR was used to confirm differential expression in select genes identified by microarray analyses and to quantify expression of other genes known to be involved in smolting. This study expands our understanding of the molecular processes that underlie smolting in Atlantic salmon and identifies genes for further investigation.

Comparative Biochemistry and Physiology, Part D: G↗
USGS2016

Transcriptome discovery in non-model wild fish species for the development of quantitative transcript abundance assays

Environmental studies increasingly identify the presence of both contaminants of emerging concern (CECs) and legacy contaminants in aquatic environments; however, the biological effects of these compounds on resident fishes remain largely unknown. High throughput methodologies were employed to establish partial transcriptomes for three wild-caught, non-model fish species; smallmouth bass ( Micropterus dolomieu ), white sucker ( Catostomus commersonii ) and brown bullhead ( Ameiurus nebulosus ). Sequences from these transcriptome databases were utilized in the development of a custom nCounter CodeSet that allowed for direct multiplexed measurement of 50 transcript abundance endpoints in liver tissue. Sequence information was also utilized in the development of quantitative real-time PCR (qPCR) primers. Cross-species hybridization allowed the smallmouth bass nCounter CodeSet to be used for quantitative transcript abundance analysis of an additional non-model species, largemouth bass ( Micropterus salmoides ). We validated the nCounter analysis data system with qPCR for a subset of genes and confirmed concordant results. Changes in transcript abundance biomarkers between sexes and seasons were evaluated to provide baseline data on transcript modulation for each species of interest.

Comparative Biochemistry and Physiology, Part D: G↗
USGS2008

Effects of ACTH, capture, and short term confinement on glucocorticoid concentrations in harlequin ducks ( Histrionicus histrionicus )

Little is known about baseline concentrations of adrenal hormones and hormonal responses to stress in sea ducks, although significant population declines documented in several species suggest that sea ducks are exposed to increased levels of environmental stress. Such declines have been observed in geographically distinct harlequin duck populations. We performed an adrenocorticotropic hormone (ACTH) challenge to evaluate adrenal function and characterize corticosterone concentrations in captive harlequin ducks and investigated the effects of capture, surgery, and short term confinement on corticosterone concentrations in wild harlequin ducks. Harlequin ducks responded to the ACTH challenge with an average three-fold increase in serum corticosterone concentration approximately 90 min post injection, and a four- to five-fold increase in fecal glucocorticoid concentration 2 to 4 h post injection. Serum corticosterone concentrations in wild harlequin ducks increased within min of capture and elevated levels were found for several hours post capture, indicating that surgery and confinement maintain elevated corticosterone concentrations in this species. Mean corticosterone concentrations in wild harlequin ducks held in temporary captivity were similar to the maximum response levels during the ACTH challenge in captive birds. However, large variation among individuals was observed in responses of wild birds, and we found additional evidence suggesting that corticosterone responses varied between hatch year and after hatch year birds.

Comparative Biochemistry and Physiology, Part A: M↗
USGS2010

Annual sex steroid and other physiological profiles of Pacific lampreys ( Entosphenus tridentatus )

We documented changes in plasma levels of estradiol 17-β (E2), progesterone (P), 15α-hydroxytestosterone (15α-T), thyroxine (T4), triiodothyronine (T3), protein, triglycerides (TGs), and glucose in adult Pacific lampreys ( Entosphenus tridentatus ) held in the laboratory in two different years. Levels of E2 in both sexes ranged from 0.5 to 2 ng/mL from September to March, peaked in late April (2–4 ng/mL), and decreased in May, with levels higher in males than in females. Levels of P were low from September through April, but then increased substantially during May (2–4 ng/mL), with levels again highest in males. Levels of 15α-T in males were around 0.75 ng/mL through the winter before exceeding 1 ng/mL in April and decreasing thereafter, whereas females showed a gradual increase from 0.25 ng/mL in November to 0.5 ng/mL in April before decreasing. Thyroxine concentrations differed between fish in each year, with most having levels ranging from 0.75 to 2.5 ng/mL in the fall and winter, and only fish in 2003 showing distinct peaks (3–4 ng/mL) in early April or May. Plasma T3 was undetectable from November through mid-March before surging dramatically in April (ca. 150 ng/mL) and decreasing thereafter. Levels of protein, TGs, and glucose decreased or were stable during the fall and winter with TGs and glucose surging in late April to early May for some fish. Our study is the first to document long-term physiological changes in Pacific lampreys during overwintering and sexual maturation and increases our understanding of the life history of this unique fish.

Comparative Biochemistry and Physiology, Part A: M↗
USGS2016

Salinity effects on plasma ion levels, cortisol, and osmolality in Chinook salmon following lethal sampling

Studies on hydromineral balance in fishes frequently employ measurements of electrolytes following euthanasia. We tested the effects of fresh- or salt-water euthanasia baths of tricaine mesylate (MS-222) on plasma magnesium (Mg 2+ ) and sodium (Na + ) ions, cortisoland osmolality in fish exposed to saltwater challenges, and the ion and steroid hormone fluctuations over time following euthanasia in juvenile spring Chinook salmon ( Oncorhynchus tshawytscha ). Salinity of the euthanasia bath affected plasma Mg 2+ and Na + concentrations as well as osmolality, with higher concentrations in fish euthanized in saltwater. Time spent in the bath positively affected plasma Mg 2+ and osmolality, negatively affected cortisol, and had no effect on Na + concentrations. The difference of temporal trends in plasma Mg 2+ and Na + suggests that Mg 2+ may be more sensitive to physiological changes and responds more rapidly than Na + . When electrolytes and cortisol are measured as endpoints after euthanasia, care needs to be taken relative to time after death and the salinity of the euthanasia bath.

Comparative Biochemistry and Physiology, Part A: M↗
USGS2018

Shifts in the relationship between mRNA and protein abundance of gill ion-transporters during smolt development and seawater acclimation in Atlantic salmon (Salmo salar)

Smolting Atlantic salmon exhibit a seasonal increase in seawater tolerance that is associated with changes in the abundance of major gill ion-transporter transcripts and proteins. In the present study, we investigate how the transcript and protein abundance of specific ion-transporter isoforms relate to each other during smolt development and seawater acclimation, and how each correlates to seawater tolerance. We show that during smolt development both mRNA and protein abundance of gill Na + /K + -ATPase α1a subunit (NKAα1a) decreased but the decrease in the mRNA was five-times greater than that of the protein. Gill NKAα1b mRNA levels increased only slightly (1.5-fold) throughout development whereas protein abundance increased 30-fold at its peak. Gill Na + /K + /2Cl − co-transporter 1 (NKCC1) increased at the mRNA and protein level (5- and 12-fold) in smolts. The abundance of a gill ion-transporter's mRNA and protein changed in the same direction through development and after seawater transfer, but the changes were not always strongly correlated: NKAα1a ( r = 0.768), NKAα1b ( r = 0.40), and NKCC1 ( r = 0.898). The maintenance of plasma chloride concentration correlated most strongly with the abundance of NKAα1a mRNA, and the ratio of NKAα1b to NKAα1a mRNA and protein. Growth performance after seawater transfer correlated most strongly with the abundance of NKAα1b protein and the ratio of NKAα1b to NKAα1a protein. Our results indicate that the abundance of ion-transporter mRNA and protein do not always correlate well and a decrease in the abundance of gill NKAα1a mRNA and increase in NKAα1b protein are strong predictors of seawater tolerance and growth performance after seawater transfer.

Comparative Biochemistry and Physiology, Part A: M↗
USGS2012

Identification of the thiamin pyrophosphokinase gene in rainbow trout: Characteristic structure and expression of seven splice variants in tissues and cell lines and during embryo development

Thiamin pyrophosphokinase (TPK) converts thiamin to its active form, thiamin diphosphate. In humans, TPK expression is down-regulated in some thiamin deficiency related syndrome, and enhanced during pregnancy. Rainbow trout are also vulnerable to thiamin deficiency in wild life and are useful models for thiamin metabolism research. We identified the tpk gene transcript including seven splice variants in the rainbow trout. Almost all cell lines and tissues examined showed co-expression of several tpk splice variants including a potentially major one at both mRNA and protein levels. However, relative to other tissues, the longest variant mRNA expression was predominant in the ovary and abundant in embryos. During embryogenesis, total tpk transcripts increased abruptly in early development, and decreased to about half of the peak shortly after hatching. In rainbow trout, the tpk transcript complex is ubiquitously expressed for all tissues and cells examined, and its increase in expression could be important in the early-middle embryonic stages. Moreover, decimated tpk expression in a hepatoma cell line relative to hepatic and gonadal cell lines appears to be consistent with previously reported down-regulation of thiamin metabolism in cancer.

Comparative Biochemistry and Physiology, Part B: B↗
USGS2022

Metabolic costs associated with seawater acclimation in a euryhaline teleost, the fourspine stickleback (Apeltes quadracus)

The cost of osmoregulation in teleosts has been debated for decades, with estimates ranging from one to 30 % of routine metabolic rate. The variation in the energy budget appears to be greater for euryhaline fish due to their ability to withstand dynamic salinity levels. In this study, a time course of metabolic and physiological responses of the euryhaline fourspine stickleback ( Apeltes quadracus ) acclimated to freshwater (FW) and then exposed to seawater (SW) was examined. There was 18% mortality in the first 3 days following exposure to SW, with no mortalities in the FW control group. Gill Na + /K + -ATPase (NKA) activity, an index of osmoregulatory capacity, increased 2.6-fold in SW fish peaking on days 7 and 14. Gill citrate synthase activity, an index of aerobic capacity, was 50–62% greater in SW than FW fish and peaked on day 7. Tissue water content was significantly lower in the SW fish on day 1 only, returning to FW levels by day 3. Routine metabolic rate was decreased within 24 h of SW exposure and was maintained slightly (8–22%) but significantly lower in SW compared to FW water controls throughout the 2-week experiment. These results indicate that elevated salinity resulted in increased SW osmoregulatory and aerobic capacity in the gill, but with a reduced whole animal metabolic rate to this euryhaline species.

Comparative Biochemistry and Physiology Part B: Bi↗
USGS2024

Dopaminergic and anti-estrogenic responses in juvenile steelhead (Oncorhynchus mykiss) exposed to bifenthrin

The frequency of detection and concentrations of bifenthrin, a pyrethroid insecticide, in the waterways inhabited by the endangered species, steelhead trout ( Oncorhynchus mykiss ), has become a significant concern for regulatory agencies. Endocrine disruption has been observed with estrogenic and anti-estrogenic responses in fish species at different life stages. Since several studies have indicated alterations in dopaminergic signaling associated with endocrine responses, juvenile steelhead were exposed to environmentally relevant concentrations of 60 or 120 ng/L bifenthrin for two weeks. Fish brains were assessed for dopamine levels and the expression of genes involved in dopaminergic and estrogenic processes, such as catechol- o -methyltransferase ( comt ) and monoamine oxidase ( mao ). Vitellogenin ( vtg ) and estrogenic receptors ( ERα1 , ERβ1 , and ERβ2 ) were also evaluated in livers of the animals. Dopamine concentrations were significantly higher in fish brains following bifenthrin exposure. Consistent with a reduction in dopamine clearance, there was a significant decrease in the mRNA expression of comt with increased bifenthrin concentration. Hepatic expression of ERα1 and ERβ2 mRNA was significantly decreased with increased bifenthrin concentration. These data support the possible mechanism of bifenthrin altering the dopaminergic pathway at low ng/L concentrations, in juvenile steelhead, which could interfere with endocrine feedback loops. These findings support the need for and importance of identifying species and life stage differences in pesticide modes of action to reduce uncertainties in risk assessments.

Comparative Biochemistry and Physiology Part C: To↗
USGS2025

Amitriptyline and nortriptyline induce ocular toxicity in early life stage zebrafish (Danio rerio)

The global use of antidepressants has steadily increased, raising concern to aquatic ecosystems due to the incomplete removal during wastewater treatment. Tricyclic antidepressants (TCAs) act on the neuronal system by inhibiting the reuptake of serotonin and norepinephrine. However, despite visual function being heavily dependent on the neuronal system, a knowledge gap remains regarding the ocular toxicity of TCAs. To bridge this knowledge gap, zebrafish ( Danio rerio ) embryos were exposed to sublethal test concentrations of amitriptyline (AMI, 0.3 to 300 μg/L nominal, 2.04 to 234 μg/L measured) and nortriptyline (NOR, 0.03 to 300 μg/L nominal, >0.107 to 20.7 μg/L measured), with the lowest test concentrations being environmentally relevant. Visual function was assessed with the optokinetic response assay, eye structure development was assessed histologically, and gene expression changes were analysed via transcriptomic profiling. Larval zebrafish (120 h post fertilization (hpf)) exposed to 4.99 and 234 μg/L of AMI exhibited a 26 % and 86 % decrease in the number of eye saccades respectively, with zebrafish exposed to 20.7 μg/L of NOR exhibiting a 65 % decrease. Histological analysis indicated a significant increase of the retinal pigment epithelium thickness after exposure to 234 μg/L of AMI and 20.7 μg/L of NOR. Transcriptomic analysis resulted in 1207 and 2742 differentially expressed genes across both AMI and NOR treatment groups respectively, including genes involved in vision, synaptic signaling, and neuronal development. These findings demonstrate that sublethal concentrations of AMI and NOR affect early life stage zebrafish visual development, which may be sensitive endpoint that could be incorporated into ecological risk assessments.

Comparative Biochemistry and Physiology Part C: To↗
USGS2004

Intra-strain dioxin sensitivity and morphometric effects in swim-up rainbow trout ( Oncorhynchus mykiss )

Inter and intra-specific differences in sensitivity of early life stage salmonids to 2,3,7,8-TCDD exposure have been reported, but intra-strain differences have not been found in the literature. Our results indicate that intra-strain variability in terms of embryo mortality (LD50) is small in Eagle Lake strain of rainbow trout, LD50 values ranging from 285 to 457 pg TCDD egg g −1 . These results confirm Eagle Lake as a less sensitive strain within rainbow trout, and do not indicate overlap with reported LD50 values for brook or lake trout. Our results also demonstrate that although generalized edema in regions including the yolk-sac are frequently associated with mortality following dioxin exposure, not all edematous fish die. We detected dose-dependent decreases in cranial length, eye diameter, mass, and total length ( P <0.05) in viable swim-up rainbow trout. These effects are presumed to indicate more subtle dose-dependent disruptions of the viteline vein vasculature and, therefore, in access to energy sources. A tendency for dose-dependent decrease in liver glycogen reserves concurred with previous results on salmonids and with the well described TCDD-induced alterations in intermediate metabolism of rats and chicken embryos (wasting syndrome). This syndrome could be contributing to the reduced growth that we observed.

Comparative Biochemistry and Physiology, Part C: T↗
USGS2004

PCB impairs smoltification and seawater performance in anadromous Arctic charr ( Salvelinus alpinus )

The impacts of polychlorinated biphenyl (PCB) exposure on smoltification and subsequent seawater performance were investigated in hatchery-reared, anadromous Arctic charr ( Salvelinus alpinus ). The fish were subjected to a 2-month summer seawater residence, after which they were orally dosed with 0 (Control, C), 1 (Low Dose, LD) or 100 mg Aroclor 1254 kg −1 body mass (High Dose, HD) in November. They were then held in fresh water, without being fed (to mimic their natural overwintering in freshwater), until they had smolted in June the next year. The smolts were then transferred to seawater and fed to mimic their summer feeding residence in seawater, followed by a period without food in freshwater from August until maturation in October. Compared with C and LD charr, the HD charr had either a transient or a permanent reduction in plasma growth hormone, insulin-like growth factor-1, and thyroxin and triiodothyronine titers during the period of smoltification. These hormonal alterations in the HD charr corresponded with impaired hyposmoregulatory ability in May and June, as well as reduced growth rate and survival after transference to seawater. Consequently, fewer fish in the HD group matured in October compared to the other two treatments. The HD fish had a liver PCB concentration ranging between 14 and 42 mg kg −1 wet mass, whereas there were similar, and very low, liver PCB concentrations in LD and C fish throughout the smolting period. Our findings suggest that PCB might compromise mechanisms important for fitness in a fish species living in an extreme environment.

Comparative Biochemistry and Physiology, Part C: T↗
USGS2005

Distinct effects of 4-nonylphenol and estrogen-17β on expression of estrogen receptor α gene in smolting sockeye salmon

Xenoestrogens such as 4-nonylphenol (4-NP) have been shown to affect the parr–smolt transformation, but their mechanisms of action are not known. We therefore examined effects of 4-NP and estradiol-17β (E2) on expression of estrogen receptor (ER) α gene in the liver, gill, pituitary and brain of sockeye salmon to elucidate molecular mechanisms of 4-NP and E2 and developmental differences in response during smolting. Fish were treated twice within a week with 4-NP (15 and 150 mg/kg BW), E2 (2 mg/kg BW) or only vehicle at three stages of smolting, pre-smolting in March, early smolting in April and late smolting in May. The absolute amounts of ERα mRNA were determined by real-time PCR. The basal amounts of ERα mRNA peaked in April in the liver, gill and pituitary. In March, E2 extensively increased the amounts in the liver, while 4-NP had no effects at this stage. In contrast, 4-NP (but not E2) decreased liver ERα mRNA in April. 4-NP also decreased the amount of ERα mRNA in the gill in April. In the pituitary, 4-NP increased ERα mRNA in March but decreased it in May. There were no significant effects in the brain. Changes in basal ERα mRNA observed in this study indicate that estrogen responsiveness of tissues may change during salmon smolting. Furthermore, 4-NP and E2 have different effects on expression of ERα gene in the liver and gill during smolting, and the response is dependent on smolt stage.

Comparative Biochemistry and Physiology, Part C: T↗
USGS2005

Responses of hybrid striped bass to waterborne and dietary copper in freshwater and saltwater

Mechanisms of copper toxicity and consequences of exposure vary due to uptake route and ionoregulatory status. The goal of this research was to develop a model fish system to assess the influence of different Cu exposure routes (waterborne or dietary) on bioavailability, uptake, and effects in hybrid striped bass ( Morone chrysops × Morone saxatilis ) acclimated to fresh- or saltwater. Initially, hybrid striped bass were exposed to dietary Cu concentrations of 571, 785, and 1013 μg Cu/g, along with a control (∼ 5 μg Cu/g), for 14 days in saltwater. Intestinal and liver Cu accumulated in a dose-dependent manner in fish exposed to increasing levels of dietary Cu. Chronic (42 days) experiments were then conducted to determine sub-lethal effects of aqueous, dietary, and combined aqueous and dietary Cu exposures to both freshwater- and saltwater-acclimated hybrid striped bass. Growth and Cu accumulation in the gill, intestine, and liver were measured. Although no significant effects were observed in fish exposed to waterborne Cu, those exposed through the diet accumulated significant liver and intestinal Cu but showed no significant change in growth. Overall, these results suggest that at the levels tested, exposure to elevated waterborne Cu did not cause significant long-term tissue Cu accumulation, whereas dietary Cu exposure caused significant liver and intestinal Cu accumulation in hybrid striped bass which was comparable in both freshwater and saltwater (15 g/L).

Comparative Biochemistry and Physiology, Part C: T↗
USGS2005

A non-lethal method to estimate CYP1A expression in laboratory and wild Atlantic salmon ( Salmo salar )

Expression of cytochrome P4501A (CYP1A) has been used as a biomarker for possible exposure to contaminants such as PCBs and dioxins in teleost fish. Using a quantitative reverse transcription-polymerase chain reaction (Q-RT-PCR) and a non-lethal gill biopsy, we estimated levels of CYP1A mRNA expression in Atlantic salmon ( Salmo salar ). Groups of ten Atlantic salmon juveniles (48–76 g) received an intraperitoneal injection of 50 μg g − 1 β-naphthoflavone (BNF) or vehicle. Their gill tissues were repeatedly sampled by non-lethal biopsies on day 0, 1, 2 and 7. Control fish expressed basal levels of CYP1A over the duration of sampling. BNF-treated salmon demonstrated similar levels of CYP1A to control fish at day 0 and higher levels over the course of each additional sampling point. Gill biopsies from wild salmon sampled from Millers River (South Royalston, Worcester County, MA, USA), known to contain PCBs, showed significantly higher CYP1A levels over an uncontaminated reference stream, Fourmile Brook (Northfield, Franklin County, MA, USA). We conclude that gill biopsies coupled with Q-RT-PCR analysis is a valuable tool in environmental assessment of wild Atlantic salmon populations and has the potential to be applied to other populations of fish as well.

Comparative Biochemistry and Physiology, Part C: T↗
USGS1997

Proximate composition and energy density of some North Pacific forage fishes

Mature pelagic forage fish species (capelin, sand lance, squid) had greater lipid concentrations than juvenile age-classes of large demersal and pelagic fish species (walleye pollock, Pacific cod, Atka mackerel, greenling, prowfish, rockfish, sablefish). Myctophids preyed on by puffins have at least twice as much lipid per gram compared to mature capelin, sand lance and squid, and an order of magnitude greater lipid concentrations than juvenile forage fish. Energy density of forage fishes was positively correlated with lipid content, and negatively correlated with water, ash-free lean dry mass (mostly protein), and ash contents.

Comparative Biochemistry and Physiology, Part A: M↗