Assessing the Quantity and Downstream Performance of DNA Isolated from Beluga (Delphinapterus leucas) Blow Samples

Richard, Justin T.; Schultz, Krystle; Goertz, Caroline E. C.; Hobbs, Rod S.; Sartini, Becky L.

doi:10.1578/am.43.4.2017.398

Cited by 4 publications

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“…As behavioural observations of belugas with known reproductive status during the breeding season are not feasible in the wild, determining the sex ratio, age structure and kinship within a population is necessary to determine the number of potential sires available and the proportion of potential sires that are reproducing. Advancing unmanned aerial vehicle sampling methodologies (Centelleghe et al 2020) could allow blow sampling for molecular purposes (Richard, Schultz et al 2017) to support these efforts, especially for young animals typically excluded from biopsy sampling (Citta et al 2018). Applying this study's approach of simultaneously monitoring behaviour and physiology to larger and more diverse social groups, made possible through the application of minimally invasive blow sampling, will also be an important component of these efforts.…”

Section: Discussionmentioning

confidence: 99%

Minimally invasive physiological correlates of social behaviour in belugas (Delphinapterus leucas) under human care

2021

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Simultaneous observations of reproductive physiology and behaviour were conducted on a group of two male and two female belugas under professionally managed care for one year to explore potential mating strategies. Weekly blow sampling for progesterone in females was used to define the breeding season by detecting three oestrous cycles in one female. Twice weekly blow sampling for testosterone and twice monthly testes measurements via ultrasonography were used to detect reproductive seasonality in both males. Female–male association frequency varied longitudinally, with 70% of all interactions occurring during the 16-week breeding season. Male–male associations did not vary seasonally. Male display behaviours towards the female occurred 14.8 times more frequently during the breeding season (0.164 ± 0.188 behaviours/min) than outside of the breeding season (0.011 ± 0.042 behaviours/min). The cycling female responded variably to male display behaviours by altering swim speed or body orientation towards the male. Although this small sample size limits broad conclusions, the frequent display behaviours, low copulation rate and lack of serious male–male aggression are consistent with predictions for pre-copulatory female mate choice developed from the current knowledge of beluga reproductive physiology. These observations, which are not feasible for wild belugas, provide important management considerations because reduced opportunities for mate choice could limit the reproductive rate, especially in small populations.

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Section: Discussionmentioning

confidence: 99%

Minimally invasive physiological correlates of social behaviour in belugas (Delphinapterus leucas) under human care

2021

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“…Furthermore, unlike feces and blubber that accumulate hormones over hours or days 29 , the hormone content in blow may reflect more rapid and short-term acute responses 30 associated with a particular event or stressor. As a more nascent sample type for hormone analysis, cetacean respiratory vapor has been shown to contain detectable levels of steroid (e.g., testosterone, progesterone, estradiol, estrone, cortisol) and thyroid hormones (e.g., thyroxine and triiodothyronine) 4 , 9 , 31 – 34 , and a vast number of volatile and nonvolatile compounds 34 – 36 , in addition to DNA from both the individual whale 37 – 39 and respiratory microorganisms 10 , 40 .…”

Section: Introductionmentioning

confidence: 99%

Quantifying hormones in exhaled breath for physiological assessment of large whales at sea

Burgess

Hunt

Kraus

et al. 2018

Sci Rep

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Exhaled breath analysis is a non-invasive assessment tool that has shown promise in human diagnostics, and could greatly benefit research, management, and conservation of large whales. However, hormone assessment of whale respiratory vapor (blow) has been challenged by variable water content and unknown total volume of collected samples. To advance this technique, we investigated urea (a compound present in narrow range in circulation) as a normalizing factor to correct for blow sample concentration. Normalized progesterone, testosterone, and cortisol concentrations of 100 blow samples from 46 photo-identified North Atlantic right whales (Eubalaena glacialis) were more biologically relevant compared to absolute estimates, varying by sex, age class, or individual. Progesterone was elevated in adult females compared with other cohorts and highest in one independently confirmed pregnant female. For both sexes, testosterone was two-fold higher in reproductively mature whales but studied adult females showed the widest variation. Cortisol was present in relatively low concentrations in blow and demonstrated variation between individual whales, suggesting potential for studies of individual differences in adrenal activity. Incorporation of methodologies that normalize sample concentration are essential for blow hormone analysis of free-swimming whales, and measurement of urea could be used to optimize non-invasive physiological assessment of whales.

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“…Therefore, the first exhale may have had the largest quantity of cells and microorganisms compared with successive exhales. A study of the effect of the number of exhales collected and DNA yield in belugas produced similar results, with declining mean and median DNA yields per exhale as the number collected increased from one to two to four ( Richard et al, 2017a ). Variation in blow sample volume in belugas ( Richard et al, 2017b ) and variation in the amount of cellular debris expelled per exhale could explain the variation in RNA yield observed in this study.…”

Section: Discussionmentioning

confidence: 75%

“…Blow samples consisting of one to six successive exhales were collected into a sterile polypropylene 50-ml conical tube (Fisher Scientific, Waltham, MA, USA, #14-432-22) held inverted ~5 cm directly above the blowhole and tilted cranially by 30–45° as described in Richard et al (2017a) . To simulate sample collection from a free-swimming beluga, no attempt was made to clear environmental water from the blowhole prior to sample collection.…”

Section: Methodsmentioning

confidence: 99%

“…Immune function gene expression studies have been performed in this species using other tissue sources ( Unal et al, 2018 ). Additionally, the utility of blow sampling for DNA analysis ( Richard et al, 2017a ) and the quantification of steroid hormones ( Thompson et al, 2014 ; Richard et al, 2017b ) have already been demonstrated in this species. In this study, a method for the isolation of total RNA from blow samples was described and the relationship between RNA yield and sample handling procedures was explored to determine the applicability of the sampling methods to field conditions.…”

Section: Introductionmentioning

confidence: 99%

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Evaluating beluga (Delphinapterus leucas) blow samples as a potential diagnostic for immune function gene expression within the respiratory system

Richard

Schultz

Goertz

et al. 2022

Conservation Physiology

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Evaluating respiratory health is important in the management of cetaceans, which are vulnerable to respiratory diseases. Quantifying the expression of genes related to immune function within the respiratory tract could be a valuable tool for directly assessing respiratory health. Blow (exhale) samples allow DNA analysis, and we hypothesized that RNA could also be isolated from blow samples for gene expression studies of immune function. We evaluated the potential to extract RNA from beluga blow samples and tested whether transcripts associated with immune function could be detected with endpoint polymerase chain reaction. A total of 54 blow samples were collected from clinically healthy aquarium belugas (n = 3), and 15 were collected from wild belugas temporarily restrained for health assessment in Bristol Bay, Alaska (n = 9). Although RNA yield varied widely (range, 0–265.2 ng; mean = 85.8; SD = 71.3), measurable RNA was extracted from 97% of the samples. Extracted RNA was assessed in 1–6 PCR reactions targeting housekeeping genes (Rpl8, Gapdh or ActB) or genes associated with immune function (TNFα, IL-12p40 or Cox-2). Fifty of the aquarium samples (93%) amplified at least one transcript; overall PCR success for housekeeping genes (96/110, 87%) and genes associated with immune function (90/104, 87%) were similarly high. Both RNA yield and overall PCR success (27%) were lower for wild beluga samples, which is most likely due to the reduced forcefulness of the exhale when compared with trained or free-swimming belugas. Overall, the high detection rate with PCR suggests measuring gene expression in blow samples could provide diagnostic information about immune responses within the respiratory tract. While further study is required to determine if quantitative gene expression data from blow samples is associated with disease states, the non-invasive nature of this approach may prove valuable for belugas, which face increasing anthropogenic disturbances.

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Assessing the Quantity and Downstream Performance of DNA Isolated from Beluga (Delphinapterus leucas) Blow Samples

Cited by 4 publications

References 0 publications

Minimally invasive physiological correlates of social behaviour in belugas (Delphinapterus leucas) under human care

Minimally invasive physiological correlates of social behaviour in belugas (Delphinapterus leucas) under human care

Quantifying hormones in exhaled breath for physiological assessment of large whales at sea

Evaluating beluga (Delphinapterus leucas) blow samples as a potential diagnostic for immune function gene expression within the respiratory system

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