Search Results (22)

The meat yield (MY) is a key economic trait in Pacific whiteleg shrimp (Penaeus vannamei) breeding, necessitating accurate genomic prediction for efficient genetic improvement. In this study, we investigated single-trait (STGMs) and multi-trait genomic models (MTGMs) for predicting MY and related traits, using two cross-validation strategies reflecting different data-availability scenarios. A total of 899 individuals from 63 full-sibling families were phenotyped for MY, net meat weight (MW), body weight (BW), body length (BL), and abdominal segment length (AL). We estimated the genomic heritability and genetic correlations of MY and related traits in P. vannamei, followed by comparing the prediction accuracy of STGMs and MTGMs for MY and MW. Two validation approaches were then applied: CV1 retained auxiliary traits in the validation sets, and CV2 excluded both target and auxiliary traits. Heritability estimates indicated that MY had low heritability (STGM: 0.160; MTGMs: 0.145–0.156), whereas MW, BW, BL, and AL showed low-to-moderate heritability (0.099–0.204). Genetic correlations revealed strong associations between MY and MW/BW/BL (r_g = 0.605–0.783), yet a low positive correlation with AL (r_g = 0.286). Across all comparisons, MTGMs consistently surpassed STGMs. For MY, MTGMs improved the accuracy by 4.8–58.8% relative to STGM (0.187), with the MY-MW model achieving the highest accuracy (0.297) under CV1. Similarly, MTGMs enhanced MW prediction by 36.6–138.2% over STGM (0.254), with the MW-BW model reaching the highest accuracy (0.605) under CV1. Notably, retaining auxiliary traits (CV1) boosted accuracy gains substantially (up to 138.2%), whereas excluding them (CV2) yielded only marginal improvements (≤8.6%). Moreover, incorporating AL as an auxiliary trait increased heritability estimates for MW, BW, and BL by 5.4–7.6%, indicating its synergistic value in MTGMs. Overall, these results demonstrate that MTGMs markedly enhance genomic prediction for carcass traits compared to STGMs, particularly when auxiliary trait data are accessible (CV1). The findings underscore the importance of maintaining auxiliary trait records in breeding populations, offering a robust framework for improving P. vannamei through multi-trait genomic prediction models. Full article

Harvest weight uniformity is a critical economic trait in the production of Pacific white shrimp (Penaeus vannamei). Social interactions among individuals can significantly influence both uniformity and productivity in aquaculture. To improve harvest weight uniformity through selective breeding, it is essential to accurately partition the genetic component of social effects, known as an indirect genetic effect (IGE), from purely environmental factors. Since IGEs cannot be estimated when all individuals are kept in a single group, a specialized experimental design, such as the grouping design with three families per group (3FAM), is required. With this experimental design, the shrimp population is divided into multiple groups (cages), each containing three families. Individuals from each family are then evenly subdivided and placed in three cages, thereby enabling the estimation of both direct and social genetic effects. Additionally, integrating genomic information instead of relying solely on pedigree data improves the accuracy of genetic relatedness among individuals, leading to more precise genetic evaluation. This study employed a 3FAM experimental design involving 40 families (36 individuals per family) to estimate the contribution of direct and indirect genetic effects on harvest weight uniformity. The genotypes of all tested individuals obtained using the 55K SNP panel were incorporated into a hierarchical generalized linear model to predict direct genetic effects and indirect genetic effects (IGE) separately. The results revealed that the heritability of harvest weight uniformity was low (0.005 to 0.017). However, the genetic coefficient of variation (0.340 to 0.528) indicates that using the residual variance in harvest weight as a selection criterion for improving uniformity is feasible. Incorporating IGE into the model increased heritability estimates for uniformity by 150% to 240% and genetic coefficient of variation for uniformity by 32.11% to 55.29%, compared to the model without IGE. Moreover, the genetic correlation between harvest weight and its uniformity shifted from a strongly negative value (−0.862 to −0.683) to a weakly positive value (0.203 to 0.117), suggesting an improvement in the genetic relationship between the traits and better separation of genetic and environmental effects. The inclusion of genomic data enhanced the prediction ability of single-step best linear unbiased prediction for both harvest weight and uniformity by 6.35% and 10.53%, respectively, compared to the pedigree-based best linear unbiased prediction. These findings highlight the importance of incorporating IGE and utilizing genomic selection methods to enhance selection accuracy for obtaining harvest weight uniformity. This approach provides a theoretical foundation for guiding uniformity improvements in shrimp breeding programs and offers potential applications in other food production systems. Full article

Snapping Shrimps (SSs) live in warm marine areas. Snapping Shrimps Noise (SSN), loud sounds generated by these underwater creatures, serves as a major source of in performance degradation by decreasing the Signal-to-Noise Ratio (SNR) for underwater acoustic communication and target detection. Thus, we propose a unified solution for SSN detection and reduction in this paper. First, Signal Presence Probability (SPP) is calculated for SSN detection, and then the SPP is provided to Non-negative Matrix Factorization (NMF) as a weight for SSN reduction. In the proposed method, SPP acts as a key factor for SSN detection and reduction. To verify the effectiveness of the proposed method, the SAVEX-15 dataset, real ocean data containing SSN, is used. As a result of SSN detection, it was confirmed that SPP presented the highest performance in the Receiver Operating Characteristics curve, and we achieved 0.014 higher Area Under the Curve compared to competing methods. In addition, Continuous Wave and Linear Frequency Modulation signals were set as target signals and combined with the SAVEX-15 data for evaluation of noise reduction performance. As a result, the performance of the SPP-weighted NMF (WNMF) presented at least 2 dB higher SNR and SDR while maintaining less LSD compared to the Optimally Modified Log Spectral Amplitude estimator and NMF. Full article

Shrimp farming is a growing industry, and automating certain processes within aquaculture tanks is becoming increasingly important to improve efficiency. This paper proposes an image-based system designed to address four key tasks in an aquaculture tank with Penaeus vannamei: estimating shrimp length and weight, counting shrimps, and evaluating feed pellet food attractiveness. A setup was designed, including a camera connected to a Raspberry Pi computer, to capture high-quality images around a feeding plate during feeding moments. A dataset composed of 1140 images was captured over multiple days and different times of the day, under varying lightning conditions. This dataset has been used to train a segmentation model, which was employed to detect and filter shrimps in optimal positions for dimensions estimation. Promising results were achieved. For length estimation, the proposed method achieved a mean absolute percentage error (MAPE) of 1.56%, and width estimation resulted in a MAPE of 0.15%. These dimensions were then used to estimate the shrimp’s weight. Shrimp counting also yielded results with an average MAPE of 7.17%, ensuring a satisfactory estimation of the population in the field of view of the image sensor. The paper also proposes two approaches to evaluate pellet attractiveness, relying on a qualitative analysis due to the challenges of defining suitable quantitative metrics. The results were influenced by environmental conditions, highlighting the need for further investigation. The image capture and analysis prototype proposed in this paper provides a foundation for an adaptable system that can be scaled across multiple tanks, enabling efficient, automated monitoring. Additionally, it could also be adapted to monitor other species raised in similar aquaculture environments. Full article

Selecting shrimp strains that perform well on a plant-based diet holds promise for solving the fishmeal (FM) shortage issue, with genetic improvement of feed efficiency (FE) being a key focus. This study compared a plant-based (fishmeal-free) diet with a 25% FM diet for the Pacific whiteleg shrimp (Penaeus vannamei) to determine whether the FE-related traits, including residual feed intake (RFI), average daily weight gain (ADG), and daily feed intake (DFI), exhibit diet-specific genetic variations and genotype-by-diet (genotype × diet) interactions. Results showed that RFI was significantly higher under the plant-based diet treatment compared to the FM-based diet treatment (0.0049 g/d vs. –0.0045 g/d, p < 0.001), while ADG was notably lower (0.043 g/d vs. 0.110 g/d, p < 0.001), indicating that a plant-based diet may lead to reduced yields and impact the profitability of shrimp culture. In the case of being unable to estimate common environmental effects, the heritability estimates of RFI under the plant-based and FM-based diets were 0.743 ± 0.157 and 0.440 ± 0.125, respectively, and those of ADG and DFI ranged from 0.314 ± 0.121 to 0.947 ± 0.158. Genetic correlations between diets for RFI and DFI were 0.646 ± 0.162 and 0.549 ± 0.163, respectively, suggesting a degree of genotype × diet interactions. In contrast, the genetic correlation between diets for ADG was only 0.296 ± 0.259, indicating a strong genotype × diet interaction. These findings highlight the importance of considering selective breeding for improved feed efficiency in a plant-based diet environment. Full article

Information about the population structure, including the age and sex composition, growth characteristics, and diet of fish, is essential for the conservation and sustainable exploitation of fish stock. The burbot, Lota lota, is the only freshwater species of the family Lotidae in the Amur (Heilongjiang) River catchment located in northeastern China. Information on the biological characteristics and data on the population structure of this fish from the Amur River are scarce. To study these factors in burbot, Lota lota, in the Amur River of China, 749 specimens from four sampling areas were taken and analyzed in October 2022, January 2023, and May 2023. The ages of the sampled fish ranged from 1+ to 7+ years. The female/male ratio was 1.04:1, and body length and mass varied from 175 to 595 mm and 73.5 to 1958.7 g, respectively. The length–weight regression parameter b value was estimated as 2.80. The parameters of the von Bertalanffy growth model were L_∞ = 596, K = 0.221, and t₀ = −0.771 for all sampled fish; L_∞ = 625, K = 0.208, and t₀ = −0.756 for females; and L_∞ = 584, K = 0.219, and t₀ = −0.980 for males. The analysis of the stomach contents showed fish to be the major source of nutrition across all areas and sampling periods. In January, the secondary prey of burbots in the Huma reach of the upper Amur River was aquatic insect larvae, while, in the Tongjiang and Fuyuan reaches of the middle Amur River, the secondary prey was shrimp. The mean stomach fullness index and rate of empty stomachs differed with the sampling area, with the greatest proportion of empty stomachs observed in the Luobei reach of the middle Amur River, and the lowest in the Tongjiang and Fuyuan reaches. The mean stomach fullness index showed the opposite trend. In the Huma reach, the contributions of fish to the diet and the mean stomach fullness index were significantly higher in May than in October and January. The study provides the first detailed information on the population age and sex structure, growth patterns, and feeding ecology of burbots from the Amur River, China. The results will aid in formulating management strategies and regulations for local populations of burbots in the Amur River, China. Full article

The genotype-by-environment interaction (G × E) might have crucial impacts on the performance and fitness of agricultural species, such as Pacific whiteleg shrimp (Litopenaeus vannamei). This study explores how enhancements in management practices can counteract G × E effects on growth traits. We analyzed a selectively bred population of whiteleg shrimp spanning the latest two generations, encompassing 259 full-sib and half-sib families with 40,862 individual shrimp, measured for body weight and total length. Our analysis revealed moderate genetic correlations (0.60–0.65) between trait expressions in pond and tank environments, a significant improvement compared to earlier generations. Employing the average information-restricted maximum likelihood (REML) approach in mixed model analysis showed significant differences in heritability (h²) estimates between the two environments; however, the extent of these differences varied by trait (h² = 0.68 in pond vs. 0.37 in tank for weight, and 0.41 vs. 0.67 for length). Our results indicate that G × E effects on growth traits in this population of L. vannamei were moderate but biologically significant. Consistent with our previous estimates in this population, genetic correlations between body weight and total length remained high (close to one) in pond and tank environments. The present findings collectively demonstrate that management improvements targeting stocking density, aeration, water quality, feeds, and feeding regimes mitigated the G × E effects on two economically significant traits in this population of whiteleg shrimp. Full article

Fisheries management requires improvement in scientific knowledge to ensure sustainable exploitation of important commercial species and population renewal. Within this context, this study focused on the reproductive biology of spot tail mantis shrimp, Squilla mantis, in the Central Mediterranean Sea, aiming to understand the reproductive pattern of this species and validate the macroscopic maturity scale through histological analysis. A multi-year sampling was performed from 2016 to 2020 by a commercial fishing fleet in the Northern Central Adriatic Sea (GSA 17), and a total of 2206 individuals were collected. The monthly average value of the total sex ratio of S. mantis was 0.524 ± 0.044 (mean ± SEM) in favor of females, which dominated the population from September to April. The proposed 5 stage macroscopic maturity scale was validated histologically through histological analysis, confirming synchronous ovarian development. The somatic indexes (GSI and K Fulton) and monthly incidence of macroscopic ovarian maturity stages highlighted a protracted reproductive season from winter to spring (January–May). Although the length-weight relationship showed a similar growth trend between genders, males reached a bigger size in terms of carapace length (C.L.) and dominated the population from 32 mm (C.L.). The macroscopic L50 estimated was 25.94 mm (C.L.). Full article

The current study aimed to provide a precise assessment of the genetic parameters associated with growth and white spot syndrome virus (WSSV) resistance traits in Pacific white shrimp (Litopenaeus vannamei). This was achieved through a controlled WSSV challenge assay and the analysis of phenotypic values of five traits: body weight (BW), overall length (OL), body length (BL), tail length (TL), and survival hour post-infection (HPI). The analysis included test data from a total of 1017 individuals belonging to 20 families, of which 293 individuals underwent whole-genome resequencing, resulting in 18,137,179 high-quality SNP loci being obtained. Three methods, including pedigree-based best linear unbiased prediction (pBLUP), genomic best linear unbiased prediction (GBLUP), and single-step genomic BLUP (ssGBLUP) were utilized. Compared to the pBLUP model, the heritability of growth-related traits obtained from GBLUP and ssGBLUP was lower, whereas the heritability of WSSV resistance was higher. Both the GBLUP and ssGBLUP models significantly enhanced prediction accuracy. Specifically, the GBLUP model improved the prediction accuracy of BW, OL, BL, TL, and HPI by 4.77%, 21.93%, 19.73%, 19.34%, and 63.44%, respectively. Similarly, the ssGBLUP model improved prediction accuracy by 10.07%, 25.44%, 25.72%, 19.34%, and 122.58%, respectively. The WSSV resistance trait demonstrated the most substantial enhancement using both genomic prediction models, followed by body size traits (e.g., OL, BL, and TL), with BW showing the least improvement. Furthermore, the choice of models minimally impacted the assessment of genetic and phenotypic correlations. Genetic correlations among growth traits ranged from 0.767 to 0.999 across models, indicating high levels of positive correlations. Genetic correlations between growth and WSSV resistance traits ranged from (−0.198) to (−0.019), indicating low levels of negative correlations. This study assured significant advantages of the GBLUP and ssGBLUP models over the pBLUP model in the genetic parameter estimation of growth and WSSV resistance in L. vannamei, providing a foundation for further breeding programs. Full article

Intertidal macroinvertebrates, such as crabs and mud shrimps, are invertebrates inhabiting the intertidal zone that are sufficiently large to be identified with the naked eye. Most intertidal macroinvertebrates typically construct burrows of various shapes in sediment to protect themselves from environmental extremes, with surface openings that reflect features of their species and body size. Especially circular burrow openings correlate with an organism’s body size; thus, measuring these openings can provide estimates of the organism’s size, weight, growth rate, and biomass. Traditional studies of these organisms have relied on field surveys, which are constrained by time, cost, and logistical limitations. This study introduces an innovative method for measuring the burrow opening diameters of intertidal macroinvertebrates using high-resolution optical images from a portable drone system. By leveraging the reflectance disparity between the sediment and burrow openings, this method facilitates the extraction and sizing of burrow openings. Our methodology was applied to three crab species known for their circular burrow opening: the red-clawed fiddler, the milky fiddler, and the ghost crab. Validation was confirmed through field data from the Mageumri and Sinduri tidal flats, South Korea. The method achieved a correlation coefficient (R²) of 0.94 and a root mean squared error (RMSE) of 1.68 mm across a diameter range of 6.21–33.59 mm. These findings suggest the potential of drone remote sensing systems as a non-invasive and efficacious approach for quantifying burrow sizes over extensive intertidal areas, thereby facilitating more accurate biomass estimations and surmounting the limitations of conventional field surveys. Future research could extend this method to additional species and further refine its precision. Full article

An 8-week feeding trial was conducted to assess the effects on growth, antioxidant capacity, digestive enzyme activity, and gene expression related to muscle growth and protein synthesis of juvenile greasyback shrimp (Metapenaeus ensis) using five experimental diets containing 29.37%, 34.30%, 39.11%, 44.05%, and 49.32% of protein. The results demonstrated that juvenile greasyback shrimp consuming 39.11%, 44.05%, and 49.32% dietary protein had a significantly higher final body weight (FBW), weight gain (WG), feed conversion ratio (FCR), and specific growth rate (SGR) than other groups (p < 0.05). The protein efficiency ratio (PER) showed a significantly quadratic pattern with increasing dietary protein levels (p < 0.05). The highest trypsin and pepsin activities were observed in the group with a protein level of 44.05% (p < 0.05). Relatively higher superoxide dismutase (SOD) activity was found in groups with protein levels of 39.11% (p < 0.05). Alkaline phosphatase (AKP) and catalase (CAT) activity showed a significantly linear increasing pattern with increasing protein intake up to 44.05%, and then decreased gradually (p < 0.05). Compared to the dietary 29.37% protein level, the expression levels of myogenic regulatory factors (mef2α, mlc, and myf5) and mTOR pathway (mtor, s6k, akt, and pi3k)-related genes were significantly up-regulated in muscle with 39.11%, 44.05%, and 49.32% dietary protein levels (p < 0.05). The AAR pathway (gcn2, eif2α, and atf4)-related gene expression levels were significantly lower in muscles with 39.11%, 44.05%, and 49.32% protein levels than in other groups (p < 0.05). Based on the broken-line regression analysis of SGR, the estimated appropriate dietary protein requirement for juvenile greasyback shrimp is 38.59%. Full article

(1) Background: Although studies have suggested that dietary interventions may have potential benefits over conventional medical treatments, research on the association between dietary patterns and hyperemesis gravidarum (HG) in pregnant women is scarce. (2) Methods: To explore the relationship between dietary patterns and the risk of HG, a cross-sectional study was conducted in Xi’an, China from April 2021 to September 2022. Dietary intake was assessed by a semi-quantitative food-frequency questionnaire, and then factor analysis was used to derive dietary patterns. HG was defined as persistent and severe nausea and vomiting with weight loss ≥ 5%, pregnancy-unique quantification of emesis (PUQE) score ≥ 13, or hospitalization due to vomiting. Logistic regression models were used to estimate ORs and 95% CIs for HG according to dietary pattern scores. Stratified analyses and tests for interaction were performed by potential confounders. (3) Results: Of the 3122 pregnant women enrolled, 2515 individuals (mean age: 31.2 ± 3.4 years) were included in the final analysis. In total, 226 (8.9%) pregnant women were identified as having HG. Five dietary patterns were identified. After adjusting for covariates, the highest quartile of the “fish, shrimp and meat” and “egg, milk and water drinking” patterns was associated with a 37% and 58% lower risk of HG compared with the lowest quartile, respectively (p-trend < 0.05). Conversely, the highest quartile of the “beverage” pattern was associated with a 64% higher risk of HG compared with the lowest quartile (p-trend = 0.02). Furthermore, significant interactions were observed between the “egg, milk and water drinking” pattern and parity, employment status and nutritional supplement use (p-interaction < 0.05). (4) Conclusions: A diet rich in eggs, milk, seafood and unprocessed poultry and animal meat may be a protective factor against HG, while a diet high in beverages may be detrimental to HG. These associations may vary by parity, employment status and nutritional supplement use. Full article

Porphyry and epithermal deposits are important sources of base and precious metals. Most actively mined deposits have been exhumed such that ore bodies are relatively close to the surface and are therefore locatable and economic to extract. Identifying and characterizing concealed deposits, particularly more deeply buried porphyry deposits, represents a far greater challenge for mineral exploration, and will become progressively more important as near-surface resources are gradually exhausted over time. We report high-precision ⁴⁰Ar/³⁹Ar dates for coarsely crystalline alunite that precipitated from magmatic steam in open fractures in Oligocene dacitic volcanic rocks, and a SHRIMP ²⁰⁶Pb/²³⁸U zircon date for one of several rhyolite dikes present at Alunite Ridge and Deer Trail Mountain, Utah. Both the magmatic-steam alunite and rhyolite dikes are related to concealed intrusions. The rhyolite dike yielded an age of 30.72 ± 0.36 Ma, which is older than a commonly cited 27.1 Ma age estimate for the Three Creeks Tuff Member of the Bullion Canyon Volcanics that is cut by the dike. ⁴⁰Ar/³⁹Ar data for samples of magmatic-steam alunite and sericite from six mines and prospects provide evidence for at least two periods of episodic hydrothermal activity at ca. 15.7–15.1 Ma and ca. 14.7–13.8 Ma, with the older and younger pulses of activity recorded at the more eastern and western sites, respectively. These two periods of hydrothermal activity are consistent with previous interpretations that Alunite Ridge and Deer Trail Mountain are underlain by two concealed porphyry stocks. ⁴⁰Ar/³⁹Ar analyses of individual bands in a sample of massive, centimeter-scale banded vein alunite yield indistinguishable ages with a weighted mean of 13.98 ± 0.12 Ma, consistent with a short-lived (≲250 ka) magmatic event with episodic vapor discharge recurring on short timescales (≲36 ka). ⁴⁰Ar/³⁹Ar geochronology of magmatic-steam alunite is a valuable tool to constrain the timing and duration of magmatic hydrothermal activity associated with unexposed intrusions and potentially porphyry deposits, and therefore may be useful in exploration. Full article

Salt mining and shrimp farming have been practiced in the Non Thai district and the surrounding areas for more than 30 years, creating saline soil problems. To solve the soil salinity problem, soil salinity prediction and mapping utilizing the electromagnetic induction method (EMI) and spatial interpolation methods were examined in the Non Thai district, Nakhon Ratchasima province, Thailand. The research objectives were (1) to predict soil salinity using spatial interpolation methods and (2) to identify a suitable spatial interpolation method for soil salinity severity mapping. The research methodology consisted of five steps: apparent electrical conductivity (ECa) measurement using an electromagnetic induction (EMI) method; in situ soil sample collection and electrical conductivity of the saturated soil paste extract (ECe) measurement; soil electrical conductivity estimation using linear regression analysis (LRA); soil salinity prediction and accuracy assessment; and soil salinity severity classification and overlay analysis with relevant data. The result of LRA showed a strong positive relationship between ECe and ECa. The correlation coefficient (R) values of a horizontal measuring mode (HH) and a vertical measuring mode (VV) were 0.873 to 0.861, respectively. Four selected interpolation methods—Inverse Distance Weighting (IDW), Ordinary Kriging (OK), Ordinary CoKriging (OCK) with soil moisture content, and Regression Kriging (RK) without covariable factor—provided slightly different patterns of soil salinity prediction with HH and VV modes. The mean values of the ECe prediction from the four methods at the district level varied from 2156.02 to 2293.25 mS/m for HH mode and from 2377.38 to 2401.41 mS/m for VV mode. Based on the accuracy assessment with the rank-sum technique, the OCK is a suitable interpolation method for soil salinity prediction for HH mode. At the same time, the IDW is suitable for soil salinity prediction for the VV mode. The dominant soil salinity severity classes of the two measuring modes using suitable spatial interpolation methods were strongly and very strongly saline. Consequently, the developed research methodology can be applied to conduct soil salinity surveys to reduce costs and save time in other areas by government agencies in Thailand. Nevertheless, to apply the EMI method for soil salinity survey, the users should understand the principle of EMI and how to calibrate and operate the EM device properly for accurate ECa measurement. Full article

Cadmium (Cd) and its compounds are hazardous environmental pollutants with renal toxicity and human carcinogenicity, with ingestion of contaminated foods representing the major mode of exposure. There have been a number of reports evaluating the Cd content in various foods; however, regarding the actual risk posed by dietary cadmium exposure, only a few reports are available in which single point evaluation (less accurate than multiple point evaluation) was employed. In this study, we used a margin of exposure (MOE) model and @RISK software (for multiple evaluation) to evaluate Cd-related health risk in the local Guangzhou residents at varying ages, through a comparison between the estimated monthly exposures and the provisional tolerable monthly intake (0.025 mg/kg body weight (b.w.)), based on the Cd contents in various food categories available locally (a total of 3964 food samples were collected from each of the 13 districts of Guangzhou between 2015 and 2019), which were determined by using inductively coupled plasma mass spectrometry. In this study, Cd was detected in 69.6% of the samples (averaged 0.120 mg/kg), and rice and its products, leafy vegetables, bivalves, and shrimp and crabs contributed most to Cd exposure (8.63, 3.18, 2.79, and 1.48 ng/kg b.w./day, respectively). The MOE values demonstrated the following tendency: the younger age group, the lower MOE, and its 95% confidence range for the (youngest) 3~6 year old group started from 0.92, indicating a health risk of young children, while that for the other age groups were all above 1.0. Our preliminary findings warrant further clarification using biomarker assays in the relevant population. Full article