Prepectoral breast reconstruction has once again appealed, which attributes to the introduction of acellular dermal matrices (ADMs) and mesh. Postmastectomy radiation therapy (PMRT), meanwhile, is crucial in the whole course of treatment for breast cancer patients with lymph node-positive. The impact of PMRT on outcomes after prepectoral breast reconstruction has not been clearly defined to date. This study aimed to compare the impact of PMRT on outcomes after prepectoral vs. subpectoral breast reconstruction.A comprehensive research on databases including PubMed, Embase, and Cochrane libraries was performed to retrieve literature pertaining to prepectoral breast reconstruction from database inception to October 2021. All included studies evaluated the impact of PMRT on outcomes after breast reconstruction. Only studies comparing patients who underwent prepectoral breast reconstruction with a control group who underwent subpectoral breast reconstruction were included. Data were analyzed using RevMan version 5.2.A total of 4 studies were included in the meta-analysis, with a total of 394 breasts. In the setting of postmastectomy radiation therapy, 164 breasts were reconstructed with a prepectoral approach, whereas the remaining 230 breasts underwent subpectoral reconstruction. Overall, outcomes between PBR and SBR was no statistical significance in the overall complications , infection , seroma , skin flap necrosis , hematoma , wound dehiscence . But, included studies lacked data about the patient quality of life and satisfaction with the outcome of the reconstructed breast.In the setting of postmastectomy radiation therapy, prepectoral breast reconstruction is a safe and effective option. Breast cancer is the most common malignancy in women around the world, subpectoral breast reconstruction had both demonstrated low complication rates and proved successful reconstructive modalities when utilized appropriately . MeanwhiPostmastectomy radiation therapy (PMRT) is an integral part of comprehensive treatment for breast cancer, due to its proven capacity to decrease local recurrence and improve survival . FindingSince the lack of randomized and prospective studies about the impact of PMRT on outcomes after prepectoral vs. subpectoral implant-based reconstruction and the limited number involved, the outcomes after reconstruction remain controversial. And there are few meta-analyses of the results of the impact of PMRT on outcomes after prepectoral vs. subpectoral breast reconstruction.The purpose of this systematic review and meta-analysis is to evaluate the impact of PMRT on outcomes after prepectoral vs. subpectoral breast reconstruction, and further assess the safety and applicability.A systematic literature search of PubMed, EMBASE and Cochrane Library databases pertaining to prepectoral breast reconstruction was conducted in October 2021. Utilizing PRISMA guidelines, databases were searched in varying combinations of keywords “prepectoral”, “subcutaneous”, “subpectoral”, “breast reconstruction”, “prosthesis”, and “Implant”. CZ and CX independently screened the title and abstract of the article to determine whether the article meets the eligibility criteria. The inclusion criteria were as follows: (1)the article described breast reconstructions with an implant placed either prepectorally or subpectorally. (2)patients who received post-mastectomy radiation therapy. (3) patient-reported outcomes were complications after breast reconstruction. (4)the article described a randomized controlled trial, a retrospective cohort study. (5) publication was from database inception to October 2021. (6) the full text was available. The full text was independently reviewed by CZ and CX, and differences were discussed by CZ, CX, and SL to reach a consensus, and the references of selected articles were screened to determine other relevant papers. Since this is an analysis of previously published articles, participants' informed consent and ethical approval are not required.Reviews, comments, letters to the editor, animal studies, conference papers, case reports, and non-English articles were excluded.Data were independently extracted by CZ and CX, including authors, publication year, study design, number of subjects, age, and follow-up period, followed by a record of the type of reconstruction and reported complications for all patients. Filtered patients were divided into two groups: the PBR group and the SBR group in the setting of postmastectomy radiation therapy (PMRT).p < 0.05 were considered to be statistically significant. To ensure a better overall understanding we have calculated the odds ratio (OR) in every single study (PMRT to SBR vs. PMRT to PBR) as well as the weighted average of the ORs with 95% confidence intervals. All p values and 95% confidence intervals (CIs) were 2-sided. Statistical heterogeneity was evaluated using the I2 test. Statistically, significant heterogeneity was defined as an I2statistic > 40%. If heterogeneity was observed, we used a random-effects model to reduce the impact of heterogeneity on the results. If heterogeneity was not observed, a fixed-effects model was used. For each meta-analysis endpoint, a visual assessment funnel plot was used to assess potential publication bias. When p > 0.05, there was no publication bias.All of the statistical analyses were performed using RevMan version 5.2. Values of n = 394) were deemed eligible for meta-analysis  are summarized in I2 = 0%, p = 0.11)  of 164 breasts in the PBR group and 37 (16.1%) of 230 patients in the SBR group were infected. There was no significant difference between the two groups  , indicatI2 = 43% was found. Thus, the random effect model is mainly considered, which shows that 16 (9.8%) of 164 breasts in the PBR group and 22 (9.6%) of 230 patients in the SBR group were infected. There was no significant difference between the two groups   , indicatI2 = 60% was found. Thus, the random effect model is mainly considered, which shows that 12 (7.3%) of 164 breasts in the PBR group and 20 (8.7%) of 230 patients in the SBR group were infected. There was no significant difference between the two groups   , indicatI2 = 39%, p = 0.15)  of 164 breasts in the PBR group and 11 (4.9%) of 230 patients in the SBR group were infected. There was no significant difference between the two groups  , indicatI2 = 0%, p = 0.63)  of 164 breasts in the PBR group and 19 (8.3%) of 230 patients in the SBR group were infected. There was no significant difference between the two groups  , indicatNo data about the patient quality of life and satisfaction with the overall outcome and breast among patients following breast reconstruction are available.The funnel plot of total complications was shown in Mastectomy results in loss of sensation in the skin of the breast and nipple-areolar complex, and loss of the breast for cosmetic, body image, and psychosocial health . All thiBreast reconstruction in the setting of postmastectomy radiation therapy(PMRT) presents a unique challenge, because radiation therapy has an impact on the skin and soft tissues of the chest wall, leading to fibrosis that increases risks of contracture, infection, pain, necrosis, and atrophy in reconstructed tissues . ConsequTraditionally, subpectoral implant-based breast reconstruction has remained the mainstay treatment choice for the majority . This apIn recent years, with the introduction of acellular dermal matrices (ADMs) and mesh , an incrThis study showed no statistical significance in the overall complications , infection , seroma , skin flap necrosis , hematoma , wound dehiscence . Recently, Patel et al.  found thThe patient quality of life and satisfaction with the overall outcome and breast among patients following breast reconstruction also is one of the key evaluation indicators. Nonetheless, no relevant data are available in this systematic review and meta-analysis.Our study is to compare the impact of PMRT on outcomes after prepectoral vs. subpectoral breast reconstruction based on a comprehensive search. However, our study has its limitations. First, the included studies were not randomized controlled trials, but retrospective comparative studies. The inclusion of patients with potential selection bias and imbalanced baseline may affect the results of the study. Some variables, such as mesh material and implant surface, could not be accurately allocated between the two groups. Second, the difference in follow-up time may lead to an inaccurate assessment of the incidence of complications. In addition, other factors that may influence overall reconstructive success need further research in the future.For women who plan to undergo breast reconstruction in the setting of postmastectomy radiation therapy (PMRT), prepectoral breast reconstruction (PBR) is a safe and effective option.


The effects of host tree identity on epiphyte lichen communities are a controversial issue, as the results obtained in different forest environments studied are not consistent. We investigated the host preferences for lichens in the laurel forest of Macaronesia. For this purpose, we analyzed the lichen communities growing on the four most common trees  Wilbur, Laurus novoca-nariensis Rivas-Mart., Lousa, Fern. Prieto, E. Días, J.C. Costa and C. Aguiar, and Ilex canariensis Poir. in Lamarck) in the laurel forest of the Canary Islands. The diversity, richness, and lichen composition showed a repetitive and common pattern with the functional traits studied. Although the existence of specificity with respect to the phorophyte species was not demonstrated, there was a clear affinity of the epiphytic lichens to the physico-chemical features of the bark (texture and pH), canopy architecture, foliar characteristics, etc. Our results highlight the importance of the natural diversity of tree species in the laurel forest. Due to the diversity and uniqueness of the lichen species that support each of the phorophytes, this fact should be taken into account in landscape protection and restoration actions, especially in those islands where the forest is highly fragmented. The Macaronesian laurel forests, also called laurisilva and monteverde, are humid to hyper humid evergreen forests of the cloud belt of the Macaronesian islands. They are a value not only in terms of biodiversity but also biogeography . Tree spCloud forests are considered one of the most vulnerable terrestrial ecosystems to climate change . Global Lichens are one of the most remarkable organisms in monitoring and indicating environmental quality, especially atmospheric pollution ,12, but Although some fungi have a broad host range and are thus not tree specific , things Earlier studies in Macaronesian cloud forests have shown that the epiphyte–host relationships vary depending on tree identity in some groups of epiphytes, such as bryophytes ,33,34,35The present research contributes to a better understanding of this highly threatened ecosystem, where basic ecological knowledge is needed for monitoring its evolution, impacts, and even for the control and improvement of conservation measures.This study was carried out in the Canary Islands. The main areas where the laurel forests are growing and are more abundant are located in Tenerife, La Gomera, and La Palma, so we restricted the fieldwork to these islands. The other islands do not have a proper representation of this forest because: (i) the island does not have the environmental conditions required for its presence, or (ii) logging over the last five centuries has eliminated this endemic forest ,37,38.Macaronesian cloud forests comprise an environmentally complex system in which different plant associations of evergreen laurel forest can be distinguished. While macroclimatic conditions in all these stands are quite similar, there are small differences in some abiotic variables, such as the altitude, exposure, topography, fog regimes, etc., so microclimatic conditions may be different and could affect the lichen communities. To minimize the effect of these differences and to homogenize our plots as much as possible, all of them were always located in the potential habitat of the humid evergreen laurel forest , the most widespread association in the archipelago following Ref. . Figure. The besThe temperatures in these areas are mild throughout the year, with no frost or snow in winter. The mean annual temperature varies from 13 to 18 °C. Annual rainfall ranges from 500 to 1200 mm with a dry period in summer . HoweverOur field campaigns were authorized by Área de Medioambiente-Cabildo Insular de Tenerife, by Área de Medioambiente-Cabildo Insular de La Palma, and by Parque Nacional de Garajonay, with permission to have access and collect data .Morella faya, two of Erica canariensis, two of Laurus novocanariensis, and two of Ilex canariensis  > 22 cm were sampled. To study lichen diversity, we used a frame of 50 × 10 cm subdivided into 5 quadrats of 10 × 10 cm. The uppermost edge of the frame was located at 1.5 m above ground level, adjusted up to a maximum of 2 m if the trunk was unsuitable at the desired height, following Refs. ,47 with At each quadrat of 10 × 10 cm, all lichens (macro- and microlichens) were collected and subsequently identified in the laboratory. All details of the identification methods were described in Ref. . VoucherThe species were classified according to the traits: (1) photobiont, (2) growth form, and (3) reproductive strategy. Additionally, the species were grouped in functional groups (4) according to their ecological requirements  according to Refs. ,51 Tabl. Functiop ≤ 0.05) and the Mann–Whitney test adjusted by the Bonferroni procedure . The dependence relationship between the traits and lichen functional groups and laurel forest selected trees was studied with the chi-square test. To test if the ecological requirements of lichens  are similar among the trees, we used the community-weighted mean trait value CWM [The statistical procedures applied are described in depth in Ref. . We usedalue CWM . We builalue CWM , and comalue CWM  to reveaalue CWM . Finallyalue CWM , and to alue CWM .® Excel. NMDS, ANOSIM, and SIMPER analyses were analyzed and represented using Community Analysis Package 3.11  v.3.12 . The CWM11 .Parmeliaceae (32 taxa), Ramalinaceae (15), Lobariaceae (12), Lecanoraceae (11), and Pertusariaceae (10). The five most frequent species were: Parmotrema perlatum (202 samples), Chrysothrix candelaris (142), Leucodermia leucomela (117), Phlyctis agelaea (84), and Bacidia absistens (73). A total of 165 epiphytic lichen taxa (grouped in 34 families and 62 genera) were identified within more than 4000 samples collected. A complete list of all lichen species recorded is presented in Erica canariensis, (1)—13.6—(28) in Ilex canariensis, (2)—12.3—(25) in Laurus novocanariensis, and (1)—9.5—(25) in Morella faya. In total, Erica canariensis had at least 65 taxa (in 49 genera and 18 families), Ilex canariensis had 89 taxa (in 53 genera and 22 families), Laurus novocanariensis had 74 taxa (in 41 genera and 22 families), and Morella faya had 66 taxa (in 32 genera and 20 families). We did not find significant differences between the phorophyte and richness —12—(20) in richness a.Erica canariensis, 14 (8.5%) in Laurus novocanariensis, 19 (11.5%) in Morella faya, and 29 (17.6%) in Ilex canariensis. The shared taxa between Laurus novocanariensis and Ilex canariensis , and Erica canariensis and Morella faya  stand out above the shared taxa among the other possible combinations . In summary, there were more exclusive taxa (75) than taxa shared between two phorophytes (61), three phorophytes (18), and all phorophytes (11) ; however, the number of species exclusive to each of the studied phorophytes was higher: 13 taxa (7.9%) in tes (11) b.Trentepohliaceae (13.2%) (mainly in Ilex canariensis) and cyanobacteria (13.9%) (cyanolichens primarily in Laurus novocanariensis and Ilex canariensis). Epiphytic lichens in all phorophytes disperse mostly sexually (54.6%); this is more noticeable in species growing on Laurus novocanariensis and Ilex canariensis than on Morella faya and Erica canariensis. Regarding asexual strategy, dispersion by soredia (25.1%) is the most frequent propagation type, followed by isidia (11.5%) and both types combined (5.8%) . Green algae clorococcoid photobiont predominates (71.5%), followed by green algae d (5.8%) .Morella faya and Erica canariensis share pH substrate categories 2 (“on acid substrate”) and 3 , whereas the epiphytic lichens of Laurus novocanariensis and Ilex canariensis are mainly associated with substrate pH level 3  and “sites with plenty of diffuse light but scarce direct solar irradiation” (category 3), followed by “sun-exposed sites, but avoiding extreme solar irradiation” (category 4), but some species present in Morella faya are typical of “sites with very high direct solar irradiation” (category 5). A similar response is observed in the epiphytic lichens of Ilex canariensis, with most lichens in categories 2 and 3, followed by category 4. The epiphytic composition of Laurus novocanariensis is mainly grouped in category 3, followed by category 4. Finally, Erica canariensis composition differs from that of other phorophytes, with lichen species typical of illuminated sites being more abundant  (very xerophytic” environment (category 5). The lichen composition of Morella faya, Laurus novocanariensis, and Ilex canariensis is shared between categories 2 (“intermediate between hygrophytic and mesophytic stands”) and 3 (“mesophytic”), and the composition of Erica canariensis is shared among categories 3 and 4 , followed, to a lesser extent, by category 2 (very high eutrophication” (category 5). The lichen composition of Laurus novocanariensis and Ilex canariensis has a similar response to eutrophication—typical of “very weak eutrophication” stand (category 2), followed by “no eutrophication” site (category 1) and “weak eutrophication” environment (category 3). The lichen composition of Morella faya is divided between category 1 (50% of the composition) and categories 2 and 3 (the other 50% of the composition). Finally, the lichen composition of Erica canariensis has a bimodal structure, with lichens typical of no eutrophication places (category 1) and species typical of “rather high eutrophication” sites (category 4) , 1 , and 2 (“in moderately disturbed areas”). The lichen composition of Ilex canariensis is typical of poleotolerance categories 1 and 2, followed by category 0. The epiphytic lichens of Morella faya are linked to category 1, and, to a lesser extent, to category 0. Finally, the lichen composition of Erica canariensis has a bimodal structure, with maxima in categories 1 and 3 (“species occurring in heavily disturbed areas”)  b. We didtegory 2 c. We didegory 4) d. With r areas”) e.Laurus novocanariensis vs. Erica canariensis, Laurus novocanariensis vs. Morella faya, Erica canariensis vs. Ilex canariensis, and Morella faya vs. Ilex canariensis), with good separation between Erica canariensis and Ilex canariensis , according to Ref [Laurus novocanariensis vs. Ilex canariensis and Erica canariensis vs. Morella faya)  .Erica canariensis vs. Morella faya (87%) and Ilex canariensis vs. Laurus novocanariensis (88%)  and the lowest among is (88%) .The SIMPER analysis revealed discriminant species. Not all species contributed equally to the differences between the phorophytes. A list of characteristic species, with percentages of contribution in brackets, of each phorophyte studied is shown in Laurus novocanariensis and Ilex canariensis , between Erica canariensis and Morella faya (two species: Chrysothrix candelaris and Platismatia glauca), and between Ilex canariensis and Morella faya (only Thelotrema lepadinum). On the other hand, two discriminant species are common to Laurus novocanariensis, Erica canariensis, and Morella faya: Parmotrema perlatum and Leucodermia leucomela. No discriminant species is common to all the studied phorophytes.Some discriminant species , all from La Palma Island, are on the right side of the NMDS plot. Finally, two samples of Laurus novocanariensis, one of them from Tenerife (plot 6) and another from La Gomera (plot 13), are on the left of the NMDS plot . The endemic element (restricted to the Macaronesian region) is not very numerous, but it is represented by species such as Lobaria macaronesica) [Most of the 10 most frequent monteverde species have a wide European distribution  ,64,65,66onesica) ,67,68,692 in Erica and Morella, 20.2/m2 in Laurus, and 24.7/m2 in Ilex. Some authors have described that trees with different bark characteristics have different epiphyte species. For this reason, a forest environment in which phorophytes with different barks coexist will accumulate a higher diversity of epiphytes than monospecific forest formations [The species richness found in the Canarian Monteverde  varies slightly among the phorophytes, with values of 18.3/mrmations ,70,71. 2 [2, 83 species of the 165 observed). It is very relevant to highlight the importance of the contribution of microlichens to the total specific richness.Although there are no previous studies in the Canarian Monteverde that allow us to compare our data with those studies, we were able to do so with the richness of epiphytic macrolichens identified in a study of the Madeiran monteverde, with a value of 1.4 epiphytic macrolichens/m2 . This vaLaurus novocanariensis, Morella faya, and Erica canariensis) in Madeira [Laurus was the species tree that hosted the most epiphytic species, which agrees with our work in this sense. On the other hand, Ref [2 for Laurus in the Monteverde of Tenerife. This value is much lower than the lichen diversity that we observed on this same phorophyte. This same pattern was observed by Ref [Studying bryophytes on different phorophytes , as well as the diversity of the genera and families present in the four phorophytes studied, there seems to be a disjunction in the Canarian Monteverde between the lichens present in analysis . Our resFunctional traits have been used as bioindicators of forest attributes of interest for forest characterization and management ,80,81. HMorella + Erica and Laurus + Ilex) are due to intrinsic characteristics, such as: tree diameter and age, presence of latex and other epiphytes, chemical composition, pH, stability, roughness, thickness and water-holding capacity of the bark, number and size of lenticels, and cortical runoff [Morella and Erica as two trees with acid pH, low stemflow funneling ratio values, small leaf area [Ilex and Laurus as two trees with neutral pH, high stemflow funneling ratio values, larger leaves [The study areas were established in homogeneous (meso)climatic zones. Thus, it is expected that the differences found between the two groups of phorophytes described  and Laurus, since both trees have a smooth bark compared to Morella and Erica, which have a rough bark. On the other hand, foliaceous species with large thalli, mainly from the Parmeliaceae family, show a high biodiversity in Erica and Morella, where they are more frequent, as well as in Laurus. The dichotomy between Erica + Morella and Laurus + Ilex, previously mentioned, is further emphasized when studying the dimorphic biotype, represented only by the genus Cladonia. It is found exclusively on Erica and Morella. Cladonia has generally been associated with acid substrates, where it predominates [Cladonia exclusively in Erica (pH = 5.2 ± 0.7) and Morella (pH = 4.5 ± 1.2) and its absence in Ilex (pH = 6.4 ± 0.4) and Laurus (pH = 6.3 ± 0.6) [Cladonia species identified are mostly found in heathlands and acid substrates, both in the monteverde and in other forest environments [Erica and Morella, there is also a high frequency of Chrysothrix candelaris, a leprose lichen that has been related to acid substrates, rough bark, and environments with high luminosity, both in the Monteverde and in other forest environments [In terms of the most frequent and diverse taxa found in the studied phorophytes, we observed that crustaceous lichens are very common in  authors ,31. Howe authors . As obse authors ,31, rougominates ,98. This3 ± 0.6) , althoug3 ± 0.6) . The Claronments ,101,102.ronments ,69. Howeronments , which dronments  and has Sticta canariensis cyanomorphotype, Crocodia aurata, Sticta, and Leptogium) and tripartite lichens  in Ilex and Laurus. However, no cyanolichens were found among the most frequent species in Erica or Morella. No representative of this group was found in Morella. Although there seem to be exceptions [Ilex, 28.5 for Laurus, 22.8 for Erica, and 12.7 for Morella) support our results. The higher richness of lichens with Trentepohliaceae on Ilex compared to the rest of the studied phorophytes could be explained by the high number of crustaceous species that develop on this tree. All lichens with this photobiont have this biotype. Finally, the predominance of lichens with chlorococoid green algae found in the monteverde is simply a consequence of their higher abundance in nature [When studying the most frequent species, as well as the most diverse genera and families, we only found cyanolichens  co-dominate. Considering that the trees in a given study plot are within a few meters of each other, the lower abundance of sorediate lichens on Ilex and Laurus compared to Morella and Erica cannot be explained solely based on the higher difficulty of dispersal of these propagules compared to spores. It has been shown that soredia can reach more than 200 m [Regarding reproductive strategies, sexual reproduction predominates in the monteverde, followed by multiplication by soredia, isidia, a conjunction of both (soredia and isidia), and fragmentation of the thallus. This pattern has also been described in other forest environments ,106,107.an 200 m . Therefoan 200 m  concludean 200 m ,111,112.The presence and abundance of some epiphytic lichens in forests are related both to the characteristics of the forest itself and to the ecological requirements of the lichen species . Thus, iEpiphytic lichens are influenced by macro-, meso-, and microclimatic factors, as well as by the physical and chemical features of the substrate where they grow. Because of these requirements, several authors have used lichens as bioindicators of both polluted and well-preserved environments ,114. MosLaurus and Ilex have an affinity for subacid to subneutral substrates, and these trees are the only ones with lichens species with an affinity for slightly basic substrates. However, the epiphytic lichens growing only on Morella and Erica are associated with acidic and subacid to subneutral substrates, with no exclusive lichen species having an affinity for (slightly) basic substrates. This is consistent with the previous comments on the pH characteristics of the four phorophytes and the occurrence of particular groups of lichens associated with this character. Due to the architecture and structure of the studied tree species, the morphology and size of leaves, the characteristics and nature of the bark affecting the water-holding capacity, runoff, etc., a differential response of lichens to solar radiation, aridity, and even pH would be expected. Regarding pH, it should be noted that lichens found exclusively on Erica, which has substantially different leaves. In the case of Ilex, Laurus, and Morella, we expected to obtain a significant proportion of poorly light-tolerant lichens, typical of shady environments, such as evergreen forests like the Canarian Monteverde  phase of the forest [In terms of the response of lichens to solar irradiation, we expected to obtain a certain homogeneity due to the similar characteristics of the canopy of the different localities and the fact that all are evergreen and broadleaf trees, with the exception of nsu Ref. ). We didnteverde . Howevere forest .Erica are by far the smallest and narrowest of the four trees analyzed [Morella, although some authors considered them small compared to those of Laurus or Ilex [Erica. This would explain the higher abundance of lichens related to shady environments with respect to Erica but less in relation to Laurus or Ilex. On the other hand, the leaves of analyzed ,96. Ther or Ilex , are mucErica epiphytes tend to tolerate a higher degree of aridity than expected , as it is a forest linked to high humidity . Erica eexpected c. This cnot high , the watIlex and Laurus are in accordance with those expected, with a high proportion of lichens occurring exclusively on old trees in ancient, undisturbed forests or natural  habitats. Consequently, they are mostly non-resistant to eutrophication or resistant to a very weak eutrophication  on various substrates [Morella and Erica forest [For the eutrophication and poleotolerance, low values were expected for the four phorophytes analyzed, as it is a primary forest. The values for hication d,e. Morebstrates , but it a forest , and in a forest . The divThe low proportion of shared lichens among all the studied phorophytes compared to the diversity found exclusively in each phorophyte, suggests a possible lichen–phorophyte specificity. The specificity of epiphytic lichens and their phorophytes is a controversial issue, and it does not seem that the same patterns occur in all forest ecosystems. Thus, while some authors report some specificity ,29, otheErica and Morella on the one hand, and between Laurus and Ilex on the other, suggests a possible affinity of lichens for the trees that comprise these two subsets. In this case, this pattern seems to be consistent with what has been reported by other authors in tropical forests. Several authors have found similar patterns, i.e., affinity of lichen species for groups of phorophytes, and have explained them based on the physical and chemical features of the trees and their bark  [Laurus novocanariensis + Ilex canariensis and Morella faya + Erica canariensis) are the same as those observed by Refs. [The high species richness shared between r, etc.) ,85,125. by Refs. ,70 when According to Ref. , the speErica + Morella and Laurus + Ilex. This differentiation is clear, considering the specific diversity and the number of lichen species shared by these two groups. These data are also supported by the patterns of functional traits of lichens that comprise these two groups. The differences found may respond to the physico-chemical features of the tree . Further studies would be necessary to elucidate the importance of each of these factors separately. The results presented will be useful to improve the ecological restoration efforts in the Canarian Monteverde.No specificity of Canarian Monteverde lichens concerning the tree species analyzed was observed. However, an affinity toward two well-defined groups of phorophytes was identified:


The automated classification of brain tumors plays an important role in supporting radiologists in decision making. Recently, vision transformer (ViT)-based deep neural network architectures have gained attention in the computer vision research domain owing to the tremendous success of transformer models in natural language processing. Hence, in this study, the ability of an ensemble of standard ViT models for the diagnosis of brain tumors from T1-weighted (T1w) magnetic resonance imaging (MRI) is investigated. Pretrained and finetuned ViT models  on ImageNet were adopted for the classification task. A brain tumor dataset from figshare, consisting of 3064 T1w contrast-enhanced (CE) MRI slices with meningiomas, gliomas, and pituitary tumors, was used for the cross-validation and testing of the ensemble ViT model’s ability to perform a three-class classification task. The best individual model was L/32, with an overall test accuracy of 98.2% at 384 × 384 resolution. The ensemble of all four ViT models demonstrated an overall testing accuracy of 98.7% at the same resolution, outperforming individual model’s ability at both resolutions and their ensembling at 224 × 224 resolution. In conclusion, an ensemble of ViT models could be deployed for the computer-aided diagnosis of brain tumors based on T1w CE MRI, leading to radiologist relief. Brain tumors (BTs) are characterized by the abnormal growth of neural and glial cells. BTs cause several medical conditions, including the loss of sensation, hearing and vision problems, headaches, nausea, and seizures ,2. ThereSeveral previous studies have attempted to automatically classify brain tumors using MRI images, starting with traditional machine learning classifiers, such as support vector machines (SVMs), k-nearest-neighbor (kNN), and Random Forest, from hand-crafted features of MRI slices ,10,11,12Lately, attention-based transformer networks  have becTherefore, to fully exploit the power of ViTs, a large amount of data is required, and it may not be possible in medical imaging domains to collect such an amount of data. To overcome this, in , severalThis section describes the dataset, the ViT architecture, the computational infrastructure for model training, hyperparameter tuning using the validation set, and testing. The ViT model ensembling and the performance metrics employed are also discussed.An openly available dataset from figshare consists of 3064 T1w CE MRI slices from 233 patients with meningiomas, gliomas, or pituitary tumors. The images are available in all sagittal, coronal, and axial directions, with spatial resolutions of either 512 × 512 or 256 × 256. More details about the dataset are available in ,41. A feMeningiomas: Meningiomas are mostly benign tumors originating from the arachnoid cap cells and often occur in older-age individuals and females. These tumors account for 13–26% of all intracranial tumors .Gliomas: Gliomas are the most frequent and primary intracranial tumors that are malignant. They represent 81% of all intracranial tumors that can cause significant mortality and morbidity .Pituitary Tumors: Pituitary tumors originate in the pituitary gland and are mostly benign. Since this gland regulates different hormones, tumors present in it may cause severe changes in the body. These tumors contribute to 10–15% of all intracranial tumors .The number of images for each tumor category and the number of images used for training, validation, and testing in a 70:10:20 ratio are described in H: height, W: width, C: number of channels). Afterward, linear embeddings are computed for these flattened image patches, and position embeddings are added to them to keep the patch positional information  head. Furthermore, these combined patches and position embeddings are fed to the transformer encoder model, which has alternating layers of multi-headed self-attention and MLP blocks .In this work, pretrained and finetuned ViT base (B) and large (L) models, B/16, L/16, B/32, and L/32 (16 and 32 indicate square patch size), on ImageNet-21k and ImageNet-1k datasets were used. Hence, the MRI images were resized to the resolutions of 224 × 224 and 384 × 384. Since these pretrained ViT models require three channels in the input and since the MRI slice has a single channel, the same grayscale MRI image is copied into the other two channels.N, which is a learnable linear projection; and l. The second block also starts with an LN layer, followed by an MLP and a residual connection with output Similar to [class] in BERT , a learnentation  during bMSA output in the transformer encoder is obtained from the concatenation of several self-attention heads within it. Mathematically, self-attention is described in Equation (5), where Q, K, and V are the query, key, and value matrices obtained after matrix multiplications with Q matrix is obtained as The MSA  are dot-wise-added and finally divided by the number of individual models (N) to obtain the final output (esoftmax) of the ensemble classifier. Two ensembling procedures are evaluated, where the first one is the ensemble of all models at 224 × 224 resolution, and the second ensemble is the combining of all individual models at 384 × 384 resolution.To evaluate the ensemble model for class prediction, the procedures described in Equations (7) and (8) are followed. The softmax outputs of each model (RMSprop/Adam/Adadelta), the learning rate (lr), the number of epochs (ne), and the mini-batch size (mbs). The optimization of the hyperparameters is conducted using the validation set. To calculate the performance metrics on the test set, the hyperparameters that gave the best accuracy values during the 5-fold cross-validation are considered.Since a multi-class classification task is carried out, sparse categorical cross-entropy is used as the loss metric, and sparse categorical accuracy is used as the performance metric during training and validation. The confusion matrix and overall sparse categorical accuracy are used as model evaluation metrics during testing. In addition, overall sensitivity and specificity calculated as means of per-class sensitivities and specificities respectively are also used as performance metrics for the ensemble models. The tuned model’s hyperparameters are the optimizer (Adam optimizer with lr = 0.0001, ne = 25, and mbs = 16. The B/16 model performed the best at this resolution, with a validation accuracy of 97.83%. Regarding the remainder of the models, their performances at different hyperparameter combinations are given in Initially, the image intensities were rescaled to produce values between -1 and 1, which is a requirement for ViT models. During training, all parameters of the ViT models were allowed to be finetuned. For the input image resolution of 224 × 224, the optimized hyperparameters with respect to the validation accuracy were the Adadelta with lr = 0.1, ne = 10, and mbs = 8. Adadelta was solely the best optimizer at this resolution. The optimized hyperparameters and validation accuracies of all other models, B/16, B/32, L/16, and L/32, were 98.10%, 98.04%, and 98.55%. Due to computational constraints, training at 384 resolution was implemented with lower mbs values.Similarly, at 384 × 384 resolution, the optimized hyperparameters for the best validation accuracy of 98.64% from the L/16 model were The test accuracy values for both the input image resolutions of 224 × 224 and 384 × 384 for all ViT models are given in The performance of the ViT models on the test set in the form of confusion matrices is given in lr, ne, and mbs, which indicates that the ViT models are robust across different hyperparameter settings; however, the Adadelta optimizer outperformed all other optimizers at 384 × 384 resolution. Nevertheless, to evaluate the performance of the models on the test set, the models that yielded the highest accuracy values on the validation set were considered, which is the standard procedure. Individual model’s performances on both the validation and test sets were slightly better at the image resolution of 384 × 384 compared to 224 × 224, which could be because the general performance of ViT models is better at higher resolutions, as evaluated by the experiments in [In this study, the ability of pretrained and finetuned ViT models is investigated both individually and in an ensemble manner for a three-class classification of brain tumors, namely, meningiomas, gliomas, and pituitary tumors, from T1w CE MRI. In general, all ViT models demonstrated the ability to classify with validation and test accuracies above 97% during most scenarios refer to . Based oments in . SimilarComparing the performances of the ViT models in this study with previous studies based on the same dataset given in During training, all the model parameters starting from the patch embedding layer were allowed to be finetuned because, based on a few experiments conducted by freezing the initial layers, including some transformer encoder block layers of the ViT models, the validation and test accuracies were around a couple of percentage points lower than the accuracy values obtained by unfreezing all model parameters. Even though the model’s performance improved at 384 × 384 resolution, training at this resolution was computationally demanding and, hence, implemented in a TPU environment. Furthermore, the performance of the ViTs at the original input image resolution of 512 × 512 may be better, and this hypothesis could be investigated in a high-level computing environment. Furthermore, the cross-validated models from this study can be finetuned for use with other brain tumor datasets. In addition, in a future study, it could be interesting to investigate the ability of other vision transformer variants, such as swin vision transformers , data-efThe performance of the ensemble model at 384 × 384 resolution is on par or better than that of previous CNN models for the classification of brain tumors from MRI, achieving an overall test accuracy of 98.7% and a specificity of 99.4%. Using the same ensemble model, the test classification accuracy for gliomas is 100%. The developed framework is made available publicly here. Therefore, the computer-aided diagnosis of brain tumors from T1w CE MRI using an ensemble of finetuned ViT models can be an alternative to manual diagnoses, thereby reducing the burden on clinical radiologists.


Nephrotic syndrome (NS) in the first year of life is called congenital (CNS) if diagnosed between 0–3 months, or infantile (INS) if diagnosed between 3–12 months of age. The aim of this study was to determine if there were clinically meaningful differences between CNS and INS patients, regarding clinical presentation, management and outcomes.Eleven Pediatric Nephrology Research Consortium sites participated in the study, using IRB-approved retrospective chart reviews of CNS and INS patients born between 1998 and 2019. Data were collected on patient characteristics, pertinent laboratory tests, provided therapy, timing of unilateral/bilateral nephrectomy and initiation of renal replacement therapy (RRT).vs. INS patients , while weekly infusions were more common in INS patients . During the 6 months preceding RRT, albumin infusions were more frequently prescribed in CNS vs. INS . Nephrectomy was performed more often in CNS (78%) than in INS . End-stage kidney disease tended to be more common in children with CNS (80%) vs. INS .The study included 69 patients, 49 with CNS and 20 with INS, with a median age at diagnosis of 1 and 6 months, respectively. Management for the two groups was similar regarding nutrition, thyroxin supplementation, immunoglobulin administration, and thrombosis prophylaxis. Within the first 2 months after diagnosis, daily albumin infusions were used more often in CNS Compared to INS, patients with CNS had a more severe disease course, requiring more frequent albumin infusions, and earlier nephrectomy and RRT. Despite center-specific variations in patient care, 20–40% of these patients did not require nephrectomy or RRT. Nephrotic syndrome (NS) is the most common glomerular disease in children, but its occurrence in the first year of life is rare. It is defined as congenital nephrotic syndrome (CNS) if the disease onset is in the first 3 months of life, or infantile nephrotic syndrome (INS) if it occurs between 3 and 12 months of life. CNS, if left untreated, is a fatal disease . ManagemTreatment may include the use of anti-proteinuric agents such as renin-angiotensin-aldosterone system (RAAS) inhibitors  and nonsDue to the consequences of urinary losses of specific proteins, many patients receive specific interventions, including L-thyroxine supplementation , 9, mediMany patients eventually require bilateral nephrectomy followed by dialysis until transplantation , 12 in oThe aims of our study were: 1) to describe and compare the clinical presentation of children with CNS and INS, and 2) to investigate practice patterns for their management across North America.Eleven sites, members of the Pediatric Nephrology Research Consortium (PNRC), participated in a retrospective chart review of patients born between 1998 and 2019. IRB approval was obtained at each participating site. Study inclusion criteria were a diagnosis of CNS or INS. Infectious etiologies  or immune-mediated nephritides, with or without hypocomplementemia, were excluded.A secure, 21.CFR compliant tool was created in Qualtrics for data entry. The data collected included age at diagnosis; serum creatinine at various time points, as eGFR is not standardized for children in the first year of life ; supportU test or Welch's t-test for continuous variables and by the chi-square or Fisher exact test for categorical variables, as appropriate. Counts and frequencies of missing values of the two groups were displayed. No imputation was made for missing data. Statistical analysis was performed with Prism version 9.0 statistical software. A 2-sided p < 0.05 indicates statistical significance. Kaplan-Meier estimates were generated, and a log-rank test was performed to calculate the probability of requiring nephrectomy (Nx) or RRT during the 2 years following diagnosis of CNS or INS.Patient demographics and clinical characteristics were reported as counts and proportions for categorical variables and medians with interquartile ranges (IQRs) or means with standard deviations for continuous variables, as appropriate. Differences between patients' characteristics and management with CNS and INS were determined by the Mann-Whitney The study included data from 69 children, 49 with CNS  and 20 with INS .p = 0.04. Negative results were noted in two patients, and an additional one result was inconclusive (94% positivity rate). Two-thirds of the patients had documented edema, half were noted as having anasarca. Patients with INS had a higher serum albumin concentration at the time of diagnosis compared to CNS, with no difference in serum creatinine (The median age at diagnosis was 1 month for CNS (IQR 1–1) and 6 months for INS (IQR 5–7.8). Genetic testing was performed in 50 (72%) patients, 39 with CNS, and 11 with INS, eatinine .About a third of all patients received high energy  and high protein (3–4 g/kg/day) diets. This intervention was more common in children with CNS .p = 0.006), whereas weekly infusions were reported in only one infant with CNS vs. 5 with INS (p = 0.001). The significant difference in albumin infusion requirements was maintained over the course of the disease, being higher in those with CNS than in those with INS in the 6 months preceding RRT, 25 with CNS vs. 3 with INS (p = 0.007).Albumin infusions were prescribed during the first 6 months following diagnosis in 64% of all patients, 34 with CNS and 10 with INS. Of note, in the first 2 months after establishing the diagnosis, albumin infusions were prescribed daily in 27 patients with CNS and only in 3 patients with INS .RAAS inhibition was prescribed in 54 patients (78%), 38 with CNS, and 16 with INS. Of those with CNS, 22 received albumin infusions during the 6 months preceding RRT, compared to 3 with INS (χn = 5) and epistaxis (n = 1).Indomethacin, the sole NSAID used, was prescribed in 19 patients, 16 with CNS, of which 15 also received RAAS inhibitors, and 3 with INS, who also received RAAS inhibition. Indomethacin was discontinued in 6 patients (32%) due to adverse effects: AKI (p = 0.124). Four patients with CNS (29%) receiving IVIG/SCIG experienced sepsis, compared to none in the INS group , including 14 with CNS and 2 with INS . Similar finding was observed in patients with INS who received thrombosis prophylaxis vs. those who did not . Also, for patients who received thrombosis prophylaxis, the thrombotic events were comparable in patients with CNS vs. those with INS .There were 12 thrombotic events reported in 7 patients with CNS, and 7 events in 3 patients with INS. Locations included venous and arterial vessels and the right atrium . Among tvs. 10; p = 0.02). Nephrectomy was unilateral, with no RRT at the time of reporting in 2 (4%), unilateral/sequential, preceding RRT, in 7 (15%), and bilateral in 39 (81%), most often after dialysis, or at the initiation of dialysis (Nephrectomy was performed in 48 (70%) patients, with a higher rate in infants with CNS compared to those with INS  in those with bilateral nephrectomy (p = 0.02)  received RAAS inhibitors and one  received indomethacin for anti-proteinuric effect. The median serum albumin in those with unilateral/sequential nephrectomy was 1.1 g/dL (range 0.5–1.4 g/dL)  = 0.02) . Histopan = 1), stable CKD (n = 7), or unknown reasons (n = 9) (Twenty-one patients 30%) did not undergo nephrectomy due to remission of proteinuria , patient demise ( (n = 9) . The ret0% did no (n = 9) . Most pavs. 55%; p = 0.09), and pre-emptive kidney transplantation in 4 children (3 with CNS and 1 with INS)  , 36 with CNS and 11 with INS (73 ith INS) . Eight o = 0.04) .p = 0.58).Dialysis was initiated at a median age of 16 months (range 1–109) for the CNS group and 18 months for the INS group (range 3–59) followed by transplantation in 42 patients  . The timvs. 1 with INS, at a median age of 70.5 months (IQR 51–85).Pre-emptive kidney transplantation was performed in 3 patients with CNS p = 0.09) . RAAS inhibition was recorded in 8 of the 10 patients (80%) with CNS vs. 6 of the 8 patients (75%) with INS.In patients with unilateral/sequential nephrectomy, 89% (8/9) required albumin infusions during at least the first 2 months following diagnosis, compared to only 50% (8/16) of those patients who did not undergo nephrectomy or RRT ( = 0.09) . Of the  = 0.09) . In compThe outcome at 12 months  was not vs. non-genetic etiologies of NS in the first year of life require a more precise diagnosis.The age-based definitions of CNS and INS have been used for decades to help guide the diagnosis, management and predict clinical outcomes in patients with NS in the first year of life. More recently, however, this rationale is being questioned because of increasing knowledge about the genetic basis of early-onset NS including the fact that a particular gene defect can present as CNS or INS . AlthougSeveral management approaches have been recommended. These include albumin infusions, NSAIDs and/or RAAS inhibitors, nephrectomy, and RRT , 12. In Our study revealed a considerable variation in the management of CNS and INS among North American centers, and even among patients at the same center. The rarity of the disease does not help consolidate single-center clinical experience or study the management outcomes, difficult tasks for all rare disorders, requiring personalized approach to care. Along with the variations in clinical presentation, physicians' personal beliefs and preferences play a role in management, in the absence of any evidence-based guidelines. Facing the same challenges, the European Reference Network for Kidney Diseases (ERKNet) and the European Society for Pediatric Nephrology (ESPN) Working Group recently published their expert opinion-based consensus guidelines for the management of CNS .Our study, which is the largest such study to date in North America, revealed that at the time of diagnosis, edema was present in two-thirds of patients, at similar rates in CNS and INS. The question of why the remainder of the patients with congenital nephrotic syndrome did not have clinically evident edema in the setting of low serum albumin, is not completely understood, although interstitial fibrosis with tubular atrophy along with low eGFR could have been responsible for the lack of edema in some patients , 7. Our Since the only available data are the number of patients who received RAAS inhibitors and NSAIDS, known as anti-proteinuric agents, and the protein/creatinine ratio is less reliable at this age because of low urine creatinine, we could not evaluate the impact of these medications on proteinuria in this special population.We also found that 50% of those with CNS required albumin infusions during the 6 months preceding RRT, compared to 15% of patients with INS, although there was no significant difference in the use of NSAIDs and RAAS inhibitors, or unilateral nephrectomy, between the groups. Although it is unclear if this approach postponed nephrectomy or renal replacement therapy, or led to a complication-free course, patients with INS who received RAAS inhibitors were less likely to receive albumin infusions during the 6 months prior to RRT when compared to those with CNS.Historically, the lack of minimal response to steroid therapy in this patient population has shifted the treatment paradigm toward symptomatic relief with emphasis on optimal nutrition and growth, until the moment RRT is needed. In our study, we have identified only 2 children with INS who were prescribed steroid therapy, because genetic testing was not available at that time. The kidney biopsy, performed as indicated by the lack of response after the conventional timeframe, identified minimal change disease in one and mesangial hypercellularity with sclerosis in the other, and was reassuring that both achieved remission. As the availability of genetic testing is increasing, steroid therapy or other immunosuppressive drugs will likely be prescribed in those cases with no identified gene mutations, who on biopsy will be found to have minimal change or focal segmental glomerulosclerosis.Although well justified, supplementation with high caloric diet was received by only a third of the patients in our study. We believe that a dietitian should be part of a complex management team for these patients, and more attention should be paid to CNS and INS infants' nutrition to improve their growth.With the goal to diminish proteinuria, NSAIDs and RAAS inhibitors were widely used by many centers. In our study, RAAS inhibitors were prescribed alone or in addition to indomethacin to prevent inadequate growth and development from the massive urine protein losses. Indomethacin had to be stopped in a third of those receiving it, due to side effects. The addition of anti-proteinuric agents has helped a significant number of these children to require less often albumin infusions in the 6 months preceding RRT, more so in INS than in CNS. However, due to the various lengths of follow-up and timing to RRT, we cannot determine the exact impact of these agents on the renal survival in this cohort. Nonetheless, based on our data, we support the initiation of therapy with low-dose anti-proteinuric agents, titrating to effect, monitoring for side effects, and not exceeding the maximal dose.NPHS1 , with a median follow-up of 1.1 years, one study reported 63 verified, and 62 suspected, episodes of sepsis , central line infections (48 and 47%), or sepsis (54 and 53%) between 17 patients managed conservatively and 25 patients with bilateral nephrectomy, 2.8 years after diagnosis  level, attributed mostly to the urinary loss of thyroid-binding globulin and elevated thyroid-stimulating hormone (TSH), with improving levels following thyroxine replacement , 23. Rouvs. bilateral nephrectomy vs. early renal replacement therapy in these rare genetic disorders.In 2019, the ESPN Dialysis Working Group published their findings on 80 children with CNS , 25. TheNPHS1 mutations  the voluntary participation does not allow us to draw the conclusion that the cohort is representative of nephrotic syndrome in the first year of life in the North American population, as there are centers caring for patients such as these, not members of PNRC, and member-centers of PRNC who did not participate in this study; b) its retrospective design does not allow us to determine outcomes related to certain therapies; c) the few missing data prevented us from reaching firm conclusions in some areas. At the same time, our study has some strengths: a) to date, it is the largest cohort of nephrotic syndrome in the first year of life in North America; b) provides data supporting the majority of the prior expert recommendations.Based on the data described, we are proposing the following diagnostic and management guidelines:Genetic testing should become a diagnostic standard for suspected CNS and INS.A high-energy diet  is an essential component of the management of CNS and INS, and a dietitian should be part of the treatment team.Albumin infusions should be based on clinical indicators, not serum albumin concentration.Therapy with RAAS inhibitors and NSAIDs should be initiated at a low dose and titrate to effect with close monitoring of side effects, not exceeding the maximal dose.The need for prophylaxis of thromboembolic events should be made on a case-by-case basis, mostly after thrombotic events or other thrombophilia indications.Thyroid replacement therapy is justifiable in the complex management of CNS and INS patients.Prevention of infection is an important part of the treatment approach for CNS and INS patients. There is a lack of strong data supporting or denying IV or SC immunoglobulin infusions, and this therapy should be decided individually.WT1 mutation are appropriate candidates for early bilateral nephrectomy.Patients with Future studies should include the creation of a prospective worldwide registry to include the diagnosis, management, complications, and long-term outcomes of infants with CNS and INS to better enable pediatric nephrologists to develop robust evidence-based approaches to improve the care of these critically ill infants.The original contributions presented in the study are included in the article/The studies involving human participants were reviewed and approved by institution-specific IRBs. Written informed consent from the participants' legal guardian/next of kin was not required to participate in this study in accordance with the national legislation and the institutional requirements.AC, TV, TM, LG, and WS: conceptualization. AC, NH, MM-L, EB, EP, MR, BW-R, and MK: data curation. AC: formal analysis and writing-original draft. TV, AC, and JN: methodology. WS and TV: supervision. TM, WS, JN, YK, and TV: validation. YK: visualization. SW, AA, AT, MR, RM, LG, KT, TV, TM, and WS: writing-review and editing. All authors contributed to the article and approved the submitted version.Texas Tech University Health Sciences Center Collaborative Seed Grant provided support for TV.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.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Despite the global reduction in under-five mortality, still many children die before their fifth birthday in Ethiopia. The main aim of this study was to identify determinants of under-five mortality using count data regression models based on 2019 Ethiopia mini demography and health survey data.The data source for this study was the 2019 Ethiopia mini demography and health survey data. Various count data regression models were applied to identify the determinants of under-five mortality.A total of 5,535 mothers with children aged 0–59 months were included in the study. Of the total, 1,277 (23.07%) women had lost at least one child by death before celebrating fifth birthday. Zero-Inflated Poisson model was found to be the best model, and it revealed that mother's age, marital status, mother's age at 1st birth, place of delivery, current contraceptive type used, type of cooking fuel, residence, region, religion, time to get drinking water, number of children at home, birth order, and birth type were significant factors that determine U5 mortality in Ethiopia.Mothers aged 15–24 years  and above 24 years  at their 1st births, mothers from rural area , mothers traveled for 1–30 min  and >30 min  to get drinking water, mothers used charcoal  and wood , children with birth order of 2nd-3rd , 4th -5th , 5th and above , and multiple born children  had higher risk of under-five mortality while mothers aged 25–34 years , unmarried mothers , mothers delivered in public health sectors , mothers used Pill/IUD , mothers who had 3 to 5  and more than 5  children at home had lower risk of under-five mortality.Mothers should be encouraged to deliver at health institutions. Mothers also should be given awareness to use Pill/IUD contraceptive type. Moreover, facilitating rural areas to get electricity and drinking water near to homes helps to reduce the burden of U5M and to be in line with sustainable development goal. •Our study was an attempt to identify the risk factors for under-five mortality in Ethiopia.•We used 2019 Ethiopian Mini Demographic and health Survey data.•Analyses were done using Statistical sofwares R, Stata, and SPSS.•Count Data Regression Models were employed to identify the risk factors of under-five mortality.•The best model that fits the data well was selected using selection criterion like AIC, BIC.•Zero-Inflated Poisson model was found to fit the data well.•The study found that mother's age, marital status of mother, age of mother's at first birth, place of delivery, place of residence, time to get drinking water, number of children at home, birth order, type of birth were statistically significant determinants for under-five mortality in Ethiopia. The deaths of under-five children have fallen from 12.5 million in 1990 to 5.3 million in 2018 [In 2019, nearly half (49%) of all U5 deaths occurred in just five countries: Nigeria, India, Pakistan, the Democratic Republic of the Congo and Ethiopia. In 2019, sub-Saharan Africa had an average U5M rate of 76 deaths per 1,000 live births. That is equivalent to 1 child in 13 dying before reaching age 5. This rate is 20 times higher than that of 1 in 264 in the region of Australia and New Zealand [In Ethiopia, as the 2019 EMDHS report shows the trends of U5M declined from 123 deaths per 1,000 live births in 2005 to 59 deaths per 1,000 live births in 2019, a 52% decrease . Even ifUnderstanding determinants of under-five mortality is essential to inform public health policies and design strategies to accelerate the reduction of under-five mortality ,6. In Et22.1The study was a retrospective design study, and the data source was the 2019 Ethiopia mini demography and health survey data. The census frame was a complete list of 149,093 enumeration areas (EAs), among them, 35,292 are in urban areas and 113,801 in rural areas. In the first stage, 305 EAs  were selected with probability proportional to EAs size and the household listing was carried out in each EA. In the second stage, 30 households per cluster were selected with equal probability selection [2.2A total of 8,663 households were successfully interviewed with a response rate of 99%. In the interviewed households, 9,012 women aged 15–49 were identified for individual interviews [2.3The outcome variable was total number of children who died under the age of 5 per woman in her lifetime measured as count 0, 1, 2, … The predictor variables in this study were mother's age, mother's education level, mother's literateness, marital status, religion, mother's age at 1st birth, place of delivery, current contraceptive type, residence, region, number of women in the home, source of water, toilet facility, time to get drinking water, age of household head, wealth index, number of household members, relationship to household head, type of cooking fuel, number of U5 children, number of children at home, birth order, birth type, and child's sex.2.4The under-five mortality data experienced excess zeros characterized by over-dispersion and heteroscedasticity. The most popular distribution for modeling such data is zero-inflated model and hurdle models. The over-dispersion has been explained as heterogeneity that has not been accounted for unobserved population which consists of several sub-populations in this case of Poisson type, but the sub-population membership is not observed in the sample. This excess variation may be occurred incorrect inference about parameter estimates, standard errors, tests, and confidence intervals. The Negative binomial model addresses the issue of over-dispersion by including a dispersion parameter to accommodate the unobserved heterogeneity in the count data. However, it cannot address the over-dispersion caused by an excessive number of zeros, in such case zero-inflated and Hurdle models are appropriate. Zero-inflated models mix a count component and a point mass at zero, allowing for over-dispersion ,12.The likelihood-ratio test is used to test the null hypothesis of no over-dispersion  against the alternative hypothesis the over-dispersion parameter is different from zero . Furthermore, log likelihood, MSE, MAE, AIC and BIC were used to compare various candidate models, and the model with the smallest AIC and BIC value was considered as a better fit . The dat33.1A total of 5,535 women were included of which 1,277 (23.07%) women had lost at least one child by death before celebrating fifth birthday whereas the remaining 4,258 (76.93%) of the mothers had not lost their U5 children by death. This indicates zero outcomes were large in number. The histograms are highly picked at the beginning . However, large number of under-five deaths per mother were observed less frequently. Additional screening of number of child deaths showed that the variance (0.57) was greater than the mean (0.35) indicating over-dispersion .Table 1N3.2At the point when the significant wellspring of over-dispersion is a dominance of zero tallies, the subsequent over-dispersion cannot be modeled precisely with the negative binomial regression model. An elective path for demonstrating this kind of data is the zero-inflated Poisson or zero-inflated negative binomial regression model which considers the excess of zeroes. And, the overall models comparison was presented in 3.3The rate of non-zero U5 death for children born to 25–34 years old mother decreased by 27%  as compared to 15–24 years old mother keeping other variables held constant in the model. Concerning mother's marital status, the risk of U5M for unmarried mothers was 0.68  times lower among U5 children born to married mothers. With regard to mother's religion, the risk of U5M for children whose mothers who follow Muslim, protestant and other religions were 1.35 , 1.18  and 1.69  times higher as compared to children whose mothers who follow Orthodox religion, respectively. Similarly, the rates of U5 death for children whose mothers were 15–24 and above 24 years old increased by 24%  and 66%  as compared to mothers younger than 15 years, respectively. Regarding place of delivery, mother who delivered in public health sector had 0.59  times lower risk U5 death rate as compared to mother who delivered in home. Similarly, the incidence rate of U5 death for mothers who used Pill/IUD was decreased by 36%  as compared to mothers who did not use any contraceptive type. In the zero-inflated part, the estimated odds that the number of U5 death becomes zero for mothers who follow Muslim and protestant decreased by 87%  and 45%  as compared to mothers who follow Orthodox religion, respectively .Table 3Z3.4The risk of U5M among children of mothers from rural area was increased by 27%  compared to urban area mothers. Mothers who living in Addis Ababa city decreases the incidence of U5M by 62%  compared to those living in Tigray region. The risks of U5M were increased by 78%, 64%, 51% and 35% for mothers living in Afar, Somali, Oromia and Benishangul-Gumuz regions compared to those living in Tigray region, respectively. The incidence rates of U5 death were increased by 62%  and 82%  for mothers who traveled for ≤30 min and >30 min to get drinking water as compared to mothers who get drinking water near to their homes (or took 0 min), respectively. Likewise, the incidence rates of U5 death were increased by 83%, 73% and 57% for children of mothers whose household heads ages were 25–34, 35–44, and above 44 years compared to 15–24 years old household head, respectively. The richest mothers had lower U5M rate  compared to poor mothers. Mothers from more than six household members had lower U5Mrate  compared to mothers from 1 to 3 household members. Regarding mothers’ cooking fuel types, incidence of U5M rates were increased by 86%, 64% and 114% for children of mothers who used charcoal, wood and other cooking fuel types compared to children of mothers used electricity fuel type, respectively .Table 4Z3.5The incidence of U5M rate was decreased by 63%  for more than three the number of 5 and under children in the household compared to only one child in the household. Likewise, the incidences of U5M rates were decreased respectively by 49%  and 73%  for three to five, and above five number of children in the homes compared to less than or equal to two home. The death rates of U5 for children with birth orders of 2nd–3rd, 4th–5th, and above 5th were 3.91, 13.14, and 38.17 times higher compared to 1st birth order child, respectively. On the other hand, multiple birth type of child was 50% increased risk of U5M  compared to a single birth type. In the zero-inflated part, the estimated odds that the number of zero U5 death for two children (5 and under) in the household was 4.09 time higher  compared to only one child in the household .Table 5Z4In this study 5,535 mothers with children aged 0–59 months were included, of which 1,277 23.07%) women had lost at least one child by death before celebrating fifth birthday. The study revealed that mother aged 25–34 years had reduced risk of under-five mortality as compared to 15–24 years old mother. This shows that a younger aged mother face higher U5 child mortality risk. This finding is consistent with other studies conducted in Ethiopia , Kenya , Nigeria.07% womeThe study also revealed that the incidence rate of under-five mortality among children whose mothers' ages at first birth were 15–24 years and older than 24 years were significantly more than among children whose mother's age at first birth was less than 15 years. This finding agrees with result of  while itAs presented in this finding, unmarried women had lower risk of under-five deaths than married counterpart. This finding is consistent with study conducted in sub-Saharan Africa countries . HoweverThis study found that children of Muslim, protestant and other religion followed mothers were having higher risk of dying before the age of five years compared to children whose mothers followed orthodox religion. This might be partly due to the fact that Muslim women tend to face oppositions regarding the use of contraceptive methods from their husbands ,31.Findings from this study also revealed that place of delivery is another significant determinant factor of under-five mortality. Children born in a healthcare facility that is in the public or private sectors were at lower risk than those born at home. This might be due to the proper health care and attention these facilities provided to them during and after delivery, and this finding which is confirmed by other studies ,29,31,32Findings of this study indicated that lower mortality rate of children dying before age of five was associated with mothers using contraceptive type. Thus, U5 mortality among children from mothers used pill/IUD contraceptive type was significantly less than children from mothers not used any contraceptive type. This result agrees with previous findings ,15,29.The incidence of U5 death was higher among children of mothers living in rural area than those living in urban area. Several researchers found similar results showing children from rural area had higher mortality rate than urban area . The posRegion was also found to be significant factor determining under-five mortality. Tigray region had lower risk of under-five mortality compared to other regions except Addis Ababa city administration. This finding is in agreement with other studies which found that region is determinant factor of infant and under-five mortality in Ethiopia ,9,27,32,Wealth index was another important determinant factor of under-five mortality. The richest women had significant reduction in under-five mortality compared to the poorest women. This result agrees with various findings ,40,42,43This study also found that the incidence rate of U5M increased with increased birth order of the child. This is consistent with other previous studies ,46,47. PFurthermore, the study found that U5M was significantly determined by the age of household head, number of household members or family size, time to get the source of drinking water, type of cooking fuel, number of children at home, and birth type. Higher U5M rate was associated with older household head. This finding is consistent with the study . The hig5This study was based on nationally representative data with a large sample size. Moreover, since it is based on the national survey data, the study result has the potential to give insight for ministry of health, policy-makers, and other concerned bodies to design appropriate intervention strategies both at national and regional levels. However, this study had limitations in that the EMDHS is mostly based on respondents' self-report and might have the possibility of recall bias. In addition, some variables like weight of child at birth ,53,54, m6This study aimed to identify risk factors of under-five mortality using count data regression model based on 2019 Ethiopia mini demography and health survey data. The ZIP regression model was found to be the best and revealed that mother's age, mother's age at 1st birth, marital status, place of delivery, current contraceptive type used, type of cooking fuel, residence, region, religion, time to get drinking water, number of children at home, birth order, and birth type were significant factors that determine U5 mortality. Moreover, mothers traveled for long hours to obtain drinking water, mothers from Afar, Somali, Oromia and Benishangul regions, mothers from rural area, mothers delivered in homes, mothers used charcoal and wood cooking fuels, children of 2nd and above birth orders, and multiple born children were associated with high incidence of U5M. Thus, Mothers should be encouraged to deliver at health institutions. Mothers also should be given awareness to use Pill/IUD contraceptive type. Moreover, facilitating rural areas to obtain electricity and drinking water near to homes helps to reduce the burden of U5M and to be in line with sustainable development goal.The authors are authorized to download Survey data from the Demographic and Health Surveys (DHS) Program. The data is publicly available and has no personal identifiers.No one funded this research.Corresponding author (ASA) had designed the manuscript, written the whole manuscript parts, edited and analyzed the data. Author GGM coded, edited and analyzed the data, edited and revised the manuscript. Finally, both authors read and approved the final manuscript.We used publicly available secondary data in this study and has no personal identifiers.Name of the registry:Unique Identifying number or registration ID:Hyperlink to your specific registration (must be publicly accessible and will be checked):alex.siffir@gmail.com) and Gizachew Gobebo Mekebo (gizmake@gmail.com)Alemayehu Siffir Argawu (Email:All relevant data and material are available to any interested researchers upon reasonable request from corresponding author.Both authors declare that they no conflict of interest.Not Applicable.Both authors have no conflict of interests.


Salmonella enterica serovar Typhimurium (ST) is a food-borne pathogen that can infect animals and humans. It is currently the most common bacterial pathogen that negatively affects the poultry industry. Although different chicken breeds have been observed to exhibit diverse resistance to ST infection, the underlying genetic mechanisms remain unclear and the genes involved in this differential disease resistance need to be identified. To overcome this knowledge gap, we used a liver transcriptome analysis to screen differentially expressed genes (DEGs) in two different chicken breeds  and commercial Guang Ming No. 2 broiler line B (GM)) before and after ST infection. We also performed weighted gene co-expression network analysis (WGCNA) to detect hub genes, and employed selection signal analysis of candidate genes. Three promising genes  were eventually identified, and were significantly and differentially expressed in the same breed under different conditions, and in the two breeds after ST infection. Hub genes, such as PPFIA4 and ZNF395, were identified using WGCNA, and were associated with the ratio of heterophils to lymphocytes (H/L), an indicator of disease resistance. the present study identified several genes and pathways associated with resistance to ST infection, and found that BY had greater resistance to ST infection than GM. The results obtained provide valuable resources for investigating the mechanisms of resistance to ST infection in different chicken breeds. Salmonella infection.As an important global poultry resource, chicken meat and eggs provide high dietary protein for humans ,2. ChinaSalmonella spp. are Gram-negative bacteria that cause gastroenteritis and enteric fever [Salmonella disease can lead to death of a large number of chickens, reduce egg production and hatching rate of laying hens, and increase the probability of egg contamination [Salmonella spp. are commonly detected in chicken farms, among which ST, a group of nonadaptive or pan-tropic Salmonella spp., has a wide range of hosts [Salmonella-infected food products can lead to gastroenteritis in humans [Salmonella spp. has important public safety significance in reducing Salmonella infection.ic fever , which amination . At presof hosts . As one of hosts , ST can n humans . TherefoMany studies have shown that widely reared livestock and poultry generally exhibit interspecies differences in resistance to various infectious diseases . In the Ethical approval on animal care and experimental procedures was obtained from the Animal Ethics Committee of the Institute of Animal Sciences, Chinese Academy of Agricultural Sciences .The two chicken breeds, Guang Ming No. 2 broiler line B  and Beijing You , exhibit completely different genetic backgrounds. All Chickens were obtained from the Changping Experimental Base of the Institute of Animal Sciences . The two breeds were raised under the same conditions for 22 days and then transferred to two isolation chambers, each consisting of 15 BY and 15 GM broilers. The chickens were divided into four groups: Beijing You chickens ST infection group (BY_ST), Beijing You chickens control group (BY_CTL), Guang Ming broilers ST infection group (GM_ST) and Guang Ming broilers control group (GM_CTL). After the chickens were raised for 28 days, ST was used to infect BY_ST and GM_ST. One day after the infection, we weighed the chickens and took liver tissues and blood for follow-up experiments.Salmonella enterica serovar Typhimurium 21484 standard strain (CICC), and the half-lethal dose (LD50) of the strains was 2.5 × 1010 colony-forming units (CFU)/mL/chicken [The ST strain used in this study was /chicken  administThe bacterial cells were cultured in Luria Bertani (LB) broth in an orbital shaker at 37 C and 150 revolutions per minute (rpm) for overnight culture, recovered, and then incubated again for 12 h.To calculate the H/L ratio, we extracted the peripheral blood of BY and GM chickens one day after ST infection, placed them in anticoagulant tubes, and prepared blood smears. The blood smears were dried and stained with Wright-Giemsa stain, and the number of heterophils (H), lymphocytes (L) and monocytes (M) were recorded using a light microscope under 100× magnification. The total number of H, L and M was controlled at about 100 [® Spectrophotometer , while the integrity and concentration of the RNA were determined using the Bioanalyzer 2100 system’s RNA Nano 6000 Assay Kit . Fragmentation buffer was added to the purified mRNA to generate short fragments, and the first strand of cDNA was synthesized using six-base random primers with fragmented mRNA as template; subsequently, buffer, dNTPs, RNaseH and DNA Polymerase I were added to synthesize the second strand of cDNA, which was purified by the QIAQuick PCR kit and eluted with EB buffer. The eluted and purified double-stranded cDNA was then subjected to end repair, addition of base A, and sequencing junction, and recovered by agarose gel electrophoresis for target size fragment and PCR amplification, thus completing the whole library preparation.RNA was extracted from the liver tissues collected from all 60 chickens. Liver samples were aseptically collected using sterile scissors and tweezers, stored in a cryovial tube, snap-frozen in liquid nitrogen and stored at −80 °C for later RNA extraction. Then, total RNA was isolated using the QIAGEN RNeasy Kit and genomic DNA was removed by using the TIANGEN DNase KIT . The purity of the RNA was determined using the kaiaoK5500After library construction, the initial quantification was performed using the Qubit 3.0 to dilute the library to 1 ng/µL, and the insert size of the library was tested using the Agilent 2100. Subsequently, the effective concentration of the library (>10 nM) was accurately quantified using a Bio-RAD  CFX 96 fluorescence quantitative PCR instrument and Bio-RAD KIT iQ SYBR GRN Q-PCR to ensure the quality of the library. The libraries with expected quality were sequenced using the Illumina  platform with PE150 sequencing strategy.Q-value < 0.05.Gene expression was normalized according to the fragments per kilobase of exon model per million mapped reads (FPKM) value of each gene, followed by DEGs analysis using DESeq2 software (Version 18.2.0), with |log2(FoldChange)| > 1 and WGCNA is a method to analyze the gene expression pattern of multiple samples, which can cluster and form modules of identically expressed genes . With spBy measuring the changes in the H/L ratio (an indicator of disease resistance) and three inflammatory factors  before and after ST infection, the resistance of the two chicken breeds to ST infection was determined. The results showed that the H/L ratio of the ST infection groups was higher than that of the control groups, irrespective of the chicken breed; however, BY_ST chickens showed a significantly higher increase in H/L ratio A.With regard to alterations in the three inflammatory factors, both IFN-γ and IL-1β concentrations in the serum of GM chickens presented a significant increase after ST infection. In contrast, only the IFN-γ concentrations differed in the control groups, with GM exhibiting significantly lower IFN-γ concentration than BY B,C. MeanQ-value < 0.05, 563 DEGs were identified between the BY_ST and BY_CTL groups, including 220 upregulated and 343 downregulated genes | > 1 and ed genes A, and 32ed genes B.FOS gene was simultaneously enriched in the four signaling pathways and although not significantly enriched, these pathways were significantly associated with immunity, including the Toll-like receptor signaling pathway, Salmonella infection, MAPK signaling pathway and apoptosis [DEGs were compared between the BY and GM groups, and 293 genes were significantly differentially expressed in the GM groups but not in the BY groups C. Subseqpoptosis   and KEGG pathways analyses were employed. The GO bar chart showed that some processes, such as cell part, cellular process and binding, were predominant in both BY and GM groups A,B, suggY groups C, whereaM groups D.The optimal soft thresholding was set at 18 and 7 for BY and GM groups, respectively A, becausPPFIA4, MKX, DYTN, FSIP1, SNCA, TSHR and NIM1K, were obtained for BY (ADORA1 (adenosine A1 receptor), MCHR2, ALAD, ELF2, PRDM4, SLC22A23, ADAMTS6, AR, ZNF395, ATAD2, KLF8, TXNRD3 and BCAR1, were obtained for GM , along with JUN and FOS genes . The count values for these three genes were ascertained , thus reducing the concentration of ST in the organism, so that there is less ST stimulation and insignificant changes in inflammatory factor concentrations. These results indicated that GM is more sensitive to ST infection, and that BY might be more resistant to ST infection than GM.In this study, we compared the H/L ratio, inflammatory factors, and liver transcriptomic data of two chicken breeds before and after ST infection to determine the breed with stronger disease resistance and screen related genes. Phenotypic changes before and after ST infection were observed by examining the H/L ratio and serum concentrations of three inflammatory factors. The H/L ratio, a recognized indicator of stress resistance ,16, cleant to ST ,20. IL-1nt to ST ,22. ST int to ST . In the This study has certain limitations, and it cannot directly prove the strength of disease resistance in the two breeds due to a lack of data on the bacterial load. However, in this study, we compared the H/L ratio, inflammatory factors and liver transcriptomic data to indirectly verify the breed with stronger disease resistance and screen related genes. In addition, according to the measurement of the bacterial load in ST-infected chickens of two breeds in the early stage of the project, it was shown that BY showed a significantly lower liver bacterial count than CB chickens (This breed has similar disease resistance to GM), implying that BY chickens were more effective in eliminating ST, further confirming the findings of this paper .Salmonella infection, were highly correlated with inflammatory response genes, including FOS, LIFR and other genes. In BY, DEGs, including JUN, GADD45G, TUBA1A, TNFSF10 and CASP14, were enriched in the apoptosis pathway. As many pathogens evade the defense system by inhibiting apoptosis (an innate defense mechanism), it can be presumed that these genes are associated with immunity in BY.Transcriptome sequencing is widely used to obtain information on all mRNA sequences in a particular tissue or organ in a given state . In thisFOS gene family was screened by KEGG pathway enrichment analysis. The FOS gene family consists of four members , which encode leucine zipper proteins that dimerize with proteins in the JUN family, thereby forming the transcription factor complex AP-1. Thus, FOS proteins are believed to regulate cell proliferation, differentiation and transformation. In some cases, FOS expression is also associated with the death of apoptotic cells. It has been reported that FOS plays an important role in avian influenza virus infection. Therefore, FOS was validated as a candidate gene.In GM, only 293 genes were significantly enriched, and the PPFIA4 is a prognostic monitoring indicator of thyroid cancer, lipoma and epilepsy, and DYTN is associated with Leiber visual atrophy, dystonia and lymphatic malformations. A total of 18 hub genes, including ADORA1, ZNF395 and RAB33A, were detected in GM. ADORA1 is involved in promoting tumor growth through bone marrow-derived suppressor cells and has been reported to support tumor growth outcomes in colorectal adenocarcinoma, human leukemia Jurkat cells, breast cancer and kidney cancer. ZNF395 is an activator of a subset of IFN-stimulated genes [RAB33A is a T-cell regulatory molecule associated with tuberculosis that has been suggested to be involved in disease processes [To examine the underlying mechanisms of disease resistance in BY and GM chickens, WGCNA was performed. A total of seven hub genes were identified in BY chickens, among which ed genes , and RABrocesses . As thesEGR1, JUN and FOS, were screened, which have been reported to have a major role in the resistance of chickens to avian influenza [EGR1, as an important transcription factor, plays a crucial role in cell survival and death as well as in the inflammatory response process [FOS is also involved in the inflammatory response to mammalian infection [JUN, encodes the transcription factor complex AP-1, which is, thus, involved in regulating cell proliferation and differentiation, as well as in activating the transcription of pro-inflammatory genes [EGR1 and JUN were significantly elevated after ST infection in BY, but were not significantly increased in GM, whereas FOS, a DEG significantly expressed in GM, was considerably elevated only in ST-infected GM. Moreover, expressions of all three genes were significantly different between the two chicken breeds, implying that these genes were associated with ST infection. Furthermore, selection signature analysis of EGR1, JUN, and FOS genes in BY and GM revealed segregation in the three gene regions, indicating that both chicken breeds were selected in all the three gene regions during the evolutionary process, and that this selection may be related to the differences in the disease resistance of the two breeds. Thus, future follow-up studies must focus on EGR1, JUN and FOS genes and identify effective genes for breeding disease-resistant poultry.After the analysis of DEGs and hub genes, a total of three genes, namely nfluenza . EGR1, a process ,30. FOS nfection ,32 and, ry genes ,34,35. ISalmonella infection pathway and apoptosis, as well as many immune genes, were found to be enriched, and all of these pathways and genes are known to play important roles in the resistance to ST infection [FOS and JUN, which constitute a pathway with the TLR4 receptor, were screened in the present study and are significantly associated with ST infection [Many studies have confirmed that different chicken breeds have diverse levels of resistance to ST infection, which may be related to the differential expression of some genes, and accordingly, many immunity-related genes have been successfully screened. In the present study, we found that local chicken breeds (such as BY) are essentially more resistant to diseases than commercial chicken breeds (such as GM). In a publication by Li et al., a comparison of the disease resistance of three chicken breeds  showed that the local breed was more resistant to disease than the commercial breed, which is consistent with the findings of this study . It has nfection . The effnfection , and canEGR1, FOS and JUN, were chosen for selection signal analysis. The results demonstrated that both BY and GM underwent selection during evolution, and that BY had higher gene polymorphism and was less exposed to selection. These findings provide a theoretical basis for future breeding of disease-resistant BY and GM.In the present study, analysis of the H/L ratio and serum concentrations of the three inflammatory factors  in two chicken breeds (BY and GM) revealed that BY has a stronger ability to resist ST infection when compared with GM. To confirm this finding and investigate the underlying genes and pathways that are responsible for resistance to ST infection, comparative transcriptome analysis and WGCNA were performed, and three genes,