Globally, all countries regard the development of economic zones around basins as the focus and main axis of national economic construction. The economic development of basin areas must consider the constraints of environmental protection, which requires local governments to adopt a coordinated development approach to the green economy. The Chengdu-Chongqing Economic Circle is located in the upper reaches of the Yangtze River Basin and will be built into a growth pole for China’s economic growth. This paper uses the panel data of 16 cities in the Chengdu-Chongqing Economic Circle from 2005 to 2019 and measures the level of coordinated development of the green economy among cities. Using fixed effect regression models, this paper identified the factors driving the coordinated development level of the urban green economy. The results show that (1) the overall trend of coordinated development of green economy in the Chengdu-Chongqing economic circle is enhanced; (2) the pulling force includes the similarity of economic agglomeration and regional openness, the resonance force includes the similarity of traffic and industrial structure, the pushing force comes from the central city; (3) in the urban agglomeration with double centers, the cooperation and competition between two “pole” cities may coexist. Globally, countries have been giving priority to the development of river basins, and many important civilizations and economic centers of gravity have flourished in basin areas ,2. In anThe Yangtze River Basin is rich in environmental and element resources . The YanEnvironmental protection and economic development are the two main themes of the construction of the CCEC, which requires the economic development model of the CCEC to consider the requirements of green development . Green dNurturing a green economy is an important means of realizing the sustainable growth of the global economy . To realIn summary, this study contributes to the realization of a green future for the CCEC. Specifically, the focus is on the ecological environment and economic development of watersheds in underdeveloped areas. Additionally, this paper studies the coordinated development of the city level from the perspective of a green economy. Lastly, this paper examines the factors driving the coordinated development of the CCEC in the past 15 years from the perspective of the coordinated development of a green economy. The results of this study will not only elucidate the periodic changes that occur over the process of collaborative development, but will also shed light on the mechanisms of influencing factors that drive these changes.Currently, the coordinated development of a green economy is an important means of realizing the development of a green economy in real conditions. The formation of a green economy requires local governments to consider the constraints of environmental protection and resource conservation while developing the economy . HoweverCurrently, the coordinated development of urban agglomerations has become the leading force of sustainable economic and social development globally. The coordinated development of urban agglomerations involves the dismantling of administrative divisions between two or more cities in the region, so that development factors and resources may flow freely. Such optimized resource allocation promotes the integration of economic and social development while providing complementary advantages to both cities . From thThe “push-pull theory” of population migration alludes to the factors that attract immigrants, with the desire to improve their lives, to reside in new places ,38. In lThe concept of “resonance” derived from the study of physics refers to the amplitude of a physical system which increases when the system is subject to a frequency close to the natural frequency of the system . This an“Pull” measures the upward supporting effect of a specific city on the coordinated development of any two cities . The theCollectively, the factors driving the coordinated development of a green economy can be divided into three categories: pull, resonance, and lift.Hypothesis 1:If the “pull” driving factor is in play, the lower the similarity of the influencing factors between the two cities, the higher the degree of collaborative development.Hypothesis 2:If the “resonance” driving factor is in play, the higher the similarity of the influencing factors between the two cities, the higher the degree of collaborative development between them.Hypothesis 3:If the “push” driving factors are in play, the higher the level of central city development, the higher the degree of collaborative development.The planning outline for the construction of the CCEC was approved by the central government. It stipulates that the CCEC include 16 cities, namely Chongqing (CQ), Chengdu (CD), Zigong (ZG), Luzhou (LZ), Deyang (DY), Mianyang (MY), Suining (SN), Neijiang (NJ), Leshan (LS), Nanchong (NC), Meishan (MS), Yibin (YB), Guang’an (GA), Dazhou (DZ), Ya’an (YA), and Ziyang (ZY). This paper used the data of the 16 cities in the CCEC, from 2005 to 2019, to constitute the balanced panel data. The data in this paper are traced back to 2005, because in 2005, China included the CCEC in the National Eleventh Five-Year Plan for the first time, marking the importance of the coordinated development of the CCEC at the national level. The data in this paper are from the Sichuan statistical yearbook, Chongqing statistical yearbook, and the statistical yearbooks of various associated cities.Current research suggests that the index system for evaluating the development level of a green economy has not been unified. In recent years, research on coordinated development has introduced the theory of complex systems, which holds that regional coordinated development is in itself a complex system . In his This paper employed the entropy weight method to measure the GED. The entropy weight method is commonly used in the academic circle to measure the development level ,59 and it year, t year, and its distribution interval is set to , which is calculated according to the entropy weight method of the green economy evaluation index mentioned above.This paper used the physics-based capacity coupling coefficient model to measure the coupling and degree of coordination of GED between two cities . The cout. The value interval is set to , such that the higher the t years, t year, the undetermined coefficients At this stage, the determination of the coefficient is controversial, and some scholars will assign both 0.5 each ,63,64. H0.5 each ,66,67. BBased on the calculation results of a green economy, this study decided to use the coupling coordination degree model according to (1)–(4) to calculate the coupling coordination degree among cities in the CCEC from 2005 to 2019. The results of the four-year modeling are shown in At the coordination level, the degree of coordination of urban coupling in CCEC and the associated regional coordination development was generally on the rise. In 2005, the average coupling coordination among cities in the CCEC was 0.408. This number increased to 0.527 in 2019. This indicates that the level of coordinated development of the green economy had significantly improved. From 2005 to 2019, compared to Chongqing, Chengdu experienced higher coupling and coordination for various cities, which likely played a strong role. This development can be attributed to the other cities that belong to the same provincial administrative unit as Chengdu. Additionally, the coordinated development within Chengdu had more advantageous policies.At the spatial level, this study selected 4 years to plot the coupling coordination values of Chengdu and Chongqing to other cities. The distribution of coordination values exhibited obvious spatial characteristics. The level of coordinated development of neighboring cities in Chengdu and Chongqing is higher than that observed in central cities. This phenomenon is recapitulated in the U-shaped curve, with Chengdu and Chongqing occupying the poles and the central cities occupying the dipping point. This observation conforms to the characteristics of coordinated development of urban agglomerations. The central cities of Chengdu and Chongqing experienced a high degree of coordinated development, while the peripheral cities suffered from poorly coordinated development. However, the CCEC is different from the unipolar urban agglomeration of two polar cities, which produces this U-shaped structure of extremes in coordinated development.To determine the factors that affect the coordinated development of GED, this paper assigned the coupling coordination value of the two cities as the explained variable in the previous stage, and established a double fixed effect model as follows to analyze the possible factors influencing coordinated development:The influencing factors that affect the coordinated development of GED can be divided into three categories: pulling force, resonance, and push. As Chengdu and Chongqing represent the two poles in the CCEC, the coordinated development of GED in any city must account for the influence of these two cities. Therefore, the per capita GDP of Chengdu and Chongqing in that year was introduced as a factor to measure the pair of two cities, To consider the distance factor in the study of regional coordinated development ,69,70, aEconomic agglomeration similarity, resource endowment similarity, transportation infrastructure similarity, regional openness similarity, market subject similarity, and industrial structure similarity were selected as explanatory variables. (1) Similarity of economic agglomeration: economic agglomeration, which reflects the degree of agglomeration of economic activities that occur within a city, was measured by the labor force per unit land area . (2) SimAs the values of explanatory variables are different based on the cities, the two-city level values of the driving factor were incorporated into the coupling degree model formula. Additionally, the similarity was calculated, and the variables were represented on a logarithmic scale. The description and statistical description of the main variables in this model are shown in To verify whether the selected influencing factors exert an impact on the coordinated development of GED, several steps were taken. First, the entire sample was subjected to fixed effect regression, as shown in R2 of models (5), (6), and (7) are 0.705, 0.705, and 0.713, respectively, indicating that the model fits well. The variables LEA, LRE, LT, LRO, LMS, and LIS are significant at the 1% level in all models, indicating that these six influencing factors are related to the level of coordinated development between the two cities’ relationship, similar to our expectations. Simultaneously, model (7) has the highest fitting degree, indicating that it is meaningful to consider the distance among the influencing factors. The variables LCD and LCQ are both significant at the 1% level in models (6) and (7), indicating that it is necessary to consider the role of bipolar cities in the urban agglomeration. This study also changed the calculation method of the explained variables for robustness testing. The specific method is to bring the GEDs of the two cities into the model (1) for calculation to obtain the explained variables. The test results are shown in Panel C. The directions of all variables have not changed, which proves that the conclusions obtained in this paper are robust and reliable.To verify whether the double fixed effect model established in this study meets the requirements of empirical analysis, this study carried out the Hausman test to select between fixed and random effects, the Wald test for heteroskedasticity, the Pesaran test for cross-sectional dependence, and a Wooldridge test for autocorrelation in panel data. The test results are shown in Panel B of According to Rene Thom’s sudden change theory, the influencing factors of coordinated development may display discontinuous characteristics near the critical point. To test this hypothesis, the entire sample was divided into several intervals according to time. Next, a fixed effect regression analysis was conducted on the influencing factors in each interval and evaluated based on discontinuity near the critical point by the coefficients of each influencing factor.Before the development and promotion of the CCEC as a national strategy, the State Council approved the Regional Plan of the Chengdu-Chongqing Economic Zone in 2011. More recently, the National Development and Reform Commission and the Ministry of Housing and Urban-Rural Development jointly issued the Development Plan of Chengdu-Chongqing Urban Agglomeration in 2016. These two documents were taken into consideration as critical developmental time points before the division of the sample time into three periods: 2005–2010, 2011–2015, and 2016–2019. The fixed effect regression was conducted on these periods and the results are shown in To observe the pulling effect of relatively strong cities on weak cities, it is vital to be able to distinguish these cities. The measurement results of the GED revealed Chengdu and Chongqing to be “strong cities” in the CCEC, while other cities were labeled as “weak cities.” By coupling Chengdu and Chongqing with other cities, the pulling effect of “strong cities” on “weak cities” could be observed. The results are shown in The similarity of the industrial structure was significant at the 1% level in models (5), (6), and (7) . As the R2 values of 0.705, 0.713, 0.705, and 0.713, for columns I, II, III, and IV, respectively, indicated that the model fits well.These findings were further supported by the upward support of a specific city to the coordinated development of any two cities. In the CCEC, only the major cities Chengdu and Chongqing play a supporting role for other cities. Hence, the current per capita GDP of Chengdu and Chongqing were incorporated into model (5) and the results are shown in In Through the fixed effect analysis of the whole sample, combined with the robustness test, this study confirmed a correlation between economic agglomeration similarity, resource endowment similarity, transportation infrastructure similarity, regional openness similarity, market subject similarity, industrial structure similarity, Chengdu per capita GDP, Chongqing per capita GDP, and the explained variables.The degree of economic agglomeration and regional openness serve as pulling forces. According to the fixed effect results of only retaining the coupling samples of Chengdu, Chongqing and other cities in  display Transportation infrastructure and industrial structure align with the theory of “resonance.” According to the fixed effect analysis results of the whole sample in , the simThe per capita GDP of Chengdu and Chongqing played a supporting role for other cities in the urban agglomeration. The addition of the per capita GDP of Chengdu and Chongqing in The “rush” of the transition of cities will hinder the coordinated development of urban agglomerations. During the three policy periods of 2005–2010, 2011–2015, and 2016–2019, the coefficient of Chengdu’s per capita GDP experienced a change from positive to negative. This study attempted to regress the data of Chengdu by adding the base period and changing the measurement method. This approach showed that the economic development of Chengdu in 2016–2019 negatively correlated with the coordinated development of a green economy in CCEC. This study speculates that this may be related to Chengdu’s economic “rush” as the city entered a stage of rapid development after 2016. For instance, Chengdu hosted Jianyang to build the Tianfu International Airport in 2016. In the following year, the talent settlement policy was launched. In 2021, the resident population exceeded 20 million. This type of rush in economic development is the result of regional competition and is not conducive to the coordinated development of urban agglomerations in the long run.Globally, all countries regard the development of economic zones around basins as the focus and main axis of national economic construction. The Yangtze River is the mother river of China. The CCEC is located in the upper reaches of the Yangtze River and has been established by China as the fourth pole of economic development. Research on the coordinated development of the green economy in CCEC will provide suggestions for economic development and environmental protection in underdeveloped and non-coastal areas. In this paper, the entropy weight method was employed to measure the GED level of the CCEC from 2005 to 2019. Based on the green economy measurement values of each city each year, the coupling coordination degree model was used to measure the coordinated development level of the green economy among cities in the CCEC. Finally, the coupling coordination degree was assigned as the explained variable. The influencing factors of coordinated development were divided into pulling force, resonance, and pushing force. The main conclusions are as follows. First, the overall trend of coordinated development of a green economy in the CCEC is improving. Second, the pulling force includes the similarity of economic agglomeration and regional openness, while the resonance force includes the similarity of traffic and industrial structure. The two major cities of Chengdu and Chongqing have played promising and facilitative roles. Third, in the urban agglomeration with double centers, the cooperation and competition between two “pole” cities may coexist. This will lead to the formation of a U-shaped economic structure with two “pole” cities at the highest points, and other regions in the drooping center.Based on the above research conclusions, this paper enlightens its readers about the process of collaborative development in future works.First, the economic development of cities must consider environmental constraints. Economic entities need to consider the impact of negative externalities of economic activities on the environment and its bearing capacity. To achieve the green development of urban agglomerations, cities are required to adopt a development model that is cooperative rather than confrontational. Future works should pay attention to the balanced development of urban agglomerations, reduce the central collapse caused by the rush of extreme cities, and avoid unbalanced development during the contemporary round of balanced development. To accomplish this, more detailed coordinated development strategies need to be introduced at the level of urban agglomeration. Additionally, economic development factors should be allocated more reasonably, and the development momentum of each city should be balanced. Simultaneously, cities should actively integrate themselves into the process of coordinated development of urban agglomerations by actively moving closer to advantageous cities, paying attention to the division of labor and cooperation, and building industrial belts in the CCEC.Secondly, attention should be paid to the differentiated development of the similarity of economic agglomeration and regional opening under the action of a pulling force. This approach will be essential to avoid homogeneous competition. Prosperous cities, such as Chengdu and Chongqing, should continue to attract talent, absorb a high-quality labor force to form economies of scale, and expand the degree of opening to the outside world. The integration of international partners into The Belt and Road Initiative construction will be beneficial. By building itself as a node city of foreign trade, Chengdu and Chongqing can promote the green and positive development of urban agglomerations.Third, attention should be paid to the similarity of traffic and industrial structure under the “resonance” theory. This will ensure cooperation and balance between regions and help identify areas with weak traffic infrastructure. At all times, priority should be given to the development of traffic infrastructure and the opening of the circulation channels of production factors. In terms of industry construction, strengthening industrial cooperation among cities and paying attention to the division of labor and cooperation will be essential in building industrial integration.


Forensic genetics is a rapidly evolving science thanks to the growing variety of genetic markers, the establishment of faster, less error-prone sequencing technologies, and the engineering of bioinformatics models, methods, and structures. In the early 2000s, the need emerged to create an international genetic database for forensic purposes. This paper describes a judicial investigation of skeletal remains to identify the subject using various methods. The anthropological examination of the remains allowed identification of the Caucasoid (European) ethnic group, a height of 156 ± 4 cm, and an age between 47 and 50 years. The genetic profiles obtained from typing several microsatellites made it possible to evaluate the compatibility between the skeletal remains and the suspected decedent. To identify the remains, the two extrapolated genetic profiles were compared. The case described highlights the central role of forensic genetics in identifying skeleton remains by means of comparison. Genetic investigations were first applied in the forensic anthropological field in the 1980s in the study of anomalous and uncontrollable migratory flows, and in particular of criminal acts, catastrophes of all kinds, and war events, and the need for forensic investigation of skeletons or unrecognizable bodies . GeneticIn the forensic field, the most commonly used genetic markers are microsatellites or STR (short tandem repeats). A comparison between two genetic profiles obtained by typing several microsatellites makes it possible to evaluate the compatibility between the two subjects ,5. The pThanks to these comparisons, forensic genetics is now the cornerstone of some judicial investigations . SequencMicrobiome and metagenomic analyses are inspired by the fact that human beings host diverse microbial communities at the level of all body surfaces, continuously interacting with and altering the surrounding environments. The latest studies aim to study the information relating to these interactions, especially for thanato-chronological purposes .In line with the advances in these technologies, in the early 2000s, the need emerged to create an international genetic database for forensic purposes . This neIt follows that with the advances made in meta-genomic technology and computational prediction, as well as the creation of international genetic databases, possible issues related to individual privacy have emerged. However, this subject will not be dealt with in depth in this manuscript.This paper describes a judicial investigation of skeletal remains to identify the subject using various methods. Personal identification was made possible by comparing the genetic profile from the skeletal remains with a genetic profile from a sample obtained from a previous sexual assault victim.The manuscript aims to highlight the complexity of forensic investigations in human skeletal remains, with particular attention to the cornerstone role of forensic genetics in attributing identity to skeletal remains, per se limited by the absence of an adequate comparator database.The first investigation carried out was that of the inspection. On that occasion, metric and photographic surveys were carried out on the remains, which were then transported to the Forensic Anthropology laboratory of the University of Bari “Aldo Moro.” At the same time, investigations by the Judicial Police were launched into the environment of neglected and homeless people, narrowing the field around an unknown subject, a prostitute and the victim of multiple episodes of abuse, including sexual violence, that had led to the collection of biological samples, appropriately stored in the laboratories of the Scientific Section of the Police.For recognition, several investigations were carried out.A 1-milligram bone sample (femur) was taken for the dating study of bone remains using an analysis of the carbonate phosphate (C/P) index by an external laboratory. The anthropological measurements were performed according to international standards of forensic anthropology ,12 and wsagittal contour;facial morphology;frontal and parietal bones;orbits;interorbital distance;nasion;nasal opening;nasal spine;zygomatic arches;palate;mastoid;nuchal crest.For the determination of the ethnic group, the following parameters of the skull were examined:morphological and craniometrical parameters of the skull;morphological parameters of the pelvis.The following skeletal segments were examined for sex determination:The remains were analyzed following the method proposed by Ozaslan, using the right humerus, radio, femur, and tibia ,12,13The Suchey-Brooks method , the metThe dental arches were reconstructed with the dental elements found, and the teeth, the upper jaw, and the mandible were then inspected with exposure to a source of UVA light . Periapical digital photographic and radiographic surveys of the dental arches and sensors for radiovideographies were made ,18,19,20To isolate the DNA, allowing personal identification of the subject whose skeletal remains had been found, two teeth  and a 1 cm piece of the femoral shaft of the skeleton were removed.The vaginal swab taken during a previous judicial investigation into a case of sexual violence, and duly filed in the refrigerators of the Scientific Police, was later used for comparative purposes to verify whether the remains found could belong to a previous victim of sexual abuse.®DNA Tissue by the company MACHEREY-NAGEL, Allentown, PA, USA, following the specific protocols indicated by the manufacturer [Tooth 37 and the vaginal swab were subjected to DNA extraction at the Forensic Genetics laboratory of the University of Bari, using the commercial kit NucleoSpinfacturer ,22.In particular, in the preliminary stages of the investigation, the DNA was extracted and examined starting from tooth 37; after the investigations conducted by the Judicial Police, it was possible to access the vaginal tampon. DNA extraction from the vaginal swab was subsequently performed by a female operator with all the PPE provided and after the decontamination of the worktops, pipette tips, and pipes, with exposure to UV rays, sodium hypochlorite, and ethanol.Skull CT was performed.The skeletal remains were found on the fourth floor of an abandoned farmhouse in the industrial area of a city in Southern Italy, partially covered by a wooden box and some wooden planks, arranged as a funeral bed .The distribution of the bone remains, with some residues of mummified soft tissues, suggested that the corpse had been laid in supine decubitus, with the upper limbs flexed and adducted to the trunk and the lower limbs in normal position.The skeleton was wrapped with two pieces of duct tape .The right shoulder bones  were wrapped in a cotton fabric.There were signs of micro- and macrofauna activity on the skeletal remains and in the surrounding environment.The carbonate phosphate index (C/P) identified the skeletal remains as recent clinical/forensic bone samples (C/P = 0.66) and not as archaeological remains. The remains were carefully cleaned and placed in an anatomical position.The skeleton appeared complete and intact, light brown in color, with bones.−The Skull: Intact and complete with a fractured right portion of the nasal bone; dental elements and partial dentures were found.−Hyoid bone: The body and the large horns were intact but disjointed.−Spine: All vertebrae were present and intact.−Clavicle and scapula: The joint complexes were intact and inter-articulated.−The ribs and the sternum were intact and inter-articulated.−Pelvis: The pelvis was intact but disjointed from the sacrum.−Upper limbs: Humeri intact and preserved, the left disjointed. Ulna and radius were present and intact. The bones of the complete right hand, the pisiform bone, the bones of the distal row of the carpus, and the distal phalanges of the IV-V radius of the left hand were missing.−Lower limbs: Femur, patella, and tibia present bilaterally. The soles of both feet were complete.To determine ethnicity, the morpho-dimensional characteristics of the skull were examined, and the distances summarized in the following table calculated :−Morphological parameters of the skull, which appeared long and not very robust, with thin and sharp supraorbital, small, rounded, and medially inclined mastoid processes, pronounced frontal and parietal bones, spherical orbit with sharp edges, forehead, regular and U-shaped palate, nuchal crest and insertion crests of the nuchal muscles, glabella, cheekbones, and minimal development of mastoids.−Pelvis, macroscopically, appeared low and wide with a broad, heart-shaped pelvic cavity. The subpubic angle was shaped like an inverted U. The preauricular sulcus was present, the auricular surface raised and stepped, the ventral arch was present, the obturator foramen was triangular, the ischio-pubic branch moderately thin and narrow, the sacrum flattened in an anteroposterior direction.To determine the sex, the following skeletal remains were examined:These elements depicted the skull and pelvis of a female subject.Measurement of the right femur yielded a maximum length of 40 cm, equal to a stature of 156 ± 4 cm.The Suchey-Brooks method was used to determine the age of the subject using morphological analysis of the modifications of the symphysis facet of the pubis to define the age of a white female cadaver. The articular surface was free of crestlines, flat, with evidence of a slight depression of the symphysis facet and lipping (presence of minute lingulae) on the dorsal margin; bony outgrowths of the ventral margin were found, with signs of bony destruction on its upper margin.The application of this method showed a presumed age of 48.1 ± 14.6 years.This method, which studies age-related changes in the auricular surface of the ileum, allowed us to highlight a skeletal age between 47 and 54.5 years.This method, which evaluates the degree of obliteration of the exocranial sutures, allowed us to detect complete obliteration of the sagittal suture, an almost complete obliteration of the coronal suture near the first and third segments, and partial obliteration of the lambdoid suture in correspondence with the first and second section. These data indicated an estimated age of 45–50 years.The confluence of the various methods applied made it possible to place the average skeletal age between 47.5 and 50.75 years.Examination of the skull made it possible to detect a displaced bone fracture of the right nasal bone (antemortem) with a small, displaced bone fragment.There was no evidence of post-traumatic alterations of the remaining bone structures of the skull.The characteristics of the skull and jaw allowed us to recognize a typical conformation of the female sex and Caucasoid ethnicity.The study of the relationships between the extension of the dental pulp and the volume of the entire tooth, obtained with radiographic images of the upper and lower canines according to Cameriere’s formula, allowed us to confirm an estimated age equal to 50.4 (±5) years .The general state of the mouth, wear of dental prostheses, and dental neglect made it possible to conclude that the subject had not undergone dental checks for at least five years.Research on DNA polymorphisms established that the DNA extracted from tooth 37 and the vaginal swab possessed the genetic characteristics summarized in the following table . Only peThe judicial investigation described in the study was aimed at identifying the individual whose skeletal remains had been found.Personal identification of the remains of an unknown subject is obtained through visual identification by showing the body or photographic reproduction of the face to people they knew in life . When, oIn an abandoned cadaver, tissue transformation is completed on average within 3–5 years. Gradually, after the destruction of the soft parts, the fibrous tendon and cartilage tissues disappear, and only the bones remain. Over time, the bones are freed from even the most minor residues of soft parts that are still adhered, dried, or encrusted but retain their organic component for a long time. Later, after many years, only mineral scaffolding made of phosphates and calcium carbonates remains. Still, although decalcification takes place with the passing of time, bones remain light, porous, brittle, and susceptible to pulverization at the slightest contact . HoweverOne of the main limitations of forensic samples, in addition to the fact that they contain only small amounts of biological material, is that very often the DNA is degraded, i.e., reduced into small fragments by chemical and physical agents that cause the rupture of the ligaments of the double helix ,28,29,30In the present case, obtaining the generic DNA profile from the two molars was possible without using extraction from the femur. In particular, one of the limitations of our study was the limitation imposed by the magistrate in charge of the case, who expressly requested we protect the most significant number of samples for any subsequent forensic investigations.A study of the genetic marker “amelogenin” identified the sex of the subject, specifically confirming what was inferred from the forensic anthropological study, namely that the individual was female.In the case of our study, once the genetic profile had been obtained from the no. 37 molar, it was isolated and then compared with the data of a profile extracted from a vaginal sample of a woman who had suffered sexual violence a few years before the discovery of these skeletal remains. This comparison was made because, during the inspection, the police had noticed the recurrence of a name written with a spray can on the farmhouse wall where the remains were found.However, after consulting the missing person reports, the police could not trace any individual with that name. The agents then extended the search to the regional register of reported episodes of violence, tracing back to a woman whose name in Polish was similar to the one written on the wall.The woman listed in the regional register was Polish and had reported sexual violence; the emergency room health workers, on that occasion, had carried out a vaginal swab from which the genetic profile was extrapolated.To identify the remains, the two extrapolated genetic profiles were compared and determined to be statistically compatible, allowing the identification of the subject with the genetic profile observed in tooth 37, excluding the markers that most likely showed the dropout phenomenon (D7S820 and TPOX).Considering the possible compatibility of the two genetic profiles, various investigations were carried out by the national and international police forces. Thanks to these investigations, it was learned that the woman, aged 51 years old, from Poland, had gone to Italy in the year preceding the discovery of the skeletal remains and was involved in a prostitution ring. From all the anthropological elements that emerged, it was possible to attribute the cause of death to a traumatic injury resulting from aggression.It is interesting to note that genetic profiling was carried out using a relatively recent low-cost technology.Forensic genetics is a rapidly evolving science thanks to the growing variety of genetic markers, the establishment of faster, less error-prone sequencing technologies, and the engineering of bioinformatics models, methods, and structures .In our case, the identification was possible thanks to “luck” in finding a clue that allowed the police to carry out in-depth research, helping us to match the genetic profile isolated from the skeletal remains with one previously collected from a victim of violence.If a match had not been found, the next step would have been to consult the international databases, as required by the Prüm agreement .In fact, for the genetic information relating to an individual to be entered into the international database, precise sampling and analysis methods of the biological find must be followed according to protocols validated at the international level and indicated by the European Network of Forensic Science Institutes (ENFSI) . The neeHowever, the addition of genetic information is allowed only in a few forensic laboratories that follow specific certification procedures controlled by external bodies. Alternatively, authorization from the Judicial Authority is required.In addition, there are restrictions on consultation so as to safeguard the individual’s privacy by allowing access to data only by a limited number of laboratories.The case described illustrates the central role of forensic genetics in identifying skeletal remains using comparison. However, surveys on an international scale still present limits to database consultation due to legal and political implications related to privacy and protected health information. The search for a missing person is challenging for law enforcement officers and requires collaboration between states. The authors propose the facilitation of access to international databases without neglecting ethical issues or the privacy of the individual. On the contrary, the individual must remain at the center of future philosophical, sociological, and legal work while introducing further provisions to make identification more accessible internationally.


The inclusion of ancestrally diverse participants in genetic studies can lead to new discoveries and is important to ensure equitable health care benefit from research advances. Here, members of the Ethical, Legal, Social, Implications (ELSI) committee of the International Genetic Epidemiology Society (IGES) offer perspectives on methods and analysis tools for the conduct of inclusive genetic epidemiology research, with a focus on admixed and ancestrally diverse populations in support of reproducible research practices. We emphasize the importance of distinguishing socially defined population categorizations from genetic ancestry in the design, analysis, reporting, and interpretation of genetic epidemiology research findings. Finally, we discuss the current state of genomic resources used in genetic association studies, functional interpretation, and clinical and public health translation of genomic findings with respect to diverse populations. Definition of terminologies used in the manuscriptAdditive genetic effect: the total effect, either over alleles at a locus or over loci, equals the sum of the effects of the individual alleles or loci.Admixture mapping: a gene mapping approach for phenotypes that exhibit ancestry differences used to identify associations between local ancestry and the phenotype in a sample of admixed individuals.Causal inference: a process to study the causal effect of a particular factor on an outcome of interest based on data by relying on assumptions, study designs, and estimation strategies.Haplotype phasing: the process of statistically inferring an individual'sgenome into maternally and paternally inherited segments.Heritability: a measure of the extent to which genetic variation explains phenotype differences.Mendelian randomization: a causal inference approach that uses genetic variation as a natural experiment to infer causal relationship between modifiable risk factors and health outcomes.Meta analysis: integrated analysis of data from different studies to answer a research question.Mixed models: statistical models that include fixed effects and random effects for which the variance matrix is specified. In GWAS, the effect of a locus of interest is often modelled as fixed and is the target of inference, while the genome‐wide additive genetic contribution to the phenotype is modelled as a random effect, with the variance specified by a measure of relatedness.Population bottleneck: a short period of low population size during the history of a population that leads to reduced genetic variation.Principal component analysis: a dimension‐reduction technique that replaces a large number of correlated variables with uncorrelated linear combinations of the original variables. In genetic studies, the first few principal components typically reflect population structure.QTL mapping: a method to identify genetic loci that are associated with a quantitative trait such as height or a measure of gene expression.Structural variants: refers to a broad range of genome variations including insertion, deletion, inversion, duplication, translocation, and copy number variation.1APOL1 variants which may have risen to high frequency in parts of Africa to provide protection against African sleeping sickness  published has grown 40‐fold in the past decade, with over 5000 unique publications and now nearly 300,000 associations reported in the GWAS catalog  committee of the International Genetic Epidemiology Society (IGES), we have worked together to provide guidance and support to our community for the conduct of research that is more inclusive. We released a short guideline as a website statement on the use and reporting of race/ethnicity/ancestry in IGES abstracts and papers to promote inclusion diversity and equity , or it can be more specific . All individuals have ancestors originating from multiple different locations at different time depth; as such, it can be more accurate to refer to proportions of genetic ancestries  , which are commonly used in genetic epidemiology research. However, this mutation rate is highly variable and can depend on genetic and epigenetic context, sex, and age of the father at the time of conception, among other factors  and of tissue‐ or cell‐specific expression are frequently performed. Other approaches include colocalization, which aims to discover genes for which a single causal genetic variant underlies two phenotypes , aggregate many genetic variants investigated in GWAS  will likely lead to further improvements in PGS performance across populations.Recently, approaches have been proposed to develop PGS for diverse populations. Simulations and real data applications have shown that including in PGS variants discovered in diverse ancestral populations led to reduced bias and higher genetic risk accuracy in admixed individuals . Future work is needed to investigate optimal usage of ancestry and admixture in genome references.A key resource for the identification and functional interpretation of disease‐associated variants in whole genome or exome sequencing analysis is the human reference genome   (Karczewski et al., ACKR1 gene with the development of neutropenia, a serious adverse event that results in stopping the clozapine treatment (Bentley et al., Pharmacogenomics has been successfully applied in clinical genetics. However, underrepresented populations have lower potential to benefit from predicted drug responses. In this regard, a recent GWAS on schizophrenia patients of African ancestry undergoing clozapine treatment showed a strong association of a regulatory variant in the Increasing the diversity of the biomedical research workforce can also contribute to improving scientific innovation derived from having different perspectives that promote creativity. Concerted efforts are being made to promote the training and support of scientists from underrepresented backgrounds working in the genomic field (Bonham & Green, 5.1Interpretation of admixture mapping findings in the context of trait differences between populations warrants careful consideration of its assumptions and limitations. Ancestry estimation in admixed populations is error‐prone because the true historic ancestral parent populations are unknown and the availability of representative ancestral reference genetic data is limited for some populations (Baran et al., Genetic differences between individuals within the same population are often higher than differences between individuals from different populations (Rosenberg, 6The field of genetic epidemiology is undergoing a transformation with the collection of data with more diverse ancestral and social origins and with research staff and participants that are gradually becoming more inclusive. This shift is fuelled in part by concerted efforts toward the collection of diverse data at the study design level and as part of large reference data sets. We must be prepared to continue to support these efforts by engaging financially and scientifically in the years ahead to achieve broad data collections that will have meaningful impacts. The benefits of this shift toward more diversified genetic research include the potential for providing better treatment options to currently underrepresented communities, more equitable power dynamics with respect to health decisions and the potential for novel genetic discoveries that will be valuable to all communities, not just those where the discoveries are made.To fully benefit from the growing admixed and diversified data collections, methodological tools should follow suit. We need to move beyond the traditional stratified datasets based on arbitrary population boundaries. We should support existing and future data analysts with documentation, training, and guidelines. As new models and approaches are being developed, our community should proceed with transparency following our hard‐earned standards of methodological rigor. Concerted efforts to evaluate and compare the performance and limitations of the various methods being developed will help to document, improve and set standards for the conduct of analyses with diverse and admixed genetic data sets.


Capsicum or chilli is an important crop in India which exhibits immense structural and genetic variations reflecting their intra- and inter-specific relationships. The aim of this study was to establish relationships amongst 54 Capsicum accessions through analysis of genetic and population structure using ISSR markers.I), expected heterozygosity (He), Nei’s gene diversity, total diversity among species (Ht), diversity within species (Hs), respectively. Further, the coefficients of gene differentiation (Gst) and gene flow (Nm) were 0.393 and 0.773, representing higher genetic variation among the population which was confirmed by analysis of molecular variance (AMOVA).Out of 19, successful DNA amplifications were shown by 7 ISSR primers and a total of 80 bands were identified ranging between 8 and 14 with an average of 11.43 bands/primer. A significant degree of polymorphic information content (PIC), discriminating power (DP), resolving power (RP), effective multiplex ratio (EMR), and marker index (MI) were identified as 0.39, 0.70, 6.40, 5.88, and 2.30, respectively, using ISSR markers in chillies. The cross-transferability ranged from 8.0 to 72.15% with an average of 52.63% among chillies. Amongst genetic information, grand mean values were 0.264, 0.180, 0.376, 0.296, and 0.180, which correspond to Shannon’s information index (ISSR markers represented a potent system for the estimation of relationships or variation studies and generated information useful for planning crop management and improvement strategies in chilli breeding. Capsicum genus represents a wide genetic diversity comprising 38 species  and incubated at 65°C for 1 h followed by chloroform:isoamyl alcohol  treatment. DNA pellet obtained in chilled isopropanol was washed with 70% ethanol. Isolated DNA samples were treated with RNase for 1 h at 37°C followed by phenol:chloroform:isoamyl alcohol (25:24:1 v/v/v) treatment then washed with 70% ethanol. DNA pellet was air-dried and dissolved in TE buffer and then stored at −80°C . The qua2, 1 μl of primer (10 pmole/ μL), 0.25 μl of dNTPs (100 mM), and 0.1 μl of Taq DNA polymerase (0.5 U). PCR amplification conditions included initial denaturation at 94°C for 3 min followed by 35 cycles which included denaturation at 94°C for 1 min followed by annealing at 45 to 51°C for 1 min depending upon primers and then extension at 72°C for 2 min with final extension at 72°C for 7 min. All amplified products were separated through agarose gel electrophoresis using 1.2% agarose gel (Himedia) in 0.5× TBE (Tris-Borate- EDTA) buffer for ∼1.5 h at 70 V. Gel was stained with ethidium bromide dye, and BioRad gel doc system was used for visualization of DNA bands and further analysis.A total of 19 ISSR primers  were examined for DNA amplifications in chilli accessions. Finally, 7 ISSR primers were selected for analysis among 54 chilli accessions due to their sharp and clear banding profiles. All PCR reactions were performed in the final volume of 10 μl each using thermal cycler . Each reaction mixture contained 1 μl of DNA template (25 ng), 1.0 μl Taq buffer (10X) with 2.5 mM of MgClClear and reproducible amplified bands obtained from the DNA amplifications profile for each ISSR primer were used for the experiment and scored as binary matrix, 1 for the presence and 0 for the absence. The efficiency of ISSR markers was calculated as described by polymorphic information content (PIC), discriminating power (DP), resolving power (RP), effective multiplex ratio (EMR), and marker index (MI) using iMEC platform . The relNa), number of effective allele (Ne), Shannon’s information index (I), expected heterozygosity (He), unbiased expected heterozygosity (uHe), analysis of molecular variance (AMOVA), and principal coordinate analysis (PCoA) were evaluated through GenALEx 6.5 program  in Dipteroniadyerana surveyed by ISSR marker [Jatropha curcas by microsatellite-based marker (ISSR and DAMD) system, Gst and Nm were reported to be 0.4053 and 0.8085, respectively [The coefficients of gene differentiation , and structure analysis which exhibited consistency of results in the characterization of Capsicum accessions. Due to varied genome size, morpho-physiological variation, and distinct agro-ecological environments, the result of the present study represented a significant genetic relationship among chilli accession. Important factors which explain these results regarding harmony and discordance among the chilli accessions are the nature of marker system used, level of polymorphism, the number of detected loci, and region coverage of genome by each marker [According to Jaccard’s similarity coefficient and UPGMA clustering analysis, the varied level of relationships revealed with low, moderate, and extensive genetic association among the different h marker , occurreh marker , 102.Thus, the effect of each factor or combined effects of multiple factors have an impact on mechanism that shapes the population genetic structure while extensively related and dissimilar genetic variations might be associated with increased and reduced amount of genetic information respectively. Therefore, evaluation of genetic diversity is an important factor for explicating the connection among various chilli accessions which is essential component of germplasm characterization. Identification and characterization of new variations from the germplasm will help to develop new cultivars with improved agronomic trait, useful for crop improvement, and breeding program in chillies.Na, Ne, I, uHe, Hs, Ht, and Nei’s statistics) which highlighted the molecular variability among different chilli accessions. Thus, the knowledge obtained through genetic variability could be used in the management of chilli germplasms. High level of coefficient of gene differentiation (Gst) represented restricted gene flow (Nm) due to genetic drift which can be correlated with high or low rate of allelic acquisition, adaptation, mating nature, interaction with different ecological conditions, and changes in morphological distinctiveness during the course of evolution. The present study provides a fundamental insight for germplasm characterization, genetic arrangement, and population structure of chilli germplasm which could be utilized in the effective management and selection of superior germplasm for breeding purposes.The study reveals valuable information about genetic polymorphism, cross-transferability, and genetic and structural plasticity among 54 accessions of chilli using ISSR markers. A significant amplification profiles were obtained which reflects marked genetic polymorphism and cross-transferability among chilli accessions indicating efficiency of ISSR markers for genetic discrimination and conservation among chilli accessions. A significant level of genetic information was revealed by the estimation of various factors (


High-density single nucleotide polymorphisms (SNPs) are the most abundant and robust form of genetic variants and hence make highly favorable markers to determine the genetic diversity and relationship, enhancing the selection of breeding materials and the discovery of novel genes associated with economically important traits. In this study, a total of 105 barley genotypes were sampled from various agro-ecologies of Ethiopia and genotyped using 10 K single nucleotide polymorphism (SNP) markers. The refined dataset was used to assess genetic diversity and population structure.The average gene diversity was 0.253, polymorphism information content (PIC) of 0.216, and minor allelic frequency (MAF) of 0.118 this revealed a high genetic variation in barley genotypes. The genetic differentiation also showed the existence of variations, ranging from 0.019 to 0.117, indicating moderate genetic differentiation between barley populations. Analysis of molecular variance (AMOVA) revealed that 46.43% and 52.85% of the total genetic variation occurred within the accessions and populations, respectively. The heat map, principal components and population structure analysis further confirm the presence of four distinct clusters.This study confirmed that there is substantial genetic variation among the different barley genotypes. This information is useful in genomics, genetics and barley breeding.The online version contains supplementary material available at 10.1186/s12863-023-01109-6. Hordeum vulgare L., 2n = 2x = 14 chromosomes, 5.1 Gb haploid genome size) is one of the main important cereal crops cultivated worldwide in a wide range of environments [Barley , scald  and leaf rust (Puccinia hordei) [Barley is the fifth most important cereal crop in Ethiopia after teff, maize, sorghum and wheat both in area of production and amount . Ethiopi−1 . It is g−1 , 7 but i−1 , 9. The  hordei) , 11. UndHordeum vulgare L.), and it is landraces are genetically unique and diverse [Ethiopia is considered as a center of diversity for barley  has collected and conserved more than 16,000 accessions sampled from various agro-ecologies across the country. It is a useful resource for genetic diversity and can play an important role in developing new barley varieties with higher yield potential and other desirable agronomic traits . KonopkaAnalyzing the molecular diversity and it is genetic relationship encompassed in crop genetic resources is a prerequisite for designing efficient selection in crop breeding programs and for developing conservation and valorization strategies. Previous studies have used DNA markers to determine the genetic diversity of barley. Markers such as amplified fragment length polymorphism (AFLP) , restricCurrently, single nucleotide polymorphism (SNP) markers are the markers of choice for genetic diversity studies , genome-A recent study by Teklemariam et al.  has obseH.vulgare genome  distribution of the 10,103 SNPs remaining after imposing a quality control threshold was plotted in 10 Mb (megabase pair) window size across the The genetic parameters included gene diversity, heterozygosity (the ratio of observed to expected heterozygosity), minor allelic frequency (MAF) and the polymorphic information content (PIC) of the 10,103 SNP markers of 105 barley genotypes were presented in Table P < 0.0001) than the variation within accessions  (Table  P < 0.0001) was recorded between the barley populations  showed that the proportion of variance within the barley populations was significantly higher  among barley populations was computed using SNP markers (Table Pairwise genetic differentiation (Frs Table . The lowrs Table . Althougrs Table . This reA high genetic distance (0.08) was recorded among barley genotypes obtained from Tigray and ICARDA, which indicates the existence of high genetic differences among the genotypes Table . The genTo quantify the genetic variation between genotypes, we perform a principal component analysis (PCA). Four clusters were observed which accounted for 47.7% and 13.5% of the total variation in PC1 and PC2 of the total variance, respectively  followed by cluster III (n = 16), cluster IV (n = 15) and cluster II (n = 14). The number of genotypes belonging to distinctive clusters varies from 14 in Clusters II to 60 in Cluster I  in the Oromia region, indicating the existence of high genetic variability between the genotypes Table . This maA higher genetic variation of 52.85% was observed within the populations, showing the variations among the barley genotypes in the regions (Table ST) was an evaluation of population variations because of genetic structure [ST = 0.019, 0.022, 0.038) genetic differentiations were shown in Table ST = 0.019) was observed between the Amhara and Oromia genotypes while distant dissimilarity (FST = 0.117) between the SNNP and Tigray genotypes (Table ST = 0.089–0.117) related to other regions of the populations, which showed geographical origins could be the basis of a genetic variant. The genotypes obtained from the Tigray region may be exploited as a source of breeding materials to improve genetic diversity in barley breeding through hybridization programs. Our study result revealed moderate genetic differentiation (FST = 0.117) for the pairwise comparison among the barley population implying a smaller amount of genetic relatedness and this indicated a lesser genetic relationship. This result was confirmed by genetic differentiation largely higher in marginal populations than in the favorable environment. Marginal stands are grouped by geographic and genetic separation because of spatial segregation and restricted gene flow [ST = 0.082) was shown in the Ethiopian barley population. On the contrary, the largest genetic differentiation (FST = 0.257) was revealed between the barley population by Allel et al. [The genetic variance fixation index , and most of the genotypes included an altitude of 2001–3000 masl Fig. , Table 57.2%, andIn the current study, the SNP data generated using advanced molecular tools provide useful information that can be utilized in breeding and genetic research in barley. Based on SNP data, the barley genotypes were genetically divergent. The AMOVA indicated high genetic variations within the populations and genotypes. This high diversity could be the foundation for developing and generating desirable new barley varieties with superior grain yield potential and wide adaptability, enhanced with abiotic and biotic resistant traits. This study identified four subpopulations, considered as four independent subpopulations in the improvement program, but was not grouped the genotypes according to collection origin and adaption zone. This information on genetic diversity and population structure of Ethiopian barley genotypes will be applied for current and future research using genome-wide association and genomic selection for economically useful traits in barley.ex-situ collection of the Ethiopian Biodiversity Institute (EBI) along with their passport data , Holetta. For two weeks old barley leaf samples were collected and pooled from the same genotype in equal amounts. Then samples were placed into 96 collection plates (96-well plate holds 12 × 8-strip tubes). During the sample preparation, the leafcutters (scissors) were sterilized with 70% alcohol before cutting the next genotype to prevent cross-contaminate. The collected leaf samples were first freeze-dried at -20 °C for 24 h. Then the freeze-dried leaf samples were shipped for genotyping to the Integrated Genotyping Service and Support (IGSS) platform located at Biosciences Eastern and Central Africa- International Livestock Research Institute Hub (BecA-ILRI Hub) based in Nairobi. Genomic DNA from 105 barley genotypes was extracted using the Nucleomag Plant Genomic DNA extraction kit following the manufacturer's instructions. The genomic DNA concentration was used in the range of 50–100 ng/μl. The quality and quantity of extracted DNA in each sample were determined by electrophoresis on 0.8% agarose gel.Hordeum vulgare_v2.0 [Genomic libraries were constructed according to Kilian et al. , complexare_v2.0 , to idenAn initial set of 14,454 single nucleotide polymorphisms (SNPs) containing monomorphic and undefined chromosomes or positions were removed from the raw dataset. Then further quality control was performed, and SNP markers with > 95% call rate, minor allele frequency (> 5%), and missing rate per sample (< 10%) and per SNP (< 30%) were retained for downstream analysis using R software version 4.1.3 . The divaov function.Genetic parameters such as polymorphism information content (PIC), genetic diversity, heterozygosity (observed and expected), minor allele frequency (MAF), genetic differentiation and genetic distance were computed using the R package snpReady . The molK-value for estimating an optimum subpopulation size for the dataset was determined based on peak ΔK values following the Evanno method [prcomp function to summarise the contributions of each part to the variation that existed in the population. The genetic diversity pattern among the population was visualized using the pheatmap R package [To explore the genetic structure of the 105 barley accessions we first undertook a cluster analysis using the agglomerative hierarchical algorithm  , Ethiopian Institute of Agricultural Research (EIAR) and Universities with the region of origins and altitude ranges.Additional file 2: Figure S1. The administrative map of Ethiopia indicates the collection points of the barley genotypes. Dots represent the barley genotypes in different colors depending on various region of origins and altitude ranges, according to the legend.


Cordia myxa Roxb.) is a potentially underutilized fruit plant in arid and semi‐arid regions. It has long been associated with health and nutrition. Morphological diversity of 75 accessions of this species was evaluated. The accessions studied showed significant differences in terms of the characters measured. Ripening date varied from late May to mid‐July. Fruit color was yellow‐cream in 54, light orange in 8, and orange in 13 accessions. The range of fruit weight was 0.71–11.83 g with an average of 3.12, while fruit flesh thickness ranged from 0.63 to 7.86 mm with an average of 2.55, and fruit jelly part thickness varied between 1.34 and 6.40 mm with an average of 2.75. Principal component analysis could describe the evaluated traits as the nine main components that were able to justify 79.04% of total variance. Hierarchical clustering showed that the accessions were placed into two main clusters using the measured traits data, exhibiting a wide range of variability. Based on the traits related to selection of the ideal genotype, such as big fruit size, high fruit flesh thickness with high yield, and longer harvesting period, 11 accessions, including Jangal‐4, Jangal‐9, Rask‐7, Jangal‐5, Jangal‐6, Jangal‐11, Rask‐1, Jangal‐1, Rask‐4, Rask‐5, and Rask‐2, were superior. It is recommended to use the best accessions selected in breeding programs. The commercial orchards of those best accessions should be extensively constructed to take advantage of the high yield of C. myxa as a crop and its medicinal properties.Spistan ( Based on the traits related to select ideal genotype, such as big fruit size, high fruit flesh thickness with high yielding and longer harvesting period, 11 accessions, including Jangal‐4, Jangal‐9, Rask‐7, Jangal‐5, Jangal‐6, Jangal‐11, Rask‐1, Jangal‐1, Rask‐4, Rask‐5, and Rask‐2, were superior. It is recommended to use the best accessions selected in breeding programs. The commercial orchard of those best accessions should be extensively constructed to take advantage of the high yield of C. myxa as a crop and its medicinal properties. There are reported to be 300 species in the genus Cordia . Common names of this species include Assyrian plum, lasura, laveda, pidar, panugeri, naruvilli, geduri, spistan, burgund dulu wanan, and ntege. It is in the form of a medium‐sized deciduous tree and its height reaches 10.50 m  was performed to evaluate the variation among the accessions based on the traits measured using SAS software  belonged to fruit stone thickness (13.88%) and then fruit stone length (14.40%). The CV of 46 out of 48 characters recorded was more than 20.00%: tendency to form suckers exhibited the highest CV (700%) followed by fruit weight (104.73%), leaf serration depth (95.95%), bunch weight (84.43%), fruit color (81.26%), and fruit flesh thickness (76.36%) Table  reportedC. myxa germplasm from India. Also, leaf serration shape was highly variable, including entire (23 accessions), undulate (23), dentate (14), and serrate (15) (Table C. myxa germplasm from India. Nagar and Fageria (Tree growth habit was highly variable and included weeping (16 accessions), spreading 26), open (5), semi‐erect (21), and erect (7). Tree growth vigor, tree height, branching, branch density, branch flexibility, and leaf density were dominantly high Table . The maj6, open (5) Table  reported Fageria  reportedC. myxa germplasm from India. Four types of fruit shape were observed, including oblate (12 accessions), round (14), oblong (31), and obovate (18). Sivalingam et al. , green‐cream (15), cream (22), brown (5), and dark brown (6). Fruit color was yellow‐cream in 54, light orange in 8, and orange in 13 accessions. Fruit taste was astringent‐sweet in 24 and sweet 51 accessions Table  reportedC. myxa germplasm from India, while Meghwal et al.  variation and partly of nonheritable. The proportion of total variation, which is due to genetic differences, is termed as heritability in a broad sense , high fruit flesh thickness with high yield, and longer harvesting period, 11 accessions, including Jangal‐4, Jangal‐9, Rask‐7, Jangal‐5, Jangal‐6, Jangal‐11, Rask‐1, Jangal‐1, Rask‐4, Rask‐5, and Rask‐2, were superior. It is recommended to use the best accessions selected in breeding programs. The commercial orchards of those best accessions should be extensively constructed to take advantage of the high yield of C. myxa as a crop.Since there are no known commercial cultivars in None.The authors declare no conflict of interest.None.None.


Next-generation sequencing technologies have made it feasible to generate large amounts of data per plant genome, and now it is even possible to generate long reads for large and complex genomes as well . Many plClimate change and population increase are the two main factors driving the demand for progressive increase in agricultural productivity. Stress matrix data on different combinations of environmental conditions have already suggested significant negative impact on agricultural production . This, iThe attempts to enhance the genetic gain across any plant species would require the understanding of the genetic structure of that species. This means the number of sub-population groups, genetic diversity available in the cultivated gene pool, ploidy, etc., for every species need to be looked into along with the genomic information available such that utilization of the available information is incorporated while designing breeding programs targeting desired haplotypes .The earliest suggestion of identifying haplotype tag SNPs (htSNPs) within haplotype-based blocks came after the human genome sequence was available with an idea of reducing the number of markers needed to capture useful information within a genomic region . Developr2 = 0.25 level was much faster in maize (1 kb) than in soybean (150 kb in euchromatic and 5 kb in heterochromatic regions) or rice (123 kb)  with diverse genotypes/populations emerged as one of the key strategies for unlocking genetic diversity . However(123 kb) . The stude novo assembly in crops with complex genomes for hundreds of individuals is still beyond the reach of individual research groups  for targeting trait enhancement. The phenotypic and genotypic data available for multiple locations and diverse genotypes, respectively, have to be sieved into two parts: 1) explaining breeding value and 2) number of loci underlying a trait. Components of variations observed explaining breeding value of a trait, and haplotype needs to be clearly dissected and then targeted for marker development and deployment such that desired haplotype(s) can be fixed as early as possible in the targeted genetic background in a breeding cycle. To achieve this, genomic data currently available and being generated need to be looked from a genetics perspective of the target trait and crop species. The progress made in understanding traits  and crops  will have to be leveraged to make trait-specific mini-core/core collections or practical haplotype graphs with suitable marker sets available for selection of the “ideal” haplotype. The design of markers having a higher prediction value and the use of only a significantly associated subset of markers in prediction and selection will ensure that genotyping costs are not prohibitive. These can then be used by crop improvement programs targeting a particular trait.


Ziziphus species particularly Ziziphus mauritiana and Ziziphus nummularia constitute an important part of genetic resources in India. They contribute economically as a fruit crop with lots of morphological and pomological variability. In current study, 48 accessions belonging to two wild Ziziphus species, i.e., Z. mauritiana and Z. nummularia, were characterized using SSR markers. In addition, external features were also examined using stereomicroscope.Present investigation was done to explore the genetic structure of North Indian jujube. In total, 23 SSR markers detected 57 SSR alleles with an average of 2.47 alleles. Highest number of alleles (4) were detected by three primers, namely BFU1178, BFU479, and ZCMS14, while lowest number of alleles (2) were detected by fifteen primers. Highest Polymorphism Information Content (PIC) was 0.500 and shown by two primers, namely BFU528 and BFU1248, while lowest PIC (0.041) was observed in primers BFU286 with mean value of 0.443. Similarly, highest value of marker index (MI) was detected by primer BFU1178 i.e. 1.969, and lowest value of marker index was observed in primer BFU286 i.e. 0.021. Dendrogram generated using SSR markers data and principal component analysis showed two major groups of the analyzed germplasm with intermixing. STRUCTURE analysis also clustered all the accessions into two groups. We did not found correlation between geographic and genetic distances.The preliminary results suggest that there is high level of gene pool mixing in these species which can be attributed to their cross-pollination habit. However, more such studies with large numbers of samples are required in future to gain concrete insights of the genetic structure in these species.The online version contains supplementary material available at 10.1186/s43141-022-00458-6. Ziziphus mauritiana and Ziziphus nummularia both jujube species are commonly known as ber (Indian jujube) in India. These are subtropical to tropical plant species, and fruits of these species are liked and eaten by large population of India. Z. mauritiana is a cultivated species, while Z. nummularia is a wild species, and both of these vary in their habits. Generally, both the species have prickly stems and branches with ovate leaves and minute flowers. The pollination is strictly cross-pollination as it also shows the protandry habit. However, plants of Z. mauritiana are larger in height having larger leaves, stems, branches and fruits, while plants of Z. nummularia show shrub-type habit with smaller leaves, profuse branching and small fruits [Ziziphus species with specific objectives to characterize the wild and cultivated genetic resources of Ziziphus in north western Indian states and to establish genetic relationships among the analyzed accessions of both the species, i.e., Z. mauritiana and Z. nummularia.l fruits , 58, 64.l fruits , 37, 40.l fruits , 62. Chal fruits , 61 and l fruits , 66, 67.l fruits . Classicl fruits . These fl fruits . Molecull fruits , 53, 55.l fruits , 49, 51.l fruits , 52. In l fruits –60, 70. Ziziphus accessions i.e. 20 accessions of Z.mauritiana and 28 accessions of Z. nummularia, were collected from different geographical locations of northwest Indian states  and were analyzed using SSR markers. Of these, thirteen samples were from Punjab, twelve from Rajasthan, twelve from Haryana, and eleven from Himachal Pradesh. A detailed description with locations of all the accessions and their altitude range is given in Table In the present study, 48 Ziziphus species from various locations of northwest India (Table 2 (25 mM), 1.0 μl 10 × PCR buffer , 0.5 μl dNTP mix , and 0.2 μl Taq polymerase (5 U/μl). The PCR conditions were as follows: 1 cycle of 5 min at 94 °C, 35 cycles of 1 min at 94 °C, 1 min at respective annealing temperature for each primer as shown in Table Thirty-one SSR primers which were developed by Wang et al.  were anaia Table . Out of Only unambiguously amplified alleles were scored manually and converted into binary data, i.e., 1 for the presence of band and 0 for the absence of band. Polymorphism Information Content (PIC) values were calculated using the formula given by Botstein et al. , 26. DisK = 2 , two major groups Figs.  and 3 weZiziphus germplasm from India is needed for its improvement in future and for the conservation of diverse and promising accessions. Genetic diversity gives the estimates of DNA polymorphism of the analyzed germplasm, and this polymorphism can be used in future for improving and manipulating the germplasm for various purposes. In the genus Ziziphus, the characterization of the cultivars had been largely based on morphological characters and their uses [Z. mauritiana) of Saudi Arabia were done using ISSR markers and it has been observed that a cultivar called Um-sulaem was paraphyletic to the other four [Z. mauritiana, some other workers also conducted research using RAPD and ISSR [Z. jujuba from China, and reported high genetic diversity (98.2%) in corresponding 3 clusters was observed using 31 primer pairs [Genetic diversity and population structure of eir uses . Howeverher four  accessioand ISSR , 47. Furand ISSR . The simand ISSR . Most reer pairs . The preZiziphus jujuba. STRUCTURE analysis and multivariate analyses (cluster and PcoA) were also done for the grouping of jujube accessions. Fu et al. [Ziziphus jujuba Mill.) for population genetics, and the average number of alleles per locus was found 12.8 which was much greater than the number of allele obtained in present study. Using 11 ISSR primers to access genetic diversity within and among 34 accessions of Z. spina-christi collected from different regions of Saudi Arabia, Saleh Alans et al. (2016) obtained 109 scorable loci, of which 93.4% were found to be polymorphic. The size of amplified bands ranged between 250 and 3000 bp. Significant variability was observed in 10 selected Z. nummularia accessions based on various quantitative and qualitative characteristics of leaves, fruits, and seeds. Akhtar et al. [Z. mauritiana and one wild accession of Z. nummularia using 18 ISSR markers; a total 167 products were detected, of which 89.96% were reported polymorphic. Cluster analysis based on UPGMA method and Bootstrap analysis separated all the 48 Ziziphus genotypes in 4 distinct clusters, and they were found to be divergent on the bases of Jaccard coefficient. Similar finding was observed in present study as dendrogram is showing divergent clustering as well as mixing of accessions. Saha et al. [Z. mauritiana which included 6 accessions of wild-type Z. mauritiana, two accessions of Z. nummularia, and 1 accession of Z. xylopyrus. The PIC values range between 0.23 to 0.46 and 0.11 to 0.36, respectively, for the RAPD and ISSR primers which are somewhat lower values as observed in present investigation using SSR markers. Thirty-eight microsatellites were isolated from Z. mauritiana to evaluate genetic diversity by Chiou et al. [To investigate the genetic relationship between the domesticated and wild jujube populations, chloroplast microsatellite markers (cpSSR) were developed by Huang et al. . Using tu et al.  used SSRr et al.  investiga et al.  studied Ziziphus species to understand the spread of species and sharing of genomes between wild and cultivated germplasm. Furthermore, identification of diverse accessions based on minute morphological differences as well as at DNA level can be done for conservation and for initiating new breeding programs in Ziziphus species.In the present study, SSR markers showed high genetic diversity and mixing of gene pools in studied accessions of both jujube species. Results showed that there are two genetic stocks contributing in analyzed accessions. We found no specific correlation between different accessions of same species on the basis of geographical locations. The results of this research work can be useful in future research works in Additional file 1: Supplementary Fig. 1 Showing correlation between Genetic and geographic distances of 48 samples used in present study.Additional file 2: Supplementary Table 1 Axis wise Eigen values.


Aedes notoscriptus, an important disease transmitting mosquito at the Mornington Peninsula, Australia. We sampled and reared Ae. notoscriptus eggs at two time points from 170 traps up to 5 km apart and generated genomic data from 240 individuals. We also produced a draft genome assembly from a laboratory colony established from mosquitoes sampled near the study area. We found low genetic structure (Fst) and high coancestry throughout the study region. Using genetic data to identify close kin dyads, we found that mosquitoes had moved distances of >1 km within a generation, which is further than previously recorded. A spatial autocorrelation analysis of genetic distances indicated genetic similarity at >1 km separation, a tenfold higher distance than for a comparable population of Ae. aegypti, from Cairns, Australia. These findings point to high mobility of Ae. notoscriptus, highlighting challenges of localised intervention strategies. Further sampling within the same area 6 and 12 months after initial sampling showed that egg-counts were relatively consistent across time, and that spatial variation in egg-counts covaried with spatial variation in Wright’s neighbourhood size (NS). As NS increases linearly with population density, egg-counts may be useful for estimating relative density in Ae. notoscriptus. The results highlight the importance of acquiring species-specific data when planning control measures.Dispersal is a critical parameter for successful pest control measures as it determines the rate of movement across target control areas and influences the risk of human exposure. We used a fine-scale spatial population genomic approach to investigate the dispersal ecology and population structure of  Operophtera brumata is used to indicate when farmers should implement control measures to mitigate the damage to plants while decreasing the use of insecticides   studies are commonly used , a container breeding mosquito native to mainland Australia and Tasmania and invasive in the Torres Strait Islands, New Zealand, New Guinea, New Caledonia, Indonesia  . We also calculate spatially-varying estimates of Wright’s neighbourhood size , Blairgowrie  and Rye (‘north-east’ and ‘south-east’)  or Roche High Pure PCR Template Preparation Kits  following the manufacturers’ instructions. We prepared double‐digest restriction site‐associated DNA sequencing (ddRADseq) libraries following the method for Ae. aegypti  and 2 μL of 10 μM standard Illumina P1 and P2 primers. These were run for 12 PCR cycles, then cleaned and concentrated using 0.8x paramagnetic beads. Each ddRAD library contained 24 mosquitoes, and each was sequenced on a single sequencing lane using 150 bp chemistry. Libraries were sequenced paired-end at GeneWiz, Inc  using a HiSeq 4000 .Process_radtags program in Stacks v2.0 , using –very-sensitive alignment settings. We filtered all alignments to paired reads that aligned concordantly, requiring the two paired reads to align to the same contig to avoid multi-mapping using Samtools . Beagle v4.1 (Browning and Browning vcftools thinned SNPs so that no two SNPs are at 500 bp distance to each other (–thin 500). After filtering, we retained 11,091 SNPs.We used the Populations was used to calculate pairwise FST values between the four sampling sites. We tested for isolation by distance between all samples and within each of the four sampling sites using the mantel.randtest function in the R  from recent releases in the area and wildtype individuals without the infection (WT). We analysed these separately to avoid bias. We down sampled each Ae. aegypti dataset to only include one sample per trap to achieve maximum comparability to the Ae. notoscriptus dataset.To contextualise these spatial genetic structure results, we compared results for mgram analysis on two sets of the Ae. notoscriptus dataset separated into the ‘north-west’ and ‘central’ as well as ‘north-east’ and ‘south-east’ samples sites to archive a more comparable trap distance distribution to the Ae. aegypti datasets.Additionally, we included the Ae. notoscriptus and coancestry between individuals of the different sampling sites, we used the program fineRADstructure . The data was used to investigate whether egg counts differed between sites and if they were consistent over both time points. The same researcher counted all eggs using a light microscope to ensure consistency. Additionally, we compared predicted egg counts throughout the sampling area with spatially-varying estimates of Wright’s neighbourhood size  to the effective density (D) of breeding adults in the local area. The relationship between NS and D is thus highly correlated, with population density increasing linearly with NS  against the natural logarithm of geographical distance  , indicating that isolation by distance at the scale of the whole sampling area is mostly driven by closely related individuals. No isolation by distance was detected when tested within each sampling site, where the mean distances between trap pairs were 522 m, suggesting no clear genetic structure for Ae. notoscriptus at this scale.We found isolation by distance in the positive relationship between genetic distance and the natural logarithm of geographical distance when calculated on the entire  m) Fig. . HoweverAe. notoscriptus samples around the range of 1300 m  as well as at 3700 m   where individual mosquitoes were effectively less related than they would be at random. By comparison, the population of Ae. aegypti from Cairns showed a signature of spatial structure that was positive and significant in the first distance class of 100 m , then decreased sharply, dropping below zero beyond 500 m . The range of trap distances for WT Ae. aegypti was 35 m to 3280 m and 36 m to 3057 m for the wMel infected population  Fig. . At dist0 m Fig. . In Ae. ion Fig. .Fig. 3SpAe. notoscriptus does not show any significant negative autocorrelation , ion Fig. , indicatPC-Relate. We also designated 8 pairs with k > 0.07 as putative first cousins. Jasper et al.  and exhibited a maximum observed distance of 1267 m. We found a mean separation distance for half-siblings of 1296 m , and 2778 m  for first cousins  in the ‘south-east’ and ‘south-west’ sites, with an increase in NS in the ‘central’ (240−265) and ‘north-west’ (260−285) sites. The estimated prediction error for ordinary kriging after cross validation indicates that, on average, an error of ~1.04 of mean NS can be expected at any given location. Spatial variation in NS was roughly consistent with egg counts  across the study area.The NS for each trap location estimated using l’ 240−26 and ‘norAedes mosquitoes have recorded how Ae. aegypti disperse within and between buildings in urban Malaysia  to around 700 m, while some half-sibling pairs were found multiple kilometres apart . Our analysis showed genetic dissimilarities between Ae. notoscriptus were only significant at greater than 4 km separation, while Ae. aegypti had significant dissimilarities from around 1 km, which suggests greater dispersal activity in Ae. notoscriptus . We detected isolation by distance in Ae. notoscriptus on the scale of the entire dataset  and calculated a neighbourhood size . The study employed gravid traps for four weeks to remove female mosquitoes and their offspring from the environment. The trial was evaluated by egg counts collected before and after the intervention and resulted in no measurable reduction in numbers of Ae. notoscriptus eggs. The success of the intervention could have been affected by several factors, including low uptake of gravid traps by residents, but was also likely affected by the size of the controlled zones. The intervention zones (~250 × 350 m) were sized under the reasonable assumption that Ae. notoscriptus dispersal in urban areas is similar in scale to Ae. aegypti. However, our results highlight that generalising ecological traits between related species can be problematic when planning species-specific interventions. The dispersal estimates we discussed above indicate that it is likely that the dispersal of Ae. notoscriptus exceeded the size of controlled zones. As a result, zones were likely to be invaded by mosquitoes from surrounding areas while control measures were in place, which could have compromised the success of lowering mosquito numbers. Controlled zones were also likely to be re-invaded quickly after the trial, given the generation time of Ae. notoscriptus of approximately one month.Our results and analyses indicate a high dispersal ability of n et al.  argue thAedes egg numbers collected by oviposition traps and numbers of adult mosquitoes collected in adult traps  Figs. , 6. ThouAe. notoscriptus at the Mornington Peninsula, Australia. Dispersal in this population seems to take place over greater scales than in the Cairns population of the congeneric Ae. aegypti, which highlights the importance of acquiring species-specific and even population-specific ecological information when planning management strategies. We showed the challenge of generalising between related species when planning interventions. Finally, we provide evidence that spatial variation in egg counts collected by oviposition traps can be linked to spatial variation in neighbourhood size and, thus also, density and may be a valid means of estimating relative densities and the impact of control measures.This study uses recently developed genomic approaches to provide evidence for high dispersal abilities of Supplementary Information