Candida auris is an emerging fungus that presents a serious global health threat. It is often multidrug-resistant and difficult to identify with standard laboratory methods. C. auris outbreaks have been associated with high mortality in healthcare settings. Rapid and accurate screening for C. auris is crucial for preventing transmission and directing treatment. This study evaluated a new fully automated, lab-developed real-time PCR assay for the detection of C. auris.C. auris detection using the Open Access™ functionality on the Panther Fusion® instrument. It provided fully automated nucleic acid extraction, amplification, and detection workflow. The primers and probe are published sequences designed by CDC which target the internal transcribed spacer 2 (ITS2) region of the C. auris ribosomal RNA gene. Analytical studies evaluated assay sensitivity, specificity, and inclusivity. An internal control is also included to ensure assay validity.Nasal swabs collected with Aptima Multitest swabs were used as a representative clinical matrix to evaluate Candida auris real-time PCR assay is 5 CFU/mL for nasal swabs. Eleven C. auris strains from CDC clinical isolates were reactive for inclusivity study. No cross reactivity or interference was observed from 60 microorganisms of closely related yeast, bacteria and viruses typically present in skin and groin samples.The limit of detection (≥ 95% detection rate) of the C. auris with internal control, utilizing primers and probe sequences published by CDC, demonstrated excellent sensitivity, specificity, and inclusivity. This lab developed test (LDT) provides a rapid, sensitive, and scalable method for detection of C. auris. The results are crucial for timely detection, transmission prevention and targeted treatment of the multidrug-resistant C. auris.A fully automated and high throughput real-time PCR test for All Authors: No reported disclosures.


Toxoplasma gondii oocysts are widespread, and their contamination can be primarily attributed to the members of the Felidae family. This study aimed to estimate the prevalence and determine the dense granule antigen 6 (GRA6) genotype of T. gondii among domestic cats in the Phayao Province, Thailand.Globally, toxoplasmosis is an important zoonotic parasite infection of many warm-blooded animals (including humans). T. gondii DNA using targeted B1 gene polymerase chain reaction (PCR) amplification, and positive samples were subsequently analyzed for their T. gondii genotype through PCR-restriction fragment length polymorphism (RFLP) analysis and sequencing of the GRA6 gene.A total of 124 fecal samples were collected from owned cats in the Muang district, Phayao, Thailand, from January to December 2020. Fecal samples were tested for the presence of T. gondii. Only 10 positive DNA samples were successfully amplified for the GRA6 marker. Subsequent PCR-RFLP and sequence analyses indicated that all T. gondii isolates from cats in Phayao belonged to GRA6 genotype I.Among the 124 samples, 46 (37.1%) were tested positive for T. gondii. Our results further suggested that infective oocysts of T. gondii are widely distributed and that environmental contamination with these oocysts will introduce more risks of disease transmission to humans and other animals.Data revealed that toxoplasmosis is remarkably distributed among (studied) domestic cats in Phayao, Thailand. Moreover, the virulent GRA6 allele was found to be circulated among domestic cats in this area. However, no significant correlation was observed between infection rates and different risk factors, which indicated that pet cats of any age, gender, or breed have similar risks of being infected with  Toxoplasma gondii, is a common zoonotic parasite infection associated with pet cat ownership  was included as an outgroup. The final tree was visualized by FigTree 1.4.4 .DNA sequences of GRA6 PCR products were assembled and processed using BioEdit version 7.2.5  . Online Bcutter2/. T. gonde.ac.at/ . The GRAT. gondii infection was determined as the proportion of infected cats to the total number of examined cats. A Chi-square test was performed to assess the correlation between the prevalence of infection and variable risk factors. Differences were considered to be statistically significant when p ≤ 0.05.The prevalence of Of the 124 cats examined, 62 were female and 62 were male. Cats were aged between 2 months and 4 years (with an average age of 11.8 months). They were further allocated into two age groups: <1 year (83) and ≥1 year (41). Based on cat breeds, there were 45 local (Thais), 42 exotic , and 37 crossbred.T. gondii infection among the 124 cat fecal samples. A 115 bp PCR product was amplified for 46 fecal samples . In addition, the prevalence of toxoplasmosis was slightly higher (41.0%) in young cats aged under 1 year than in older cats (29.3%) aged over 1 year. Up to 43.5% of female cats were found to be positive for T. gondii infection, whereas the latter only reached 30.7% in male cats. Regarding cat breeds, a higher percentage of infection was recorded for local (44.4%) followed by crossbred (35.1%) and exotics (31.0%). Regarding T. gondii infection, no significant association could be found among age groups, genders, or cat breeds. The prevalence of T. gondii infection based on each parameter is summarized in The overall rate of All PCR-positive samples (n = 46) were subsequently analyzed using GRA6 genotyping. Amplified fragments with an expected size of 344 bp were obtained through nested PCR amplification. Only 10 samples successfully generated GRA6-RFLP profiles. All samples exhibited a type-I genotype-specific pattern displaying two bands of 258 and 86 bp . No unusT. gondii sequences showed great similarity to 344 bp GRA6 gene region sequences of T. gondii, which were available on GenBank and ToxoDB databases. Among the 10 GRA6-positive samples, three sequences were identical. Such sequences exhibited the highest similarity (with 100% homology) to that of T. gondii RH-88. Sequence and blast analyses of other positive samples further revealed 99.13–99.71% homology to T. gondii RH-88 (genotype I) compared with the 98.26–99.13% homology to T. gondii ME49 (genotype II) and T. gondii VEG (genotype III).Sequence analysis of GRA6-positive samples (with 98.55–100% sequence homology) was performed on a 344 bp amplicon. For all isolates, the obtained MseI sequence digestion revealed one MseI recognition site (TTAA) at nucleotide position 258/259 within the 344 bp T. gondii GRA6 sequence, which resulted in two fragments (258 and 86 bp). The MseI-RFLP profile for T. gondii GRA6 sequences was consistent with the results obtained in the PCR-RFLP assay.Analysis of T. gondii isolates and each clonal lineage based on their representatives  [et al. [T. gondii oocyst was found in cat feces in Malaysia using modified Kato–Katz and Sheather’s sugar methods. Toxoplasma oocysts were identified by morphological characteristics, but the researchers did not include the molecular identification of T. gondii.The present study revealed a high prevalence of oocyst shedding in cats, in which 37.1% 46/124) of Phayao cats have d (4.7%)  and neigd (4.7%)  and Indo of Phaya [et al.  recordedT. gondii oocysts are similar, which are often misdiagnosed with another coccidian [T. gondii DNA.Microscopic examination of fecal oocysts is a low-cost and rapid method for direct examination. It also has limited sensitivity. This method also requires laboratory experience in identifying parasite oocysts as occidian . Thus, toccidian . In the T. gondii oocysts are not frequently found during routine stool examinations with only 1% of infected cats reportedly excreting oocysts in a given period [T. gondii-infected cats in this area could contribute to oocyst burden in the environment, although infected cats only excrete oocysts early on after infection [Toxoplasma prevalence area can be exposed to oocysts and may be reinfected, resulting in the re-excretion of more oocysts.In general, n period , 28. Basnfection , 29. Nevnfection , and thenfection . These cnfection , 32–34. T. gondii oocyst shedding among cats, the lack of statistical significance regarding age groups, breeds, and gender indicated that these factors were insignificant factors for T. gondii infections in cats. In addition, the role of age in the epidemiology of feline toxoplasmosis among cats in many regions has been determined, showing that the seroprevalence of T. gondii increased with age [T. gondii oocyst excretion in cats [T. gondii oocysts.In the present study of with age –37. Howe in cats , 39. ThuT. gondii oocyst excretion are unclear [T. gondii oocyst shedding, most previous reports suggested no effect, which was consistent with the current findings [et al. [However, the effects of domestic feline breeds or gender on  unclear . Althoug unclear . With refindings . By cont [et al.  reportedT. gondii strain typing [T. gondii isolated from humans, animals, and meat products [T. gondii [T. gondii GRA6 in domestic cats in South Thailand [T. gondii as the predominant genotype for feline definitive hosts in China [T. gondii types I and II have been detected via GRA6 marker PCR-RFLPs in oocyst-contaminated soil samples from North Iran [Several genetic markers  have been utilized for n typing . Among tproducts –44. The . gondii . The preThailand  and OkinThailand . Other sin China . Furtherrth Iran .T. gondii DNA in fecal samples obtained from pet cats in Phayao, Thailand. Analyses of PCR-RFLP patterns and nucleotide sequences confirmed the presence of type-I T. gondii. These results indicated the circulation of type-I T. gondii GRA6 in this area, although such results were based on a single locus. However, data on circulating T. gondii genotypes from different hosts in Thailand are limited; thus, further investigations are necessary. In particular, epidemiological data of other animal species related to the life cycle of T. gondii and analyses of population genetics were required to accurately identify the circulating genotype of T. gondii in Thailand and comprehensively understand the relationship of toxoplasmosis in humans and animals.The present study demonstrated a high occurrence of OJ, CS, and KP: Conceived and designed the study. OJ and CP: Collected the samples. OJ, CS, and CP: Performed the experiments. OJ: Conducted molecular work and performed sequence analyses. OJ and KP: Analyzed the data. OJ and CS: Drafted and revised the manuscript. All authors have read and approved the final manuscript.


Solid pulmonary nodules are different from subsolid nodules and the diagnosis is much more challenging. We intended to evaluate the diagnostic and prognostic value of radiomics and deep learning technologies for solid pulmonary nodules.Retrospectively enroll patients with pathologically-confirmed solid pulmonary nodules and collect clinical data. Obtain pre-treatment high-resolution thoracic CT and manually delineate the nodule in 3D. Then, all patients were randomly divided into training and testing sets at a ratio of 7:3, and convolutional neural networks (CNN) models and random forest (RF) models were established. Survival analyses were performed for patients with solid adenocarcinomas.Totally 720 solid pulmonary nodules were enrolled, 348 benign and 372 malignant. The CNN model with clinical features achieved the highest AUC  with a sensitivity of 0.778, specificity of 0.788 and accuracy of 0.783. No significant differences were observed between the CNN and radiomics models. There were 295 solid adenocarcinomas in survival analysis. Different disease-free survival was observed between the low-risk and high-risk groups divided according to the radiomics Rad-score. However, the groups based on deep learning signatures showed similar survival. Cox regression analysis indicated that the radiomics Rad-score  was an independent predictor of recurrence.The radiomics and deep learning models can well predict the malignancy of solid pulmonary nodules. Radiomics signatures also demonstrate prognostic value in solid adenocarcinomas.The online version contains supplementary material available at 10.1186/s12885-022-10224-z. Lung cancer screening with low-dose computed tomography among high-risk individuals can reduce lung cancer modality , 2. HoweRecently, radiographic assessment of disease is being improved by advanced computational analyses. On the one hand, radiomics approach can digitally decode radiographic images into quantitative features , and therefore classify the medical image into a predefined group . On the In previous studies, researchers have investigated the diagnostic performance of radiomics and deep learning technologies for solid pulmonary nodules –19. HoweTherefore, the current study intended to establish CNN and radiomics models for solid pulmonary nodules without restricting the nodule size and pathology. Furthermore, survival analyses were performed for patients with solid adenocarcinomas.This retrospective study was approved by the institutional review board of the West China Hospital of Sichuan University. We collected possible cases by reviewing discharge records of patients in West China Hospital from January 2010 to July 2017. The following terms were used to extract the data: lung cancer, lung adenocarcinoma, lung squamous carcinoma, non-small cell lung cancer, small cell lung cancer; inflammatory lung nodule, benign lung nodule, benign lung tumor, lung hamartoma, lung sclerosing hemangioma, lung tuberculosis, lung granuloma. Then, the patient was enrolled based on the following criteria: (a) there was an untreated, pathologically confirmed, 5–30 mm noncalcified solid nodule detected on chest CT; (b) the slice thickness of CT was less than or equal to 1 mm. Otherwise, patients were excluded if (a) there were multiple pulmonary nodules, or pleural effusion, atelectasis, lymph node enlargement was observed; (b) it wasn’t a primary lung tumor.N = 315, 90.5%) and CT guided percutaneous lung biopsy , while the malignant nodules was confirmed by surgery , CT guided percutaneous lung biopsy  and transbronchial lung biopsy , respectively.Totally, the current study enrolled 720 patients with 720 nodules, 348 benign and 372 malignant. The pathology of benign nodules was confirmed by surgery  and compared with Chi-square test.The classification performance of the models was evaluated on sensitivity, specificity, accuracy, receiver operating characteristic curves (ROC) and values of area under the ROC curve (AUC). Calibration curves were also plotted to evaluate the accuracy of risk estimate. Additionally, Brier scores were calculated that quantitatively measure the distance in the probability domain and a lower score means better prediction. Differences in the AUC values were assessed by Delong test .http://tissuearray.org/) . Similarly, studies predicting risk of solid nodules of 6 mm to 15 mm found the CNN and radiomics model demonstrated an AUC of 0.93 and 0.97, respectively , 17. Wu In the radiomics models, most selected predictors were texture and histogram distribution features. The texture features can measure the spatial inter-dependency or co-occurrence of information across adjacent voxels . SpecifiWe also evaluated the prognostic value of radiomics and deep learning technologies for patients with solid adenocarcinomas. It was reported that the recurrence rate of early-stage non-small cell lung cancer is still substantial about 15–38.5% . For adeThere were some limitations need to be considered when interpreting our results. Firstly, this was a single center study and models weren’t externally validated. Previous studies have shown that when revalidated with external data, the performance of models may be reduced due to heterogeneous acquisition protocols and patient populations , 37. SecOn the one hand, it was found that the CNN models and radiomics models demonstrated good performance in predicting the malignancy of solid nodules, superior to the model based on clinical variables alone. On the other hand, radiomics features demonstrated potential to predict the DFS of patients with solid adenocarcinomas.Additional file 1: Table S1. Distribution of the CT manufacturer in the training and testing sets. Table S2. Distribution of the CT slice thickness in the training and testing sets. Fig. S1. ROC curves of different classification models in the training set. RF: random forest, CNN: convolutional neural network. Fig. S2. Kaplan-Meier survival curves in the training set based on radiomics features. Fig. S3. Kaplan-Meier survival analysis among stage I patients (testing set). The disease-free survival between the low-risk and high-risk groups were statistically different (P < 0.05). Fig. S4. Random forest of the multivariate Cox regression analysis among stage I patients. Rad-score  was the only independent predictor of disease-free survival. Fig. S5. Kaplan-Meier survival curves with malignancy-score derived from the CNN model with clinical features in the training (A) and testing (B) set.


Trichogramma wasps are minute egg parasitoids that are one of the most used biocontrol agents to manage lepidopteran pests. Given the great success in industrializing a mass-rearing method for the cost-effective production of Trichogramma dendrolimi, this parasitoid wasp has been used extensively to control agricultural and forestry pests in China. So far, while the biology and ecology of T. dendrolimi have been intensively studied to provide successful biological control, the limited genome information constrains the interpretation of the molecular mechanism underlying its host recognition and parasitism. Here, we assembled a high-quality reference genome for T. dendrolimi and identified the repetitive sequences and protein-coding genes. A phylogenomic tree constructed for T. dendrolimi and 24 hymenopteran species places the family Trichogrammatidae sister in the remaining Chalcidoidea species. We found that significantly expanded and contracted gene families are involved in the development, regulation and transport processes in T. dendrolimi, respectively. Additionally, 24 odorant binding proteins, 100 olfactory receptors, 27 gustatory receptors, 27 ionotropic receptors and 87 venom genes were identified in the genome of T. dendrolimi. Our study provides a foundation for future studies on the molecular mechanism underlying host recognition and parasitism in the Trichogramma species.Trichogramma dendrolimi is one of the most successfully industrialized Trichogramma species used to control agricultural and forestry pests in China. However, the molecular mechanisms underlying its host recognition and parasitism remain largely unknown, partially due to the limited genome information of this parasitoid wasp. Here, we present a high-quality de novo assembly of T. dendrolimi through a combination of Illumina and PacBio sequencing technologies. The final assembly had a length of 215.2 Mb and contains 316 scaffolds with a scaffold N50 size of 1.41 Mb. Repetitive sequences with a length of 63.4 Mb and 12,785 protein-coding genes were identified. Significantly expanded gene families were identified to be involved in the development and regulatory processes, while remarkably contracted gene families were involved in the transport processes in T. dendrolimi. The olfactory and venom-associated genes were identified in T. dendrolimi and 24 other hymenopteran species, using uniform methods combining BLAST and HMM profiling. The identified venom genes of T. dendrolimi were enriched in antioxidant activity, tricarboxylic acid cycle, response to oxidative stress and cell redox homeostasis. Our study provides an important resource for comparative genomics and functional studies to interpret the molecular mechanisms underlying host recognition and parasitism of Trichogramma species. Trichogramma (Hymenoptera: Trichogrammatidae) wasps are minute egg parasitoids (<1 mm in length) that are one of the most used biocontrol agents to control lepidopteran pests [Trichogramma species, these egg parasitoid wasps have been widely utilized to control Asian corn border (ACB) Ostrinia furnacalis (Lepidoptera: Crambidae) [Chilo suppressalis (Lepidoptera: Crambidae) [Grapholita molesta (Lepidoptera: Tortricidae) [Mythimna separata (Lepidoptera: Noctuidae) [Heortia vitessoides  [Trichogramma dendrolimi is one of the most successfully industrialized Trichogramma species to control agricultural and forestry pests because of its broad distribution in China, the wide host range and optimized mass-rearing method on eggs of the Chinese oak silkworm, Antheraea pernyi (Lepidoptera: Saturniidae) [T. dendrolimi mass-reared on eggs of A. pernyi have been successfully applied to control ACB through wide-scale inundative releases, resulting in 70% of ACB eggs being parasitized in the first generation, which has become one of the widely used biocontrol agents in integrated pest management programs [T. dendrolimi has increased nine times over the past decade, accounting for 35% of the corn production area in northeast China [an pests ,3. In Chambidae) , rice stambidae) , orientaricidae) , orientactuidae)  and mothralidae) . Trichogrniidae) ,9. In noTrichogramma species has been fruitful on many applied aspects, such as host location, host suitability, parasitic ability and breeding behavior to improve its biological control and mass-rearing efficiency [T. dendrolimi showed that there were different types of sensilla on the antenna, eyes, mouthparts, wings, legs and external genitalia, which provided a fundamental understanding of the sensory mechanisms in T. dendrolimi [Trichogramma species identified several olfactory receptors (ORs), gustatory receptors (GRs), ionotropic receptors (IRs) and odorant binding proteins (OBP)-related genes, providing genetic bases to further study the olfactory systems of the Trichogramma species [Trichogramma species. Many studies have shown that Trichogramma females usually showed high fitness on young host eggs, which may provide more available essential nutrients to support the development of the parasitoid offspring or have less competitive host embryos [Trichogramma females parasitize their highly suitable hosts [T. dendrolimi, T. ostriniae has also been widely used as a biological agent to control ACB in most of corn-growing regions in China. However, the selection of the best Trichogramma species to control ACB in the field remains a practical challenge that cannot be well addressed with the current knowledge. Compared with T. ostriniae, T. dendrolimi is a less effective biocontrol agent because it prefers to parasitize young ACB eggs (0–8 h old), while T. ostriniae can parasitize 0–48 h-old ACB eggs [T. dendrolimi on ACB eggs pre-parasitized by irradiated T. ostriniae females could be significantly enhanced, indicating that it is likely that the host-specific venom from T. ostriniae is more effective to counteract the immune response of ACB eggs [Trichogramma species for decades has achieved great success in providing an environmentally friendly and cost-effective approach to reduce the abundance and damage of lepidopterous pests in the field, the molecular mechanisms underlying the host location and parasitism in the Trichogramma species remain largely unknown, partially due to limited genome information of this parasitoid wasp.The biocontrol research of the ficiency . For insndrolimi . Recent  species ,12,13. I species ,14. The  embryos ,15. Howele hosts . BesidesACB eggs . InteresACB eggs . So far,Nasonia species’ genomes initiated a new era of genomic studies of parasitoid wasps [Trichogramma genomes have been published to date, including T. pretiosum and T. brassicae [T. pretiosum showed that its specialized egg parasitoid lifestyle and adaptation to miniaturization may be due to the rapid evolution of its genome [T. pretiosum and T. brassicae revealed that around 10% of ortholog clusters were specific to each of the two Trichogramma genomes [Trichogramma species  available on NCBI is highly fragmented. Recent studies have shown that parasitoid wasps exhibit exceptional levels of variations in gene content and gene regulation among different species [T. pretiosum genome or the T. brassicae genome for interpreting data derived for species-specific features in a T. dendrolimi genetic background. The whole genome sequence of the T. dendrolimi is not only critical to fully exploit the various biological process in T. dendrolimi, but also enriches the genetic resources of Trichogramma species to perform the comparative genomic analysis.Whole-genome information could advance a deep understanding of the genetic and evolutionary bases for physiological biology and parasitism strategies of parasitoid wasps, facilitate mapping and cloning the key quantitative traits loci and provide insights to further increase the efficiency and effectiveness of biocontrol by utilizing parasitoid wasps as pest insect control agents. Since the first three id wasps , an incrid wasps . To the rassicae ,21. The s genome . Compara species ,23. Even species ,25,26. TT. dendrolimi through a combination of Illumina short-read sequencing and PacBio long-read sequencing technologies. Phylogenetic analysis revealed the genetic relationships among the Trichogramma species, 18 representative Chalcidoidea species and 3 outgroup hymenopteran species with publicly available genomes. Significantly expanded and contracted gene families in the T. dendrolimi genome were investigated. The olfactory and venom-associated genes were identified in T. dendrolimi and 24 other hymenopteran species with uniform methods combining BLAST and HMM profiling. Our study provides an important resource for comparative genomics and functional studies to interpret the molecular mechanism underlying host recognition and parasitism in Trichogramma species.In this study, we present a high-quality reference genome of T. dendrolimi were collected from the corn fields in Yitong, Jilin Province, China  in 2015 and were confirmed through the morphological characteristics of male genital capsules. The wasp population was reared on the eggs of Antheraea pernyi . The eggs were kept at 26 °C ± 1 °C, 70% ± 5% relative humidity (RH) with a 16:8 h (L:D) photoperiod.T. dendrolimi and the need for a large amount of DNA for sequencing, genomic DNA was extracted from more than 10,000 haploid male pupae of T. dendrolimi dissected from the eggs of A. pernyi using a DNeasy Blood and Tissue Kit (Qiagen). We utilized the Illumina HiSeq 2000  and PacBio platforms  to sequence the genome of T. dendrolimi. Large insert (20 Kb) SMRTbell libraries were prepared and sequenced using the PacBio Sequel system according to the recommended protocol, yielding a total of 16.86-Gb high-quality long reads. DNA extracted from the same sample was used to build Illumina sequencing libraries. Five paired-end libraries with average insert sizes of 250 bp, 450 bp, 2 kb, 5 kb and 10 kb were constructed for genomic survey and sequenced with pair-end 150 bp on the Illumina HiSeq 2000 platform to yield 44.04 Gb (~200-fold coverage) short reads for error correction.The line used for sequencing was created from an individual mother and kept in the laboratory for more than three years without outcrossing with other lines. Given the tiny body size of The PacBio SMRT subreads were error corrected and assembled using Canu v1.8  using default parameters . To imprThe genome completeness and quality were assessed using the benchmarking universal single-copy orthologs (BUSCO) v3.02 (BUSCO: RRID:SCR_015008) . The assT. dendrolimi genome were identified using a combination of de novo and homology searches for known repeat databases. Briefly, the homology searches were carried out by screening the RepeatMasker v4.0.5 (RepeatMasker: RRID:SCR_012954) [www.repeatmasker.org, accessed on 12 December 2021). For de novo predication, the repeat library was constructed using RepeatModeler v1.0.7 [T. dendrolimi genome assembly to the de novo library for identifying repetitive sequences. Tandem repeats were also searched using Tandem Repeats Finder v4.09 (Tandem Repeats Finder: RRID:SCR_022193) [Repetitive sequences of the _012954)  against _012954)  and the r v1.0.7  (RepeatM_022193) . All repT. dendrolimi genome. For homology-based predication, protein sequences from five sequenced insect species, including Nasonia vitripennis, Apis mellifera, Apis cerana, Trichogramma pretiosum and Drosophilia melanogaster, were aligned against the T. dendrolimi genome using TBLASTN v2.8.1  [−5) and the genome sequences were aligned against the matching protein sequences using Genewise v2.4.1 (Genewise: RRID:SCR_015054) [T. pretiosum. For the transcriptome-based method, our previous published RNA-seq reads [T. dendrolimi by Tophat2 (Tophat: RRID:SCR_013035) [−5). The annotation information from different sources was combined for each gene in the final gene annotation set.A combination of protein-homology-based predication, ab initio prediction and transcriptome-based methods were utilized to predict the protein-coding genes in the _004870)    to defin_015054)  and SNAP_015054)  were pereq reads  were ali_013035)  to ident_013035) . The de _013035)  with def_013035)  to build_013035)  to build_013035) , InterPr_013035) , Swiss-P_013035)  and NCBINoncoding RNAs, including transfer RNA (tRNA), microRNA(miRNA), ribosome RNA (rRNA) and small nuclear RNA (snRNA), were identified using the following method: rRNA, miRNA and snRNA were annotated by mapping against the Rfam database (Rfam: RRID:SCR_007891)  using BLApis mellifera (Amel) (GenBank assembly accession: GCA_003254395.2), Orussus abietinus (Oabi) (GenBank assembly accession: GCA_000612105.2), Athalia rosae (Aros) (GenBank assembly accession: GCA_000344095.2), Copidosoma floridanum (Cflo) (GenBank assembly accession: GCA_000648655.2), Trichogramma pretiosum (Tpre) (GenBank assembly accession: GCA_000599845.3), Trichogramma brassicae (Tbra) (GenBank assembly accession: GCA_902806795.1), Trichogramma evanescens (Teva) (GenBank assembly accession: GCA_902732785.1), Ormyrus nitidulus (Onit) (GenBank assembly accession: GCA_900474335.1), Ormyrus pomaceus (Opom) (GenBank assembly accession: GCA_900474385.1), Eurytoma adleriae (Eadl) (GenBank assembly accession: GCA_900480045.1), Eurytoma brunniventris (Ebru) (GenBank assembly accession: GCA_900475205.1), Torymus auratus (Taur) (GenBank assembly accession: GCA_900474315.1), Torymus geranii (Tger) (GenBank assembly accession: GCA_900474355.1), Megastigmus dorsalis (Mdor) (GenBank assembly accession: GCA_900490025.1), Megastigmus stigmatizans (Msti) (GenBank assembly accession: GCA_900490015.1), Eupelmus urozonus (Euro) (GenBank assembly accession: GCA_900480035.1), Eupelmus annulatus (Eann) (GenBank assembly accession: GCA_900480025.1), Cecidostiba fungosa (Cfun) (GenBank assembly accession: GCA_900474305.1), Cecidostiba semifascia (Csem) (GenBank assembly accession: GCA_900474235.1), Pteromalus puparum (Ppup) (GenBank assembly accession: GCA_012977825.2), Trichomalopsis sarcophagae (Tsar) (GenBank assembly accession: GCA_002249905.1), Nasonia vitripennis (Nvit) (GenBank assembly accession: GCA_009193385.2), Nasonia giraulti (Ngir) (GenBank assembly accession: GCA_000004775.1) and Nasonia longicornis (Nlon) (GenBank assembly accession: GCA_000004795.1). OrthoFinder v2.5.1(OrthoFinder: RRID:SCR_017118) [A. mellifera and O. abietinus were 235–240 million years ago (Mya) [The protein sequences of 25 hymenoptera genomes were downloaded from InsectBase 2.0  and NCBI_017118) . The bes_017118) , using d_017118) . The mingo (Mya) .T. dendrolimi genome with Viterbi p-values < 0.05, using the Bioconductor package topGO (version 2.32.0) (topGO: RRID:SCR_014798) [CAFÉ  v4.2.1 (CAFÉ: RRID:SCR_005983) was used to analyze the gene family expansion and contraction ,59. The _014798) . In the Apis mellifera and several parasitoid wasps, including Pteromalus puparum, Nasonia vitripennis, Chelonus inanitus, Leptopilina boulardi and Leptopilina heterotoma from NCBI GeneBank and Hidden Markov models (HMMs), were used as references for gene identification. BLASTP v.2.7.1  (BLASTP: RRID:SCR_001010) and HMMER v3.3.2 (HMMER: RRID:SCR_005305) [T. dendrolimi and 24 other hymenopteran species. The annotated candidate gene sequences were aligned using MAFFT v7.453 [The gene families of olfactory receptors (ORs), odorant binding proteins (OBPs), gustatory receptors (GRs), ionotropic receptors (IRs) and venom proteins were manually annotated. The known orthologous protein sequences of _005305)  were useT v7.453  was usedT. dendrolimi through single-molecule real-time (SMRT) sequencing with the PacBio Sequel platform and paired-end sequencing with the Illumina HiSeq platform. The assembly of 16.86 Gb PacBio long reads, representing ~75-fold sequencing coverage of the T. dendrolimi genome, resulted in a 215.2 Mb assembly containing 316 scaffolds with a scaffold N50 size of 1.41 Mb , 1.0% (13) and 5.6% (78) of the 1367 BUSCO genes within the insecta_odb10 BUSCO set were present in the assembled T. dendrolimi genome as complete, fragmented and missing genes, respectively   . Such lo8–39.9%) .T. dendrolimi gene models except average intron length were smaller than those of T. pretiosum , which was predicted using the frequency of k-mers [Our pipeline identified that 29.46% of the  63.4 Mb . The mosf k-mers .T. dendrolimi, including transfer RNA (tRNA), microRNA (miRNA), ribosome RNA (rRNA) and small nuclear RNA (snRNA). There were 1140 non-coding RNAs identified in the T. dendrolimi genome, which covered 0.26 Mb and represented 0.12% of the genome  (including single-copy core OGs and variable-copy core OGs), dispensable OGs, species-specific OGs and singleton genes were estimated . The significantly expanded gene families of T. dendrolimi were overrepresented for several GO terms related to development and regulatory process: juvenile hormone biosynthetic process, negative regulation of hemocyte proliferation, methylation and caste determination  had the largest the number of OBPs (68 OBPs) and other Chalcidoidea species had 28–59 OBPs  to activate olfactory receptor neurons and to stimulate behavioral responses. The four –59 OBPs a. Phylog–59 OBPs b. The OBNasonia, Ormyrus and Cecidostiba species were found to contain more than 200 ORs and the other Chalcidoidea species had 100–200 ORs, except T. brassicae, which only harbored 30 ORs , contained 27–45 GRs. Other Chalcidoidea species had 27–95 GRs, in which the Nasonia, Ormyrus, Cecidostiba and Eurytoma species were found to contain more than 50 GRs   family, which have been identified to be involved in not only olfactory functions but also taste, temperature and humidity perception . Most of 16 IRs) a. The IR 16 IRs) e. IntereT. dendrolimi and 24 other hymenopteran species through BLAST searches against publicly available venom proteins sequences of Apis mellifera (Lepidoptera: Apidae) [Pteromalus puparum  [Nasonia vitripennis  [Chelonus inanitus (Hymenoptera: Braconidae) [Leptopilina boulardi and Leptopilina heterotoma  [Trichogramma species contained the minimum number of venom genes (86–112 venom genes), while seven species of Pteromalidae and Apis mellifera had the maximum number of venom genes (139–190 venom genes). The rest of the hymenopteran species had about 120 venom genes. Further analysis revealed that majority of the variations in venom gene numbers were from the hits matched against venom gene sequences from Apis mellifera, Nasonia vitripennis and Pteromalus puparum   and sevemalidae) , Nasoniamalidae) , Chelonuconidae) , Leptopigitidae)  using a  puparum , indicatTrichogramma are one of the most used biocontrol agents worldwide through inundative releases [Trichogramma in the last decades, China is leading the world in Trichogramma-based biological control programs such as diapause manipulation, optimal use of factitious hosts and new application methods [Trichogramma species, T. dendrolimi is one of the most successfully industrialized Trichogramma species used to control agricultural and forestry pests because of its broad distribution, the wide host range and optimized mass-rearing method on eggs of the Chinese oak silkworm, A. pernyi [T. dendrolimi, leading to successful biological control in the field [Trichogramma species is lacking and restricts further exploration of the molecular mechanism underlying its host recognition and parasitism of T. dendrolimi.Egg parasitoids in the genus releases . Due to  methods . Among t. pernyi ,9. So fahe field ,6,9,74. Trichogramma, have focused on the identification of traits that contribute to successful biological control, while the underlying molecular mechanisms remain poorly understood, which is partially due to the lack of a comprehensive genome data for this species. So far, three Trichogramma genomes  are publicly available on InsectBase 2.0 [Trichogramma genomes, our genome was sequenced using a combination of Illumina and PacBio platforms and the final genome had higher continuity and completeness. Notably, the three Trichogramma species with publicly available genomes are not commonly used in China. Therefore, our genome will serve as the first whole genome sequence of the Trichogramma species in China and provide comprehensive genomic information to identify the species-specific features.To date, the whole genomes have been obtained for many parasitoid wasp species , most ofBase 2.0  and NCBIT. dendrolimi and the three other Trichogramma species have moderate-size genomes (213.67–235.41 Mb) among the hymenopterans with sequenced genomes (180–340 Mb) [Trichogramma species (T. dendrolimi and T. pretiosum) is nearly the same as that of most chalcidoids. Copidosoma floridanum contains about 31.12% repetitive sequences in its genome (data from NCBI Copidosoma floridanum Annotation Release 101) and Nasonia species were also found to harbor about 30% of repetitive sequences in their genomes [T. dendrolimi, which is within the range of 12,000–20,000 genes reported for hymenoptera genomes [–340 Mb) ,75,76. T genomes 0–340 Mb  genomes . Hymenop genomes , and tha genomes .T. pretiosum and three other Trichogramma species increased from 6.4 Mya to 16.9 Mya.Our phylogeny analyses found that the family Trichogrammatidae was clustered in its own clade as the basal lineage, sister to all the other species of Chalcidoidea, which agrees well with previous finding that this family Trichogrammatidae is one of the earliest branching families of Chalcidoidea . A previTrichogramma species harbored significantly different numbers of olfactory-related genes and the gene number was correlated with the host range of these Trichogramma species [T. pretiosum identified 22 OBPs and 105 ORs, while we found 29 OBPs and 146 ORs in T. pretiosum, using methods combining BLAST and HMM profiling. Different versions of the whole genome sequences and methods might result in different gene numbers.The number of olfactory-related genes showed dramatic variations among the studied hymenopteran species, supporting the differences among their chemosensory systems and biology. The four  species , indicatT. pretiosum and N. vitripennis were reported to show high similarities [Nasonia lineage [OBPs are some of the most abundant proteins in insect olfactory organs, where they are involved in the first step of odorant detection and carrying odorant molecules to the olfactory receptors. Subsequently, the three transmembrane transporters, ORs, GRs and IRs, function as olfactory receptors to receive and discriminate these signals. We found an obvious contraction of OBPs, ORs, GRs and IRs in Trichogrammatidae compared with other species of Chalcidoidea, which may result from their very tiny body size and spending most of their lifetime inside the host eggs. The Pteromalidae family was found to harbor a high number of OBPs and olfactory receptors relative to other species of Chalcidoidea, which could be related to the need of this family to detect and discriminate between a number of diverse odors for food and reproduction or to avoid harmful substances . The amilarities , indicat lineage .T. dendrolimi and 24 other hymenopteran species through a blast method using very stringent criteria, the further analysis showed that such a blast method was very biased because it could only identify venom genes that had homologs in databases with predicated functions. Nowadays, an increasing number of studies reported that there exist variations in venom compositions between different parasitoid species, even closely related species [Trichogramma species inject into the host.The roles of the venom in parasitoids have evolved in parallel with their parasitic lifestyle. In ectoparasitoids, the venom often induce a long-term paralysis of the hosts , while e species ,73. TherT. dendrolimi, an important biocontrol agent to control lepidopteran pests, through a combination method of Illumina short-read sequencing and single-molecule real-time (SMRT) long-read sequencing technologies. We analyzed the genetic relationships among T. dendrolimi, 3 Trichogramma species, 18 representative Chalcidoidea species and 3 outgroup hymenopteran species with publicly available genomes. We also investigated the expanded and contracted gene families in T. dendrolimi and found that a number of contracted gene families were involved in transport processes. In addition, we identified olfactory and venom-related genes in T. dendrolimi and 24 other hymenopteran species through a uniform blast method. Our study provides an important resource for future studies in comparative genomics, functional studies, parasitoid-host interaction and biological pest control.In this study, we provide a high-quality genome assembly of


Oral squamous cell carcinoma (OSCC) is a common malignant tumor of the oral cavity. Shelterin complex gene (SG) has an important role in regulating telomere structure and length. SG is considered promising as a novel prognostic marker for cancer and a potential target for tumor therapy. However, SGs have not been systematically studied in OSCC. We analyzed SGs based on public data from OSCC patients and showed that SGs are closely associated with the prognosis of OSCC patients. Two different subtypes of SGs were identified in the TCGA and GEO cohorts, and LASSO regression analysis was used to further construct an SGs-related prognostic model. Randomized cohorts and different clinical subgroups validated the model's accuracy. The assessment of clinical characteristics, tumor mutational burden (TMB), and tumor microenvironment (TME) between high- and low-risk scores groups showed lower TMB, more abundant immune cell infiltration, and better prognosis in the low-risk group. According to the IPS analysis, patients in the low-risk group were more responsive to immunotherapy. This study establishes a foundation for research on SG and confirms that risk scores can predict prognosis and guide clinical treatment in OSCC patients. Telomeres are specialized nucleotide arrays located at the ends of linear chromosomes. As a result of shelterin being able to lengthen telomeres and protect the ends, malignant cells have bypassed senescence, which results in genomic instability –3. The DOne of the most common oral cancers is OSCC , with more than 350,000 diagnoses each year, corresponding to roughly 2% of all tumor diagnoses , 8. OSCCShelterin complex genes (SGs) have been implicated in cancer development in previous studies , 12. Li In order to predict prognosis and guide treatment, it was of primary importance to comprehensively evaluate the expression patterns of SGs in OSCC and develop a prognostic risk scoring model for SGs. Risk scores were used to assess tumor mutational burden, tumor microenvironment, immunotherapy response, drug sensitivity, and clinical prognosis in OSCC patients. Findings from these studies may provide new perspectives on how to better understand and treat SGs in OSCC.https://portal.gdc.cancer.gov/) and GEO databases (https://www.ncbi.nlm.nih.gov/geo/). TCGA-OSCC data  were obtained from the TCGA-HNSCC project. In the subsequent analysis, FPKM values were transformed using log2 (FPKM +1) [https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE41613) database, Platform: GPL570. An RMA normalization was carried out on the GSE41613 datasets. From GSE103322, we derived the data on single cells RNA-seq from oral cancer (http://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE103322). Cells from 18 patients with oral tumors made up 5902 single cells [OSCC patient-related data were retrieved from TCGA (FPKM +1) , 16. Genle cells . Public P > 0.05) between normal tissue and tumor tissue [https://metascape.org/gp/index.html#/main/step1) contains gene annotations and analyses [By analyzing TCGA-OSCC and GSE41613 samples, we aimed to determine which SGs differed in expression between normal and tumor tissues. The SVA package was used to normalize RNA expression profiles and eliminate batch effects on TCGA and GEO data . Researcr tissue . The Metanalyses ; this stWe identified the distinct patterns of SG expression in OSCC patients by using consensus clustering based on their expression levels to classify them further. The above steps were performed through the R package “ConsensusClusterPlus” and repeated 1000 times to ensure clustering stability , 22. Thehttps://www.gsea-msigdb.org/) was used to obtain the gene set (c2.cp.kegg.v7.2.symbols.gmt).Functional enrichment analysis of SGs clusters was conducted using the “GSVA” R package . ClusterSG-related patterns are differentiated by differently expressed genes (DEGs). Subsequently, DEGs with P < 0.05 were also included in the univariate Cox regression test utilizing the selection operator (LASSO) algorithm for dimensionality reduction and least absolute shrinkage –26. Basehttps://cancer.sanger.ac.uk/cosmic) and then gathered genomic mutation data of TCGA-OSCC for further analysis [In order to summarize OSCC patients' mutations, we used the COSMIC , we downloaded immunophenotype scores (IPS) from the TCGA-OSCC project. The higher the IPS, the better the accuracy of the more accurate result , 30.P values for all statistical analyses were two-sided, and a significance level of P < 0.05 was considered.Statistical analyses were carried out using R version 4.1.0, GraphPad Prism 8, and SPSS 23.0. We used the cluster profile package to examine GO, KEGG, and functional annotation enrichment. The ROC curve analysis was conducted using the “timeROC,” “survminer,” and “survival” R packages. The volcano and heat map were developed by R software's “ggplots” package. The shelterin complex consists of six proteins , termed We obtained 5902 single-cell data from 18 oral tumor patients from GSE103322 to determine the expression distribution of SGs in different cell types. Using TISCH (Tumor Immune Single-cell Hub) database, we visualized them. Findings revealed that these cells were divided into 20 clusters . Primaryk) from 2 to 9, we found the highest correlation between OSCC patients and other groups when k = 2  and risk scores to validate the accuracy of risk scores further and identify their role Figures . A signiAccording to an increasing amount of evidence, TMB may be a catalyst for tumor progression. Using the COSMIC database, we analyzed the mutation status of OSCC. The most common mutation types in OSCC were missense, G > A, and C > T mutations Figures . AccordiThe application of the ssGSEA algorithm allowed us to estimate immune cell infiltration in the high-risk and low-risk groups. Results revealed that the low-risk group had more immune cells present . Using ridge regression, we estimated the half-maximal inhibitory concentration (IC50) of samples. We calculated the prediction accuracy using the R package “pRRophetic” . As a result of the functional enrichment analysis, they were largely enriched in pathways related to telomeres . Several cancer types require telomere-associated proteins to maintain normal telomere function , 37, 38.We developed prognostic models based on these DEGs. In order to expand the use of SGs in prognosis and clinical treatment for OSCC patients, we tested their stability and accuracy among randomized cohorts and different clinical subgroups. As a result, the model accurately predicted patients' prognoses. Notably, patients in the low-risk group tended to have lower TMB and more abundant immune-infiltrating cells, including Mast cells resting, T cells CD4 memory activated, T cells follicular helper, T cells regulatory (Tregs), plasma cells, T cells CD8, and B cells naive. Previous studies have suggested that highly abundant TME may be more sensitive to immunotherapy. In order to identify high- and low-risk immunotherapy groups, we calculated IPS scores. We found that low-risk groups tended to have higher IPS scores, representing the low-risk groups more sensitive to immunotherapy. The drug sensitivity analysis revealed that various chemotherapeutic drugs are more or less sensitive to high- and low-risk groups of patients. The findings of our study can help treat the OSCC patients and update the study of SGs. We believe that the risk scores can be used to predict OSCC prognosis and help predict patients' clinical response to immunotherapy.However, there are several limitations to this study. The majority of the data we used came from public databases, so more experiments are required to verify the findings. In addition, studies reporting on SGs' role in OSCC are few, and our study only provides theoretical foundations for future experimental testing.In summary, we systematically examined the expression levels and prognostic relevance of SGs in OSCC. In addition, a risk score model for SGs was constructed to evaluate the prognosis and TME of OSCC. Using risk scores, it is possible to predict OSCC prognosis and assess TME and immunotherapy responses. Risk scores allow for more individualized clinical treatment and will assist in guiding medical practice.


Acquired cystic kidney disease (ACKD) is rarely massive in size. The great majority is asymptomatic and incidentally found on imaging studies for unrelated causes. We reported a case of an 85-year-old male with bilateral multiple huge acquired renal infected cysts, initially presenting with bowel obstruction-like symptoms. The computed tomography (CT) scan later aided in an accurate diagnosis. Symptomatic huge ACKD has, to our knowledge, scarcely been described. In addition, retroperitoneal lesions resulting in abdominal pain remain unusual and underrecognized in general practice. A cystic kidney disease is a group of conditions that cause fluid-filled sacs to form in or around the kidneys . It is a3 with neutrophil predominance (79%). His serum creatinine was 2.13 mg/dL without proteinuria. Urinary analysis showed leukocyturia (a urine white blood cell count of 30–50 cells/high power field) with nitrite positive. After he was diagnosed with bilateral multiple infected acquired renal cysts, ciprofloxacin was administered intravenously. Four days later, his fever and abdominal pain had largely diminished. The hemoculture was reported negative and urine culture grew Escherichia coli. After his clinical conditions improved, ciprofloxacin was switched to oral form, and he was referred to a urologist for further evaluation and considering an evaluation for kidney transplantation.An 85-year-old male was evaluated at this hospital because of acute abdominal pain. His comorbidities included essential hypertension, old myocardial infarction, chronic kidney disease, and gouty arthritis. No family history of early-onset chronic kidney disease was documented. Two years earlier, he had the first episode of urinary tract infection (UTI), which was resolved with a ten-day course of oral antibiotics at another hospital. One week after treatment, he underwent a kidney ultrasound examination to evaluate the possibility of complicated UTI. However, it revealed no abnormalities. The patient had been in his usual state of health until ten days before admission to our hospital, when acute abdominal pain, constipation, early satiety, and nausea developed. Abdominal pain was generalized, dull, 6 out of 10 in severity, and gradually worsened. He denied other organ-specific symptoms, except for a low-grade fever. He had never undergone abdominal surgery. On examination, the temperature was 38.4°C, the blood pressure was 127/73 mm·Hg, the heart rate was 86 beats per minute, and the respiratory rate was 18 breaths per minute. His abdomen was soft and moderately distended with slight but diffuse tenderness to soft touch and normal bowel sounds. The remainder of the examination was normal. A digital rectal examination revealed hard, brown stool with no blood or mucus. Plain film of the abdomen demonstrated a large well-defined high-density shadow on the left upper quadrant abdomen, deviating left colon to the central abdomen, without significant bowel dilatation . He thenA spectrum of adult renal cystic diseases was categorized into hereditary and non-hereditary conditions. The common causes in patients with multiple renal cysts remain autosomal dominant polycystic kidney disease (ADPKD), ACKD, and simple renal cysts , 3. Our The four cardinal symptoms of bowel obstruction are pain, vomiting, obstipation (or constipation), and distention . HoweverACKD is a condition commonly found in patients with advanced chronic kidney disease, presenting with multiple bilateral cysts, which are usually between 0.5 and 3 cm in diameter . The preEscherichia coli, Klebsiella pneumoniae, Salmonella enteritidis, Citrobacter diversus, Enterococcus faecalis, and Enterococcus spp. [According to his cyst infection, common pathogens included cus spp. . Lipid-scus spp. , 28. Beccus spp. . The ACKcus spp. . Moreovecus spp. . In paticus spp. . Althougcus spp. , 34.Our case exemplifies the importance of constructing a wide differential diagnosis in patients without prior abdominal surgery presenting with bowel obstruction-like symptoms. Retroperitoneal lesions, although rare, can cause abdominal pain and bowel obstruction-like symptoms. Careful radiographic examination and thoughtful evaluation of clinically relevant information is of importance to establish an accurate diagnosis and provide proper treatment.


Toxoplasma gondii is an obligate intracellular zoonotic pathogen capable of infecting almost all cells of warm-blooded vertebrates. In intermediate hosts, this parasite reproduces asexually in two forms, the tachyzoite form during acute infection that proliferates rapidly and the bradyzoite form during chronic infection that grows slowly. Depending on the growth condition, the two forms can interconvert. The conversion of tachyzoites to bradyzoites is critical for T. gondii transmission, and the reactivation of persistent bradyzoites in intermediate hosts may lead to symptomatic toxoplasmosis. However, the mechanisms that control bradyzoite differentiation have not been well studied. Here, we review recent advances in the study of bradyzoite biology and stage conversion, aiming to highlight the determinants associated with bradyzoite development and provide insights to design better strategies for controlling toxoplasmosis. Toxoplasma gondii is a cyst-forming obligate intracellular protozoan. Due to its wide host range, T. gondii is considered one of the most successful parasitic pathogens  and dihydrofolate reductase (DHFR), respectively, are widely used because of their significant activities against tachyzoites  has a distinct crescent shape and is approximately 2 × 6 μm in size . Tachyzo2+ agonists due to reduced Ca2+ levels and attenuated Ca2+ signaling (Bradyzoites (approximately 1.5 × 7 μm) contain significantly more micronemes than tachyzoites, and the nucleus is located at the posterior end of the parasite . High leignaling . The sizignaling . In the ignaling .Tachyzoites and bradyzoites differ not only in morphology but also at the molecular levels , the difin vivo , indicating that hsp30/bag1 is a specifically expressed gene in bradyzoites  are structurally related to SAG1 and all are confirmed to be expressed only on the surface of tachyzoites  sugar residues in a mucin-like domain that binds Dolichos biflorus agglutinin/lectin (DBA) (N-acetylglucosamine (GlcNAc)-decorated cyst structures , which are required for the formation of a mechanically stress-resistant cyst wall  . This glructures . The mucyst wall . One of on cysts , 2017. Son cysts . Among ton cysts . Like CSon cysts . In addion cysts . MAG2   synthesis, FASI and FASII pathways are performed in the cytosol and apicoplast of T. gondii, respectively , and TPK is critical for in vitro tachyzoite survival due to its high negative fitness scores based on the genome-wide CRISPR screen in T. gondii (5) to initiate CoA biosynthesis are shown to be essential for tachyzoite growth (Intracellular tachyzoites mainly utilize host-derived glucose as a carbon source and then catabolize glucose for biogenesis pathways . In the pathways . Pyruvatse (LDH) . Ldh1 isted mice . In the ted mice . These sectively . Parasitectively . Howeveranalyses . Tachyzoanalyses . To assi. gondii . In addie growth .eno2 promoter, while the promoter activity of eno1 is highly induced   activity cannot be detected, indicating that bradyzoites lack a functional TCA cycle and respiratory chain . However induced . Disruptntiation . LDH2 thntiation . On the ntiation . Bradyzontiation . Throughntiation . However in vivo . Using 1e (PPDK) . Furthere (PPDK) . Site-die (PPDK) . The studyzoites .Toxoplasma gondii tachyzoites can infect almost all nucleated cells of warm-blooded vertebrates  are infected with yoblasts . The brayoblasts . Overexproblasts . These fT. gondii can cross the BBB and invade various cell types, including neurons and astrocytes, as well as microglia  is a protective niche that restricts the entry of pathogens into the central nervous system (CNS). However, icroglia . Analysior cysts . Howeveror cysts . Thus, mT. gondii infection, several host-derived anti-parasitic mechanisms have been developed to limit parasite growth. The work focuses on how host microenvironment adjustments control T. gondii infection. This section lists host-sensing tools related to parasite control and bradyzoite differentiation/activation, including immune response and metabolite supply.After primary T. gondii infection in different hosts   pathway is an effective strategy primarily used to control nt hosts . In the  (IL-12) . The IL- (IL-12) . Unlike n humans . The hieT. gondii in C2C12 SkMCs, IFN-γ stimulation significantly inhibits parasite growth via inducing IRGs and NO, suggesting that IFN-γ also plays a pivotal role against T. gondii in skeletal cells  signaling and ROP18 virulence complex (ROP5/ROP17/ROP18/GRA7/GRA60)-dependent phosphorylation of IRGs contribute to parasite evasion from immune-mediated destruction, resulting in the conversion of tachyzoites to bradyzoites . Pathogenesis is associated with host CD4+ T cell impairment and TgROP18-mediated phosphorylation  reverses CD8+ T cell dysfunction, indicating that CD4+ T cells help to reduce CD8+ T cell exhaustion and that both are considered to be critical for improving parasite control to prevent fatal TE in infected mice -derived cholesterol. Tachyzoites increase LDL uptake in CHO cells, whereas deprivation of LDL-derived cholesterol induces bradyzoite conversion  condition, indicating that pyrimidine deprivation  triggers bradyzoite differentiation  via a GPI-anchored cell surface glycoprotein-CD73, which is highly expressed on CNS-resident cells  factors found in plants, Apicomplexa express several plant-like proteins called ApiAP2 factors. These ApiAP2 carry DNA-binding domains that cooperate with histone-modifying machinery (acetylation or deacetylation) . T. gond0 ApiAP2 , but onlT. gondii and thus exhibits the ability to sequence-specific DNA-binding to regulate gene promoter activities  and bag1 promoters to inhibit the bradyzoite transcriptome in competition with transcription activator AP2IV-3  is associated with stress-induced response in eukaryotic cells for selectively enhancing translation of relevant mRNAs and downregulating global protein expression . In T. g latency . The eIF latency . ER stre latency , 2. Comp latency . A recennfection , 2. Two nfection . TgIF2K-rylation , but theT. gondii PTM is histone acetylation, which is catalyzed by two GCN5-family lysine acetyltransferases (TgGCN5a and TgGCN5b). GCN5b cannot be knocked out, suggesting that it is essential for T. gondii viability  inhibits TgCARM1 activity and induces cyst formation in vivo  is recruited to target H3 [R17] along with GCN5, which perfectly targets H3 [K18] . AMI-1 , including strain-specific T. gondii effectors , may influence the transmission and pathogenicity of these T. gondii strains  or to treat parasites with an alkaline state in the absence of host cells  reproduces spontaneous differentiation of T. gondii into mature bradyzoite cysts (>14 days), which is a valuable tool for studying the effect of parasite differentiation on neuron biology  monolayer culture systems that are relatively easy to maintain and widely used in different experimental procedures (T. gondii life cycle stages is challenging due to the lack of specialized cells and limited animal models. Emerging 3D culture systems have been developed to fill the knowledge gap (in vitro model about T. gondii-infected human brain organoids supplemented with a Matrigel matrix is successfully used to simulate cyst formation, and parasites in infected organoids retain similar pathogenicity, indicating that this physiologically relevant system helps elucidate the mechanisms of T. gondii stage conversion and parasite–host interactions in the brain (These ocedures . Studyinedge gap . A novelhe brain .T. gondii transmission, and the reactivation of persistent bradyzoites in immunocompromised hosts may lead to severe zoonotic toxoplasmosis. Despite significant progress over the past decade in dissecting the mechanism of transition from tachyzoites to bradyzoites and vice versa, there is still much uncertainty about how host sensing and T. gondii self-regulation contribute to bradyzoite differentiation and reactivation. Many unknowns lead to currently ineffective treatment for underlying T. gondii infections.The conversion of tachyzoites to bradyzoites plays a cardinal role in in vitro. In particular, the regulation of polysaccharide–amylopectin metabolism by CDPK2 is essential for bradyzoites, as abnormal accumulation of amylopectin puts parasites at risk (in vitro conditions, host metabolites that support parasite growth directly contribute to T. gondii stage conversion (in vitro. However, the effect of critical metabolites on bradyzoite development in muscle cells and brain cells remains unclear. Since sulfonamides and pyrimethamine, which inhibit folate synthesis, are used as effective drugs against toxoplasmosis in animals (First, tachyzoites and bradyzoites have different metabolic characteristics to meet their carbon requirements . Associa at risk , so dissnversion . These s animals , the nexT. gondii (T. gondii antagonism of host immunity is achieved in the chronic phase. Since a subset of neurons can clear intracellular parasites via immunity-regulated GTPases (T. gondii-infected neurons actively harbor cysts while others do not? Single-cell RNA sequencing approaches may be necessary for revealing cell status to detect cell-to-cell differences. In immunocompromised hosts, bradyzoites can dedifferentiate into tachyzoites, depending on host CD4+ T cell and CD8+ T cell exhaustion (T. gondii? How much do they contribute to the recrudescence of toxoplasmosis in immunodeficiency individuals? In the tachyzoite–bradyzoite interconversion, unknown detail about how to maintain host immune surveillance and how T. gondii disrupts the balance of immune microenvironment needs to be explored.Additionally, despite host IFNγ-dependent defensive strategies, some tachyzoites escape immune-mediated destruction and convert to bradyzoites . Efficie. gondii . Thus, a GTPases , is therhaustion  and TgROhaustion . So, areT. gondii effectors are involved in regulating the self-cellular environment to optimize its persistence (T. gondii stage conversion. Meanwhile, efficient proximity labeling is a powerful approach to study the global identification of T. gondii effectors secreted into the host cell nucleus to regulate bradyzoite differentiation (T. gondii vacuole and cyst and their correlations need further study.In response to stress-induced responses, sistence . Among tsistence . Consequntiation . In addintiation , raisingT. gondii feedback. After that, studies focusing on the factors that control T. gondii stage conversion in an intermediate host help to understand the detailed mechanism of these processes and are promising for future drug design and vaccine development. Ultimately, better prevention and treatment can ensure that we win the tug-of-war between T. gondii and its hosts, reducing the public health burden of this parasite.In short, the underlying molecular mechanisms of bradyzoite differentiation and reactivation remain unclear, except for a convincing causal explanation for host stress and S-YH conceived and designed the review. MP, C-CG, Y-MF, Q-WJ, and BS drafted the first version of the manuscript. All authors provided feedback on the manuscript, read, and approved the final version.


Bacillus amyloliquefaciens provides a strong prospect as a biocontrol agent against Alternaria sp. on pepper plants.Plant pathogenic fungi cause serious damage in agriculture, resulting in major losses in the yield and the quality of different economic crops. Chemical fungicides are dangerous to human health and the environment. They have many harmful side effects on non-target organisms. Moreover, their residues have been found in human food. Endophytic bacteria could be a valuable and safe alternative method for the biological control of phytopathogenic fungus. The endophyte Bacillus amyloliquefaciens and B. velezensis and the production of several antimicrobial metabolites associated with the biocontrol of Alternaria sp., which cause serious diseases and affect important vegetable crops in Egypt. Twenty-five endophytic bacteria isolates were obtained from different plants cultivated in El-Sharkia Governorate, Egypt. Dual culture technique was used to evaluate the bacterial isolates’ antagonistic potentiality against Alternaria sp. and Helminthosporium sp. The most active bacterial isolates obtained were selected for further screening. The antifungal activity of the most active endophytic bacterial isolate was assessed in vivo on pepper seedlings as a biocontrol agent against Alternaria sp. A significant antifungal activity was recorded with isolates C1 and T5 against Alternaria sp. and Helminthosporium sp. The bacterial endophyte discs of C1 and T5 showed the highest inhibitory effect against Alternaria sp. at 4.7 and 3.1 cm, respectively, and Helminthosporium sp. at 3.9 and 4.0 cm, respectively. The most active endophytic isolates C1 and T5 were identified and the 16S rRNA sequence was submitted to the NCBI GenBank database with accession numbers: MZ945930 and MZ945929 for Bacillus amyloliquefaciens and Bacillus velezensis, respectively. The deformity of pathogenic fungal mycelia of Alternaria sp. and Helminthosporium sp. was studied under the biotic stress of bacteria. The culture filtrates of B. amyloliquefaciens and B. velezensis were extracted with different solvents, and the results indicated that hexane was the most efficient. Gas Chromatography-Mass Spectrometry revealed that Bis (2-ethylhexyl) phthalate, Bis (2-ethylhexyl) ester, and N,N-Dimethyldodecylamine were major constituents of the endophytic crude extracts obtained from B. amyloliquefaciens and B. velezensis. The in vivo results showed that Alternaria sp. infection caused the highest disease incidence, leading to a high reduction in plant height and in the fresh and dry weights of pepper plants. With B. amyloliquefaciens application, DI significantly diminished compared to Alternaria sp. infected pepper plants, resulting in an increase in their morphological parameters. Our findings allow for a reduction of chemical pesticide use and the control of some important plant diseases.Endophytic bacteria are plant-beneficial bacteria with a broad host range. They provide numerous benefits to their hosts, helping them tolerate several biotic and abiotic stresses. An interest has recently been developed in endophytic bacteria which are producing bioactive compounds that contribute to the biological control of various phytopathogens. This research paper aimed to investigate the potentiality of new local strains of endophytic bacteria such as  Fungal diseases are a massive threat to crop yields and global food security. They destroy one-third of all food crops each year, causing economic losses and affecting global poverty levels ,2. Many The biological methods of controlling diseases that affect plants is an eco-friendly alternative, especially applicable when there is pesticide resistance and the need for environmental protection toward sustainable agricultural methods ,5. MicroIn pathological research, an endophytic bacterium suppressing plant diseases has received much attention . EndophyFusarium oxysporum, Sclerotium rolfsii and Rhizoctonia solani, are inhibited by endophytic bacteria [Capsicum annuum have an antagonistic effect against several phytopathogens. Also, infection by fungal pathogens results in stronger reductions in plant biomass and survival compared to uninfected plants [Bacillus was mentioned in numerous research papers due to its prevalence in many plants, its antibacterial properties [Capsicum annuum) as a biocontrol agent against Alternaria sp.Endophytic bacteria can be isolated from almost all plant species ,16. Planbacteria . Amaresad plants ,18,19. Boperties ,21,22, aoperties ,23,24,25Solanum melongena, S. lycopersicum, Allium cepa, Coriandrum sativum, Pisum sativum, Portulaca oleracea, and Brassica oleracea  for 10 min, rinsed three times in sterile distilled water, and dried on sterile filter paper. Sterilized segments were placed on a 9 cm Petri dish containing potato dextrose (PDA) medium with rose bengal 30 mg/L and 250 mg/L streptomycin. Plates were incubated for 10 days at 30 °C. Fungal colonies were purified  and idenAlternaria sp. and Helminthosporium sp. using a dual culture technique. A ten-millimeter disc of a seven-day fungal pathogen culture was inoculated in the middle of Petri plates. The bacterial endophyte was streaked on the opposite side of the agar (PDA) plates. The plates inoculated with the pathogenic fungal disc were considered as the controls [The endophytic bacterial isolates were tested for antagonistic activity against controls , and thecontrols .PercentThe width of the inhibition zone was evaluated as + for 2–5 mm; ++ for 5–10 mm; and +++ for > 10 mm .Alternaria sp. and Helminthosporium sp. separately poured into a Petri dish of 90 mm in diameter. The plates were allowed to solidify and were then seeded with: (i) cell-free culture (150 µL) obtained by the cultivation of bacterial isolates in nutrient broth for 48 h and 150 rpm, and a millipore filter (0.45 μm) was used to filter-sterilize the culture supernatant using the well-diffusion method [PDA (20 mL) was inoculated with one mL of fungal spore suspension of n method ; and  (accessed on 1 September 2021).The DNA of the most active isolates was extracted using standard bacterial procedures . PCR wasA phylogenetic tree was created using MEGA 6.0 . The seqAlternaria sp. and Helminthosporium sp.) on PDA plates was examined. The hyphal strands from the confrontation lines at the end of the fungal colony were extracted and compared to the control plates under a light microscope  for anomalies [The morphological deformation caused by the most bioactive endophytic bacterial strains on the mycelia of each pathogenic fungus , and hexane separately in a separating funnel by shaking vigorously for 15 min. The mixtures were allowed to settle until two different layers appeared: the upper solvent and the lower aqueous layer. The extraction was repeated three times [Alternaria sp. and Helminthosporium sp. were put separately on PDA media and the solvents were used as a control. Gas Chromatography and Mass Spectrometry (GC-MS) were used to analyze the crude extracts.The two selected endophytic bacterial strains were cultured by placing agar blocks of actively growing pure culture (10 mm in diameter) in three Erlenmeyer flasks (1 L), each containing 300 mL of sterile nutrient broth for each endophytic bacterial strain, and incubated at 32 ± 2 °C for 24 h with continuous shaking at 150 rpm/min. After the incubation period, stationary growth cultures were centrifuged at 4000× The bioactive components in the crude extracts of two endophytic bacterial strains were identified by GC-MS . The cruB. amyloliquefaciens against Alternaria sp. was evaluated in a pot experiment in a greenhouse of the Botany and Microbiology Department, Faculty of Science, Zagazig University, with a temperature range of 23–30 °C and a relative humidity of 60–85% in a completely randomized design.The antifungal activity of B. amyloliquefaciens was grown at 30 °C for 48 h in nutrient broth. The bacterial suspension was obtained containing 107 CFU/mL.On a rotary shaker (180 rpm), Alternaria sp. was conducted by pipetting individual droplets of fungal suspension (105 cfu/mL) on the surface of healthy leaves after gently removing the leaf wax of control and infected pepper leaves using a brush. Also, pepper plants pipetted with tap water droplets were used as control plants. After pathogen inoculation, the inoculated plants were kept under polyethylene bags for 24 h to ensure the infection process and maintain high humidity conditions. Then, they were exposed to greenhouse conditions. There were ten replicates (n = 10) for each particular treatment. Four weeks after inoculation, disease symptoms were recorded.Plastic bags (12 cm diameter and 22 cm high) containing sterile field clay soil (2 kg/ag) were collected from an agricultural field in Minia Al-Qamh, El-Sharkia Governorate. Experimental treatments were applied to 45–50 days old pepper seedlings procured from the local market. Initially, bacterial suspension, according to Widnyana and Javandira , was useB. amyloliquefaciens application, pepper plants from the Alternaria sp. infected and non-infected treatments were uprooted and washed with tap water. The total heights of the pepper plants were measured. The total fresh weights (TFW) of the pepper plants for each treatment were taken, and then placed in the oven at 70 °C for two days. Their total dry weight (TDW) was recorded.After four weeks of Alternaria sp. and for the Alternaria sp. pepper plants infected and treated with B. amyloliquefaciens, was determined by the following formula:The incidence of the disease for the pepper plants that were only infected with p < 0.05 was used. The software package statistics 10.1 was utilized for statistical analysis.One-way ANOVA was used to analyze the data. To compare the means of the treatments, Duncan’s multiple range test at Solanum melongena, five isolates  were identified, while four isolates  were isolated from S. lycopersicum leaves. On the other hand, only one isolate (C1) was isolated from Brassica oleracea. These isolates were examined to see whether they had high anti-Alternaria sp. and anti-Helminthosporium sp. effects using the dual culture technique , S. lycopersicum (isolates T4 and T5), S. melongena (isolates E3 and E5), and B. oleracea (isolate C1) had exhibited inhibitory activities against Alternaria sp. and Helminthosporium sp. Moreover, the bacterial endophyte discs of C1 and T5 isolated from the leaves of B. oleracea and S. lycopersicum, respectively, showed the highest inhibitory effects against Alternaria sp.  and Helminthosporium sp.  as compared to the other isolates.The results in The morphological and biochemical characteristics of the antagonistic bacteria are listed in The most active endophytic bacterial isolates were selected and verified using the 16S rDNA gene sequence. The obtained partial sequence of the 16S rDNA gene was deposited in the GenBank database under accession numbers; MZ945930 and MZ945929, respectively, as shown in B. amyloliquefaciens and B. velezensis caused abnormal mycelial growth and significant morphological changes in Alternaria sp. and Helminthosporium sp., primarily manifesting as contraction, collapse, deformation, deformity of the conidium, and globular swellings at the tips of hyphal strands  was investigated using the filter paper diffusion method against both Alternaria sp. and Helminthosporium sp. Our results showed that all solvents (control) had no inhibitory effects on Alternaria sp. and Helminthosporium sp. as seen in B. amyloliquefaciens and B. velezensis had inhibitory effects on both fungal pathogens.The biological activity of the crude extracts of endophytic bacterial strains , Dibutyl phthalate, Methyl palmitate, and Ethyl hexadecanoate (The bioactive components of tography . The dettography . There wecanoate .B. amyloliquefaciens) was selected to act as a biocontrol agent via a greenhouse experiment. Alternaria sp. was inoculated into pepper plants either in the presence or absence of B. amyloliquefaciens. The positive effect of the application of B. amyloliquefaciens on pepper TFW, TDW, and plant heights were confirmed , TDW (1.4299 g/plant), and plant height (28 cm/plant). Also, Alternaria sp. exhibited the highest DI (80%) in the control plants, while with B. amyloliquefaciens inoculation, DI was greatly reduced in Alternaria sp. infected pepper plants (40%), as seen in B. amyloliquefaciens reduced the disease symptoms; therefore, B. amyloliquefaciens exhibited strong antagonism toward Alternaria sp. infection and improved the growth of the infected pepper plants.The most active bacterial endophyte in our study  acting against Sclerotinia sclerotiorum. Earlier investigations documented that antibiotics, such as mycosubtilins, iturins, and bacillomycins, are active metabolites with antimicrobial activities produced by B. subtilis [Some studies consider bacterial endophytes as potential biocontrol agents for various hazardous fungi ,7. Selime et al.  isolatedsubtilis ,51,52.Bacillus spp. can be used to develop effective microbial biopesticides in the form of biological control agents [Bacillus sp. forms beneficial relationships with plants directly or indirectly. B. velezensis and B. amyloliquefaciens are Gram-positive bacteria that have been used to promote the growth of numerous plants directly or indirectly as they are efficient in plant colonization and commercialized around the world [B. velezensis and B. amyloliquefaciens are the same, whether through direct antibiosis or plant-mediated induced disease resistance [B. velezensis and B. amyloliquefaciens produce several antimicrobial compounds against various phytopathogens [Helicobasidium purpureum, F. oxysporum, and Rhizoctonia solani, are inhibited by the B. velezensis strain FKM10. [B. velezensis can also cause the development of systemic resistance in plants [B. velezensis was found to produce several metabolites related to disease resistance, including NH3, antimicrobial proteins, polyketides, and siderophores [l agents . Olanrewl agents  reportedhe world ,23,54. Tsistance ,56,57. Aathogens . Some fun FKM10. ,60. B. vn plants . In somerophores ,58,60,61Alternaria sp. and Helminthosporium sp. revealed that treatment with B. amyloliquefaciens and B. velezensis caused abnormal mycelial growth. These anomalies showed a problem with fungal cell wall formation [F. oxysporum, Magnaporthe grisea, and Alternaria sp. when interacting with endophytes. The B. velezensis strain FKM10 destroyed the cell wall and cell membrane upon interacting with F. verticillioides [B. velezensis LM2303 affected the cell membrane permeability of F. graminearumon [B. velezensis CE 100 in mitigating phytophthora root rot, which suppressed mycelial growth, causing hyphae to enlarge and distort.The optical microscopic examination of ormation . Zhao etormation  noticed llioides . Furthernearumon . Moon etnearumon  investigB. amyloliquefaciens and B. velezensis were analyzed using GC-MS chromatography. The antifungal actions of these extracts could be related to various chemical classes, including esters, fatty acids, aldehydes, tertiary amines, alkaloids, and ketones. Among these bioactive compounds, the two bacterial strains had significant peaks in cell-free extracts, indicating that they played a substantial role in antibacterial and antifungal activities. These compounds include: Bis (2-ethylhexyl) phthalate followed by Bis (2-ethylhexyl) ester, N, N-Dimethyldodecylamine (Tertiary amine), Dibutyl phthalate, Methyl palmitate, and Ethyl hexadecanoate. Phthalates have antimicrobial and antifungal activities [The bioactive metabolites of tivities ,67,68,69tivities  reportedtivities  reportedtivities ,73.Bacillus atrophaeus strain XEGI50 species was a promising candidate as a biocontrol agent. The GC-MS analysis of cell-free extracts showed that numerous compounds had antimicrobial activity, including Bis (2-ethylhexyl) phthalate and Bis(2-ethylhexyl) ester. The antimicrobial activity of N, N-Dimethyldodecylamine, N-Methyl-N-benzyltetradecanamine, and N, N-Dimethyltetradecylamine (tertiary amine) against bacteria, yeasts, fungi, and enveloped viruses has been reported. [N-dimethyl-dodecyl amine against Sclerotinia sclerotiorum. The antimicrobial activity of these amine oxides was related to their interaction with biological membranes; the permeability of cellular membranes changed, and membrane-dependent activities were inhibited, resulting in cell death. Furthermore, the amine oxides caused K+ leakage from cells and lysis of osmotically stabilized protoplasts, which inhibited glycolysis [The high percentage of Bis (2-ethylhexyl) ester in both bacterial extracts had an antifungal activity. Mohamad et al.  suggesteeported. ,75. Masseported.  reportedycolysis ,75.Aspergillus spp. Moreover, ethyl hexadecanoate had antimicrobial, antioxidant, and pesticidal activities [Methyl palmitate (fatty acid methyl esters) detected in both bacterial extracts exhibited antibacterial and antifungal activities, damaging microbial cellular membranes . Chandrativities . The leativities ,78,79.Bacillus strains produce volatile organic compounds (VOCs) that may act as antifungal agents against various soil-borne diseases and limit fungal growth [B. amyloliquefaciens may inhibit F. oxysporum mycelial development and spore germination [B. velezensis suppressed the growth of B. cinerea by different numbers of VOCs. Also, Reda et al. [B. amyloliquefaciens S5I4 strain produced bioactive antimicrobial compounds.Some l growth ,48. Thesl growth ,81. For mination ,82. Gao mination  and Jianmination  reporteda et al.  indicateB. amyloliquefaciens for pepper plant growth is in coherence with Shahzad et al. [B. amyloliquefaciens RWL-1 and GGA inoculation significantly enhanced the growth traits of tomato and garlic plants under both diseased and non-diseased conditions of F. oxysporum and S. cepivorum. The extensively distinguished mechanisms for plant growth promotion caused by B. amyloliquefaciens are through phytohormones, providing the essential nutrients, N2 fixation, and phosphate solubilization [B. amyloliquefaciens on pepper plants.The enhancing capacity of d et al.  and Rashd et al. , who doclization . In addilization . Shahzadlization  discoverAlternaria, which can cause serious diseases to important vegetable crops in Egypt. Bacillus species, as biocontrol agents, could inhibit potential plant pathogens. The B. amyloliquefaciens MZ945930 and B. velezensis MZ945929 strains in this study, shared the same antifungal mechanisms by direct antibiosis against Alternaria sp. and Helminthosporium sp. Moreover, suppressive effects were associated with a variety of secondary metabolite secretions. The resulting bacterial crude extracts from both bacterial strains were promising as they have shown the highest antifungal activities. Also, the in vivo results emphasized the significance of the effect of B. amyloliquefaciens on pepper growth under both the control and diseased conditions caused by Alternaria sp. Therefore, the present study encourages the use of these bacterial strains as biocontrol agents in agriculture.Controlling plant pathogen diseases in a safe, effective, and alternative manner has become increasingly crucial for improving the quality of agricultural products. Compared to chemical control, biological control using antagonistic microorganisms, such as bacteria, is a long-term approach to inhibiting plant pathogens. The novelty of this work is the isolation of new local strains of endophytic bacteria and the production of several antimicrobial metabolites associated with the biocontrol of


Trypanosoma cruzi, which is transmitted mainly through the feces/urine of infected triatomine bugs. The acute phase lasts 2–3 months and is characterized by high parasitemia and nonspecific symptoms, whereas the lifelong chronic phase features symptoms affecting the heart and/or digestive tract occurring in 30–40% of infected individuals. As in humans, cardiac abnormalities are observed in T. cruzi-infected dogs and cats. We reviewed the technological advances in the serological diagnosis of CD in dogs and cats.Chagas disease (CD) is caused by A review of the published literature during the last 54 years (1968–2022) on the epidemiology, clinical features, diagnosis, treatment and prevention of CD in dogs and cats was conducted.T. cruzi transmission and thus the risk for human infection. Although dogs and cats are reputed to be important for maintaining the T. cruzi domestic transmission cycle, there are no commercial tests to detect past or active infections in these animals. Most published research on CD in dogs and cats have used in-house serological tests prepared with native and/or full-length recombinant antigens, resulting in variable diagnostic performance. In recent years, chimeric antigens have been used to improve the diagnosis of chronic CD in humans with encouraging results. Some of them have high performance values (> 95%) and extremely low cross-reactivity rates for Leishmania spp., especially the antigens IBMP-8.1 to IBMP-8.4. The diagnostic performance of IBMP antigens was also investigated in dogs, showing high diagnostic performance with negligible cross-reactivity with anti-Leishmania infantum antibodies.Using predefined eligibility criteria for a search of the published literature, we retrieved and screened 436 publications. Of these, 84 original studies were considered for inclusion in this review. Dogs and cats are considered as sentinels, potentially indicating an active T. cruzi infections in dogs and cats is urgently needed. The use of chimeric recombinant T. cruzi antigens may help to fill this gap and is discussed in this review.The development of a commercial immunodiagnostic tool to identify past or active The online version contains supplementary material available at 10.1186/s13071-022-05476-4. Trypanosoma cruzi. Recent estimates indicate that 6–7 million people are infected worldwide, with 10,000 deaths attributable to CD annually in 21 Latin American countries , they may be more susceptible to T. cruzi [Some cats can mount an effective immune response to T. cruzi , but thiT. cruzi infection in cats  showed 30.8% of 679 cats had anti- methods . Recent  methods  and Loui methods , respectrgentina . The autrgentina .T. cruzi antibody production reaches detectable titers and can be identified by indirect immunoassays. In dogs, serum immunoglobulin M (IgM) begins to decrease markedly about 3 months after infection, whereas the opposite is true for immunoglobulin G (IgG), which increases up to 15 months and then gradually decreases up to 2 years and then appears to stabilize over the years [In endemic areas, the presence of the above-mentioned clinical signs and clinicopathological abnormalities can lead veterinarians to suspect CD in dogs and cats. In the acute phase, parasitological   or moleche years .Fig. 7ScT. cruzi antibodies in cats. Although both tests use protein A to detect IgG antibodies, there are no studies validating their use in cats.Table T. cruzi epimastigotes have traditionally been used as complex antigen mixtures to detect anti-T. cruzi antibodies. Although these combinations have been shown to provide sufficient sensitivity to detect even low antibody levels [T. cruzi strain in the study area and the epimastigote strain used in the tests [T. cruzi has a high antigenic variation that can lead to false-negative or false-positive results [Whole-cell homogenates or fractionated lysates of y levels , difficuy levels –132. Thihe tests , 134. An results , dependiT. cruzi contain tandem repeat amino acid sequences consisting of 5–68 amino acids [T. cruzi strains [In the last 2 decades, advances in DNA recombination technology have enabled the use of recombinant proteins in immunoassays (primarily ELISA and chemiluminescence assays), as large quantities of purified antigens can be produced in transformed prokaryotic cells grown in bioreactors . Approxino acids –139. Thino acids , 141 andno acids , 142. Inno acids –145. Howno acids . The hig strains . To over strains –151. In  strains . More re strains .T. cruzi proteins have been proposed to improve the accuracy of immunoassays for the diagnosis of human CD [T. cruzi-positive and -negative samples compared to the homogenate of the whole parasite [T. cruzi proteins . This antigen was evaluated and proven to be highly sensitive for the diagnosis of human CD [T. cruzi antibodies regardless of parasite strain or gene expression intensity. In addition, these findings support the utility of performing immunochemical assays with hybrid, chimeric single-molecule antigens rather than peptide mixtures or recombinant proteins.In recent years, synthetic chimeric recombinant antigens consisting of conserved repetitive amino acid fragments of different antigenic human CD , 153. Inhuman CD . High dihuman CD . The disparasite . Similarhuman CD . A chimehuman CD . These rT. cruzi antigens have been proposed for the diagnosis of chronic CD in humans: IBMP-8.1, IBMP-8.2, IBMP-8.3 and IBMP-8.4 . The diagnostic potential of these proteins for the detection of CD in humans has been extensively studied, using different diagnostic methods and platforms such as indirect ELISA [T. cruzi proteins, as described in Fig. T. cruzi antibodies.Recently, four chimeric recombinant ct ELISA , 156–161ct ELISA , lateralct ELISA , double-ct ELISA , Westernct ELISA . IBMP anT. cruzi-positive from -negative human samples was evaluated, and the area (AUC) under the receiver-operator curve was determined for each molecule. The determination of AUC values is used as the global accuracy of immunoassays [T. cruzi human positive and -negative samples from different geographic endemic regions of Brazil and other endemic countries using indirect ELISA [Leishmania spp. was extremely low in patients with American cutaneous and visceral leishmaniasis [T. cruzi and Leishmania spp. are co-endemic [T. cruzi-positive and -negative individuals regardless of geographic origin or clinical presentation. AUC values were 100%, demonstrating an outstanding diagnostic accuracy. The study showed that the lateral flow assay based on these antigens is a promising method for screening CD [The ability of IBMP antigens to discriminate noassays  and can noassays . Accordinoassays . These rct ELISA –158 and ct ELISA . Sensitict ELISA , 168. Crmaniasis . In ligh-endemic . In 2020-endemic  proposedening CD . In 2020ening CD . Recentlening CD .T. cruzi antibodies. Recently, two recombinant T. cruzi proteins (IBMP-8.1 and IBMP-8.4) were tested as diagnostic platforms using a rapid immunochromatographic assay  [Staphylococcus aureus protein A or Streptococcus pyogenes protein G as gold-labeled reagents to visualize the precipitin band formed between immunoglobulin (Ig) G-specific antibodies and the recombinant antigen immobilized on the nitrocellulose strip used in the assay. Protein A and protein G were based on the fact that these microbial molecules bind with different affinity and specificity to immunoglobulins of different species, including dogs. The authors found that the intensity pattern of the bands was directly proportional to the serological titer in IFAT. The sensitivity was 94% and the specificity was 91%. The agreement obtained was considered substantial by kappa analysis (84%). Of the T. cruzi-positive hemoculture samples, 88.9% were positive with TR-Chagas Bio-Manguinhos. The assay was efficient in detecting infections with five of the six T. cruzi discrete typing units . Cross-reactions were not observed in infections with Leishmania infantum, Trypanosoma rangeli, Trypanosoma caninum and Dirofilaria immitis, but were observed in sera from dogs infected with Crithidia mellificae, Anaplasma spp. and Erlichia spp. However, the authors used a convenient serum panel for cross-reactivity analysis, many of which had only a single sample per disease. Therefore, further studies should be conducted to confirm or refute these results. This test provides rapid preventive measures in areas at high risk for Chagas disease occurrence in a safe, reliable, cost-effective and immediate manner without the need for more complex laboratory testing. In 2020, the diagnostic performance of a rapid test based on trypomastigote small surface antigen (TSSA) was evaluated (namely Chagas Sero K-SeT). However, low sensitivity for the diagnosis of Chagas disease in dogs was observed , indicating the need for further studies to improve test performance [With the exception of IBMP antigens, all chimeric recombinant proteins discussed here  have been evaluated only for human diagnostics. As mentioned previously, there are no commercial tests for the diagnosis of CD in dogs and cats. In 2019, a phase I study investigated the diagnostic performance of IBMP antigens in dogs . AUC val Brazil) . Recombiformance .T. cruzi antibodies is possible in any mammalian species, serological tests may give discrepant results in different situations. This is mainly due to the high genetic and phenotypic intraspecific diversity of T. cruzi [T. cruzi-infected individuals [T. cruzi in dogs and cats would be useful from both veterinary and public health perspectives. Such a test should be able to detect antibodies regardless of the geographical region and the circulating DTU, with high sensitivity, specificity and accuracy, as well as with low risk of cross-reactivity . Furthermore, the test should be rapid (rapid diagnostic test), inexpensive and easy-to-use under field conditions (point-of-care test). Data show that the chimeric recombinant antigens combine all the necessary characteristics for a test with good applicability for epidemiological surveillance in veterinary clinical practice and in animal blood centers.Although the detection of anti-T. cruzi , 174, thT. cruzi , the varT. cruzi , 176 andividuals . The devAdditional file 1. Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA).