Several free-roaming dogs can be easily found in the public areas of Bali. They go out in search of foods and friends and defecate everywhere. In general, these groups of dogs do not receive good healthcare from their owners and are generally threatened by some disease-causing organisms, especially helminths. This study was conducted to identify and measure the prevalence of soil-transmitted helminths (STHs) that cause infection in free-roaming dogs in Bali Province, Indonesia.A total of 1611 fresh dog fecal samples were collected from all areas of Bali Province and subjected to qualitative fecal examination using flotation techniques to obtain STH eggs.Ancylostoma spp., Ascaris spp., and Trichuris spp. Ancylostoma spp. had the highest prevalence of 37.8%, followed by Ascaris spp. and Trichuris spp. at 6.02% and 0.87%, respectively. Multiple infections of these worms were also recorded. The polyparasitism prevalence of Ancylostoma spp. and Ascaris spp. was 3.85%, followed by that of Ancylostoma spp. and Trichuris spp. at 0.5% and that of Ascaris spp. and Trichuris spp. at 0.06%. There were no altitude-wise differences in the prevalence of STH infection.The incidence of STHs in free-roaming dogs was 38.36%. However, three types of STHs were identified, including The prevalence of STHs was high in free-roaming dogs. This finding necessitates more serious attention as it affects both animal and public health. Dogs cannot be separated from human life, and they are often referred to as the Best Friend of Man. Petting a dog, apart from being a hobby, can provide several benefits for the owner. According to some research, a dog used as a pet can have a positive impact on psychology and health . In BaliIn contrast to the benefits provided, the maintenance patterns of allowing them to roam tend to have adverse effects on the health of the community as well as other animals . As a reAncylostoma spp., Ascaris spp., and Trichuris spp. [Ancylostoma ceylanicum, Ancylostoma caninum, and Toxocara canis [Moreover, parasitic worms that commonly infect dogs belong to the nematode species . These tris spp. . They arris spp. . Some tyra canis . STHs inra canis .T. canis was reported to infect Kintamani dogs in the Sukawana Village area with a prevalence of 22.22% [Ancylostoma spp. in tourist areas was reported as 34% [In Bali, several studies have reported the prevalence of worms that infect dogs. For instance, f 22.22% . The pred as 34% . HoweverThe study only used the dog fecal samples which were collected from the residential alley, street, or other public places. Hence, ethical approval was not necessary.This research was conducted from February to November 2019 in the province of Bali, Republic of Indonesia.An observational design with a cross-sectional method was used in this study, and the subjects were the free-roaming dogs in the province of Bali. The study sample was fresh dog feces collected from the subjects. Fresh stools were collected in the morning around the residential alley, street, or other public places and were stored in 10% formalin before being examined at the laboratory . The BalStool examination was performed by a floating concentration method using a saturated NaCl substance . The conThe following formula was used to measure the prevalence of STH , and theThis study demonstrated that the prevalence of STH infection in free-roaming dogs was high (38.36%) . To the Ascaris spp. and Trichuris whose respective prevalence rates were 6.02% and 0.87% , Ascaris spp. (6.02%), and Trichuris spp. (0.87%). Moreover, polyparasitism occurs in multiple infections caused by Ancylostoma spp. and Ascaris spp. In addition, the difference in altitude does not affect the prevalence of STH infection. Therefore, it is reasonable to conclude that the high prevalence of STH infection in free-roaming dogs in Bali is consistent with the life cycle of the parasite in the investigated area.The prevalence of STH infection in free-roaming dogs in Bali was high (38.36%). The STHs causing infection in dogs in Bali were KKA: Designed the study, performed the laboratory work, analyzed the data, and wrote the manuscript. MSA: Designed the study, collected the samples, performed laboratory work, analyzed the data, and wrote the manuscript. NAANS: Collected the samples, performed laboratory work, analyzed the data, and wrote the manuscript. WARW: Collected the samples, performed the laboratory work, analyzed the data, and wrote the manuscript. JKA: Collected the samples and performed the laboratory work. WNFG: Analyzed the data, and wrote the manuscript. IBMO: Performed the laboratory work, and analyzed the data. MS: Performed the laboratory work, and data analysis. NKB: Analyzed the data, and wrote the manuscript. All authors read and approved the final manuscript.


Spodoptera frugiperda first emerged in China in 2019, and therefore the information on indigenous natural enemies of S. frugiperda has been limited in China. In this study, we reported that a dipteran species was observed to infest S. frugiperda collected from maize fields in four different regions of China. Further morphological and molecular recognition identified all the flies as Megaselia scalaris. The findings of this study will improve our understanding on natural enemies to S. frugiperda and potentially provide new ideas for integrated pest management strategies in China.The invasive pest Spodoptera frugiperda first emerged in China in January 2019 and has, to date, migrated to 29 provinces and municipalities in China, causing heavy crop damage in large areas. As a response to this invasive species from the environment, some indigenous natural enemies have been discovered and reported after S. frugiperda invasion. In this paper, parasitic flies were collected and identified from S. frugiperda collected in the Yunnan, Guangxi, and Henan provinces and the Chongqing municipality in China. By using both conventional and molecular approaches, we were able to show that all the parasitic flies of S. frugiperda identified in the four regions were Megaselia. scalaris, and that they attacked the pest larvae and pupae. This is the first report on an indigenous Chinese Megaselia species that has parasitic ability against the invasive pest S. frugiperda, potentially providing new ideas for pest control in China.The invasive pest  Spodoptera frugiperda originated in the Americas and migrated to the African continent in 2016. It was first discovered and reported in China in January 2019 . T. TS. fru results  construc results A and COIlts (COI B gene seS. frugiperda invaded China, Chinese researchers have conducted investigations to identify natural enemies that are able to parasitize S. frugiperda. Egg parasitoid wasps [S. frugiperda larvae were found and reported [S. frugiperda were observed and caught in four regions of China. Morphological and DNA barcode identification confirmed that all the fly specimens collected in the four regions were of the species of M. scalaris.Since id wasps ,21 and sreported ,13. In tM. scalaris is active in a wide range of geographical regions and usually feeds on decaying organic materials [M. scalaris. It has been reported that adult M. scalaris can break into the hive and lay eggs in the hive; the larvae of M. scalaris then hijack the food of bee larvae, therefore hindering the growth and development of bee larvae [Parastagmatoptera tessellata and Triatoma brasiliensis could be parasitized by M. scalaris [M. scalaris can feed and grow in the host’s body. In our study, a similar phenomenon of M. scalaris parasitizing S. frugiperda in various regions of China was observed. The emergence of eggs on the epidermis of S. frugiperda larvae and pupae (S. frugiperda pupae (S. frugiperda larvae (S. frugiperda. The emergence of flies (S. frugiperda. All these results indicate that the indigenous species M. scalaris can parasitize the new invasive pest S. frugiperda in the wild in China. To our knowledge, this is the first record of M. scalaris infesting wild S. frugiperda in China. There have been a few previous reports of this fly infesting other insects in the wild [M. scalaris may play various roles in the ecosystem in addition to being saprophages, such as being parasitic to insects.aterials . It has aterials , and thee larvae . Koch anscalaris ,26; the nd pupae B–D, and da pupae E and thea larvae F demonstof flies A in the the wild ,27,28,29S. frugiperda, several native Chinese insects are reported to be parasitic and predatory to the new invasive pest S. frugiperda. However, their potential applications as biological control agents remain unclear and deserve further investigation. In terms of developing effective IPM strategies for controlling S. frugiperda in China, biological control, protection, and utilization of natural enemies should be given priority for reducing the use of pesticides. Therefore, there needs to be further investigation and exploitation of natural enemies in China to aid the development of subsequent IPM strategies.In consideration of biological control to S. frugiperda in the wild in China. Both conventional and molecular methods identified the dipterans as M. scalaris. This is the first report on the native species M. species infesting S. frugiperda in China. Further investigations on the infestation of S. frugiperda by M. scalaris are warranted and such studies can potentially provide new directions for the integrated pest management strategies in China.In this study, we reported that some dipterans were able to infest invasive pest


Xylocopavirginica , has been recorded from Canada, albeit restricted to southern Ontario and Quebec. However, a single female specimen identified by Hurd in 1954 as X.varipuncta Patton, 1879 from British Columbia is in the C.A. Triplehorn Insect Collection at The Ohio State University (OSUC), suggesting that this species was accidentally introduced into coastal western Canada. As wood-nesters, many large carpenter bees are likely capable of expanding their range great distances by natural and unnatural transport methods while nesting inside suitable substrates, the presumed mode of transport into western Canada, and likely elsewhere. The ease at which the nests are transported has likely contributed to the nomenclatural and distributional ambiguity surrounding this species due to morphological similarities of specimens from North America, Hawaii, and several South Pacific islands.Only one species of large carpenter bee, X.varipuncta from continental North America. Furthermore, these DNA barcode sequences also match those of specimens identified as X.sonorina Smith, 1874 from the French Polynesian and Samoan Islands, thus fully supporting the opinion of X.sonorina, a species described from and likely introduced to Hawaii is the oldest name available, X.varipuncta is here placed into synonymy. Additional research will be needed to trace the timing and pathway of introduction and establishment of X.sonorina; it is presumed that the species is native to the southwestern United States but has been established in Hawaii since the mid-1800s. It is also established in French Polynesia, the Samoan Islands, and likely other south Pacific islands, with additional records of occurrence from Java, New Zealand, and now Canada.By comparing DNA barcodes of specimens from the western United States to specimens from Hawaii, we confirm the early opinion of P.H. Timberlake  that spe Xylocopa Latreille, 1802 , are large bumble bee sized bees that typically excavate nesting cavities into wood, bamboo, or the woody stems of plants , a single female identified as X.varipuncta Patton, 1879, collected from Capilano Canyon near Vancouver, British Columbia by R.C. Osburn in 1949 . This unpublished Canadian record was also not included in the recent review of the subgenusNeoxylocopa Michener, 1954 . It is assumed that finding multiple specimens from the same collecting event increases the likelihood that the material was not mislabeled.To confirm that the specimen of Xylocopa in the Barcode of Life Data (BOLD) System  System , includi) System . Sequenc Islands  using vaSmith, 1874720FBA9B-6BE1-5A96-804E-1A2E9609DE79XylocopasonorinaHolotype ♀. HAWAII, Sandw[ich]. Isl[and].” , and two specimens of Trichiosomatriangulum Kirby, 1837 . The latter species at least is widespread in North America, including ranging into British Columbia.We are confident that the specimen identified as cta Fig.  from BriX.varipuncta generated in this study from western North America matched those from Hawaii, but also those from specimens in BOLD identified as X.sonorina Smith from the islands of Huahine-It and Mo’orea in French Polynesia, and Apia in the Samoan Islands, further supporting the opinion of DNA barcodes from specimens of X.sonorina has a long and interesting history, largely impacted by its arrival and subsequent establishment in Hawaii; in fact, the type locality of this species is Hawaii , south into Central and South America, and the West Indies, and also two additional subspecies, aeneipennis  in Arizona, and cubaecola Lucas, 1857 in California. The latter two subspecies of X.aeneipennis is now considered a valid species from South America (Suriname), and X.cubaecola is a valid species endemic to Cuba  previously identified as X.aeneipennis by X.varipuncta from the continental United States, an opinion shared by T.D.A. Cockerell  into synonymy with X.b.sonorina.Though na Smith  and X.ssonorina , a specisonorina , though X.sonorina, and subsequent misidentifications of material from Hawaii has had serious implications. Though there is only one species of Xylocopa known from Hawaii (see X.sonorina named five years previous) as X.aeneipennis, the species known from Suriname mentioned above. This error is likely why X.aeneipennis for Hawaiian material  which share DNA barcodes with specimens identified as X.sonorina from the islands of Huahine-It and Mo’orea in French Polynesia, and Apia in the Samoan Islands  are presumed native to the Galápagos  are probably more suitable for long distance travel than larger bees .It is entirely possible that a nest with at least one specimen of ood e.g.  from thepossible . In oceapossible  providedpossible , with lopossible  or greatpossible  with decpossible . HoweverXylocopa being intercepted at other locations; X.virginica was collected in Nottingham England from nests built in wood. Megachilexylocopoides Smith, 1853 (Megachilidae)) have been intercepted at the Canadian border, but have not been recorded as established (Much faster travel time of materials containing bee nests is obtained via commerce, which is likely the main means of introduction today. Recently, a carpenter native to Japan and China was found in California , suggestablished .X.sonorina is now known from British Columbia, no subsequent work has confirmed its establishment in Canada (see Once arriving to a new location, some bee species can succeed even with very low numbers of colonizers. nada see . Considenada see . Interesnada see  and morenada see . Donovan


Lawsonia intracellularis, an obligately intracellular enteric bacterium, infects intestinal epithelial cells, but may also be found within macrophages in the intestinal lamina propria of affected pigs. Macrophages play an important role in host defense against infectious agents, but the role of this cell in L. intracellularis infection is not well understood. The aim of this study was to evaluate the permissibility of macrophages to L. intracellularis infection in vitro. Pure culture of L. intracellularis was added to swine peripheral blood monocyte-derived macrophages. Viability of intracytoplasmic L. intracellularis was evaluated at different time points by transmission electron microscopy (TEM). Potential replication of L. intracellularis in macrophages was also evaluated by qPCR. By TEM, phagocytosis L. intracellularis within of phagolysosomes were observed 1-hour post-infection (hpi) and bacterial structures in binary fission at 48 hpi. The number of intracellular bacteria was determined at 1, 4, 24, 48, and 72 hpi by qPCR in infected macrophages and compared to the number of intracellular bacteria from culture in McCoy cells. In both cell lines, the amount of L. intracellularis was decreased at 4 hpiand increased at 24 hpi. The number of intracellular bacteria continued to increase in McCoy cells over time. This is the first study showing interaction, survival and propagation of L. intracellularis in macrophages. These findings are critical to establish an experimental model for future studies of the pathogenesis of porcine proliferative enteropathy and the potential persistence of L. intracellularis in macrophages during chronic infections. Macrophages play an important role in innate and adaptive immune responses against invading pathogens. They contribute to the elimination of microorganisms and are capable of presenting antigens to T cells. In addition, macrophages may serve as an escape route for some bacterial species, as intracellular bacteria may remain viable and potentially proliferate and/or disseminate in the host , 2.Lawsonia intracellularis is a microaerophilic, obligately intracellular bacterium that causes porcine proliferative enteropathy (PPE)  findings, we hypothesized that L. intracellularis can survive and propagate in macrophages [L. intracellularis organisms within phagolysosomes in cultured macrophages, free in the macrophage cytoplasm, and in binary fission. These results indicate that L. intracellularis has the ability to survive and propagate inside macrophages. However, despite the capacity of L. intracellularis to survive within the macrophages as shown in the present study, only fortuitous cases of L. intracellularis infection are associated to extra intestinal lesions [L. intracellularis indicate that the survival of L. intracellularis in macrophages is not related to the dissemination of the bacterium to extra intestinal tissues, as already described for other intestinal gram-negative bacteria [L. intracellularis to survive intracellularly whether in epithelial cell or macrophages are as still required.nd birds . L. intr in 1993 . Due the in 1993 , researcprevious  e 23 finrophages . The preprevious  e 23 fin lesions . These rbacteria , 28. DirRhodococcus equi and Coxiella burnetii [R. equi survive and proliferate within intracellular environment by blocking the acidification of the phagolysosome [C. burnetii delays the fusion process between the phagosome and the lysosome [Listeria monocytogenes [Franscisella tularensis [Edwardsiella tarda [L. intracellularis pathogenesis.The survival of bacteria in phagolysosomal compartments has been observed in other species, such as burnetii , 29. R. lysosome . On anotlysosome . Binary lysosome . Howeverytogenes , Franscilarensis , and Edwla tarda , are knoL. intracellularis survival within the macrophages may also be related to the persistence of the microorganisms in the host, which results in the chronic presentation of the disease. Therefore, the elimination of L. intracellularis for long periods, as previously demonstrated by Guedes and Gebhart and Stege et al, may be a source of dissemination of the pathogen in the herd [the herd , 35.L. intracellularis has the capacity to proliferate at low levels within the macrophages, but other methods should be used in future studies to confirm this hypothesis. Our findings do not corroborate Lawson et al. [L. intracellularis needs mitotically active cells for its proliferation. L. intracellularis is a unique bacterial species, with several peculiarities obligately intracellular microorganism, requires specific microaerophilic atmosphere that make it difficult to grow and manipulate under laboratory conditions. Therefore, evaluation of L. intracellularis survival and its quantification inside the cells is not possible using conventional assays used for studying other bacterial species [L. intracellularis potential to survive and to replicate within macrophages, therefore, it represents the first step towards the understanding of the persistence of the organism in chronically-infected animals.To assess the bacterial numbers in the macrophage cultures over time, qPCR was performed. This technique detects DNA sequences with good sensitivity and specificity but lacks the ability to differentiate metabolically active and inactive microorganisms. In addition, the chronological evaluation was performed by using samples from different wells, which can result in numerical variations from well to well. Nevertheless, from the results obtained, we can hypothesize that n et al.  findings species , 37. TheL. intracellularis is able to internalize and survive within phagolysosomal environment of swine macrophages. In addition, this is the first study to report the binary fission of L. intracellularis free in the cytoplasm of swine macrophages. These findings are critical to establish an experimental model for future studies the pathogenesis of porcine proliferative enteropathy and the potential persistence of L. intracellularis in macrophages during chronic infections.In conclusion, our results showed that S1 FigL: lysosome; N: nucleus; rER: rough endoplasmic reticulum.(TIF)Click here for additional data file.


Dear Editor,We appreciate the comments of Dr. Naem and Dr. Al-Kurdy. This case report emphasizes that local methotrexate injection may become the treatment of choice for cervical ectopic pregnancies in the near future. Injection of methotrexate in a cervical pregnancy with advanced gestational age (11 w) and CRL of 41 mm makes this case unique. This is the most advanced cervical pregnancy in the literature where methotrexate has been administered and resolved the pregnancy without further intervention . The comAs Dr. Naem and Al-Kurdy mentioned, it is usually the main concern of the patient to preserve her fertility . ConservOur patient had almost a symptom free follow up after methotrexate injection, so on the 7th day of her procedure she was offered to discharge with the intention of decreasing hospital stay. The pattern of the hCG decline in our patient, where hCG values normalized after 90 days from the injection, was similar with the other studies ,5.Success of systematic methotrexate injection is related to absence of fetal cardiac activity, level of hCG , gestational age smaller than 9 weeks and a crown-rump length(CRL) of <10 mm . Yet, weOverall, local methotrexate injection is a safe method to practice in cervical pregnancies with minimum complication risk, regardless the gestational age, presence of fetal cardiac activity and CRL.None.None.None. Our paper is in the format of letter to editor.None. Our paper is in the format of letter to editor.Our paper is in the format of letter to editor. It is a response to the letter to editor which referred to our article “Conservative management of 11 weeks old cervical ectopic pregnancy with transvaginal ultrasound-guided combined methotrexate injection: Case Report and Literature Review” written by the authors listed below:Ipek Betul Ozcivit, MD.Ismail Cepni, Prof. MD.Kubra Hamzaoglu, MD.Hakan Erenel, MD.Rıza Madazlı, Prof. MD.N/A.Ismail Cepni, Prof. MD.


Anisakis. In Europe, Anisakis pegreffii has been detected as the causative agent, although the sibling species Anisakis simplex sensu stricto (s.s.) is also known to cause the disease in other parts of the world, and discrepancies exist regarding their respective pathogenic potential. In Spain a high number of cases has been recorded, with marinated anchovies being the main source of infection, although no specific diagnosis has been documented in humans. In this study, we analyzed three cases of anisakiosis in patients from Barcelona (Spain) who had consumed undercooked hake. All patients described epigastric pain and several larval nematodes were removed endoscopically from their stomachs. Larvae were morphologically characterized as third-stage larvae of Anisakis simplex sensu lato (s.l.) and molecularly identified as A. simplex (s.s.) by means of PCR RFLP of the ITS region of the rDNA and sequencing of the elongation factor1 alpha1 (EF1 α-1) nDNA gen. This study represents the first specific identification of Anisakis larvae in clinical cases of anisakiosis reported in Spain. Specific molecular diagnosis is of crucial importance for assessing the health risk of Anisakis sibling species. Hake consumption stands out as a risk factor for anisakiosis, since this fish species can be highly parasitized.Anisakiosis is a fish-borne disease with gastrointestinal and/or allergic symptoms caused by the consumption of raw or undercooked fish parasitized with nematode larvae of the genus  Anisakis, although Pseudoterranova and sporadically Contracaecum species can produce pathology, in this case known as anisakidosis [Anisakiosis is caused by the consumption of raw or undercooked fish and cephalopods parasitized with larvae of anisakid nematodes. The main causative agents of the disease are species of akidosis . Dietaryakidosis . Anisakiakidosis ,4,5,6,7.akidosis ,5,6,8. Fakidosis ,10. The akidosis .Anisakis species, in consumed fish can also trigger allergic reactions of different degrees of pathogenicity, due to IgE-Anisakis hypersensitivity, the symptoms ranging from moderate, such as urticaria and angioedema, to severe, including anaphylactic shock [The most frequent forms of anisakiosis are gastric and intestinal. In these cases, third-stage larvae L3) adhere to and penetrate the mucosal layer of the stomach or intestine, causing epigastric and/or abdominal pain and usually nausea and vomiting ,12. The ic shock . In gast adhere tic shock ,12. A. simplex (s.l.): A. simplex (s.s.) and A. pegreffii. The two species have a wide defined distribution, however their range of localization overlaps in some basin waters, such as the Spanish Atlantic coast and the Japanese Sea. The third species within the A. simplex (s.l.) complex, Anisakis berlandi, which has a discontinuous distribution in the Austral region, has not been implicated in the disease [The main species implicated in human anisakiosis belong to the complex  disease .Anisakis species, such as those of the A. simplex (s.l.) complex [Anisakis species can cause anisakiosis in humans [A. simplex (s.s.) is more resistant to gastric acid than A. pegreffii and has a higher rate of penetration in the stomach, small intestine or colon wall [Anisakis genus level or as A. simplex (s.l.) [In the last two decades, the use of molecular methods as PCR RFLP, DNA sequencing or, more recently DNA microsatellites, has improved the specific identification of larvae of morphologically non-differentiable  complex ,13,14. Tn humans . Specifin humans . Some aulon wall . Despitex (s.l.) ,17,18,19Anisakis larvae obtained from three human cases of gastric anisakiosis, caused by the consumption of undercooked hake, were characterized and for the first time in Spain the causative species were molecularly identified.The aim of the present study was to perform a specific diagnosis of anisakiosis and thereby shed light on the species-related health risk for humans. Two women aged 45 and 49 years and one man aged 21 suffering acute epigastric pain were attended in three hospitals in the Mediterranean city of Barcelona, in the North-East of Spain, in January 2019. The three patients declared they had consumed fresh undercooked hake at home within 24 h before their hospital admission. In all cases the fish was purchased in a supermarket chain, although the capture zone is not known. During the medical examination, 12, 5, and 1 nematode larvae were detected adhered to the mucosal layer of the stomach of the three patients, respectively, and in the former a thickening of the gastric antrum was observed. The larvae were endoscopically removed, and the symptoms remitted spontaneously. All nematodes were preserved in 70% alcohol, and 7 specimens from the first patient, 5 from the second and 1 from the third were available for analysis.HinfI and HhaI [The morphology of the larvae was studied microscopically . For moland HhaI . The eloand HhaI .A. simplex (s.l.), based mainly on the presence of a mucron at the caudal end and an elongated ventricle with an oblique posterior end. The specific patterns obtained by PCR-RFLP, using HinfI and HhaI, showed that all the larvae studied belonged to the species A. simplex (s.s.), differentiating them from the other two sibling species A. pegreffii and A. berlandi (A. simplex (s.s.) (KT825685) [The larvae from the three patients were morphologically identified as L3 larvae of berlandi . In addiT825685) . SequencAnisakis-specific IgE [Several cases of gastric, intestinal and allergic anisakiosis have been reported in Spain in recent years, the first one being documented in 1991 . Most ofific IgE ,24,25.A. simplex (s.s.) has been frequently detected in fish species commonly consumed in Spain caught in the North-East Atlantic, including the Alboran Sea region, and shows only a sporadic presence in Mediterranean fishes, specifically those caught near the African coast [A. pegreffii, on the other hand, is endemic to the Mediterranean Sea, but has also been reported in fish caught off the North-East Atlantic coast [A. simplex (s.s.) as the causative agent suggests that the hake consumed was caught in the North-East Atlantic region. In fact, in the epidemiological studies on hake from North-East Atlantic and Mediterranean, A. simplex (s.s.) was not found in the Mediterranean hosts [None of the documented clinical cases in Spain report specific identification of larvae. an coast . The sibic coast ,26. Compic coast . All patan hosts .A. pegreffii larvae have been molecularly identified in human anisakiosis cases [A. pegreffii is endemic [A. simplex (s.s.) being the main agent in Japan and A. pegreffii in Korea [Anisakis species observed in fish between capture zones [Elsewhere in Europe, in Italy and Croatia, only is cases ,28. In tin Korea ,29. The re zones .A. simplex (s.s.). Nevertheless, the presence of A. pegreffii in Mediterranean fishes widely consumed in Spain implies that this parasite species would be also involved in the disease. The health risk posed by both sibling species should be assessed by more frequent use of molecular specific diagnosis in clinical cases of anisakiosis. The results of this study also highlight hake consumption as a risk factor for the disease, as this species of fish can be heavily parasitized [The results presented here constitute the first species-specific identification of larvae implicated in human anisakiosis in Spain. In the three studied cases all the larvae were identified as asitized ,30. Cons


Scientific Reportshttps://doi.org/10.1038/s41598-019-43515-w, published online 06 May 2019Correction to: In the original version of this Article, Supplementary Video 1 was published in an incorrect format.In addition, the email address for Anna Z. Komisarczuk quoted in the original version of this Article has been changed. Correspondence and requests for materials should be addressed to a.z.komisarczuk@ibv.uio.no.These errors have now been corrected in the HTML and PDF versions of the Article, and in the Supplementary Information that now accompanies the Article.


Picramnia and Alvaradoa, with the recent addition of a third genus, Nothotalisia, described in 2011. Only a few species of the family have been the subject of DNA-related research, and fewer than half of the species have been included in morphological phylogenetic analyses. It is clear that Picramniaceae remains a largely under-researched plant group. Here we present the first molecular phylogenetic tree of the group, based on both chloroplast and nuclear markers, widely adopted in the plant DNA barcoding. The main findings are: The family and its genera are monophyletic and Picramnia is sister to two other genera; some clades corroborate previous assumptions of relationships made on a morphological or geographical basis, while most parts of the molecular topology suggest high levels of homoplasy in the morphological evolution of Picramnia.Picramniaceae is the only member of Picramniales which is sister to the clade )) in the rosidsmalvids. Not much is known about most aspects of their ecology, geography, and morphology. The family is restricted to American tropics. Picramniaceae representatives are rich in secondary metabolites; some species are known to be important for pharmaceutical purposes. Traditionally, Picramniaceae was classified as a subfamily of Simaroubaceae, but from 1995 on, it has been segregated containing two genera,  Picramnia Sw. and Alvaradoa Liebm., were separated as Picramniaceae family [Most flowering plants families described by the end of the 18th century have proven to be robust, stable groups. However, molecular tools have also been used to find less stable assemblages. One example of this is Simaroubaceae, a family in the Sapindales which was long suspected of being “non-natural” [e family .Picramniaceae Fernando and Quinn is a rosid family ofca. 50 known species. They are dioecious, neotropical plants, with alternate, pinnately compound leaves, minute flowers with stamens opposite the petals, and a syncarpous gynoeceum with an inconspicuous style . It is tDue to their small, actinomorphic flowers, bitter bark, and pinnately compound leaves, they were initially placed in Simaroubaceae . HoweverPicramnia [Alvaradoa [Picramnia is small or mid-size trees that grow either in the lower stories of moist tropical forests  or in more dry habitats, from Mexico and Caribbeans to Southern Brazil and Northern Argentina. Some of them (like the northernmost Picramnia pentandra Sw. from South Florida) frequently grow as shrubs. Gross morphology of Picramnia is rather uniform and non-distinct, this was probably a reason of some notorious mistakes. One of examples is that Hispaniola generic endemic Casabitoa perfae described as a plant with putative affinities to Phyllanthaceae, is actually identical to Picramnia dictyoneura (Urb.) Urb. and Ekman [Picramniaceae was formerly circumscribed with two genera, icramnia  and Alvalvaradoa . Picramnnd Ekman .Alvaradoa is significantly more xerophilous and shrubby, with wind-dispersed fruits . The range of distribution of the genus is conspicuously disjunct: Mexico and Caribbeans , and Bolivia, Argentina, and South Brazil, as depicted by Thomas [y Thomas .Nothotalisia W.W.Thomas [TalisiaAubl. (Sapindaceae). Nothotalisia, like some species of Picramnia, includes mid-size forest trees that exhibits a broad distribution, from Panama to Peru, and likely associated with wet tropical forests of Choco and Amazonia. Interestingly, Nothotalisia appears not to be rare in collections: For example, it is abundant in herbarium samples collected in the Darien province of Panama . In all, we believe that diversity of Picramniaceae is far from being exhausted, and we can always expect new discoveries.Recently, the third genus, W.Thomas , was disGumillea Ruiz and Pav. [Gumillea auriculata Ruiz and Pav. reminds Picramnia but the plant bears stipules and bisexual flowers. Even though there is no recent specimen of that plant nor any molecular data, the morphology of inflorescence, flowers, and leaves suggests a relationship to Picramniales. The structures described and illustrated as “stipules” look like pseudostipules found in some species of Picramnia  Jans.-Jac.) where petiole is reduced and modified basal leaflets deflected to protect axillary buds [We should mention here also the monotypic and Pav.  which miand Pav. . This geary buds ,16.Only a few species of Picramniaceae have been the subject of DNA-related research, and there are no molecular phylogenetic analyses so far. No generic subdivision was established; even the total number of species is not absolutely clear. Apparently, Picramniaceae remains an under-researched group.To fill this gap, we performed a broad sampling of the group, aiming at collecting as many tissue samples as possible to amplify several widely used DNA barcoding markers in order to cover the species diversity in the group. We believe that our reconstruction of phylogenetic trees may become a basis for future studies in the group.We collected 276 tissue samples  from repOur strategy was developed with an idea of the broadest sampling. We typically obtained multiple samples per species and attempted to extract DNA and sequence our markers multiple times until we reach satisfactory results. As we work with under-studied group, it was also especially important to provide photographic vouchers for each sample, and to be sure of the correct identifications, so either determination from known experts or our own determinations were preferred. At the end of this process, only a few species were missed from our samples, and most of them are either rare or local. As of today, our sequencing list contains 41 species, which is about 80% of the supposed group diversity. Most of them are newly sequenced material, only eight species represented with information in the public databases (GenBank and BOLD). In all, we were able to increase the amount of available information five-fold. Due to the apparent problems with identification, we always trusted our samples first. To improve the quality of databases, we constructed the working classification of Picraminiaceae , which nDNA extraction of the Picramniaceae herbarium samples was not always straightforward. Less than 20% of herbarium samples, fresh or old, yielded the DNA of the appropriate quality. We believe that the main reasons are the conditions of their collection and preparation. We were able also to extract DNA from living samples (collected in silica gel or just air-dried) taken from 8 species, and in all these cases the DNA extraction, amplification and sequencing were successful.The DNA extraction was made using NUCLEOSPIN Plant II Kit , which we believe is a good trade-off between efficiency and simplicity. We improved the protocol to increase the lysis time to 30 min and employ thermomixer on the slow rotation speed (350 rpm) instead of a water bath. We used Nanodrop 1000 Spectrophotometer , to estimate concentration and purity (the 260/280 nm ratio of absorbance) of DNA.trnL-F [rbcL [We employed three of the most widely used barcoding DNA markers, chloroplast trnL-F  and rbcL-F [rbcL , and nuc-F [rbcL . The latTo make this research portable and expandable, we automated most of the steps with the“Ripeline” workflow . This isLeitneria floridana Chapm. (currently included in Simaroubaceae), Kirkia acuminata Oliv. (Kirkiaceae) and Talisia nervosa Radlk. (Sapindaceae), or with L. floridana alone. To stabilize the tree and to increase branch support, we used the hyper-matrix approach [With the help of Ripeline, we were able to create the super-matrix, which includes all three DNA markers and obtain maximal parsimony (MP) and Bayesian (MB) phylogeny trees. Maximum parsimony analyses were run with the help of R phangorn package  using paapproach  and concNo morphological matrix is available for the whole family, so one of the most straightforward ways to assess its morphological diversity was to use the hierarchical components of identification keys . To see Datasets, scripts, trees for individual markers and other information used in preparation of this publication are available as one zip archive from the first author’s Open Repository . We encoPicramnia and Nothotalisia + Alvaradoa, with sufficient (close to 100%) support.With the outgroups mentioned above, Picramniaceae formed a stable clade. First branches  are PicrAlvaradoa subovata Cronquist and A. puberulenta (Monach.) Sleumer form the first clade whereas West Indian species—the second clade. Alvaradoa amorphoides Liebm., which grows in Mexico and Florida, is basal to West Indian species.This part of the tree is strongly supported . South ANothotalisia peruviana (Standl.) W.W. Thomas and N. cancellata W.W. Thomas form the stable clade to  P. andrade-limae Pirani + P. elliptica Pirani and W.W. Thomas + P. bahiensis Turcz. + P. grandifolia Engl. (latter two with less confidence); (3) P. dictyoneura Planch. + P. sphaerocarpa Planch. and (4) P. glazioviana Engl. + P. parvifolia Engl.; (5) P. oreadica Pirani + P. ramiflora Planch. + P. campestris + P. juniniana J.F.Macbr. However, some lower supported groups also make sense, especially (6) P. excelsa Kuhlm. ex Pirani + P. gracilis Tul. + P. nuriensis Steyerm. +P. tumbesina Cornejo.Generally, there is much less stability in that part of the tree. Many clades are not reliably supported. The most supported are the following groupings: (1) Picramnia are still missing in our molecular dataset. To test the possible placement of these species, we used the hierarchical component of identification keys together with phylogeny trees and employed k-nearest neighbor machine learning with bootstrap to point on possible sister species. Since super-species level classification is not developed, we decided to return trios of putative neighbor species which the highest bootstrap confidence  and bicarpellate (part of Picramnia) are likely advanced character states. There is no visible pattern for the position of inflorescence : anciens sessile flowers and fruits, although pedicellate flowers occur in some specimens. It is a widespread, mostly Amazonian species that reach Central America; therefore, its closer relationship to Central American to Andean species  makes sense on a geographical basis. P. ramiflora, however, is a quite disjunct species from the Brazilian Atlantic Forest.Pirani ,16 propoP. thomasii )) is morphologically heterogeneous, with a mixture of species bearing pentamerous flowers (P. latifolia and P. thomasii) and species with trimerous or tetramerous flowers (the other two). This fact implies that even though the numbers of sepals and petals have a consistent relevance for species recognition within the genus, often used in keys  nested with P. hirsuta W.W. Thomas and P. teapensis Tul. (3–4-merous); (b) P. elliptica and P. grandifolia (5-merous) closely related to P. bahiensis and P. andrade-limae (3-merous); (c) P. excelsa (3-merous) nested within a clade with P. sellowii and P. guianensis (both 5-merous), along with P. sphaerocarpa which is particularly polymorphic producing 3–4–5-merous flowers.The clade formed by , ,36, P. tPicramnia, as well as to generate assumptions of relationships, concern to the inflorescence type and position  Planch., which bears profusely branched thyrses. In the same way, racemose P. elliptica and P. grandifolia here emerged within a clade with P. andrade-limae and P. bahiensis; both provided with thyrses.Other morphological features that have long been used in the taxonomy of n e.g., ,31); the; thePicrP. glazioviana and P. parvifolia proposed by Pirani [P. parvifolia as is P. excelsa [Araucaria forests of Southern Brazil, did not emerge in that same clade. Such reasoning may also be pointed to P. glazioviana, which is quite similar and sympatric to P. ciliata in the Brazilian coastal moist forests, but this latter emerged near the base of the molecular tree. P. andrade-limae and P. bahiensis also have trimerous flowers with peculiar obcordate petals and assumed as related to the aforementioned Atlantic Forest group with P. glazioviana [Nonetheless, some assumptions of relationships previously made on morphological grounds are here corroborated by the molecular data. For example, the close relationship between y Pirani , based o excelsa , both syzioviana , but emeP. oreadica did not emerge together, but the fact that each of them is currently ascribed to distinct allopatric subspecies may be considered here [cerrados of Central Brazilian Plateau [P. guianensis would be nested in the clade including P. oreadica and P. campestris, because they share a special type of indumentum on the ovary (peculiar clavate trichomes), and are all mostly related to sandy or rocky substrates. These species live mostly as heliophytic shrubs, contrasting with the typical understory habit prevailing in the genus. However, the fact the P. guianensis emerged in another clade, that feature may have appeared independently. P. ferrea Pirani and W.W.Thomas, unsuccessfully sampled in this analysis, could probably belong to the P. guianensis clade on a biogeographical basis, or the P. campestris clade on morphological grounds .Astonishingly, the two samples of red here . These a Plateau , known sPicramnia. On the other hand, as we pointed out previously, it is remarkable that parts of the topology seem to be more or less geographically structured since two clades are mostly composed of Bolivian to extra-Amazonian Brazilian species. Another clade is mostly composed of species ranging from Darien (Panama) to Peru extending to the Amazonian region. In contrast, the next two clades are composed of a mixture of species, either showing the latter distribution pattern or species ranging from Central American to the West Indies. Even though we are aware that the definition of these areas of distribution is somewhat artificial, they are mostly in agreement with biogeographical regions and provinces previously proposed for the Americas [Picramnia may reflect some of the geomorphological, climatic, and ecological grounds that helped to shape or drive the history of the neotropical biota.Hence, there is some evidence which corroborates previous assumptions of relationships made on a morphological basis, while most parts of the molecular topology suggest high levels of homoplasy in the structural evolution of Americas . The spaEven with extensive sampling and continuous sequencing efforts, our data is still incomplete. Only 20% of our sequencing efforts ended with reliable sequences. GenBank data generally lack sequences of Picramniaceae. Some rare species are still not sampled.Nevertheless, we think that our phylogeny results might serve as a backbone for future research. Our best hope is that this work represents the beginning of the second, molecular stage in Picramniaceae research. One of the most apparent next steps is to check how morphology and DNA go together, the molecular “weight” of traditional morphological characters.


Resilience is defined as a personal quality that enables growth in knowledge, understanding and self-actualization in the face of adversity and life disruptions. Low levels of resilience can dispose older adults to higher risks for negative health outcomes in the aftermath of traumatic events. However, we have limited knowledge of resilience among minority aging populations. This study aims to examine the presence and levels of resilience and its sociodemographic correlates among U.S. Chinese older adults. Data were drawn from the Population Study of Chinese Elderly, an epidemiology study of U.S. Chinese older adults in the Greater Chicago area. Guided by a community-based participatory research approach, a total of 3,036 Chinese older adults aged 60 and above participated in face-to-face interviews from 2015 to 2017. Spearman’s rank-order coefficient was utilized to test correlation. A 10-item validated Chinese version of the Connor-Davidson resilience scale was used to assess resilience. In our sample, 59.7% were female, and the average age was 75. The mean resilience score was 26.9, ranging from 1 to 40. U.S. Chinese older adults who were younger, male, married, had higher education and income, fewer children, better health status and quality of life, and improved health and have lived fewer years in the U.S. reported higher levels of resilience. Future longitudinal research is needed to investigate the protective effects of resilience among older Chinese Americans against mental and physical distress.


Crotalus genus stand out. Out of more than 40 species in the country, nine are found in Zacatecas: C.aquilus, C.atrox, C.basiliscus, C.lepidus, C.molossus, C.polystictus, C.pricei, C.scutulatus and C.willardi. Although these reptiles are important, due to their relevance in terms of ecology, cultural use and public health, their conservation is impacted by multiple factors, such as habitat fragmentation and indiscriminate killing. Thus, most species within this genus are found in some type of risk category at both the national and international level. The purpose of this study was to determine the potential distribution and diversity of rattlesnakes at the municipal level in the understudied state of Zacatecas. To do this, we analyzed and described the global distribution of nine rattlesnake species by building species distribution models, which determined their potential distribution based on a set of ecological variables and presence records. The resulting models were used to assess the diversity of rattlesnake species potentially present in each municipality within the state. Thirty-nine (67.24 %) out of fifty-eight municipalities registered at least one rattlesnake species. Fresnillo, Sombrerete and Valparaíso were some of the municipalities showing greatest diversity. Moreover, C.atrox, C.lepidus, C.molossus and C.scutulatus were the most widely found species in the state. On the other hand, C.basiliscus, C.polystictus, C.pricei and C.willardi were rarely spotted and so, information on their distribution patterns within Zacatecas is limited. Finally, the areas having the largest potential for the distribution of these species were defined. These findings should make field work much more time- and cost-effective, facilitating the collection of in situ data that are useful for management and conservation plans of these species in Zacatecas.Mexico is home to a large number of reptile species and has one of the greatest diversities of venomous snakes, among which the rattlesnakes pertaining to the  Crotalus genus are noteworthy. As of 2018, this genus is comprised of 47 species, with 42 of them found in Mexico, among which 27 are endemic (Uetz 2018). Recently, there have been several studies on the herpetofauna in the central and northern regions of Mexico, in which Zacatecas is located. This state is one of the largest and shares its borders with eight other states , their natural history (LC) risk category. Conservation threats mainly originate from the fragmentation of their habitat derived from land use changes, as well as hunting and illegal trade . These are based on the assumption that the distribution of a given species is the result -at least within a short time frame- of a balance between undisturbed factors, i.e., a (pseudo)-equilibrium between the biotic entities and the physical characteristics To determine the potential distribution of rattlesnakes in Zacatecas, we gathered records on their global presence. In order to do this, we conducted queries in the databases of the Global Biodiversity Information Facility: aquilus  and C.bacatecas . Finallystrada 2041) by elCrotalus genus reported in Zacatecas, two groups of study were established: 1) species endemic to Mexico and 2) species non-endemic to Mexico . The second polygon comprised the entire Mexican territory together with the southernmost states of the United States of America, namely, Arizona, Arkansas, California, Colorado, Kansas, Louisiana, Nevada, New Mexico, Oklahoma, Texas and Utah. This was done to include those species whose distribution range is also present in this portion of the USA .According to sdm, version 1.0.46, implemented in the R software, which assembles and evaluates models using 15 algorithms: BIOCLIM, BIOCLIM.DISMO, BRT, CART, FDA, GAM, GLM, GLMNET, MARS, MAXENT, MAXLIKE, MDF, RF, RPART and SVM. This provides the potential distribution of a given species or community. For each species, we used all available presence points, in addition to several background points created randomly within the modelling extent, so that the proportion of presence points was 30 % for all species , to keep only informative variables in the models. Finally, any non-significant variables that might remain in the model after this process were eliminated , which is considered a standard method to evaluate the discrimination capacity (i.e. differentiating the locations where presence and non-presence has been recorded) of predictive distribution models, avoiding the subjectivity of choosing a classification threshold. The value of AUC depends on the ratio of presence and the size of the area to be modeled, as pinpointed by . Thus, the field work in this study comprised a total of 48 localities throughout 22 municipalities within the state. These 22 municipalities were chosen randomly to include various regions throughout the state. Moreover, in these municipalities, we selected localities situated within the areas of greatest suitability as predicted by the models. A map of Mexican localities was overlapped to the potential distribution maps to conduct this geographical selection . Field wC.scutulatus to 84 records for C.willardi throughout their distribution range. The latter species, along with C.basiliscus and C.pricei, had the lowest number of available presence records, as opposed to C.molossus, which was the species with the greatest number of records in Zacatecas , bio_14 (precipitation of driest month), bio_19 (precipitation of coldest quarter), altitude, land cover and slope. Anthropogenic influence, which intuitively includes the direct influence of human beings on ecosystems , impacted the presence of C.atrox, C.lepidus, C.molossus and C.scutulatus. Rock types and edaphology were less relevant at the time of model preparation. On the other hand, variables such as bio_06 (min temperature of coldest month) or bio_10 (mean temperature of warmest quarter) were not selected in any of the distribution models.In the species distribution modeling, the most informative environmental variables for the models were C.aquilus  exhibits an extensive potential distribution within Zacatecas  and one more unpublished record for C.aquilus were added. According to the compilation of total records, Sombrerete and Valparaíso were the municipalities exhibiting the highest rattlesnake diversity, with six different species being reported, followed by Fresnillo, with five different species. The municipalities of Chalchihuites, Concepción del Oro, El Plateado de Joaquín Amaro, Genaro Codina, Guadalupe, Mazapil, Monte Escobedo and Pinos reported four species each. The municipality of Zacatecas, where the state capital is located, has the presence of three species of this genus. Although there are no occurrence records for rattlesnakes in 19 municipalities, according to the SDM, all municipalities have the minimum conditions for the presence of rattlesnakes. It is noteworthy that C.molossus has reported presence in 24 municipalities and C.lepidus and C.scutulatus in 23 municipalities, as opposed to C.pricei and C.willardi, reported in only two municipalities.Thirty-nine (67.24 %) out of the fifty-eight municipalities Table  have at C.aquilus, C.pricei and C.willardi.The model building area spanned the entire known distribution range of these species, so as to cover all possible habitats where the species might be found . HoweverC.aquilus exhibited presence records in Huanusco, Nochistlán de Mejía and Pinos. Likewise, the SDM identified the municipalities of Cuauhtémoc and Genaro Codina as potential areas for the occurrence of this species and the municipality of Pinos was also indicated as a potential area of occurrence, which is demonstrated with the presence record reported for this area.C.basiliscus was pointed out as a frequently spotted species in the region. Given that this organism is easily mistaken for C.molossus, we only considered records backed up with specimens and the knowledge that C.basiliscus prefers zones around 1,000 meters above sea level, which are the sites where georeferencing data were obtained. This is in agreement with the information reported by During our post-modelling field work, further occurrence records were collected. Moreover, when interviews were applied to individuals from different communities, C.atrox, C.lepidus, C.molossus, C.scutulatus, C.pricei and C.willardi, which is consistent with the literature and data obtained, since the first four species do not seem to have any problem to occupy habitats in close proximity to both urban and rural areas. In relation to this, presence records of C.atrox and C.molossus were obtained within populated areas, sightings of C.lepidus and C.molossus were also registered in the hills of Los Alamitos and La Virgen, which are located next to the largest and most populated urban area in the state. Sightings of C.lepidus in the aforementioned regions occurred in steep slopes and predominantly rocky soils, being the first variable (i.e. slope) among the ones selected for the construction of the SDM. Human presence is not an obstacle for the sighting of C.atrox and C.scutulatus, or at least for their transit in the vicinity of noisy urban areas with dense housing and transportation infrastructures. Indeed, some of the documented records for C.atrox are in paved and dirt roads. Moreover, C.atrox and C.scutulatus as two of the most important species in medical terms in Mexico, due to the large amount of snakebites caused by them. Those two species presented the greatest number of presence records and their distribution range is extensive across the USA and Mexico, including the state of Zacatecas.Human influence was one of the most widely used variables for the construction of potential distribution models for C.pricei in Zacatecas are scarce and SDM points out regions in northeastern Zacatecas, specifically the municipalities of Concepción del Oro, El Salvador and Mazapil, as those with a high distribution potential. This may be due to the fact that these municipalities have mountain ranges whose altitudes are higher than 2,900 meters above sea level  was confirmed in the state of Zacatecas: C.aquilus, C.atrox, C.basiliscus, C.lepidus, C.molossus, C.polystictus, C.pricei, C.scutulatus and C.willardi. In addition to the confirmed records, there is available indirect information on their presence in many areas, including personal communications, unverified sightings and detailed descriptions of specimens matching the physical characteristics and ecological requirements of the species. However, it is necessary to confirm their presence in these zones.Based on the obtained records and the field work, the distribution of nine rattlesnake species (We recommend applying greater field work efforts in areas where no occurrence records have yet been identified for these species, as well as for organisms that showed the lowest number of records, particularly in the areas with a higher potential for presence. Likewise, we strongly advise conducting field work during the July-October period, since this will increase the sighting probability. We also suggest updating models once additional occurrence records have been registered.Crotalusmolossus and C.atrox, species, which were found in these areas.Results obtained from the distribution modeling of these species generally agree with field work verification, making it possible to prioritize field efforts more effectively in different localities. During field work verification, multiple night sightings of rattlesnakes were noted, both alive or run over by vehicles on the roads, being the latter one of the major causes of death of these species in the state. This piece of information was merely gathered for This paper represents substantial contribution to the knowledge on rattlesnakes in Zacatecas. Their occurrence records are shown and zones with greatest diversity within the state are inferred. Moreover, this study indicates the potential distribution areas of these organisms. This material will be of major help for the implementation of strategies on public health issues, as well as for the proposal of management and conservation plans for these species.We believe that this study could be used by individuals wishing to conduct future projects related to rattlesnake conservation in Zacatecas. We also encourage the publication and incorporation of new occurrence records that contribute to gather further information on these organisms in the state.


In the current supplement, we are proud to present seventeen relevant contributions from the 6th International Work-Conference on Bioinformatics and Biomedical Engineering (IWBBIO 2018), which was held during April 25-27, 2018 in Granada (Spain). These contributions have been chosen because of their quality and the importance of their findings. IWBBIO 2018 Conference seeks to provide a discussion forum for scientists, engineers, educators and students about the latest ideas and realizations in the foundations, theory, models and applications in the field of Bioinformatics and Biomedical Engineering. One of the main objectives of the conference is that research in the bioinformatic field can reach the medical application. The conference sought to focus on diverse fields to create multidisciplinary researches integrating areas like biomedical engineering, computer since, mathematics, artificial intelligence, bioinformatics, statistics or biomedicine [Computational proteomics. Analysis of protein-protein interactions. Protein structure modelling. Analysis of protein functionality. Quantitative proteomics and PTMs. Clinical proteomics. Protein annotation. Data mining in proteomics.Next generation sequencing and sequence analysis. De novo sequencing, re-sequencing and assembly. Expression estimation. Alternative splicing discovery. Pathway Analysis. Chip-seq and RNA-Seq analysis. Metagenomics. SNPs prediction.High performance in Bioinformatics. Parallelization for biomedical analysis. Biomedical and biological databases. Data mining and biological text processing. Large scale biomedical data integration. Biological and medical ontologies. Novel architecture and technologies  for Bioinformatics.Biomedicine. Biomedical Computing. Personalized medicine. Nanomedicine. Medical education. Collaborative medicine. Biomedical signal analysis. Biomedicine in industry and society. Electrotherapy and radiotherapy.Biomedical Engineering. EComputer-assisted surgery. Therapeutic engineering. Interactive 3D modelling. Clinical engineering. Telemedicine. Biosensors and data acquisition. Intelligent instrumentation. Patient Monitoring. Biomedical robotics. Bio-nanotechnology. Genetic engineering.Computational systems for modelling biological processes. Inference of biological networks. Machine learning in Bioinformatics. Classification for biomedical data. Microarray Data Analysis. Simulation and visualization of biological systems. Molecular evolution and phylogenetic modelling.Healthcare and diseases. Computational support for clinical decisions. Image visualization and signal analysis. Disease control and diagnosis. Genome-phenome analysis. Biomarker identification. Drug design. Computational immunology.E-Health. E-Health technology and devices. E-Health information processing. Telemedicine/E-Health application and services. Medical Image Processing. Video techniques for medical images. Integration of classical medicine and E-Health.The list of topics in the successive Call for Papers has also evolved, resulting in the following list for the present edition:Prof. Joaquin Dopazo, Fundacion Progreso y Salud, Clinical Bioinformatics Research Area, Sevilla, SpainProf. Luis Rueda, Professor, School of Computer Science, Pattern Recognition and Bioinformatics Lab, Windsor Cancer Research Group, University of Windsor.Dr. Anagha Joshi, Bioinformatics Group Leader, Developmental Biology Division, The Roslin Institute, University of Edinburgh, UKProf. FangXiang Wu, P.Eng, SMIEEE Professor, Division of Biomedical Engineering, Professor, Department of Mechanical Engineering, College of Engineering, University of Saskatchewan, 57 Campus Dr., Saskatoon, SK Canada.Prof. Jiayin Wang, Professor, Xian Jiaotong University, China.In this edition of IWBBIO, we are honored to have the following invited speaker:These plenary lectures strengthened the aim of this conference for the diffusion and the discussion of high quality researches from some of the most recognized scientists in these fields.During IWBBIO 2018 several Special Sessions will be carried out. Special Sessions will be a very useful tool in order to complement the regular program with new and emerging topics of particular interest for the participating community. Special Sessions that emphasize on multi-disciplinary and transversal aspects, as well as cutting-edge topics are especially encouraged and welcome, and in this edition of IWBBIO 2018 a total of eleven special sessions have been presentedThe IWBBIO 2018 has continued as a two-track conference, increasing the number of sessions to a total of 25 oral and 1 poster session. It received more than 210 contributions which were reviewed by at least 3 referees from our estimated program and steering committees. The conference continues accepting both full and abstract submissions for presentations. However, it still maintained a high rate of full contributions against abstracts. IWBBIO 2018 received more than 180 attendees from diverse European nationalities  but also overseas countries like United Stated, Korea, China or India.Those contributions which were considered more relevant taking into account the evaluation and opinion of reviewers and chairmen were then invited to participate in this supplement for the BMC Bioinformatics journal . In the present issue of BMC Bioinformatics journal, it is a pleasure to present you these contributions that provide a clear overview of the thematic areas covered by the IWBBIO conference, ranging from theoretical/review aspects to real-world applications of bioinformatic and biomedical engineering.The first paper authored by Xin Guan et al. , presentThe articule by Osama Hamzeh et al.  has usedThere is an important effort of the scientific community in the development of methods to predict new disease genes from protein-protein interaction networks (PPIs). However, PPIs change during the life of the cells (dynamic) and, therefore, only the use of static PPI networks can affect the performance of the algorithms. In the paper presented by Ping Luo et al. , the autIdentification of conserved interactions between proteins and ligands that are reused across a protein family it is a key factor in understanding molecular recognition processes, also facilitating tasks such as the design of effective drug. The advancement in the promotion of computational algorithm to support our understanding of the ligand-receptor recognition process is of fundamental importance in the biological processes. The contribution by Vagner S Ribeiroy et al. , presentJan Fostier  proposedIn the contribution by Yixuan Wang et al. , authorsIn the manuscript by Michael G Sadovsky et al. , a clustFollowing, the paper by Sara Nasiri et al.  address The paper authored by Renzo Angles et al.  presentsFollowing, the paper presented by Hayman Saddik et al.  suggestsThe paper by G.V. Zhikhareva et al.  presentsWenting Wu et al.  address The subject of the paper by Ayca Kirimtat et al. - exploriThe next article, by Ana Cernea et al. , is devoSimulation of microfluidic devices for the analysis of blood samples is a great tool for optimizing these devices, requiring a sufficient degree of simulation accuracy. Accuracy is ensured by measuring appropriate values which inform about the course of the simulation and can also be measured in a real experiment. The paper from Hynek Bachraty et al.  presentsIn the paper by Annarita Fanizzi et al.  an automFinally, Cristina Soguero-Ruiz et al.  address The articles presented in this special issue provides insights related to Bioinformatics and Biomedicine Engineering. As Guest editors, we would like to express our thankfulness to all the authors contributing with their high quality researches to the achievement of this supplement. Also, we are very grateful to expert scientists that have actively collaborated with their recommendations and suggestions to review and improve these contributions. We specially thank to Mr. Omar El Bakry for his excellent and constant support with the publication and edition of this supplement. It has been an honor for us to participate in it. We finally invite authors and readers of this supplement to submit their recent works to future editions of IWBBIO, which will be announced at http://iwbbio.ugr.es. We wish the readers can benefit from insights of these relevant papers, and contribute to these rapidly and dynamics growing areas.


Spartina alterniflora is an invasive plant on the coast of China that replaces native vegetation and has a serious negative impact on local ecosystems. Monitoring the spatial distribution of S. alterniflora and its changes over time can reveal its expansion mechanism, which is crucial for the management of coastal ecosystems. The purpose of this study was to map the distribution of S. alterniflora in Zhejiang Province from 1985 to 2015 using a time series of Landsat TM/OLI images and analyze the temporal and spatial patterns of expansion of this species.S. alterniflora in Zhejiang Province was extracted, and the temporal and spatial changes in the distribution of S. alterniflora were analyzed. The classification accuracy was 90.3%. S. alterniflora has expanded rapidly in recent decades after being introduced into southern Zhejiang. Between 1985 and 2015, S. alterniflora increased its area of distribution by 10,000 hm2, and it replaced native vegetation to become the most abundant halophyte in tidal flats. Overall, S. alterniflora expanded from south to north over the decades of the study, and the fastest expansion rate was 463.64 hm2/year, which occurred between 1995 and 2005. S. alterniflora was widely distributed in the tidal flats of bays and estuaries and expanded outward as sediment accumulated.After analyzing the distribution of coastal vegetation, the vegetation index was calculated based on Landsat images for 4 years . According to a threshold determined based on expert knowledge, the distribution of S. alterniflora cover in Zhejiang and can contribute to the control and management of this invasive plant.This study reveals the changes over time in  Spartina alterniflora Loisel. is a perennial halophyte that is native to the Atlantic and Gulf coasts of North America and predominates in local salt marshes [S. alterniflora is generally considered beneficial in ecological restoration because of its well-developed underground structure, high salt tolerance, high reproductive capacity and rapid growth [S. alterniflora was intentionally introduced as an ecological engineering species into China for sediment accumulation, land reclamation and saline soil amelioration [ marshes . S. alted growth . For thiioration .S. alterniflora can perform ecological functions and provide economic benefits [S. alterniflora was employed to solve river-shore crumbling near a sluice in Zhejiang Province in 1986. The planting successfully solved this problem, costing only 800 Yuan, and withstood typhoons and floods [S. alterniflora marsh on the canal bank prevented more than 100,000 m3 of sediment per year from migrating downstream [S. alterniflora is used as animal fodder and fish feed [Experiments have shown that benefits , 4–6. Cod floods . The S. wnstream  proved tish feed , 8, 9 anS. alterniflora has greatly expanded in distribution in the coastal salt marshes of China [S. alterniflora quickly expands on bare beaches and competes with native plants, invading their habitats and replacing them. Areas of Scirpus mariqueter Wang et Tang and Phragmites australis Trin. ex Steud. and young mangrove swamps have been invaded by S. alterniflora. Research shows that the species richness of benthic macroinvertebrates in S. alterniflora swamps is reduced because its invasion alters the physicochemical properties of the sediment [S. alterniflora communities is too high and the altered habitats are no longer suitable for them [S. alterniflora, and biodiversity is reduced [S. alterniflora impedes the development of local aquaculture and tourism and hinders water-based transportation [S. alterniflora was listed as one of the top 16 invasive alien species by the State Environmental Protection Administration of China in 2003 [Since its introduction, of China , 10. An of China –14. Oncesediment . Some enfor them . Therefo reduced , 18. In ortation , 20. Theortation , 21, 22. in 2003 .S. alterniflora threatens local ecosystems and causes extremely large losses to the regional economy [S. alterniflora can be found in most of China’s coastal areas [S. alterniflora has become a hot topic for ecologists and biologists at home and abroad.Biological invasion has a strong influence on Earth’s ecosystems and is considered one of the three most pressing environmental issues –26. S. a economy . At presal areas , 27. Accal areas  stated tS. alterniflora in China’s coastal areas. However, there are few studies on the monitoring of changes in S. alterniflora distribution, with most such studies limited to monitoring the distribution of S. alterniflora in a given year. The lack of long-term sequential monitoring of S. alterniflora not only limits our understanding of the invasion mechanism but also restricts the decision-making of government departments. To better understand the expansion mechanism of S. alterniflora and prevent its further invasion, it is necessary to map its distribution and monitor its dynamic changes. Therefore, this study employed a time series of Landsat Thematic Mapper (TM)/Operational Land Imager (OLI) images to monitor the dynamic changes in S. alterniflora in the coastal areas of Zhejiang Province, China. Specifically, the spatial distribution of S. alterniflora in Zhejiang Province from 1985 to 2015 was mapped, the spatial and temporal heterogeneities of S. alterniflora expansion were analyzed, and the changes in S. alterniflora in bays and estuaries were analyzed. This study documents the temporal and spatial changes in S. alterniflora invasion on the eastern coast of China, providing important data for the ecological study of S. alterniflora.Over the years, scholars have conducted many studies on the invasion of 2. The tidal flats are mainly distributed in Hangzhou Bay, Sanmen Bay and Yueqing Bay. In 1983, S. alterniflora was first planted in the tidal flats of Yuhuan County and then introduced to other places along the coast. Over the past 30 years, S. alterniflora has rapidly expanded and become the main salt marsh vegetation along the coast of Zhejiang Province.Zhejiang Province is located in eastern China Fig. a. It hashttps://glovis.usgs.gov/app). The principles of remote sensing image acquisition included (1) selecting images taken between May and November, which is the growth season of S. alterniflora and (2) selecting high-quality images without cloud cover. Table The data collected in this study included remote sensing images, vector data such as administrative boundaries, field survey data and literature data. Remote sensing is an effective tool for monitoring changes in Earth’s surface and is suitable for coastal wetland monitoring and vegetation monitoring –30. Sincincluded  selectinS. alterniflora community as determined visually in the field was marked on a Google Earth map with polygons. In addition, the positions of S. alterniflora plants were recorded using GIStar 710, a hand-held geographic positioning system device . Field surveys provide important location information for visual interpretation and classification of remote sensing images. A large number of Chinese and English research papers and statistical yearbooks were reviewed to obtain information about the distribution of S. alterniflora. In addition, detailed consultations were conducted with local people through field investigations.From May to November in 2015, field investigations were conducted in coastal areas of Zhejiang Province. The All remote sensing images were preprocessed with ENVI 5.3 software, including radiometric calibration, atmospheric correction and image-to-image registration. The Fast Line-of-Site Atmospheric Analysis of Spectral Hypercubes  method wS. alterniflora grows only in intertidal zones with high salinity and does not exist inland. In addition, there are dikes along the coast of Zhejiang Province that can prevent waves, which form an obvious artificial boundary between tidal flats and inland areas. S. alterniflora , which can express vegetation characteristics, was calculated. Analysis revealed that during the growing season, the NDVI of S. alterniflora was higher than that of S. mariqueter, which was quite low. Therefore, according to their NDVI differences and previous studies [S. alterniflora. Then, Google Earth images were used for postprocessing, unambiguously misclassified land types were modified, and the final spatial distribution of S. alterniflora was obtained. Images from 2010 and other periods were processed in the same way. According to the field survey, 300 verification points were randomly selected for verification, a confusion matrix was used to evaluate the performance of the classification, and overall accuracy was used to assess the classification accuracy.The growth environments and spectral characteristics of  studies , 29, an S. alterniflora from all images by ENVI 5.3 software, a distribution thematic map was constructed using ArcGIS 10.2 software, and the distribution changes were analyzed using the overlay function. The area of S. alterniflora within the administrative area of each city in each period was determined using ArcGIS 10.2 software. To better understand the expansion of S. alterniflora, based on the classification results of the previous step, the change in S. alterniflora was detected. In this study, the percentage (%) of the area and total area of S. alterniflora in different regions were calculated for various years. The total expansion area per decade was calculated, and the annual expansion area was defined as (Xj − Xi)/(j − i), where Xi and Xj are the areas of S. alterniflora in the prior year i and the subsequent year j, respectively.After extracting S. alterniflora in different areas along the coast of Zhejiang Province. Bays are typically surrounded by land on three sides, exhibiting a U-shape or arc shape. The bays in Zhejiang mainly include Hangzhou Bay, Yueqing Bay and Sanmen Bay  to the southernmost point (Wenzhou) along the coastline  of the Landsat TM imagery and because vegetation with an area of less than 900 m2 could not be accurately identified from the images.Table S. alterniflora had expanded to 1827.63 hm2, mainly concentrated in Wenzhou, with an area of 1096.74 hm2, accounting for 60.01% of the total area. In 2005, the total area of S. alterniflora was 6464.07 hm2, and the species was mainly distributed in Ningbo, with an area of 3706.47 hm2. By 2015, the area of S. alterniflora reached 10,038.15 hm2, and the species was mainly distributed in Ningbo and Taizhou.By 1995, the area of S. alterniflora spread widely along the coast from 1985 to 2015, with the area increasing by 10,000 hm2 and an average annual growth area of 334.61 hm2  images to extract areas of S. alterniflora with high precision. However, the cost of purchasing high-resolution images is high, and much work is needed to extract S. alterniflora areas on a regional scale.Landsat data are the most commonly used data for monitoring vegetation on a regional scale. However, the resolution of the data is relatively coarse (30 * 30 m), there are many mixed pixels, and vegetation areas less than 1000 mS. alterniflora and need to be used according to the data and actual conditions.Different classification methods also lead to certain uncertainties. It is widely believed that object-oriented methods are more accurate than pixel-based methods, but it is difficult to determine the optimal scale for object-oriented segmentation. Variables such as spectrum, texture, shape, and expert knowledge all contribute to the extraction of S. alterniflora. S. alterniflora was concentrated in intertidal zones that are periodically submerged by seawater, and its area was affected by tidal water. In Fig. S. alterniflora was almost submerged in tidal water, and (b) and (d) are images of low-tide periods, when S. alterniflora grew well and was widely distributed. Remote sensing satellites have a defined visitation cycle and cannot obtain images of the same location every day. For example, the Landsat satellite takes images of the same place every 16 days. At the same time, if the weather conditions are not good, the images covered by clouds are almost impossible to use. When the study area is large (covering multiple scenes) and multiperiod changes are of interest, it is difficult to ensure that the acquired images are from the same low-tide period.In addition, tides are an important factor in the uncertainty of mapping Spartina alterniflora rapidly expanded in tidal flats and was removed in some areas by local residents [S. alterniflora in the same year vary among studies, as do S. alterniflora mapping results in the same year. Lu et al. [S. alterniflora in China, but their results differed because of the study differences in the types of data acquired and in the study periods. It is difficult to say whose research results are more accurate because S. alterniflora has been changing dynamically and lacks reliable historical monitoring data. Future research may require more high-resolution data or the use of multiple data sources, as well as new methods such as machine learning, to more accurately monitor the invasion of S. alterniflora.esidents . Therefou et al. , Liu et u et al.  and Mao u et al.  monitoreS. alterniflora. According to the literature, we summarize the main factors affecting the expansion of S. alterniflora. First, S. alterniflora has high tolerance to salt, flooding and temperature. S. alterniflora is a typical halophyte, with a salinity suitable for growth of 1% to 2%; this species can tolerate high salinity of up to 6%. In a saline environment, S. alterniflora adopts a salt-repelling strategy; it can also secrete salt [S. alterniflora are densely covered with stomata, and the highly developed aeration tissue transports oxygen to the underground part to relieve the oxygen deficiency caused by flooding [S. alterniflora can accumulate a large amount of soluble sugars.Many factors affect the invasion of ete salt . The leaflooding . This spS. alterniflora facilitate its rapid expansion [S. alterniflora can grow axillary buds on the nodes of its stem and on underground stems, which can emerge from the soil surface and form new plants under suitable conditions [Second, this species has a high reproductive capacity. The high seed yield and high germination rate of xpansion . In addinditions . Clonal nditions , 38.S. alterniflora. There are no special growth requirements in terms of soil, and S. alterniflora can grow in clayey, loamy and silty soils. In China, silt and muddy tidal flats in coastal areas are rich in nutrients and are widespread. The tidal power, geology, climate, soil conditions and seawater salinity in these areas are quite suitable for S. alterniflora growth and reproduction. At present, there are no local, natural enemies that can control the growth and spread of S. alterniflora in the tidal flats of China. The imbalance of natural ecological competition has led to the uncontrolled proliferation of S. alterniflora. Due to the lack of natural control mechanisms, S. alterniflora can rapidly expand due to its strong growth ability [Third, the suitable habitat and lack of natural control mechanisms in this region promote  ability .S. alterniflora in this region. S. alterniflora has played an important role in reducing coastal erosion and promoting reclamation, and for this reason, its introduction was extended from Zhejiang, Jiangsu and Fujian Provinces to all coastal areas. Natural media such as tides and winds also influence the dispersal of S. alterniflora [S. alterniflora can be unintentionally transferred through a variety of human activities, such as shipping and certain forms of land transport [Finally, intentional introduction and natural media promote rniflora . In addiransport .S. alterniflora. Summer currents flow from south to north, carrying the plants and seeds of S. alterniflora northward. Once S. alterniflora reaches new tidal flats, it can quickly settle and occupy the flats, grow extensively, and invade the native vegetation community.Muddy tidal flats are widely distributed along the coast of Zhejiang Province, providing suitable growth environments for S. alterniflora has serious consequences for local ecosystems. Accurate monitoring of the invasion is essential for coastal ecological protection. In this study, we proposed a simple and effective method to map the spatial distribution of S. alterniflora in Zhejiang Province in China, using a time series of Landsat images from 1985 to 2015. The results showed that the total area of S. alterniflora in Zhejiang was approximately 10,038.15 hm2 in 2015 and that the species was mainly distributed in bays, such as Hangzhou Bay, Yueqing Bay and Sanmen Bay, and estuaries, such as the Oujiang, Aojing, and Feiyun River estuaries. In recent decades, S. alterniflora has expanded rapidly, with the highest expansion rate between 1995 and 2005, reaching 463.64 hm2/year. During the invasion from south to north, the extent of expansion varied among regions, with Ningbo exhibiting the greatest expansion. This study provides multitemporal distribution data for the study of S. alterniflora invasion in Zhejiang, revealing the temporal and spatial dynamics of the invasion process and helping the government control S. alterniflora.The rapid invasion of China’s coastal areas by


Glycyrrhiza L. (Leguminosae), is a medicinal herb with great economic importance and an intriguing intercontinental disjunct distribution in Eurasia, North Africa, the Americas, and Australia. Glycyrrhiza, along with Glycyrrhizopsis Boiss. and Meristotropis Fisch. & C.A.Mey., comprise Glycyrrhiza s.l. Here we reconstructed the phylogenetic relationships and biogeographic history in Glycyrrhiza s.l. using sequence data of whole chloroplast genomes. We found that Glycyrrhiza s.l. is sister to the tribe Wisterieae and is divided into four main clades. Clade I, corresponds to Glycyrrhizopsis and is sister to Glycyrrhiza sensu Meng. Meristotropis is embedded within Glycyrrhiza sensu Meng, and these two genera together form Clades II–IV. Based on biogeographic analyses and divergence time dating, Glycyrrhiza s.l. originated during the late Eocene and its most recent common ancestor (MRCA) was distributed in the interior of Eurasia and the circum-Mediterranean region. A vicariance event, which was possibly a response to the uplifting of the Turkish-Iranian Plateau, may have driven the divergence between Glycyrrhiza sensu Meng and Glycyrrhizopsis in the Middle Miocene. The third and fourth main uplift events of the Qinghai-Tibetan Plateau may have led to rapid evolutionary diversification within Glycyrrhiza sensu Meng. Subsequently, the MRCA of Clade II might have migrated to North America (G. lepidota) via the Bering land bridge during the early Pliocene, and reached temperate South America  by long-distance dispersal (LDD). Within Clade III, the ancestor of G. acanthocarpa arrived at southern Australia through LDD after the late Pliocene, whereas all other species (the SPEY clade) migrated to the interior of Eurasia and the Mediterranean region in the early Pleistocene. The MRCA of Clade IV was restricted in the interior of Eurasia, but its descendants have become widespread in temperate regions of the Old World Northern Hemisphere during the last million years.The liquorice genus,  Glycyrrhiza L. is commonly known as the liquorice genus . Later, taxonomists treated both Meristotropis and Glycyrrhizopsis within Glycyrrhiza (Glycyrrhiza s.l.” Previous molecular phylogenetic studies supported that Glycyrrhiza s.l. was an early branching clade of the inverted repeat-lacking clade (IRLC) of Papilionoideae  and, therefore, did not represent strong molecular evidence. Thus, the infra-generic phylogeny of Glycyrrhiza and its relationship to Glycyrrhizopsis and Meristotropis require further investigation.Despite its large economic and cultural importance, generic and species delimitation in  from 13  to 36 , through the Russian Far East, Mongolia and northern China (as well as Sichuan and Yunnan of southwestern China) (G. acanthocarpa J.M.Black), including the states of Queensland, South New Wales, Victoria, South Australia, and Western Australia   to southustralia . Only twd States  and G. af Turkey  and Centf Turkey , respectGlycyrrhiza: Eurasian-North American disjunction  genomes to (1) reveal the infra-generic relationships of Glycyrrhiza and test the phylogenetic positions of Glycyrrhizopsis and Meristotropis and (2) estimate the divergence times of lineages and reconstruct their biogeographic history. This study will provide a robust phylogenetic framework for resolving taxonomic problems in Glycyrrhiza as well as for sustainably using and developing consumable products from the genus.In the present study, we inferred the phylogeny of Glycyrrhiza s.l., including all the species, except for the rare Glycyrrhizopsis syriaca Turrill, accepted by Glycyrrhiza s.l. Outside of Glycyrrhiza s.l., within the IRLC clade, we sampled eight species representing five genera within tribe Wisterieae sensu Hylodesmum podocarpum (Candolle) H.Ohashi & R.R.Mill as the outgroup. Most sequences for the study were obtained from field-collected or herbarium specimens and were new to this study  J.Compton & Schrire, Glycyrrhiza astragalina – 1 and Wisteria floribunda (Willd.) DC. were obtained from the DNA and Tissue Bank, Kew1. In addition, we downloaded sequences of Lotus japonicus (Regel) K.Larsen and Robinia pseudoacacia L. – 2 from GenBank  Poir. were removed. We also extracted 75 protein coding sequences  from the annotated genome and concatenated them with Geneious Prime  analyses using IQ-TREE v.1.6 . The analysis with priors only is important for revealing the actual marginal distributions of priors, which may differ from those set for the analysis due to complex interactions with other calibrations and the tree process, i.e., birth-death model, herein . In the Glycyrrhiza s.l., we used the Dispersal-Extinction-Cladogenesis model , forming a clade  with Wisterieae . This large clade was sister to all other sampled IRLC.We present the phylogenetic tree with Bayesian posterior probabilities (PP) and ML bootstrap (LBS) values on the BI tree branches . The phyGlycyrrhiza s.l. (clades I–IV), all of which were highly supported with PP = 1, LBS = 100%. Clade I contained Glycyrrhizopsis flavescens  and Glycyrrhiza asymmetrica Hub.-Mor., and was sister to all other Glycyrrhiza s.l., which correspond to Glycyrrhiza sensu Meng . Within Glycyrrhiza sensu Meng, Clade II was comprised of two well supported species, Glycyrrhiza astragalina  and G. lepidota . Within Clade III, G. acanthocarpa was sister to all other species, comprising G. echinata, G. macedonica Boiss. & Orph., G. pallidiflora Maxim., G. squamulosa Franch., and G. yunnanensis Cheng f. & L.K.Tai ex P.C.Li, which formed a clade  and was regarded as the “SPEY clade” hereafter. Clade IV consisted of 13 species and included a nested clade comprising Meristotropis bucharica (Regel) Kruganova and M. kulabensis Masl. , which were sister to the rest of Clade IV . The next diverging clade within Clade IV comprises G. foetida Desf., Meristotropis triphylla Fisch. & C.A.Mey, and M. xanthioides Vassilcz  and the remaining eight species constituted the “GAU clade” .We recovered four main clades within Glycyrrhiza s.l.-Wisterieae clade  during the late Eocene, and Clade I and Glycyrrhiza sensu Meng diverged at 17.05 Ma . The split of the ancestor of Clades II and III and that of Clade IV was dated to 5.36 Ma  around the Miocene-Pliocene boundary, followed by the divergence of Clade II and Clade III at 3.53 Ma . Within Clade II, the South American Glycyrrhiza astragalina and North American G. lepidota split at 2.55 Ma . On the other hand, at around 2.82 Ma , the most recent common ancestors (MRCA) of the Australian G. acanthocarpa separated from the SPEY clade within Clade III. The crown group of Clade IV began to diverge in the late Pliocene . Subsequently, the FOT clade and the GAU clade diverged at 2.83 Ma . More recently, the FOT clade and the GAU clade started to split at 0.67 Ma  and 0.93 Ma , respectively.All the PP values on our BEAST tree are 1.0, except for the Glycyrrhiza s.l. . The MRCA of was Clade I was restricted in circum-Mediterranean region (coding: C), and that of Glycyrrhiza sensu Meng spanned from Russian Far East and northern China to the Caucasus region . Glycyrrhiza sensu Meng split into the MRCA of Clades II and III, spanning eastern Asia and North America , and that of Clade IV (Node 6), which was distributed in the interior of Eurasia (coding: B). Subsequently, the MRCA of Clade II expanded to South America , while the ancestor of Clade III remained in eastern Asia  and subsequently migrated to Australia . As for Clade IV, most of taxa were confined in the interior of Eurasia (coding: B), while a few groups, including the MRCA of the FOT clades (Node 8), and the MRCA of G. glabra and G. glandulifera Waldst. & Kit., spread back to the circum-Mediterranean region (coding: BC). Also within Clade IV, G. uralensis, dispersed to eastern Asia (coding: AB).Our DEC analysis  indicateGlycyrrhiza belongs to the IRLC  within Glycyrrhiza  occurred at 8–7 Ma, leading to a cooler climate and aridification of inland Asia  and subsboundary : the MRCboundary .Glycyrrhiza sensu Meng underwent rapid diversification and reached the New World and Australia within a short period during Pliocene. The ancestor of each of Clades II and III separated at ca. 3.53 Ma, and the crown groups of Clades II and III diverged at ca. 2.55 Ma and ca. 2.82 Ma, respectively  rapid uplift of the QTP. The uplift yielded aridification of the Asian interior and the formation of the Loess Plateau, which may have facilitated speciation of the Eurasian Glycyrrhiza sensu Meng  seems to favor an LDD hypothesis rather than one of gradual range expansion. This is especially true due to intervening habitats across the Americas that may have been unsuitable for the ancestor of this group and would have served a barrier to gene flow along with distance following an LDD, also supporting the rapid speciation event . A more common pattern was observed for SPEY clade, for which the MRCA was located in eastern Asia  and westwards to Mediterranean region (G. glabra and G. glandulifera) within the last million years  can be found in the article/LD and JW designed the research. H-FC financially supported this study. LD, EA, KE, PL, H-FC, and HH collected the samples. Z-RZ carried out the experiment. AH and CS performed the data analysis. LD, AH, and JW discussed the results and wrote the draft. All authors agreed on the contents of the final manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.