Crocodylus siamensis) and least-concern saltwater crocodile (C. porosus) pose a significant challenge to conservation efforts due to human activities. Previous studies used microsatellite and mitochondrial DNA data to identify these hybrids, but potential biases and genetic drift within the populations could influence the results. To address these limitations and gain a clearer understanding of the genetic dynamics, we employed DArT sequencing to identify genome-wide single nucleotide polymorphisms (SNPs) in both species and confirm hybrid scenarios. We compared a population of Australian saltwater crocodiles to assess species-specific SNP distribution. Various analytical approaches were used to diagnose hybridization levels, especially in cases with potential backcrossing. Approximately 17.00–26.00% of loci were shared between Siamese and saltwater crocodile genomes. We identified 8051 species-specific SNP loci for Siamese crocodiles and 1288 for saltwater crocodiles. Using a PCR-based approach, three SNP loci were developed as markers, successfully enabling the distinction between species and various levels of hybridization. By combining mitochondrial and nuclear genetic information with species-diagnostic DNA markers, we developed a novel method for conservation prioritization, aiding long-term species survival through reintroduction and management programs.Hybrids between the critically endangered Siamese crocodile (Crocodylus siamensis) and least-concern saltwater crocodile (C. porosus) in captive populations represent a serious challenge for conservation and reintroduction programs due to the impact of anthropogenic activities. A previous study used microsatellite and mitochondrial DNA data to establish the criteria for identifying species and their hybrids; however, the results may have been influenced by biased allelic frequencies and genetic drift within the examined population. To overcome these limitations and identify the true signals of selection, alternative DNA markers and a diverse set of populations should be employed. Therefore, this study used DArT sequencing to identify genome-wide single nucleotide polymorphisms (SNPs) in both species and confirm the genetic scenario of the parental species and their hybrids. A population of saltwater crocodiles from Australia was used to compare the distribution of species-diagnostic SNPs. Different analytical approaches were compared to diagnose the level of hybridization when an admixture was present, wherein three individuals had potential backcrossing. Approximately 17.00–26.00% of loci were conserved between the Siamese and saltwater crocodile genomes. Species-diagnostic SNP loci for Siamese and saltwater crocodiles were identified as 8051 loci and 1288 loci, respectively. To validate the species-diagnostic SNP loci, a PCR-based approach was used by selecting 20 SNP loci for PCR primer design, among which 3 loci were successfully able to differentiate the actual species and different hybridization levels. Mitochondrial and nuclear genetic information, including microsatellite genotyping and species-diagnostic DNA markers, were combined as a novel method that can compensate for the limitations of each method. This method enables conservation prioritization before release into the wild, thereby ensuring sustainable genetic integrity for long-term species survival through reintroduction and management programs.Hybrids between the critically endangered Siamese crocodile ( Crocodylus spp.) : 0.002 ± 0.000), and the He values ranged from 0.000 to 0.079 (mean ± SE: 0.046 ± 0.001) , whereas the pairwise He values showed differences between one pair (CSI-CPOTH)  was −0.037, and the mean relatedness values of CSI, CPOTH, and CPOAU were −0.035 ± 0.036 (−0.971–0.071), −0.043 ± 0.024 (−0.478–0.043), and −0.035 ± 0.030 (−0.517–0.098), respectively. Significant differences (p < 0.05) were observed in the estimates of FST between all crocodile populations after 110 permutations  . A signis t-test . A compaI-CPOTH) . The oveutations . StatistK-values of 2, 3, 5, 10, 15, and 25; however, the optimized population structure patterns were assigned a K-value of 3 based on Evanno’s ΔK  with increasing nno’s ΔK . SaltwatThe PCoA revealed that the first, second, and third principal components accounted for 95.45%, 3.54%, and 0.96% of the total variation, respectively, and supported three tentatively differentiated crocodile groups  . This waUsing three different datasets of sequence depth, species were delimited between Siamese and saltwater crocodiles using the GMYC and bPTP methods, which showed that all Siamese and saltwater crocodiles were classified into the same respective species. The two hybrid individuals (CSI05 and CSI06) were not identified as Siamese crocodiles using a species delimitation approach. The saltwater crocodile group included one hybrid individual (CPO09), which was previously identified as a hybrid by microsatellite genotyping  Figure .t loci, Siamese and saltwater crocodiles shared 25.8% identical SNPt loci. Within the Siamese crocodile population, 79.8% of the SNPt loci were identical, whereas, within saltwater crocodiles from both the Thai and Australian populations, 78.3% of the SNPt loci were conserved. In the dataset with sequencing depths exceeding 10X and 15X, similarity was observed among all values, and they conformed to the same trend, as demonstrated in At sequencing depths of >5X, of the 9339 SNPt loci of Siamese and saltwater crocodiles shared sequence homology with several other species, including the American alligator, false gharial, gharial, and chicken. In Siamese crocodiles, 2033 out of 4534 SNPt loci were identified by global BLAST analyses using NCBI databases, and they were primarily associated with the short hematopoietin receptor family 2, conserved site, DNA repair metallo-beta-lactamase, tenascin, EGF-like domain, and other pathways. In saltwater crocodiles, however, 264 of the 557 SNPt loci were primarily associated with the methyltransferase small domain, mixed , peptide chain release factor, mixed (including granulin), and iron-binding zinc finger cdgsh type pathways  were partially validated . Two prit loci were obtained from our DArTseq study, which allowed for the mapping of markers back to the crocodile genomes. Based on sequencing depths of three different datasets , approximately 17.00–26.00% of the SNPt loci were conserved between Siamese and saltwater crocodile genomes, suggesting the sharing of SNPt loci after their divergence approximately 20 million years ago [inPIC similar to that of other species, such as the Ring-tailed Dragon (Ctenophorus caudicinctus) (0.19) and Plains brown tree frog (Litoria paraewingi) (0.18) [Siamese crocodiles have experienced a population decline as a result of habitat loss and illegal collection in Thailand . Moreove) (0.18) .F values were observed in the Thai populations of Siamese and saltwater crocodiles, which may have been because the specimens were collected from different farms with distinct historical origins as a Wahlund effect [FST) was observed among all crocodile populations, notably between CPOTH and CPOAU, even within the same species. This suggests that a substantial genetic exchange or gene flow between the two saltwater crocodile populations did not occur. However, the gene pool of the two saltwater crocodile populations remains consistent in the STRUCTURE analysis. A comparison of the genetic diversity parameters from the SNP analysis and microsatellite genotyping from a previous study [In addition, positive d effect ,35. Thisd effect . Populatus study  of Siameus study ,38,39,40Evidence of the genetic structure of Siamese crocodiles as one group and saltwater crocodiles from Thai and Australian populations as another group was provided via clustering analysis using STUCTURE, DAPC, and PCoA. Species-diagnostic SNPs were detected through molecular testing found for Siamese and saltwater crocodiles, most of which were associated with the methyltransferase, palmitoyl transferase and gonadal family pathway, etc. Similar gene pools were observed in two distantly isolated extant populations of saltwater crocodiles, confirming their genetic differentiation from Siamese crocodiles. The results were consistent with the two species delimitation methods. No indication of selection as a differentiating factor was found; however, the dataset or number of remaining Australian populations in the wild may be too small to detect genetic drift . Thus, tt data were compared to reliably diagnose the level of hybridization  in the presence of the admixture. CSI05 and CSI06 were tentatively identified as F1 or F2 hybrids, which was consistent with previous microsatellite genotyping results [K values in the Structure analysis. This could potentially be accounted for by the presence of a substantial gene pool within the extensive Siamese crocodile population in Thailand, where the captive population exceeds one million crocodiles [1 and BC2, have been suspected in captive crocodile populations in Southeast Asia [Hybridization between Siamese and saltwater crocodiles has previously been detected in the three individuals CSI05, CSI06, and CPO09 using a combination of mtDNA D-loop and microsatellite markers ,4. Great results ,4. Howevocodiles ,44,45. Tocodiles ,4. Moreoast Asia ,46. CrocRemoving suspected hybrids from a collection can protect the genetic integrity of the species, particularly if the collection is used to supply reintroduction candidates or augment the genetic diversity of wild populations. Moreover, removing suspected hybrids can prevent the creation of hybrid swarms, which may have a negative impact on conservation management . Our SNPThe genomic purity of over 1 million Siamese crocodiles residing in crocodile farms throughout Southeast Asia remains controversial and potentially hinders their use for conservation and reintroduction purposes ,43,44,45K levels with predictions made by various algorithms, the potential shared gene pool between species is evaluated using a 0.05 threshold for posterior probability at the K level, clustering is examined through both PCoA and the STRUCTURE plots to assess specimen groups, and maternal lineage is confirmed via mtDNA D-loop sequences. Species-diagnostic DNA markers can quickly confirm the hybridization status of Siamese crocodile individuals, thereby complementing microsatellite genotyping analysis and aiding in conservation prioritization within a reasonable timeframe before the animal’s release into the wild. However, if clear-cut interpretation cannot be provided by species-diagnostic DNA markers, such as the absence of a DNA band as in CSI05, genome-wide SNP analyses should be conducted to confirm the tested individuals. It is also recommended that Siamese crocodile candidates be genetically screened for reintroduction within each farm to select individuals with high heterozygosity, low relatedness, and low inbreeding coefficients, which will help to improve genetic diversity in the wild. Our study defined five significant guidelines that will directly contribute to the conservation of critically endangered Siamese crocodiles in Thailand: (i) Siamese crocodiles should be donated to reintroduction programs through collaboration with crocodile farms under the TCFA; (ii) our criteria should be applied to detect hybrids and introgression, which remains a major goal for the conservation management of Siamese crocodiles; (iii) individuals from the test panel should be selected for reintroduction programs by collaborative partners; (iv) habitat suitability analysis and habitat improvement of crocodile reintroduction programs should be conducted simultaneously; and (v) solutions should be sought for human–crocodile conflict interactions. Collaboration with national sectors is necessary to monitor the strategy’s long-term effects on Siamese crocodile diversity and embed biodiversity into policy. Such actions will facilitate recovery and protection and enable the evaluation of intervention success.By combining nuclear microsatellite genotyping, mtDNA D-loop sequencing, and SNPs, the maximum population genomic diversity and minimum human-induced genetic introgression can be ensured among individuals intended for release into the wild. Reliability issues may arise when using genome-wide SNP analyses to identify hybrids when a small number of crocodiles are involved in reintroduction programs ,4. HowevThis study aimed to determine the impact of hybridization on the differentiation of evolutionary lineages and identify the associated conservation consequences and policy challenges. Through comprehensive sampling and genomic analyses of genome-wide DNA polymorphism data, a complex evolutionary history involving different lineages, evolutionary trajectories, species diversification, and distinct hybrids was identified. SNP panel genotypes have numerous applications in Siamese crocodile conservation management, including the identification of two divergent crocodiles and their hybrid lineages that include critically endangered parental species ,4. A cri


Trehalose-6-phosphate phosphatase 7  has been identified as the genetic determinant for anaerobic germination (AG) and coleoptile elongation during flooding. We evaluated the coleoptile length of a diverse rice panel under normal and flooded conditions and investigated the Korean rice collection of 475 accessions to understand its genetic variation, population genetics, evolutionary relationships, and haplotypes in the OsTPP7 gene. Most accessions displayed enhanced flooded coleoptile lengths, with the temperate japonica ecotype exhibiting the highest average values for normal and flooded conditions. Positive Tajima’s D values in indica, admixture, and tropical japonica ecotypes suggested balancing selection or population expansion. Haplotype analysis revealed 18 haplotypes, with three in cultivated accessions, 13 in the wild type, and two in both. Hap_1 was found mostly in japonica, while Hap-2 and Hap_3 were more prevalent in indica accessions. Further phenotypic performance of major haplotypes showed significant differences in flooded coleoptile length, flooding tolerance index, and shoot length between Hap_1 and Hap_2/3. These findings could be valuable for future selective rice breeding and the development of efficient haplotype-based breeding strategies for improving flood tolerance.Early season flooding is a major constraint in direct-seeded rice, as rice genotypes vary in their coleoptile length during anoxia.  The priOsTPP7 and their impact on AG tolerance. In order to address this gap, we analyzed 475 Korean rice accessions and conducted genetic and haplotypic analyses of OsTPP7 (Os09g0369400) and its genetic backgrounds. By understanding the population genetic structure and evolutionary relationships for OsTPP7 within and among populations, this study will be helpful in future breeding programs aimed at developing economically desirable rice varieties.Although the AG1 gene is known to have a significant role in rice breeding, there is still much to learn about its genetic variation, population genetics, and evolutionary relationships within species or populations. This lack of knowledge has limited our understanding of the functional genetic variations in 22.1japonica (279 accessions), tropical japonica (26 accessions), indica (102 accessions), aus (9 accessions), aromatic (2 accessions), and admixture (3 accessions) (We used a core set of 475 accessions from the Korean World Rice Collection (KRICE), consisting of 421 cultivated and 54 wild accessions . These 4essions) . We deteessions) . The datessions) . The resessions) . On the essions) . All acc2.2in vitro studies have shown that this period can vary from 4-15 days depending on the purpose of investigation  and flooded germination. Twenty-one days is a commonly used duration for AG evaluation under flooded soil conditions . Howevertigation . We sele2.3OsTPP7 gene. The sequencing data of the OsTPP7 gene were deposited in the NCBI GenBank database with the accession numbers MZ682675-MZ683149  using TASSEL5 . The res toolset . This co toolset . To expl toolset  in RStudOsTPP7 gene among various plant species, we downloaded function-annotated protein sequences from 19 different species, available at (http://plants.ensembl.org/info/data/ftp/index.html)  using VCFtools 0.1.13. First, we extracted variant files for the gene region of each group using VCFtools and compared them for both SNPs and InDels. Only sites with a minor allele frequency (MAF) greater than 0.05 and no missing data were included in the analysis. For nucleotide diversity and Tajima’s D tests , Tajima’s D, and population differentiation . VCFtools was used to create a FASTA file from the VCF file, and sequences were aligned using MEGA7 . The results are presented as the mean ± standard error (SE). One-way ANOVA and Sheffe’s test were used to detect significant differences. Further statistical comparisons were conducted to determine the associations between the identified functional haplotypes and AG tolerance using the phenotypic data from the cultivated accessions. The major haplotypes were used for comparison, and the mean phenotype was calculated for each group of accessions carrying a similar haplotype. The difference between the means was compared by the Student’s t-test using the ‘t.test’ function in the R.33.1Japonica accessions displayed higher coleoptile lengths (CLs) than indica accessions under both normal and flooded conditions. The average CLs for indica, temperate japonica, and tropical japonica ecotypes under normal germination were 1.92 cm, 2.07 cm, and 1.99 cm, respectively. These values increased in response to AG, resulting in average CLs of 2.26 cm, 3.02 cm, and 2.89 cm, respectively. Consequently, the FTI values were also higher in japonica compared to indica , intermediate (1.5-2.5 cm), long (2.5-3.5 cm), and very long (> 3.5 cm)  is located on chromosome 9 at 12,251,875.12,254,061 (+ strand) and consists of seven exons with a length of 2186 bp. OsTPP7 gene region of the 475 rice accessions. Among the cultivated group, temperate japonica had 8 SNPs, followed by six SNPs in indica. In contrast, no genetic variation was observed in the aromatic group, which matches the Nipponbare reference sequence. A total of 163 genetic variations  were identified in wild rice.The 3.3OsTPP7 gene region using population structure and PCA  values based on the OsTPP7 gene using weighted methods (FST (0.533) was observed between the temperate japonica and wild groups, while the lowest was observed between the temperate japonica and aromatic groups , followed by temperate japonica (0.261), while lower genetic differentiation was observed for the aromatic and admixture groups. The FST values between the tropical japonica and admixture groups (0.318) and between the temperate japonica and admixture groups (0.274) indicated a higher degree of genetic differentiation among the cultivated population  to 0.459 (between bred and wild). These results revealed that pairwise FST values between cultivated groups were lower than those involving the wild group, indicating a closer genetic relationship for OsTPP7 within the cultivated varietal types  for the OsTPP7 gene and assessed the differences between the expected and observed segregation numbers due to selection. Here, indica type showed a positive value (0.97190), followed by the admixture (0.6003) and tropical japonica (0.21739) ecotypes, while negative values were observed in temperate japonica, aus, and wild types  SNPs (referred to as fSNPs hereafter). One of these fSNPs was a C/A substitution at position 12,253,191 in exon 5, resulting in serine to arginine change found in Hap_2. This haplotype was present in 50 accessions, including 44 indica, one aus, two admixture, and three wild accessions. Another haplotype, Hap_3, was detected in six indica and four temperate japonica accessions. Hap_3 was characterized by a non-functional C/T SNP at position 12,252,673 in exon 3, a functional C/A SNP at position 12,253,191 in exon 5, and a 20-bp insertion in the 3’ UTR region. Additionally, Hap_5, found in one temperate japonica accession, had an fSNP G/A at position 12,252,716, causing a substitution from glycine to arginine in exon 3. The most prevalent haplotype, Hap_1, was present in temperate japonica (274 accessions), indica (52 accessions), aus (seven accessions), wild (33 accessions), admixture (one accession), tropical japonica (26 accessions), and aroma rice (two accessions) (Within the coding region of essions) Figure 6Among the wild group, haplotype analysis identified 15 haplotypes, nine of which contained fSNPs. Hap_10 had one heterozygote functional allele, while the remaining eight fSNPs  were unique to the wild type. Hap_9, present in three wild accessions, had two fSNPs (C/G and G/C), while Hap_18, found in two accessions, had a G/A fSNP causing a valine to methionine substitution in exon 4 Figure 6OsTPP7 gene polymorphisms in a large number of accessions. A total of 112 haplotypes were identified, consisting of 105 polymorphisms, including 100 SNPs and 5 InDels. Among these, 72 SNPs were fSNPs located in the coding region, while the 5 InDels (two insertions and three deletions) were present in the intron and 3’ UTR  data to evaluate the d 3’ UTR 6. The p3.7indica-specific haplotype with 44 indica accessions, displayed lower FCL and FTI values than Hap_1. However, there was no significant variation in NCL among the haplotypes  , Hap_2 (23 accessions), and Hap_3 (six accessions). AG phenotyping was carried out based on the protocol of Septiningsih et al.  as different rice genotypes exhibit varying coleoptile lengths during anoxia. Although rice is semi-aquatic and adapted to a wide range of hydrological conditions, it has a certain degree of tolerance to insufficient oxygen for AG and anaerobic seedling development. Under limited oxygen conditions, rice can use the starchy reserves available in the endosperm, and rice varieties expressing the TPP7 gene are more efficient at transporting sugars from the source (endosperm) to the sink (embryo and coleoptile) compared to other crops  in japonica than indica, but higher diversity in wild rice   gene, which plays a critical role in rice anaerobic germination (AG) tolerance. To gain a better understanding of its gene function, we analyzed various diversity indices across different subpopulations of rice species, including haplotype diversity, nucleotide diversity, Tajima’s D, fixation index, population structure, and PCA. Through the genetic diversity and evolutionary analysis of OsTPP7, we gained insights into its domestication signature during gene evolution. The results revealed evidence of different directional selections, as indicated by positive and negative Tajima’s D values in cultivated rice, suggesting selective sweeps in this gene. A total of 18 haplotypes were identified in the Korean rice collection, of which three major haplotypes. Further phenotypic performance of major haplotypes showed significant differences in flooded coleoptile length, flooding tolerance index, and shoot length between Hap_1 and Hap_2/3. Specifically, the majority of japonica accessions belonging to Hap_1 exhibited higher AG tolerance compared to Hap_2/3, which were predominantly associated with indica accessions. The functional haplotypes identified in the Korean rice collection and 3K_RG data could be a valuable resource for haplotype-based breeding to improve anaerobic germination tolerance in rice. These findings provide valuable information for future selective rice breeding programs and develop more efficient and effective breeding strategies.This study aimed to investigate the The datasets presented in this study can be found in online repositories. The names of the repository/repositories and accession number(s) can be found in the article/Y-JP and S-HC acquired the funds and resources, designed the experiment, and administered and supervised the project; KA and Y-JP conceptualized the project; KA, WO, M-HM, and AS designed the methodology and performed the experiments; KA, TM, and JN curated the data, and performed the formal analysis; KA and BN wrote the draft manuscript; S-HC and K-WK Resources; K-WK, BN, S-HC, and C-YL writing -review and editing. All authors contributed to the article and approved the submitted version.


Metasequoia glyptostroboides. Using both nuclear microsatellites (nSSRs) and chloroplast DNA (cpDNA) fragments, we sampled across the narrow distribution range of this species and determined its conservation units by exploring its genetic structure and historical demography. cpDNA haplotypes were classified into two groups, but mixed in space, suggesting that the existent wild trees of M. glyptostroboides cannot be divided into different evolutionarily significant units. However, using nSSRs, we detected strong spatial genetic structure, with significant genetic differentiation and weak gene flow between the samples in the east of the species' distribution range and other samples. The divergence between the two nSSR groups was dated to the Last Glacial Maximum (c. 19.6 kya), suggesting that such spatial genetic structure has been maintained for a long term. Therefore, these two nSSR groups should be considered as different conservation units, that is, management units, to protect intergroup genetic variations, which is likely to be the outputs of local adaptation. Our findings highlight the necessity to reveal small‐scale genetic structure and population demography to improve the conservation strategies of evolutionary potential of endangered plants.Identifying conservation units is crucial for the effective conservation of threatened species. Previous cases are almost exclusively based on large‐scale but coarse sampling for genetic structure analyses. Significant genetic structure can occur within a small range, and thus multiple conservation units may exist in narrowly distributed plants. However, small‐scale genetic structure is often overlooked in conservation planning especially for wind‐pollinated and wind‐dispersed trees, largely due to the absence of dense and elaborate sampling. In this study, we focused on a representative endangered relict plant,  nSSRs‐based genetic structure and population demography analysis show two spatially segregated genetic groups with limited intergroup gene flow. Therefore, two management units should be set up to protect intergroup genetic variations. It was thought to have been extinct in the early Pleistocene but was rediscovered in the 1940s in Three Gorges Mountain Region (TGMR), China. Fossil records indicate that genus Metasequoia originated in the early Late Cretaceous (c. 100 million years ago) and once distributed over most areas of the Northern Hemisphere , China, with a census size of about 5500 . Our previous studies using random amplified polymorphic DNA markers detected a low genetic diversity in both its wild and planted populations  separated by Hanpo mountain . After testing for homogeneity among sequences using a partition‐homogeneity option implemented in PAUP* v.4.0b10  for 45 s, and 72°C for 45 s; and a final extension at 72°C for 10 min. PCR products were scanned on an ABI 3130 genetic analyzer  using GeneScan 500(‐250) LIZ as the internal lane standard (Applied Biosystems). Amplified fragments were called and binned using GeneMapper 4.0 software (Applied Biosystems).All of the 467 samples were genotyped using eight reciprocally independent microsatellite loci  developed for i et al. . PCR amp2.3H), haplotype diversity (h), and nucleotide diversity (π) using DNASP v. 6.12.01 , Taiwania cryptomerioides , and Cunninghamia lanceolata  were used as outgroups. The best model for nucleotide substitutions (the HKY + I + G model) was selected by the Akaike information criterion using jModeltest 2.1.4 ; and (3) the MRCA of C. lanceolata and other species of Taxodiaceae  ; number of effective alleles (Ne); observed heterozygosity (HO); expected heterozygosity . We allocated all sampled individuals into 18 populations according to the locations of sampling sites to perform the linkage disequilibrium test and null allele frequency analysis . Admixture model and correlated allelic frequencies were used to analyze the data set. We used the ΔK, which measures the rate of change in the log probability of the data between successive K values ) to obtain standardized measures of genetic differentiation using AMOVA  using 10,000 permutations across allele size using the program SPAGeDi  was converted to the migration rate (m) divided by mutation rate  procedure  over time; (2) the population growth model (PGM) where the ancestral effective population size Na is assumed to increase exponentially (experiencing t generations ago) with a growth rate of G, that is, Na = N × exp (G × t), to reach its current effective size (N); and (3) the size reduction model (SRM) which describes a population with an ancestral effective population size Nb experienced population decline from instantaneously t1 generations ago to reach its current effective size . Details for prior distribution of parameters for each model are shown in Table In the analysis of population size change, we calculated the average and standard deviation of the number of alleles, heterozygosity, and allele size range using ARLSUMSTAT v.3.6  as 5 × 10−4 with the prior distribution for each locus drawn randomly from gamma distribution with shape and rate parameters  ranging from 0 to 1 for eight independent loci. All priors were generated by R v.4.0.3  between the two clusters were calculated using ARLSUMSTAT, setting the other parameters using the same statistics of the best population dynamics model. The divergence model assumed the divergence between different genetic groups from their ancestral population with effective population size NANC at time T. Both unidirectional and bidirectional migration rates were set between any two genetic groups, forming three population divergence models. Each model has basic structural parameters: effective population size of a genetic group (such as NCUR1 and NCUR2), ancestral effective population size , divergence time (T), and migration rate (m). NCUR1 and NCUR2 were fixed according to the results in single population size model (Table T to the present. Prior distribution of divergence time (T), ancestral effective population size (NANC), and migration rate (m) were drawn from uniform distribution from 1 to 104, from 102 to 104, and from 10−5 to 0.3, respectively. Simulations and model choice were conducted using the same approaches in the analysis of population dynamics. For the optimal population divergence model, we conducted 105 simulations, with posterior distribution of parameters estimated using neural network regression by the R package ABC v.2.1  were identified according to the definition from Moritz  and nucleotide diversity (π) were 0.926 and 0.90 × 10−3, respectively  and Fu and Li's test  did not detect significant deviation from neutrality.Based on the cpDNA data in 172 sampled trees, we identified a total of 47 distinct haplotypes (GenBank accession: OQ415011‐OQ415039), containing 17 variants including 10 SNPs and seven indels Table . The oveM. glyptostroboides haplotypes and three taxa  for constructing phylogenetic trees. Using the Bayesian optimization approach, we found that all haplotypes of M. glyptostroboides were divided into two clades with a high posterior probability (1.00)  in Neogene  Figure , and the) Figure . In addie Figure .φST′ = 0.241), and AMOVA did not revealed a high intergroup differentiation , showing a low level of inbreeding  and FST′ (0.213)), and AMOVA also showed a significant genetic differentiation between the two groups explaining 18.34% of total genetic variance . Moreover, we detected weak but significant recent gene flow between the two groups, as of the lower boundaries of 95% confidence interval of migrate rates per generation (m) revealed by BayesAss were larger than 0, with the m from the west group to the east one being 0.0077 (95% CI: 0.0010–0.0208) and the m in the reverse direction being 0.0085 (95% CI: 0.0012–0.0192). A similar pattern of historical gene flow was also found using Migrate‐n, showing very low values of m (from the west to the east group: 0.0002 (95% CI: 0.0001–0.0003) and the reverse direction: 0.0004 (95% CI: 0.0002–0.0005)).We found high genetic differentiation between the east and the west groups  was the optimal model, which had the highest posterior probabilities for both groups. Average classification error rates of compared models were 0.295 and 0.315 of the west and east groups, respectively Table . This reNCUR1), the east group (NCUR2), and their ancestral population (NANC), respectively  from the east group to the west one was very low (1.738), with a migration rate of 0.0013  pattern ; data curation ; funding acquisition (lead); investigation ; resources ; visualization (lead); writing – original draft (lead). Min‐Yan Cui: Formal analysis ; investigation . Xiao‐Wei Le: Formal analysis ; investigation . Jun Gong: Formal analysis ; investigation . Kai Jiang: Formal analysis ; writing – original draft . Xin Tong: Formal analysis ; writing – original draft . Qian Zhang: Formal analysis ; writing – original draft . Jia‐Hui Li: Formal analysis ; investigation . Hong‐Yue Li: Formal analysis ; investigation . Ling Lu: Formal analysis ; investigation . Jie Zou: Formal analysis ; writing – original draft . Rong Wang: Conceptualization ; writing – review and editing . Xiao‐Yong Chen: Conceptualization ; methodology ; resources ; writing – review and editing .The authors declare that they have no conflict of interest.Data S1Click here for additional data file.


Breeding of fruit crops is a challenging process that must consider the need to preserve the characteristics of elite cultivars and the urgent need to obtain new cultivars with high resilience and high productivity. Changes in climate and the progressive limits on the use of agrochemicals may require the development of new genetic stocks in relatively short periods of time. Classical and innovative genetic engineering approaches may help achieve these goals. The second volume of this Research Topics aimed to present an update on the genetic tools available for the breeding of fruits crops for new traits. The articles of this Research Topic represent well the opportunities offered by genetic engineering to future fruit crop breeding.Yuan et al. describes the genetic basis of the white flesh phenotype in strawberry. The comparison by transcriptome and metabolome analysis of two strawberry cultivars, one with red skin and red flesh and the other with red skin and white flesh, allowed to demonstrate that mutation of the transcription factor FaMYB10 hinders its binding to the promoter of the FaUFGT gene that encodes for flavonol-O-glucosyltransferases, a key enzyme in the anthocyanin biosynthetic pathway.One of the first important issues for breeding is the identification of genes responsible for phenotypic traits of commercial interest and the use of molecular tools to discover the mode of action of these genes. The work of FaMYB10 and that the white flesh phenotype can be obtained with a single mutant allele. This kind of study provides useful information about gene variants that may benefit molecular breeding, and understanding the action mechanisms informs genetic manipulation as well as the fundamental basis of pigment production.These authors also provide evidence that the skin and flesh color of strawberry fruits is related to the number of fully functional copies of Cheng et al. The purpose of this work is to use phylogenetic analysis to support the breeding of Elaeagnus L. a genus of plants that is commonly grown as ornamental, and it is underutilized as a fruit crop, although its red berries have potential nutritional and perhaps medical properties. Phylogenetic analysis of Elaeagnus L species considered both DNA markers in the nuclear genome, such as the ribosomal DNA internal transcribed spacer (ITS), and the chloroplast matK gene, and integrated these data with morphological metrics clustering. The results allowed the identification of 10 wild species distributed in different parts of China that possess useful characteristics for fruit crop development. This article demonstrated the power of molecular analysis to provide the theoretical basis for crop domestication and genetic improvement.A second interesting issue is how to guide and accelerate the progression of wild species to highly valued cultivars with broad consumer interest. A case study is offered in the article by via Agrobacterium genetic transformation to generate mutants with improved agronomic traits. In these species, DNA introduced by T-DNA transfer is then eliminated by sexual crossing obtaining non-transgenic mutants. Application of this methodology is problematic in fruit crops because they are mostly clonally propagated to maintain cultivar-specific phenotypic features. Therefore, T-DNA free technologies are needed to apply genome editing in fruit crops.The development of genome editing technologies has recently opened a new scenario in plant genetic engineering, allowing specific editing of genes without the persistence of exogenous DNA material. In several herbaceous crops, genome editing has been successfully employed Scintilla et al. and Alquezar et al.This Research Topic is taken into account by the articles of Vitis vinifera cv. Crimson seedless and Sugraone) were then regenerated. This approach has successfully produced DNA-free edited grapevine plants, but it requires efficient protocols for regenerating plants from protoplasts, which is difficult in most fruit crop species and cultivars.Scintilla and colleagues obtained DNA-free edited grapevine plants by applying a single-cell based approach and transfecting CRISPR/Cas editing machinery. They produced protoplasts form embryogenic calli and edited the protoplasts by transfecting them with a ribonucleoprotein complex consisting of the Cas9 protein and an sgRNA for the target gene. From the transfected protoplasts edited non-chimeric plants of two table grape cultivars . The herbicide resistance was then used for selection of edited shoots. This article addresses another important issue for fruit crop engineering, the replacement of antibiotic selection with screening methods based on endogenous genes. The use of IMZ selection can be useful for cisgenic and intragenic modification of fruit crops.The Research Topics covered in this volume are particularly relevant as they demonstrate that new genetic techniques can bring important benefits to particularly important crops. This information is essential to demonstrate the potential of new technologies to generate consumer and regulatory enthusiasm for application in variety improvement.Current issues focus on discussion within the European Community on the acceptance of biotech plants obtained using site directed nucleases, like Cas9. These approaches are referred to as New Genetic Technologies (NGT). The European Court of Justice has already banned these technologies but thank to pressure of worldwide acceptance and EU-scientist objections, new proposals are being considered.The text proposes a legal framework for plants obtained by genetic editing and cisgenesis and for food and feed obtained from them. Different types of NGT plants are identified and it is proposed to regulate plants derived from genetic editing/cisgenesis differently than traditional transgenic (GMO) approaches. This is because genetic modifications are indistinguishable from those obtained by crossbreeding or random mutagenesis. The aim is to maintain a high level of protection of human and animal health and the environment, enable innovation in the agri-food system and contribute to the goals of the European Green Deal and the “Farm to Fork” strategy.However, the solution currently proposed in the EU is very restrictive. Gene editing is considered acceptable only if additions or deletions are less than 20 nucleotides considerably reducing the potential applications of this technology . This tyHowever, virtually internationally, these plants remain excluded from the organic certification systems which forbids crops derived from crossbreeding with gene edited and cisgenic plants.This Research Topic highlights the utility of this technology, so it is appropriate to chart the resistance it still faces before application. Continued work in this area may be valuable in swaying public opinion and lowering regulatory hurdles so that NGT may be employed to benefit the farmer, the food insecure, the environment and the consumer.TP: Writing – original draft, Conceptualization, Writing – review & editing. BM: Writing – original draft, Conceptualization, Writing – review & editing. KF: Conceptualization, Writing – original draft, Writing – review & editing.


Shisa represents a type of single-transmembrane adaptor protein containing an N-terminal cysteine-rich domain and a proline-rich C-terminal region. Nine shisa subfamily genes have been proposed in most vertebrates; however, some might be species-specific. The number of shisa genes present in zebrafish remains unclear. This study aimed to investigate the evolutionary relationships among shisa family genes in zebrafish (TU strain) using phylogenetic and syntenic analyses. The function of shisa-2 was preliminarily examined via CRISPR/Cas13d-mediated knockdown. Following identification in zebrafish, 10 shisa family genes, namely shisa-1, 2, 3, 4, 5, 6, 7, 8, 9a, and 9b, were classified into three main clades and six subclades. Their encoding proteins contained a cysteine-rich N-terminal domain and a proline-rich C-terminal region containing different motifs. A specific syntenic block containing atp8a2 and shisa-2 was observed to be conserved across all species. Furthermore, all these genes were expressed during embryogenesis. Shisa-2 was expressed in the presomitic mesoderm, somites, and so on. Shisa-2 was identified as a regulator of the expression of the somite formation marker mesp-ab. Overall, our study provides new insights into the evolution of shisa family genes and the control of shisa-2 over the convergent extension cells of somitic precursors in zebrafish. To maintain their development and growth, cells must coordinate and integrate multiple functional modules, including controlling cytoskeleton remodeling, adapting to environmental alterations, and gene expression . Functioshisa family proteins are recently discovered adaptor proteins  P×Y) motif, the PDZ (postsynaptic density protein) domain binding motif ([ST]×[VLI]), and the GRID domain . However, shisa-1 is found only in zebrafish (Danio rerio) and X. laevis [shisa-8 was not found [shisa family genes in zebrafish is still unknown. This study aimed to examine the presence of shisa family genes and their expression patterns in zebrafish and determine the potential functions of one of them during embryonic development. The findings could provide a basis for understanding the evolution of the shisa family genes and the function of related genes.To date, nine entified . Most ve. laevis . All of ot found . The zebshisa candidates were found and named shisa 1bl, 2, 2a, 3, 4, 5, 6, 7l, 8b, 9a, and 9b . Several cysteine residues were present near the C-termini of the predicted TMs in zebrafish Shisa-1–5, which were not found in Shisa-6–9 , shisa-4, shisa-7, and shisa-8b were not located on those chromosomes based on the present genomic data.The chromosomal locations of the Zebrafish shisa-2 chromosome distribution among Homo sapiens, Mus musculus, Gallus gallus, X. tropicalis, and Danio rerio. We found that a specific syntenic block containing atp8a2 (ATPase phospholipid transporting 8A2) and shisa-2, arranged in reverse order, was conserved across all species , except shisa-9b. The expression of shisa-3 was weak at 8 hpf, then increased. After 16 hpf, the expression decreased. Shisa-9b was expressed from 12 hpf. The transcripts of shisa-1, shisa-2, shisa-4, shisa-5, shisa-6, shisa-7, shisa-8, and shisa-9a were high or weak in all test stages.The temporal expression profiles of tive PCR . The resshisa-2 was analyzed using in situ hybridization in zebrafish. At 11 hpf, shisa-2 was expressed in the presomitic mesoderm, both in the medial and lateral regions of the somites and at the base of the eye vesicle , shisa-2 (shisa 2), shisa-3 (shisa 3), shisa-4 (shisa 4), shisa-5 (shisa 5), shisa-6 (shisa 6), shisa-7 (previously named shisa-7-like), shisa-8 (predicted name shisa-8b), and shisa-9 (shisa 9a and shisa 9b). A specific syntenic block containing atp8a2 and shisa-2 was found to be conserved across all species. We also found that shisa-2 was a regulator of the convergent extension cell movement of the somitic precursors during embryonic development.As adaptor proteins, shisa-1 was first confirmed in zebrafish. Vertebrates have at least nine shisa genes. shisa-2–9 were discovered in all tested species. Shisa-1 was not presented in amniotes and has been reported only in X. laevis, zebrafish, and Oncorhynchus mykiss [Shisa-1 and shisa-2 might have been produced from genome duplication as they were in the same subclade. African clawed frogs and fish are known to have gone through tetraploidization or whole genome duplication [shisa-1 subfamily was separated from subfamily shisa-2, which implied that shisa-1 did not originate from tetraploidization or whole genome duplication. However, putative shisa-1 genes in birds and the elephant shark Callorhinchus milii had been predicted in their genomic sequences [shisa-1 gene degenerated in mammals. As a result, the occurrence of shisa-1 and shisa-2 genes might be due to the gene duplication–complementation–degeneration model [atp8a2 and shisa-2 conserved across all species. The atp8a2 is considered an evolutionarily conserved gene [shisa-2 might be the original subclade gene of the shisa-1 and shisa-2 subfamilies.In this study, s mykiss ,17. Shislication ,17 and oequences . It is mved gene ; therefoShisa-8 is considered a mammalian-specific gene duplication of shisa-9 [shisa-8 and shisa-9 might have originated from genome duplication. In zebrafish, subfamily-specific gene duplications have been observed in two shisa-9s (shisa-9a and shisa-9b). In fish, many genes are found with duplicated gene phenomena, including pax3a/3b and pax7a/7b in zebrafish, medaka (Oryzias latipes), tetraodon (Tetraodon nigroviridis), fugu (Takifugu rubripes), and olive flounder  [shisa-9 genes are more likely to originate from fish-specific genome duplication. shisa-9 ; howeverivaceus) ,21,22,23ivaceus) . Two difshisa family proteins in the N-terminus, several cysteine residues were present near the C-termini of the predicted TMs  [shisa family genes might be involved in different development processes; however, the actual participation process should be determined through spatial expression and functional analysis in the future.rocesses . The expts begin , and thets begin ; therefo1.7 hpf) . Whethershisa-2 was expressed in the presomitic mesoderm (PSM), somites, optic vesicles, gill arches, and head regions. The expression is similar to that of X. laevis, chicken, and mouse shisa-2 [shisa-2 might be involved in the somite formation. Indeed, the convergent extension cell movement of somitic precursors and the maturation of somitic precursors are affected by shisa-2 in X. laevis [shisa-2 on the somite formation was preliminarily analyzed through knockdown. In wild-type zebrafish, mesp-ab is expressed as one or two stripes in the front compartment of potential somites in the anterior presomitic mesoderm [shisa-2 knockdown zebrafish, the space between the left and right stripes of the mesp-ab expression was wider than that in the control, and the expression was scattered and not clustered as a stripe. This result indicated that the convergent extension cell movement of somitic precursors was delayed after shisa-2 expression was downregulated. Therefore, shisa-2 may be crucial in the convergent extension cell movement of somitic precursors in zebrafish. The function of shisa-2 might be conserved between zebrafish and African clawed frogs. In X. laevis, Shisa-2 regulated proper segmental patterning through individual inhibition of Wnt and FGF signaling [shisa-2 in zebrafish need to be explored in the future.The results of in situ hybridization demonstrated that zebrafish  shisa-2 ,15,17,29. laevis . The effmesoderm . In the ignaling , while iignaling . The exashisa family genes suggested that all of them might be crucial in embryonic development. We could not confirm whether they have a tissue- or cell-specific expression pattern and function. As a result, the spatiotemporal expression should be investigated using other methods, such as in situ hybridization, in the future. The function of shisa-2 was only preliminarily analyzed through the recently established Cas13d-mediated gene knockdown in the embryonic stage. The method still has shortcomings, such as a lack of efficiency detection and specificity analysis methods. Thus, effective and specific detection methods should be discovered in the future. A homozygous mutant with a shisa-2 knockout should also be used to analyze its function in the future.The expression of the Wild-type zebrafish (TU strain) were cultured in a recirculation culture system at the institute aquarium . Fish were fed twice daily with commercial particulate food and once daily with brine shrimp. Fertilized eggs were obtained by mixing one male and two female fish in the morning. After washing with cycling water several times, fertilized eggs of the zebrafish were collected in a Petri dish ready for microinjection or sample collection.For sample collection, the embryos were incubated in an incubator at 28.5 ± 1 °C, obtained at different developmental stages. Approximately 30 embryos were collected in a centrifugation tube for each sample, immediately frozen in liquid nitrogen, and stored at −80 °C until RNA extraction. The wild-type zebrafish embryos and the embryos injected with Cas13d mRNA and gRNA at 12 hpf were fixed in 4% paraformaldehyde ) overnight at 4 °C. The embryos were dehydrated and stored in 100% methanol for in situ hybridization.® One-step gDNA Removal and cDNA Synthesis SuperMix kit  with OligdT.Each sample was placed in a centrifugation tube containing 200 μL of TRIzol Reagent  and homogenized using a grinding rod. Total RNA was extracted according to the manufacturer’s instructions. The quality of isolated RNA was checked using 1.5% agarose gel electrophoresis, and its concentration and purity were determined using a NanoDrop 2000 spectrophotometer . For each sample, 1 μg of RNA was transcribed into cDNA using the TransScriptshisa family genes were downloaded from the annotated zebrafish genome in the National Center for Biotechnology Information (NCBI) database (https://www.ncbi.nlm.nih.gov/ (accessed on 15 February 2022)). Primers were designed based on the predicted ORF sequences ). For Synteny analysis, TBtools [H. sapiens, M. musculus, G. gallus, X. tropicalis, and D. rerio, and all of the gene information, such as gene type and location, were visualized using the ggplot2 tool [http://smart.embl-heidelberg.de/ (accessed on 19 October 2022)) combined with the Pfam, SignalP, and TMHMM databases. The predicted structural domains were visualized using the IBS software .Phylogenetic analysis was performed using the amino acid sequences of the D. rerio . A Maxim TBtools  was usedot2 tool  of R proot2 tool . The motshisa-2 was knockdown using Cas13d mRNA and gRNAs as described by Kushawah et al. [Sal I and BamH I sites of the psp64 polyA vector using the EZ clone and named Cas13d/psp64. The Cas13d/psp64 was linearized using Pvu II  and purified by phenol/chloroform as a template to synthesize the Cas13d mRNA using mMESSAGE mMACHINE™ T7 Transcription Kit . Four guide RNAs (gRNA 1–4) were designed using online software (https://cas13design.nygenome.org/ (accessed on 6 September 2021)) [pfu enzyme and primers ) ,35. The At one cell stage, the optimized concentration of Cas13d mRNA (200 ng/μL) and gRNAs  together, Cas13d mRNA (200 ng/μL), or gRNAs  in 0.2M KCl were microinjected into the cell of zebrafish embryo using a microinjector .At 48 hpf, after the chorion was removed, the embryos were observed in water under a microscope  and photographed with a WeiTu HTC2000 digital camera; the number was counted under a stereoscope .shisa-2 was cloned using primers  and purified by phenol/chloroform as a template to synthesize the flounder shisa-2 RNA using mMESSAGE mMACHINE™ Sp6 Transcription Kit (ThermoFisher Scientific).Flounder shisa-2 mRNA (24 ng/μL) in 0.2 M KCl were microinjected into the cells of zebrafish embryos.At one cell stage, the optimized concentration of Cas13d mRNA (200 ng/μL) and gRNAs  with or without flounder At 48 hpf, the embryos were treated, observed, photographed, and counted as knockdown .shisa genes were designed using Primer3 (https://www.primer3plus.com/index.html (accessed on 2 June 2022)) and Primer-BLAST of NCBI (https://www.ncbi.nlm.nih.gov/tools/primer-blast/ (accessed on 2 June 2022)). For the semi-quantitative PCR, ef-1a was used as an internal reference gene. The total PCR volume was 25 μL and included different amounts of cDNA templates, 12.5 μL of 2 × Taq Master Mix , 1 μL of forward primer, 1 μL of reverse primer, and 9.5 μL of ddH2O. The volume of cDNA templates in the other gene response systems was 2 μL, except for shisa-3, shisa-4, and shisa-9b, which had an amount of 4 μL because their expressions were very weak. The PCR protocol was as follows: 95 °C for 3 min, 94 °C for 15 s, 55 °C for 15 s, 72 °C for 15 s, 35 cycles, and finally, 72 °C for 5 min. All RT–PCR experiments were performed in triplicate. The corresponding plasmids obtained during gene isolation  for 5 min. The embryos were soaked in the prehybridization buffer for 4 h at 65 °C for prehybridization and incubated in a hybridization buffer with each RNA probe (100–200 ng) overnight. A nitroblue tetrazolium chloride (NBT)/5-bromo-4-chloro-3-indolyl phosphate (BCIP) substrate was added to detect alkaline phosphatase until a color developed, and the reaction was stopped by rinsing with PBST. The embryos were photographed in glycerol under a microscope (Leica DM LB2) with a WeiTu HTC2000 digital camera.Whole-mount in situ hybridization Dienhart . Briefly


This work evaluates whether wheat with the Fhb1 quantitative trait locus (QTL), the most widely deployed FHB resistance locus in wheat breeding programs, provides reliable disease resistance at elevated CO2. Near-isogenic wheat lines (NILs) derived from either a highly FHB susceptible or a more FHB resistant genetic background, with or without the Fhb1 QTL, were grown in growth chambers at ambient (400 ppm) and elevated (1000 ppm) CO2 conditions. Wheat was inoculated with Fusarium graminearum and evaluated for FHB severity. At elevated CO2, the NILs derived from more FHB-resistant wheat had increased disease spread, greater pathogen biomass and mycotoxin contamination, and lower rates of DON detoxification; this was not observed in wheat from a FHB susceptible genetic background. The Fhb1 QTL was not associated with increased disease severity in wheat grown at elevated CO2 and provided reliable disease resistance.Fusarium head blight (FHB) is a destructive fungal disease of wheat that causes significant economic loss due to lower yields and the contamination of grain with fungal toxins (mycotoxins), particularly deoxynivalenol (DON). FHB disease spread and mycotoxin contamination has been shown to worsen at elevated CO Fusarium graminearum species complex [F. graminearum produces deoxynivalenol (DON), a mycotoxin that causes significant plant cell death and leaves grains withered and contaminated [Fusarium head blight (FHB) is a pervasive fungal disease of cereal crops which causes significant economic damage ,2. In no complex ,4. Funga complex . As the aminated ,6. Yieldaminated .2) concentrations, prompting a search for disease-resistant cultivars with climate resilience [The likelihood and severity of FHB infection are highly dependent on environmental conditions and field management practices. Crop residues, particularly corn stover and wheat chaff, can significantly increase the likelihood of FHB infection by providing a refuge for the pathogen to overwinter and support the formation of fungal fruiting bodies, perithecia, as the environment becomes favorable ,9. Changsilience ,17.Fhb1, Fhb2, Fhb5, and Fhb7; with Fhb1 being the most widely deployed. The Fhb1 QTL was first identified from the Sumai 3 wheat cultivar on the short arm of the 3B chromosome [Fhb1 QTL [Fhb1 helps slow the spread of the pathogen, it is not involved in the formation of D3G; however, the ability to detoxify DON has also been mapped near Fhb1 on the short arm of chromosome 3B [Resistance to FHB is often categorized by the trait effect: Type I entails resistance to initial infection, Type II is resistance to fungal spread, and Type V is the de-toxification of mycotoxins, e.g., glycosylation of DON to DON-3-glucoside, by the host ,19,20. Hromosome ,22,23. NFhb1 QTL . While Fosome 3B ,26. Fhb1 has not been resolved [Fhb1 locus, a pore-forming toxin-like gene (PFT) was identified and found to confer FHB resistance [PFT gene was also found in FHB-susceptible wheat and may be associated with general plant defense responses to biotic and abiotic stresses [Fhb1 may also be due to a deletion mutation in TaHRC, a histidine-rich calcium-binding-protein gene [Fhb1 locus, TaLAC4, is associated with increased FHB resistance through the lignification and thickening of secondary cell walls. This gene was predicted to encode a laccase protein that catalyzes lignin polymerization in the wheat rachis [TaNAC032, a transcription factor within the Fhb1 locus, regulates TaLAC4 and other lignin biosynthesis genes [Functional validation of resolved ,28,29,30sistance . This gestresses . FHB resein gene ,32. Anotis genes .2 [Fhb1 was not associated with the disproportionate loss in grain protein content [2 was found to be more susceptible to FHB spread though this effect was plant cultivar and pathogen strain specific [The Sumai 3 cultivar has multiple major and minor QTLs associated with FHB resistance and extensive breeding efforts have been conducted to incorporate these resistance factors into elite wheat breeding lines; however, producing lines with acceptable crop performance has been exceedingly difficult ,35,36. C2 ,38. Howespecific . Fhb1 QTL is widely utilized for FHB Type II resistance in wheat breeding and wheat has been observed to become more susceptible to FHB spread with rising CO2, it is vitally important to determine whether wheat with Fhb1 retain disease resistance efficacy at elevated CO2. While changes in temperature and precipitation will affect the likelihood of FHB infection based on geographic location, the rising atmospheric CO2 concentration will impact all crops globally. It is crucial for breeders to know if key loci, targeted for disease resistance, will remain reliable under future atmospheric CO2 concentrations. Since the Fhb1 QTL  derived from either a highly FHB susceptible or a more resistant genetic background, with or without the Fhb1 QTL , were grFhb1 QTL significantly reduced disease severity in wheat, maintaining a consistent percent reduction in disease spread and mycotoxin contamination regardless of genetic background or atmospheric CO2 , had significantly greater disease severity when grown at elevated CO2; the genotypes displayed greater disease spread .Increased pathogen biomass was observed in all but one M genotype at elevated COated CO2 . Increasmulation a,c. Not 2, with the sole exception of Wheaton, which had an increase in relative fungal biomass (Fhb1+), had less disease spread and DON accumulation at elevated CO2 when compared with plants grown at ambient CO2  had no increased susceptibility to FHB when grown at elevated CO biomass b. Surpriient CO2 a,c. Howe2 but the M genotypes had more severe FHB infections when grown at elevated CO2 (Fhb1 QTL had 63% more disease spread (p < 0.0001), 89% more fungal biomass (p = 0.0003), and 109% more DON accumulation (p < 0.0001) at elevated CO2 compared to ambient. While the M+ genotypes had less overall disease, due to the presence of the Fhb1 QTL, they had 70% more fungal biomass (p < 0.0001) and 52% more DON (p = 0.0065) at elevated CO2 , disease spread or area under the disease progression curve (AUDPC) by 45% (p < 0.0001), and pathogen fungal biomass (Fg) by 46% (p < 0.0001). While the Fhb1 QTL reduced disease severity in wheat, the genetic background had a greater impact on disease outcome; the M genotypes had 57% less disease spread (p < 0.0001), 71% less fungal biomass (p < 0.0001), and 77% less DON (p < 0.0001) compared to susceptible cultivars. Furthermore, in a permutational multivariate analysis of variance to determine what factors contributed most to the variance in disease severity, wheat genetic background accounted for 34%, while Fhb1 accounted for only 10% ; however, both DON and fungal biomass accumulation remained significantly lower in M− genotypes .When analyzing differences between wheat groups, S genotypes were not impacted by atmospheric COated CO2 . The M- ated CO2 . The Fhbonly 10% . Even wimination . Only at2. The proportion of DON to D3G was significantly less for most of the M genotypes grown at elevated CO2 (Fhb1 QTL (Fhb1−) and A73 (Fhb1+) differed significantly in percent D3G (p < 0.0001), despite being derived from the same parental genetic background. The percent of deoxynivalenol-3-glucoside (D3G) was evaluated to determine whether differences in glycosylation rate could account for altered FHB severity at elevated COated CO2 a. The deFhb1 QTL b. Intere2, was not the principal cause of increased FHB severity.While reduced plant detoxification of DON into D3G could account for some of the increased disease severity, percent D3G was poorly correlated with disease severity metrics in the M genotypes . Only inFhb1 QTL provided reliable FHB resistance in wheat, even at elevated CO2. Furthermore, Fhb1 provided a consistent percent reduction in disease severity regardless of the wheat genetic background: 45–47% reduction in disease progression, DON contamination, and pathogen biomass. This was consistent with previously reported FHB disease resistance provided by the Fhb1 region in the Sumai 3/Stoa NIL lines [Fhb1 reduced FHB spread in S+ genotypes compared with the highly susceptible S− genotypes, mycotoxin contamination was still substantial in all wheat from a susceptible genetic background. Our results demonstrate that the IL lines . However2 unlike wheat from susceptible genetic backgrounds. However, the Fhb1 QTL was not associated with increased relative disease susceptibility at elevated CO2 as M- genotypes, which lacked Fhb1, had more severe FHB infections and contamination when grown at elevated CO2. FHB disease resistance in wheat has been associated with the loss of grain protein and mineral content at elevated CO2 [F. graminearum genes associated with virulence and pathogenicity [F. graminearum strain used in this study (9F1) has been previously observed to increase mycotoxin biosynthesis in response to the loss of wheat nutritional content [2 concentration and the genetic background of the host, though that study employed a whole-head inoculation method rather than the single floret inoculation method used in this study [F. graminearum had greater disease spread, greater fungal biomass, and more DON accumulation in the moderately resistant wheat cultivar Alsen—harboring Fhb1 and derived from Sumai 3—when it was grown at elevated CO2.M genotypes from the Sumai 3/Stoa RIL 63–4//MN97448 pedigree suffered increased disease susceptibility to FHB spread at elevated COated CO2 ,38. The genicity ,40. Funggenicity . The F.  content . Furtheris study . As we oFhb1 QTL from Sumai 3 is the most widely studied and deployed, there are numerous other wheat QTLs associated with FHB resistance; most are considered minor contributors to plant defense but few have been validated and incorporated into breeding programs [Fhb1, was bred out of the NILs. Combining the highly effective Fhb1 QTL with the putative resistance traits within the M genetic background resulted in the lowest disease severity, even with worsening FHB infection and DON contamination at elevated CO2. We did find a significant Fhb1*genetic background interaction , W4  and are more than 97% homozygous with the recurrent parent genome. NIL genotypes from the susceptible check genetic background are referred to as S− (for those without the Fhb1 QTL) and S+ (for those with the Fhb1 QTL). Two sets of NILs with varying inherent FHB resistance were used to determine the impact of elevated COeviously . Brieflyeviously , Wheatoneviously , and Apoeviously ,58. Apogr marker  was usedFhb1 [Fhb1, i.e., mapping the genomic region harboring Fhb1. For the purpose of clarity, this group of NILs is designated genetic background ‘M’. Genotypes that possess a functional Fhb1 QTL were classified as moderately resistant (M+) while those without Fhb1 were classified as moderately susceptible (M−). The NIL set was derived from a single F7 plant that was heterozygous for Fhb1 and all NILs have the pedigree Sumai 3/Stoa RIL 63–4//MN97448. The genotypes are designated 260-2, HR 45, HR 56, HR 58, HR 123, and 260-4. All NIL lines were examined previously for agronomic performance at elevated CO2 [The second set of NILs was developed using a moderately susceptible spring wheat background that has been used for the fine mapping of Fhb1 . The choated CO2 .Fhb1 QTL is diminished at elevated CO2, the NIL wheat genotypes . Growing conditions were similar to those reported previously [2  and three set to elevated CO2 . The elevated CO2 treatment was chosen based on the predicted atmospheric CO2 concentration at the end of the century in the highest emissions scenario (SSPS-8.5) [−2 s−1 photosynthetic photon flux density). Plants were well watered, typically daily, and received additional biweekly fertilization with Peters 20-20-20 nutrient supplement  until anthesis. To evaluate whether FHB resistance from the enotypes  were groeviously . BrieflySPS-8.5) . For eacFhb1 disease resilience at elevated CO2, the F. graminearum isolate 9F1 (NRRL 37676) was used to inoculate wheat cultivars  and bag(s) were removed after 3 d. Disease progression and the area under the disease progression curve (AUDPC) was determined by visual assessment of the number of diseased florets (identified as bleached or necrotic plant tissues) on days 7, 10, 14, 17, and 21 after inoculation. At day 21, the infected wheat heads were collected and stored at −80 °C before being lyophilized and ground for mycotoxin and pathogen biomass analysis.Fhb1 impacted mycotoxin accumulation in wheat heads at ambient and elevated CO2, we measured both deoxynivalenol (DON) and DON-3-glucoside (D3G) in infected wheat heads collected 21d after inoculation. Briefly, 0.5 g of ground infected wheat heads was extracted with 10 mL of acetonitrile/water . An aliquot of the extract was analyzed for glycosylated deoxynivalenol  content using liquid chromatography/mass spectrometry coupled with electrospray ionization tandem mass spectroscopy (LC-ESI-MS/MS) on an Agilent 1100 HPLC  linked with a SCIEX 3500 TripleQuad MS . Samples were separated on a Phenomenex  Kinetex C18 column  following a previously reported methodology [To determine how hodology . A separFg DNA to wheat DNA via quantitative polymerase chain reaction (qPCR) according to the methods reported previously [Fg DNA/wheat DNA) was calculated by dividing the geometric mean of initial DNA concentration (N0) from the Fusarium probes by the geometric mean of N0 from the wheat probes. Finally, the amount of DON production per unit biomass was estimated by dividing the µg g−1 DON by the relative pathogen biomass, as quantified by qPCR.The assessment of relative fungal biomass in diseased wheat heads was determined using the ratio of eviously . Three t2. Subsequently, a post hoc analysis was performed evaluating group differences via a multiple pairwise comparison using a non-parametric Wilcoxon/Kruskal–Wallis test (α = 0.05). Pearson correlations between independent variables were determined by a multivariate analysis, α = 0.05 (JMP V15.0). A principal component analysis was performed in JMP V15.0. Additionally, a permutational multivariate analysis of variance was performed in R 4.2.1  to analyze sources of variation in disease severity treatments. Details on pairwise comparisons can be found within the table and figure legends.Results were evaluated using a non-parametric Van der Waerden test (α = 0.05) to determine significant differences due to elevated COFhb1 QTL was not associated with increased disease susceptibility at elevated CO2 and provided reliable FHB resistance. However, wheat from the M genetic background suffered greater FHB disease progression, DON contamination, and pathogen biomass accumulation at elevated CO2. Furthermore, there was a reduction in the detoxification of DON into D3G but this alone did not account for the increased disease severity. The wheat NILs derived from susceptible genetic backgrounds did not have greater disease severity or altered rates of DON detoxification at elevated CO2. Inserting Fhb1 into susceptible cultivars can significantly reduce disease, but alone, the QTL does not provide robust protection; a combination of resistance factors will likely be required to strongly suppress FHB infection. The use of FHB resistant cultivars and traits, including those derived from Sumai 3, will likely remain one of the most effective strategies to reduce disease and mycotoxin contamination in wheat. However, it is essential to develop climate resilient disease resistant wheat to ensure global food safety and security.The


To explore the differences in body-weight traits of five goose breeds and analyze their genetic diversity and historical dynamics, we collected body-weight data statistics and used Sanger sequencing to determine the mitochondrial DNA of 100 samples of five typical goose breeds in China and abroad. The results indicated that Lion-Head, Hortobagy, and Yangzhou geese have great breeding potential for body weight. Thirteen polymorphic sites were detected in the corrected 505 bp sequence of the mitochondrial DNA (mtDNA) ND6 gene, accounting for approximately 2.57% of the total number of sites. The guanine-cytosine (GC) content (51.7%) of the whole sequence was higher than the adenine-thymine (AT) content (48.3%), showing a certain GC base preference. There were 11 haplotypes among the five breeds, including one shared haplotype. We analyzed the differences in the distribution of base mismatches among the five breeds and conducted Tajima’s D and Fu’s Fs neutral tests on the historical dynamics of the populations. The distribution of the mismatch difference presented an unsmooth single peak and the Tajima’s D value of the neutral test was negative (D < 0) and reached a significant level, which proves that the population of the three species had expanded; the Lion-Head goose population tends to be stable. The genetic diversity of Lion-Head, Zhedong White, Yangzhou, and Taihu geese was equal to the average diversity of Chinese goose breeds. The Hortobagy goose is a foreign breed with differences in mating line breeding and hybrid advantage utilization. China has numerous high-quality domestic waterfowl breeds, including 31 swan genetic resources in the China Livestock and Poultry Genetic Resources Poultry Chronicle, 26 local goose breeds, and 1 cultivated breed. This high-quality genetic resource provides a solid foundation for the sustainable and balanced growth of the goose industry , the genND6) gene has a relatively short sequence length, a simple structure without introns, a moderate evolution rate, abundant sequence variations, and a significantly higher mutation rate than cell nuclear DNA and has been widely used in the population genetic structure analysis of domestic cattle [Mitochondrial DNA (mtDNA) is considered to be the best molecular genetic marker to study the origin, evolution, and classification of species because of its maternal inheritance, rich base polymorphisms, and simple structure . mtDNA gc cattle , domestic cattle , and insc cattle .D-loop region and the COI gene. For these five large, medium, and small goose breeds from home and abroad, research on the genetic diversity and evolution of mitochondrial ND6 has not been reported.The Lion-Head goose is the only large goose species in China, possessing the advantages of fast growth and resistance to rough feeding. Studies indicate that this breed is an ideal model to explore bird growth and development. The fat deposition in the liver and leg muscles of the Lion-Head goose is significantly higher than that of other goose species . The TaiND6 gene sequences with genetic diversity and historical dynamic analyses of goose breeds was used to provide a theoretical basis for the conservation, hybrid selection, development, and utilization of local goose breeds in China.In this study, three domestic large, medium, and small geese and two foreign medium-sized goose breeds, the Zhedong White goose and Hortobagy goose, were used as samples. These breeds were studied through statistics of their body weight at different ages and the determination of mtDNA All animal experiments were approved by the Institutional Animal Care and Use Committee of the Yangzhou University . All procedures were performed strictly in accordance with the Regulations for the Administration of Affairs Concerning Experimental Animals  and the Standards for the Administration of Experimental Practices .According to the pedigree records, 100 blood samples were collected from unrelated geese . AdditioTM MagneticKit , respectively. Data collection and analyses were conducted at the College of Animal Science and Technology, Yangzhou University, Jiangsu Province, China.Blood samples (approximately 3 mL) were collected from the wing veins of the geese in individual vacuum blood collection tubes. Genomic DNA was extracted using a genomic DNA extraction kit . RNA was purified to remove genomic DNA (gDNA), miRNA, and rRNA using an RNeasy Micro kit , an RNase-Free Dnase kit (Qiagen), and a RiboZeroThe 505 bp (goose) segment of the mtDNA control region was PCR-amplified using the primers 5′-GTTTTTTTTAGGGATTTGCT-3′ (F-Primer) and 5′-AACCGCCCGAATAGCAC-3′ (R-Primer), which were designed according to the goose sequence (accession number NC_012920.1) in GenBank . The PCRND6 sequence was edited and corrected using MEGA 11 software [The weight difference data recorded during the experiment were analyzed using IBM SPSS 25.0 software for the analysis of variance . Sequencsoftware . DNA SP software ,21. The software . Finallysoftware .p < 0.05) (p < 0.05). The body weight of Yangzhou geese was significantly higher than that of Hortobagy, Zhedong White, and Taihu geese at 7 d of age (p < 0.05). The body weight of Yangzhou geese was significantly higher than that of Hortobagy, Zhedong White, and Taihu geese at 14 d of age (p < 0.05). At the age of 35 days, the Lion-Head, Hortobagy, and Yangzhou geese were significantly heavier than the Zhedong White and Taihu geese (p < 0.05). At the age of 70 days, the body weights of Yangzhou, Hortobagy, and Zhedong White geese were significantly higher than those of Taihu geese (p < 0.05).The weight of Lion-Head geese was significantly higher than that of the other four breeds at the egg stage and ages of 1, 7, 14, 35, and 70 days ( < 0.05) . At 1 d ND6 fragment of mtDNA. Electrophoresis detected a clear band consistent with the size of the expected target fragment . The G+C content (51.7%) of the sequence was slightly higher than the A+T content (48.3%), showing a certain GC base preference, which was consistent with the characteristics of the bird mitochondrial DNA base composition. Eleven haplotypes were detected, of which Holdobagy geese had only one haplotype and no polymorphic sites. The polymorphic loci of the Zhedong White geese, Yangzhou geese, and Taihu geese were greater than three and the number of haplotypes ranged between two and three. The haplotype diversity was greater than 0.1 (Hd > 0.1), the acid diversity was greater than 0.001 (Pi > 0.001), and the nucleotide difference between Zhedong White geese and Taihu geese was greater than 0.5 (K > 0.5).ND6 gene in the five goose breeds is shown in The distribution of the unique haplotype variable sites of the mtDNA ND6 haplotypes in the five goose breeds are shown in The distribution and frequency of unique mtDNA ND6 gene sequence, we used DNA SP 6 software to conduct a neutrality test analysis and a base mismatch analysis of the Lion-Head goose, Zhedong White goose, Yangzhou goose, and Taihu goose populations. The results showed that the neutral test-Tajima’s D value of Taihu, Yangzhou, and Zhedong White geese were negative and the Fu’s Fs value was positive, all of which were 0.10 > p > 0.05   , indicatltimodal . This diBody-weight gain is an important indicator of animal growth and development ,25. In tND6 sequence, five parameters-including the number of polymorphic sites, the number of haplotypes, the GC content, the haplotype diversity index, and the nucleotide diversity index were used to study the genetic diversity of goose breeds. Partial sequences of the ND6 genes of the five geese breeds were obtained by PCR amplification and the average GC content (51.7%) was slightly higher than that of AT (48.3%), indicating a more obvious base bias. This was because the nucleotides of the protein-coding gene that cause mutations were under less natural selection pressure at the codon site. This was consistent with the results reported by Grzegorczyk et al. [D-loop sequences of 26 goose breeds and six Lande geese, among which the average Hd of the Chinese goose breeds was 0.1384 and the Pi was 0.00029 [Measuring differences in the species genetic structure depends on indicators such as the average nucleotide difference (K), nucleotide diversity (Pi), and haplotype diversity (Hd). In general, the higher the haplotype diversity (Hd) and nucleotide diversity (Pi) values, the greater the genetic variation. The population with the greater genetic variation generally has a greater selection potential . Speciesk et al. . Haplotyk et al. . Li et a 0.00029 . In the  0.00029 . Therefo 0.00029  and artip > 0.10), indicating that the Lion-Head goose population was in line with the neutral mutation result and that the Lion-Head goose population was in a stable state. A unimodal curve pattern that did not fit the population expansion was consistent with the neutral detection results. This indicated that the population was not undergoing species expansion or continuous growth [Analyses of population phylogenetic trees are increasingly used to infer the process of the historical evolution of populations. Population history evolution is usually detected using two methods: a population-based neutrality test such as Tajima’s D and Fu’s Fs analyses, or a base difference analysis. In the current study, the neutrality test (Tajima’s D) values of Taihu, Yangzhou, and Zhedong White geese were negative (D < 0) and the distribution of base difference mismatches showed a Poisson distribution , indicats growth ,40 and tLion-Head, Zhedong White, and Yangzhou geese had greater potential for body-weight selection in this study. The Holdobagy goose is a foreign breed that is genetically distant from the other four goose breeds. The population genetic diversity of Lion-Head, Zhedong White, Yangzhou, and Taihu geese was higher than the average level of Chinese geese and was relatively stable. Poultry breeding utilizes variations within and between populations to improve traits of interest. The upcoming era will promote the diversification of varieties and different characteristics within varieties and may shift from high-input to low-input production systems. Therefore, there is an urgent need to plan and protect the genetic resources of Chinese goose breeds and develop appropriate planning strategies to protect them against existing genetic variations.


Dasyurus hallucatus) as a case study, we applied this approach to address the need for conservation decision‐making tools that promote resilient metapopulations of this threatened species in the Pilbara, Western Australia, a multiuse landscape that is a hotspot for biodiversity and mining. Habitat and connectivity were predicted by different landscape characteristics. Whereas habitat suitability was overwhelmingly driven by terrain ruggedness, dispersal was facilitated by proximity to watercourses. Although there is limited evidence for major physical barriers in the Pilbara, areas with high silt and clay content  showed high resistance to dispersal. Climate subtlety shaped distributions and patterns of genetic turnover, suggesting the potential for local adaptation. By understanding these spatial–environmental associations and linking them to life‐history and metapopulation dynamics, we highlight opportunities to provide targeted species management. To support this, we have created habitat, connectivity, and genetic uniqueness maps for conservation decision‐making in the region. These tools have the potential to provide a more holistic approach to conservation in multiuse landscapes globally.Landscape‐scale conservation that considers metapopulation dynamics will be essential for preventing declines of species facing multiple threats to their survival. Toward this end, we developed a novel approach that combines occurrence records, spatial–environmental data, and genetic information to model habitat, connectivity, and patterns of genetic structure and link spatial attributes to underlying ecological mechanisms. Using the threatened northern quoll ( Incorporating complex genetic information into real‐world conservation planning represents a key challenge for global efforts, leading to growing concern that fundamental evolutionary concepts are not being adequately captured in management strategies  are an example of how complex modeling has been incorporated into conservation through user‐friendly decision‐support tools . These models have helped inform survey design , a declining dasyurid marsupial listed as endangered by the IUCN, and Australian federal and state legislation. Northern quolls are vulnerable to land clearing, changed fire regimes, grazing, predation by feral cats (Felis catus), and poisoning by the invasive cane toad (Rhinella marina)  subregions .We developed hypotheses regarding important environmental variables and the mechanisms that drive broad‐scale spatial associations over the life cycle of the northern quoll Figures  & 2 throNorthern quoll occurrence records were downloaded on 6 September 2020 from the Atlas of Living Australia, with ALA4R , and a minimum minor allele frequency of 0.02. We used BLAST Appendix  to removPanmixia and IBD were tested explicitly or as null models within hypotheses of IBB, IBR, and IBE. Unless otherwise stated, we used the SNP data set without HWE filtering because most analyses made no assumption of HWE.K) in the Pilbara  to balance sampling. Genetic differences among individuals were visualized using principal coordinate analysis (PCoA) in dartr  were not included in population genetic summary statistics. Summary diversity statistics were calculated for each genetic cluster in GenAlEx  in the ecodist package   with ecodist   to calculate the percentage of iterations in which each surface was the top‐ranked model  , keeping the variable that explained the greatest model deviance. Next, we ran a multivariate GDM, which included geographic distance (IBD). To avoid overfitting, we used permutation‐based backward elimination to remove nonsignificant variables that contributed least to explained deviance (100 permutations)  contributions to the explained model deviance. Variable importance in the final, reduced‐variable GDM was determined with 500 permutations.We used GDMs to determine whether gene flow was attenuated by environmental factors (IBE). We generated univariate GDMs for a range of environmental variables, testing hypotheses relating to local adaptation Figure , and excWe used variation partitioning .We found low levels of genetic structure consistent with 2 genetic clusters of northern quolls in the Pilbara, rejecting a null hypothesis of panmixia Figure . The firn = 53, Ho = 0.241 [SE 0.003], He = 0.256 [0.003]; cluster 2: n = 115, Ho = 0.244 [0.003], He = 0.259 [0.003]), the inbreeding coefficient was low (FIS = 0.050 [0.002]), and genetic differentiation among populations was low (FST = 0.020 [4.309 × 10–4])   . Heterozygosity estimates were similar between populations (cluster 1: Appendix . We did Appendix .c), the 7 resistance surfaces (and composites) explained more variation in the pairwise genetic data than Euclidian distance (IBD) alone (with the exception of the single VRM surface)  (Table According to the model selection (ranked by AICAppendix . The IBDAppendix . There w4) Table . This mo4) Table . DistancUnivariate GDMs of optimal versus suboptimal habitat (distance to water and terrain ruggedness) explained <5% of the deviance, suggesting local adaptation to these habitat types was not driving genetic turnover in northern quolls. Following variable reduction steps, the final multivariate GDM explained 58.3% of the deviance in genetic turnover in northern quolls across the Pilbara; the relationship between the composite environmental distance and observed genetic distance was linear Figure . The minWe found that complex, rocky terrain was a crucial component for northern quoll critical habitat and that watercourses facilitated connectivity between habitat patches. There was no evidence for physical barriers to dispersal, although areas with high silt or clay content show higher landscape resistance, and areas with low connectivity tended to show lower genetic diversity. There was a subtle effect of climate on genetic turnover, suggesting local adaptation could arise, though this is unlikely given high levels of gene flow across the Pilbara. The spatial‐genetic maps derived from these findings illuminate dispersal corridors and critical habitat, presenting an opportunity to focus on ground actions, such as targeted threat management, to enhance metapopulation connectivity (Dickson et al., Rocky areas are frequently identified as refugia for vulnerable species due to their thermal, hydric, and pyric characteristics (Reside et al., In contrast with our hypothesis, terrain ruggedness did not facilitate functional connectivity. Hohnen et al.  also fouDistance to water is consistently the top predictor in SDMs for terrestrial animals (Bradie & Leung, In general, genetic analyses revealed that northern quolls have high capacity for dispersal with a weak pattern of IBD, a relationship also observed in the Kimberley region (Hohnen et al., Our results add to the body of work demonstrating the value of individual‐based methods with high sensitivity to contemporary gene flow for understanding the granular drivers of connectivity (Shirk et al., Temperature and precipitation are key variables limiting animal distributions and driving adaptation to arid environments (Bradie & Leung, Combining SDMs with landscape genetics to inform conservation is a recent approach in conservation science that is being used to inform corridor design in fragmented forests (Klinga et al., We identified landscape features important for targeted conservation management to support northern quolls in the Pilbara. First, it is important to protect large areas of critical rocky habitat, especially female denning habitat, although feral cat control or habitat restoration of surrounding areas could help expand their range (see Palmer et al., The Pilbara is a vast, remote, multiuse region with only 6% protected within the conservation reserve system (Government of Western Australia, https://github.com/RobynSh/NorthernQuolls_LifeHistory2Landscape and https://doi.org/10.5281/zenodo.7005247.This article has earned Open Data and Open Materials badges. Data and materials are available at Additional supporting information may be found in the online version of the article at the publisher's website.Click here for additional data file.