Genetic Diversity and Population Structure of Mesoamerican Scarlet Macaws in an Ex Situ Breeding Population in Mexico

Given the interest in the conservation of the Mesoamerican scarlet macaw (Ara macao cyanoptera), the Xcaret Park formed an initial reproductive population about 30 years ago, which has progressively grown to a considerable population in captivity. In this work, we focus on the evaluation of the genetic diversity of the captive population, taking two groups into account: its founding (49) and the current breeding individuals (166). The genetic analysis consisted of genotyping six nuclear microsatellite loci that are characterized by their high variability. Tests for all loci revealed a Hardy–Weinberg equilibrium in four loci of the founders and in no loci of the breeding groups. The results showed that the genetic variation in the Xcaret population was relatively high (founders He = 0.715 SE = 0.074, breeding pairs He = 0.763 SE = 0.050), with an average polymorphism of 7.5 (4–10) alleles per locus in founders and 8.3 (4–14) in breeding pairs. No significant differences in the evaluated genetic diversity indexes were found between both groups. This indicates that the genetic variability in Xcaret has been maintained, probably due to the high number of pairs and the reproductive management strategy. Bayesian analysis revealed five different genetic lineages present in different proportions in the founders and in the breeding pairs, but no population structure was observed between founders and breeding individuals. The analyzed captive individuals showed levels of genetic diversity comparable to reported values from Ara macao wild populations. These data indicate that the captive population has maintained a similar genetic diversity as the metapopulation in the Mayan Forest and is an important resource for reintroduction projects, some of which began more than five years ago and are still underway.

Rediscovery of the rare Phrynomedusa appendiculata (Lutz, 1925) (Anura: Phyllomedusidae) from the Atlantic Forest of southeastern Brazil

The genus Phrynomedusa Miranda-Ribeiro, 1923 comprises rare and little known phyllomedusid species from southern Atlantic Forest, Brazil. Phrynomedusa appendiculata (Lutz, 1925) is known from three localities since its description and considered a “lost species” because it was last sighted 51 years ago. This pervasive lack of knowledge raised a significant concern about its threat status. Here, we present the rediscovery of P. appendiculata from a breeding population in the Atlantic Plateau forests of the state of São Paulo. This new record allowed the gathering of novel ecological, acoustic and morphological data for this species. Most of the novel data agreed with the variation historically reported for the species, but we found subtle divergences that we interpret as intraspecific variation. Moreover, this record also allowed a reassessment of geographic distribution of the species, and the first inference of its phylogenetic relationships based on molecular data (mitochondrial and nuclear DNA). The resulting phylogeny corroborated the generic placement and evolutionary distinctiveness of P. appendiculata, evidencing the species as sister to the clade P. marginata + P. dryade. Based on novel and historical data, we discuss some putative factors influencing the rarity of P. appendiculata and its congeners, and provide conservation perspectives. We expect that the novel data can support further assessments of threat status for this rare species, as well as initiatives aiming its conservation.

Exploiting High-Throughput Indoor Phenotyping to Characterize the Founders of a Structured B. napus Breeding Population

Phenotyping is considered a significant bottleneck impeding fast and efficient crop improvement. Similar to many crops, Brassica napus, an internationally important oilseed crop, suffers from low genetic diversity, and will require exploitation of diverse genetic resources to develop locally adapted, high yielding and stress resistant cultivars. A pilot study was completed to assess the feasibility of using indoor high-throughput phenotyping (HTP), semi-automated image processing, and machine learning to capture the phenotypic diversity of agronomically important traits in a diverse B. napus breeding population, SKBnNAM, introduced here for the first time. The experiment comprised 50 spring-type B. napus lines, grown and phenotyped in six replicates under two treatment conditions (control and drought) over 38 days in a LemnaTec Scanalyzer 3D facility. Growth traits including plant height, width, projected leaf area, and estimated biovolume were extracted and derived through processing of RGB and NIR images. Anthesis was automatically and accurately scored (97% accuracy) and the number of flowers per plant and day was approximated alongside relevant canopy traits (width, angle). Further, supervised machine learning was used to predict the total number of raceme branches from flower attributes with 91% accuracy (linear regression and Huber regression algorithms) and to identify mild drought stress, a complex trait which typically has to be empirically scored (0.85 area under the receiver operating characteristic curve, random forest classifier algorithm). The study demonstrates the potential of HTP, image processing and computer vision for effective characterization of agronomic trait diversity in B. napus, although limitations of the platform did create significant variation that limited the utility of the data. However, the results underscore the value of machine learning for phenotyping studies, particularly for complex traits such as drought stress resistance.

Genetic Variation of a Lentil (Lens culinaris) Landrace during Three Generations of Breeding

Genetic differentiation between 40 lentil genotypes was tested using molecular markers. The genotypes were produced from a Greek landrace of commercial interest via the honeycomb breeding methodology, i.e., single-plant selection in the absence of competition, across three successive pedigree generations. The selected genotypes from each generation were examined for genetic relationships using 15 SSR molecular markers with HRM analysis. As expected, low variation among consecutive generations at the level of 2.5–7.7% was detected. Analysis of molecular variance (AMOVA) revealed that partitioning of this variation was at higher percentage within each generation’s population than between them. Population structure analysis indicated that ongoing selection could effectively shift the allelic composition in each generation. The applied honeycomb breeding methodology that effectively improved progeny yield and seed quality increased the percentage of favorable alleles altering allelic composition but not eliminating genetic variation of the breeding population.

Breeding of a Wild Population of South Pacific Bonito Sarda chiliensis chiliensis (Cuvier 1832) Broodstock under Laboratory Conditions in Pisagua, Northern Chile

The wild population of South Pacific bonito Sarda chiliensis chiliensis, which has a wide distribution in northern Chile, is considered of importance in Chilean aquaculture. The biological feasibility of cultivation of any marine species begins with the establishment of an initial broodstock population to obtain eggs, larvae, and juveniles. In this work, 22 South Pacific bonito fishing campaigns were carried out in Pisagua, Chile, between spring in November 2011 and the summer in January 2012. At least 74 specimens were obtained of which 24 survived the capture and transport processes. Fish were stocked in a recirculating land-based aquaculture system, and at 14 months under captivity, fish began spawning. Eggs were collected, to describe some stages of development, and were placed in incubators at 20 °C and on the third-day eggs hatched. Larvae reached a total length between 1.435 and 1.7 mm, which were accurately characterized during their first morphological changes. This is the first work that describes the capture, transport, and acclimatization in captivity of a breeding population of wild Pacific bonito in Chile.

Reviving ghost alleles: Genetically admixed coyotes along the American Gulf Coast are critical for saving the endangered red wolf

The last red wolves were captured along the Gulf Coast in 1980, where they hybridized with coyote, to establish the captive breeding population. However, red wolf ancestry persists in local coyotes and could be leveraged by genomic innovations to support species persistence. We assessed genomic ancestry and morphology of coyotes in southwestern Louisiana, and find they carried 38-62% red wolf ancestry acquired in the last 30 years, which is enriched on land with minimal coyote hunting. These coyotes were also similar in ancestry to canids captured in the 1970s that initiated the red wolf captive breeding program. Further, we reported that coyotes with higher red wolf ancestry are larger in size. Our findings evidence the importance of hybrids as a reservoir of endangered species ancestry for contemporary conservation efforts. Admixed genomes are at the forefront of innovative solutions, with red wolf survival a prime candidate for this new paradigm.

Phenomics-Assisted Selection for Herbage Accumulation in Alfalfa (Medicago sativa L.)

The application of remote sensing in plant breeding is becoming a routine method for fast and non-destructive high-throughput phenotyping (HTP) using unmanned aerial vehicles (UAVs) equipped with sensors. Alfalfa (Medicago sativa L.) is a perennial forage legume grown in more than 30 million hectares worldwide. Breeding alfalfa for herbage accumulation (HA) requires frequent and multiple phenotyping efforts, which is laborious and costly. The objective of this study was to assess the efficiency of UAV-based imagery and spatial analysis in the selection of alfalfa for HA. The alfalfa breeding population was composed of 145 full-sib and 34 half-sib families, and the experimental design was a row-column with augmented representation of controls. The experiment was established in November 2017, and HA was harvested four times between August 2018 and January 2019. A UAV equipped with a multispectral camera was used for HTP before each harvest. Four vegetation indices (VIs) were calculated from the UAV-based images: NDVI, NDRE, GNDVI, and GRVI. All VIs showed a high correlation with HA, and VIs predicted HA with moderate accuracy. HA and NDVI were used for further analyses to calculate the genetic parameters using linear mixed models. The spatial analysis had a significant effect in both dimensions (rows and columns) for HA and NDVI, resulting in improvements in the estimation of genetic parameters. Univariate models for NDVI and HA, and bivariate models, were fit to predict family performance for scenarios with various levels of HA data (simulated in silico by assigning missing values to full dataset). The bivariate models provided higher correlation among predicted values, higher coincidence for selection, and higher genetic gain even for scenarios with only 30% of HA data. Hence, HTP is a reliable and efficient method to aid alfalfa phenotyping to improve HA. Additionally, the use of spatial analysis can also improve the accuracy of selection in breeding trials.

Aleutian Tern Onychoprion aleuticus abundance estimates at four globally significant colonies

Summary Aleutian Tern Onychoprion aleuticus numbers have been in steep decline at known Alaskan breeding colonies in recent decades (IUCN recently uplisted to ‘Vulnerable’). Available data suggest that most of the species may currently breed in Russia. Efforts to document global abundance and trends have been hampered by remoteness of colonies, lack of a formal monitoring programme, and the absence of reproducible population estimates with quantifiable errors, especially for large colonies. We surveyed four historically large colonies in Russia (2018) and Alaska (2019), which together may comprise 30–50% of the global breeding population. At each colony we obtained high resolution aerial photographs using a small Unmanned Aircraft System (sUAS). The large size of the colonies and the minimum altitude required to identify terns made it impractical to collect imagery of the entire colony. Instead, we employed a sampling approach, with sample locations selected based on spatially balanced acceptance sampling. Statistically sampled, low altitude sUAS images provided a fast, reproducible, and rigorous count of abundance for geographically large colonies, with low disturbance, and were generally consistent with concurrent ground-based observations. Concurrence among observers in photo counts indicated high precision in counts of attending birds and unattended nests, although species attribution in mixed tern colonies remains a source of significant uncertainty. Our results indicate that the four colonies surveyed here together supported <2,500 pairs of Aleutian Terns in the survey years. None of the colonies approached their peak size reported previously, likely due to recent predation, long-term decline, cold early season weather, or other factors. If these reduced colony sizes are representative of the current conditions, the implications for the global population would be dire.

Status, distribution and conservation of Kentish plover Charadrius alexandrinus (Aves, Charadriiformes) in Sicily

The Kentish Plover Charadrius alexandrinus is a sensitive species whose national breeding population has been undergoing a sharp decrease, up to 50%, in 2010 compared to the previous decade. Due to the lack of updated data and fragmented knowledge about its current status and distribution in Sicily, a specific and accurate monitoring of this species was carried out during the years 2018 and 2019. About 230 km of coastline and 64 UTM squares 10x10 km were investigated during the breeding seasons, and a total of 205-287 pairs have been estimated for the whole region, distributed in 41 UTM squares (including the island of Favignana). Pairs are mainly concentrated in the complex of “Saline di Trapani and Stagnone di Marsala” and along the sandy coast of Gulf of Gela, with a higher abundance along the southeast coast of the region. Breeding population showed a decrease of 18-28% in number of pairs in comparison to the last estimation of 250-400 pairs reported for the years 2009-2010. After the investigation of 108 km of coastline and 46 UTM squares, the Sicilian wintering population has been estimated at 376- 563 ind., with wintering flocks mainly concentrated in the wetlands of “Mazara del Vallo” and along the coasts of Syracuse and the Gulf of Gela. These updated estimates place Sicily as the second most important Italian region for the wintering of this species, just after Sardinia. For the first time, the data collected have made it possible to draw up qualitative-quantitative maps of C. alexandrinus distribution in Sicily, providing an important instrument for the management and planning of specific conservation actions, which should be based on habitat protection and implementation of appropriate artificial ecological niches.