FgPfn participates in vegetative growth, sexual reproduction, pathogenicity, and fungicides sensitivity via affecting both microtubules and actin in the filamentous fungus Fusarium graminearum.

Fusarium head blight (FHB), caused by Fusarium graminearum species complexes (FGSG), is an epidemic disease in wheat and poses a serious threat to wheat production and security worldwide. Profilins are a class of actin-binding proteins that participate in actin depolymerization. However, the roles of profilins in plant fungal pathogens remain largely unexplored. Here, we identified FgPfn, a homolog to profilins in F. graminearum, and the deletion of FgPfn resulted in severe defects in mycelial growth, conidia production, and pathogenicity, accompanied by marked disruptions in toxisomes formation and deoxynivalenol (DON) transport, while sexual development was aborted. Additionally, FgPfn interacted with Fgα1 and Fgß2, the significant components of microtubules. The organization of microtubules in the ΔFgPfn was strongly inhibited under the treatment of 0.4 µg/mL carbendazim, a well-known group of tubulin interferers, resulting in increased sensitivity to carbendazim. Moreover, FgPfn interacted with both myosin-5 (FgMyo5) and actin (FgAct), the targets of the fungicide phenamacril, and these interactions were reduced after phenamacril treatment. The deletion of FgPfn disrupted the normal organization of FgMyo5 and FgAct cytoskeleton, weakened the interaction between FgMyo5 and FgAct, and resulting in increased sensitivity to phenamacril. The core region of the interaction between FgPfn and FgAct was investigated, revealing that the integrity of both proteins was necessary for their interaction. Furthermore, mutations in R72, R77, R86, G91, I101, A112, G113, and D124 caused the non-interaction between FgPfn and FgAct. The R86K, I101E, and D124E mutants in FgPfn resulted in severe defects in actin organization, development, and pathogenicity. Taken together, this study revealed the role of FgPfn-dependent cytoskeleton in development, DON production and transport, fungicides sensitivity in F. graminearum.

The effect of Ca2+-calcineurin signaling pathway on the antifungal activity of Pd-D-V against Botrytis cinerea.

Gray mold, caused by Botrytis cinerea is an intractable fungal disease that causes extensive damage to agricultural products. In the search for novel antifungal active ingredients, we discovered a linear pyranocoumarin Pd-D-V was effective against B. cinerea in both in vitro and in vivo assays. Furthermore, this study investigated the effects of Ca2+ and the Ca2+-calcineurin signaling pathway on its antifungal activity against B. cinerea. The results indicated that Pd-D-V reduced the concentration of Ca2+ in the mycelia of B. cinerea; CaCl2, the Ca2+ channel blocker verapamil, or the calcineurin inhibitor cyclosporin A could affect the sensitivity of Pd-D-V against B. cinerea; the expression of genes (Bccch1, Bcmid1, BccnA, Bccnb1, Bcpmc1, and Bcpmr1) of the Ca2+-calcineurin signaling pathway decreased after Pd-D-V treatment. In summary, Pd-D-V is compound for developing fungicides against B. cinerea. Pd-D-V can reduce intracellular Ca2+ concentration and disturb Ca2+ homeostasis. The Ca2+-calcineurin signaling pathway is important in the antifungal activity of Pd-D-V against B. cinerea.

Bioactivity and Resistance Risk of Fluxapyroxad, a Novel SDHI Fungicide, in Didymella bryoniae.

Gummy stem blight, caused by Didymella bryoniae, is an important disease in watermelon in China. Fluxapyroxad, a new succinate dehydrogenase inhibitor fungicide, shows strong inhibition of the mycelia growth of D. bryoniae. However, its resistance risk in D. bryoniae is unclear. In this research, the sensitivities of 60 D. bryoniae strains to fluxapyroxad were investigated. The average EC50 value and MIC values of 60 D. bryoniae strains against fluxapyroxad were 0.022 ± 0.003 µg/ml and ≤0.1 µg/ml for mycelial growth, respectively. Eight fluxapyroxad-resistant mutants with medium resistance levels were acquired from three wild-type parental strains. The mycelial growth and dry weight of mycelia of most mutants were significantly lower than those of their parental strains. However, four resistant mutants showed a similar phenotype in pathogenicity compared with their parental strains. The above results demonstrated that there was a medium resistance risk for fluxapyroxad in D. bryoniae. The cross-resistance assay showed that there was positive cross-resistance between fluxapyroxad and pydiflumetofen, thifluzamide, and boscalid, but there was no cross-resistance between fluxapyroxad and tebuconazole and mepronil. These results will contribute to evaluating the resistance risk of fluxapyroxad for managing diseases caused by D. bryoniae and further increase our understanding about the mode of action of fluxapyroxad.

Loss of Y chromosome in leukocytes can be regarded as a male-specific age predictor for age group estimation in forensic genetics.

Age prediction is an important field in forensic and aging research. Traditional methods used DNA methylation, telomere shortening, and mitochondrial DNA mutations to conduct age prediction models. Sex chromosomes, like the Y chromosome, have a significant role in aging as previously reported in hematopoietic disease and many non-reproductive cancers. Until now, there is no age predictor based on the percentage of loss of Y chromosome (LOY). LOY has been previously revealed to be correlated with Alzheimer's disease, short survival, and higher risk of cancer. The possible correlation of LOY between normal aging was not fully explored. In this study, we conducted age prediction by measuring LOY percentage by droplet digital PCR (ddPCR), based on 232 healthy male samples, including 171 blood samples, 49 saliva samples, 12 semen samples. The age group of samples ranges from 0 to 99 years, with two individuals in almost every single age. Pearson correlation method was performed to calculate the correlation index. The result indicated a correlation index of 0.21 (p = 0.0059) between age and LOY percentage in blood samples, with the regression formula being y = - 0.016823 + 0.001098x. The correlation between LOY percentage and age is obvious only when the individuals were divided into different age groups (R = 0.73, p = 0.016). In the studied saliva and semen samples, p-values of the correlation are 0.11 and 0.20, respectively, showing no significant association between age and LOY percentage in these two biological materials. For the first time, we investigated male-specific age predictor based on LOY. The study showed that LOY in leukocytes can be regarded as a male-specific age predictor for age group estimation in forensic genetics. This study might be indicative for forensic applications and aging research.

Antifungal activities of a novel triazole fungicide, mefentrifluconazole, against the major maize pathogen Fusarium verticillioides.

Fusarium ear rot (FER) is a serious fungal disease occurring the late growth stage of maize. FER not only reduces the yield of maize but also causes mycotoxin contamination, which affects the quality of maize and threatens human and animal health. Fusarium verticillioides is the predominant causative pathogen of FER worldwide. At present, there is no registered fungicide for use against maize FER in China. The novel isopropyl alcohol-triazole fungicide mefentrifluconazole (MFZ) has been shown to be effective against several Fusarium spp., but little is known about its specific activity against F. verticillioides. MFZ exhibited strong antifungal activities against 50 strains of F. verticillioides collected from the major maize-growing areas in China. MFZ inhibited mycelial growth, conidium production, germination and germ tube elongation of F. verticillioides. MFZ treatment significantly reduced fumonisin production and the expression levels of fumonisin biosynthetic genes. Genome-wide transcriptional profiling of F. verticillioides in response to MFZ indicated that the expression of genes involved in ergosterol biosynthesis, including fungicide target genes (cyp51 genes), was significantly downregulated by MFZ. MFZ treatment resulted in reduced ergosterol production and increased glycerol and malonaldehyde production as well as relative conductivity in F. verticillioides. A 2-year field experiment showed a significant reduction in FER severity in maize after spraying with MFZ at the tasseling stage. This study evaluated the potential of MFZ to control FER in maize and provides insights into its antifungal activities and mechanism of action against F. verticillioides.

The ATP Synthase Subunits FfATPh, FfATP5, and FfATPb Regulate the Development, Pathogenicity, and Fungicide Sensitivity of Fusarium fujikuroi.

ATP synthase catalyzes the synthesis of ATP by consuming the proton electrochemical gradient, which is essential for maintaining the life activity of organisms. The peripheral stalk belongs to ATP synthase and plays an important supporting role in the structure of ATP synthase, but their regulation in filamentous fungi are not yet known. Here, we characterized the subunits of the peripheral stalk, FfATPh, FfATP5, and FfATPb, and explored their functions on development and pathogenicity of Fusarium Fujikuroi. The FfATPh, FfATP5, and FfATPb deletion mutations (∆FfATPh, ∆FfATP5, and ∆FfATPb) presented deficiencies in vegetative growth, sporulation, and pathogenicity. The sensitivity of ∆FfATPh, ∆FfATP5, and ∆FfATPb to fludioxonil, phenamacril, pyraclostrobine, and fluazinam decreased. In addition, ∆FfATPh exhibited decreased sensitivity to ionic stress and osmotic stress, and ∆FfATPb and ∆FfATP5 were more sensitive to oxidative stress. FfATPh, FfATP5, and FfATPb were located on the mitochondria, and ∆FfATPh, ∆FfATPb, and ∆FfATP5 disrupted mitochondrial location. Furthermore, we demonstrated the interaction among FfATPh, FfATP5, and FfATPb by Bimolecular Fluorescent Complimentary (BiFC) analysis. In conclusion, FfATPh, FfATP5, and FfATPb participated in regulating development, pathogenicity, and sensitivity to fungicides and stress factors in F. fujikuroi.

Inference of population structure and admixture proportion from Y chromosomal data of Chinese population.

In the past two decades, Y chromosome data has been generated for human population genetic studies. These Y chromosome datasets were produced with various testing methods and markers, thus difficult to combine them for a comprehensive analysis. In this study, we combine four human Y chromosomal datasets of Han, Tibetan, Hui, and Li ethnic groups. The dataset contains 27 microsatellites and 137 single nucleotide polymorphisms these populations share in common. We assembled a single dataset containing 2439 individuals from 25 nationwide populations in China. A systematic analysis of genetic distance and clustering was performed. To determine the gene flow of the studied population with worldwide populations, we modeled the ancestry informative markers. The reference panel was regarded as a mixture of South Asian (SAS), East Asian (EAS), European (EUR), African (AFR), and American (AMR) populations from 1000 Genomes data of Y chromosome using nonlinear data-fitting. We then calculated the admixture proportion of these four studied populations with 26 worldwide populations. The results showed that the Han and Hui have great genetic affinity, and Hui is the most admixed ethnic group, with 61.53% EAS, 34.65% SAS, 1.91% AFR, 1.56% AMR, and 0.04% EUR ancestry component (the AMR is highly admixed and thus should be ignored). All the other three ethnic groups contained more than 97% EAS ancestry component. The Li is the least admixed population in this study. The combined dataset in this study is the largest of this kind reported to date and proposes reference population data for use in future paternal genetic studies and forensic genealogical identification.

Development and validation of a forensic multiplex InDel assay: The AGCU InDel 60 kit.

Marker sets based on insertion/deletion polymorphisms (InDels) combine the characteristics of both short tandem repeats (STRs) and single nucleotide polymorphisms and have served as effective complementary or stand-alone systems for human identification in forensics. We developed a novel multiplex amplification detection system, designated the AGCU InDel 60 kit, containing 57 autosomal InDels, 2 Y-chromosomal InDels, and the amelogenin locus and validated the kit in a series of studies, which included tests of the PCR conditions; tests for sensitivity, species specificity, reproducibility, stability, and mock case samples; degradation studies; and a population study. The results indicated that the AGCU InDel 60 kit was accurate, specific, reproducible, stable, and robust. Complete DNA profiles were obtained even with 125 pg of human DNA. In tests of artificially degraded samples, we found that the number of alleles detected by the validated kit was considerably greater than that detected by the STR-based AGCU 21+1 kit, even as the degree of degradation increased. Additionally, 564 unrelated individuals from three Han groups were investigated using this novel system, and the values of combined power of discrimination and combined power of exclusion were not less than 1-4.9026 × 10-24 and 1-3.1123 × 10-5 , respectively. Thus, the results indicated that the novel kit was more powerful than the previous version of the InDel kit (the AGCU InDel 50 kit). Our results suggest that the AGCU InDel 60 kit can serve as an efficient tool for human forensics and a supplementary kit for population genetics research.

Validamycin A Enhances the Interaction Between Neutral Trehalase and 14-3-3 Protein Bmh1 in Fusarium graminearum.

In agriculture, Trehalase is considered the main target of the biological fungicide validamycin A, and the toxicology mechanism of validamycin A is unknown. 14-3-3 proteins, highly conserved proteins, participate in diverse cellular processes, including enzyme activation, protein localization, and acting as a molecular chaperone. In Saccharomyces cerevisiae, the 14-3-3 protein Bmh1could interact with Nth1 to respond to specific external stimuli. Here, we characterized FgNth, FgBmh1, and FgBmh2 in Fusarium graminearum. ΔFgNth, ΔFgBmh1, and ΔFgBmh2 displayed great growth defects and their peripheral tips hyphae generated more branches when compared with wild-type (WT) PH-1. When exposed to validamycin A as well as high osmotic and high temperature stresses, ΔFgNth, ΔFgBmh1, and ΔFgBmh2 showed more tolerance than WT. Both ΔFgNth and ΔFgBmh1 displayed reduced deoxynivalenol production but opposite for ΔFgBmh2, and all three deletion mutants showed reduced virulence on wheat coleoptiles. In addition, coimmunoprecipitation (Co-IP) experiments suggested that FgBmh1 and FgBmh2 both interact with FgNth, but no interaction was detected between FgBmh1 and FgBmh2 in our experiments. Further, validamycin A enhances the interaction between FgBmh1 and FgNth in a positive correlation under concentrations of 1 to 100 µg/ml. In addition, both high osmotic and high temperature stresses promote the interaction between FgBmh1 and FgNth. Co-IP assay also showed that neither FgBmh1 nor FgBmh2 could interact with FgPbs2, a MAPKK kinase in the high-osmolarity glycerol pathway. However, FgBmh2 but not FgBmh1 binds to the heat shock protein FgHsp70 in F. graminearum. Taken together, our results demonstrate that FgNth and FgBmh proteins are involved in growth and responses to external stresses and virulence; and validamycin enhanced the interaction between FgNth and FgBmh1in F. graminearum.

Fear of childbirth and its predictors in re-pregnant women after cesarean section: a cross-sectional multicenter study in China.

BACKGROUND: Since the implementation of China's two-child policy in 2016, the number of re-pregnant women after cesarean section has increased significantly. These women are more prone to fear of childbirth compared with primiparas due to their history of scarred uterus leading to a more complicated delivery process, which poses a great threat to their physical and mental health. However, there is currently limited research on the problem in China. The aim of this study was to assess fear of childbirth and its predictors in re-pregnant women after cesarean section in China. METHODS: A cross-sectional multicenter study was conducted in three hospitals from June 7 to December 7, 2020, in Changsha, China. Study hospitals were selected using a random sampling technique. Participants were selected using a convenience sampling technique. Three hundred fifty-eight women during the third trimester of pregnancy who were older than 18 years, having a history of CS(s), and not having major physical or mental health problems were included. Fear of childbirth and its predictors were evaluated using the Childbirth Attitude Questionnaire, the short form of the 32-item Chinese Childbirth Self-Efficacy Inventory, the Social Support Rating Scale, and the demographic-obstetric data sheet. After checking for completeness, data were exported to statistical software for analysis. Both univariate analysis and multiple linear regression analysis were computed to assess fear of childbirth and its predictors. Statistical significance was declared at a P-value of < 0.05. RESULTS: The average score of fear of childbirth was 43.76 (standard deviation = 5.27, range 17-58). Number of cesarean sections, experience with previous cesarean section, childbirth self-efficacy and social support were significantly associated with fear of childbirth (P < 0.05). CONCLUSIONS: In this study, re-pregnant women after cesarean section in China had moderate fear of childbirth, and the number of cesarean sections, experience with previous cesarean section, childbirth self-efficacy and social support were predictors of fear of childbirth. It is important for healthcare professionals to find re-pregnant women after cesarean section at high risk of fear of childbirth and provide appropriate services during pregnancy.

Activity and cell toxicology of fluazinam on Fusarium graminearum.

Fusarium graminearum is an important plant pathogen and the causal agent of Fusarium head blight (FHB). At present, the principal method of controlling FHB is through fungicides. Fluazinam is an agent with strong broad-spectrum antifungal activity and has been used to control many diseases. However, there are no reported uses of fluazinam for controlling FHB. This study reports the activity and cell toxicology mechanisms of fluazinam on the filamentous fungus F. graminearum and its effect on fungal growth and development. The activity of fluazinam was tested for 95 wild-type field strains of F. graminearum. The EC50 values (the 50% effective concentration) of fluazinam for inhibition of mycelial growth and spore germination ranged from 0.037 µg/ml to 0.179 µg/ml and from 0.039 µg/ml to 0.506 µg/ml, respectively. The fluazinam sensitivity of these strains varied in 4.9 and 13.0 folds, implying that the target of the fungicide remained unchanged. After treatment with 0.3 µg/ml (≈EC90) fluazinam, the production of conidia was reduced, and the cell wall and cell membrane had shrunked; the cell nucleus and septum morphology, cell membrane permeability, and sexual development were not affected. When treated with 0.1 µg/ml (≈EC50) or 0.3 µg/ml fluazinam, the mycelial respiration and deoxynivalenol (DON) synthesis of F. graminearum were decreased. Confocal images showed that the formation of toxisomes was disturbed after fluazinam treatment, suggesting that fluazinam reduces DON synthesis by inhibiting toxisome formation. Infection of wheat coleoptiles revealed that fluazinam had a strong protective activity against F. graminearum. At 250 µg/ml fluazinam the control efficacy of protective treatments reached 100% and controlled strains resistant to carbendazim. These results contribute to the understanding of the mode of action of fluazinam and its application.

Molecular genetic survey and forensic characterization of Chinese Mongolians via the 47 autosomal insertion/deletion marker.

The Mongolians are mainly distributed in the modern state of Mongolia, China, Russia, and other countries. While the historic and archaeological records of the rise and fall of the Mongol Empire are well documented, little has been known about the genetic legacy of modern Mongolian populations. Here, 611 Mongolian individuals from Hohhot, Hulunbuir, and Ordos of China were genotyped via the 47 Insertion/Deletion markers. Forensically statistical parameters indicated that this InDel system could be applied to forensic investigation in Mongolian populations. The comprehensive population comparisons indicated that targeted Mongolian populations are a homogeneous population, which kept close genetic proximity with geographically northern East Asians. The findings of the model-based clustering analysis revealed a southern East Asian-specific ancestral component, which was maximized in Hainan Li, and Mongolian populations harbored relatively less Hainan Li-related ancestry and more northern East Asian-related ancestry compared with reference Tai-Kadai, Austroasiatic and Sinitic people.

Application and Prospect of RNA Profiling Analysis in Forensic Body Fluid Identification.

In forensic physical evidence identification, the accurate identification of the individual origin and their body fluid composition of the biological samples obtained from the crime scene play a critical role in determining the nature of a crime. In recent years, RNA profiling has become one of the fastest developing methods for body fluids identification. Due to the characteristics of tissue or body fluid specific expression, various types of RNA markers have been proven to be promising candidate markers for body fluids identification in previous studies. This review summarizes the research progress of RNA markers in body fluids identification, including the RNA markers that have been effectively verified in current research and their advantages and disadvantages. Meanwhile, this review prospects the application of RNA markers in forensic medicine.

Functional Roles of α1-, α2-, ß1-, and ß2-Tubulins in Vegetative Growth, Microtubule Assembly, and Sexual Reproduction of Fusarium graminearum.

The plant pathogen Fusarium graminearum contains two α-tubulin isotypes (α1 and α2) and two ß-tubulin isotypes (ß1 and ß2). The functional roles of these tubulins in microtubule assembly are not clear. Previous studies reported that α1- and ß2-tubulin deletion mutants showed severe growth defects and hypersensitivity to carbendazim, which have not been well explained. Here, we investigated the interaction between α- and ß-tubulin of F. graminearum. Colocalization experiments demonstrated that ß1- and ß2-tubulin are colocalized. Coimmunoprecipitation experiments suggested that ß1-tubulin binds to both α1- and α2-tubulin and that ß2-tubulin can also bind to α1- or α2-tubulin. Interestingly, deletion of α1-tubulin increased the interaction between ß2-tubulin and α2-tubulin. Microtubule observation assays showed that deletion of α1-tubulin completely disrupted ß1-tubulin-containing microtubules and significantly decreased ß2-tubulin-containing microtubules. Deletion of α2-, ß1-, or ß2-tubulin had no obvious effect on the microtubule cytoskeleton. However, microtubules in α1- and ß2-tubulin deletion mutants were easily depolymerized in the presence of carbendazim. The sexual reproduction assay indicates that α1- and ß1-tubulin deletion mutants could not produce asci and ascospores. These results implied that α1-tubulin may be essential for the microtubule cytoskeleton. However, our Δα1-2×α2 mutant (α1-tubulin deletion mutant containing two copies of α2-tubulin) exhibited normal microtubule network, growth, and sexual reproduction. Interestingly, the Δα1-2×α2 mutant was still hypersensitive to carbendazim. In addition, both ß1-tubulin and ß2-tubulin were found to bind the mitochondrial outer membrane voltage-dependent anion channel (VDAC), indicating that they could regulate the function of VDAC. IMPORTANCE In this study, we found that F. graminearum contains four different α-/ß-tubulin heterodimers (α1-/ß1-, α1-/ß2-, α2-/ß1-, and α2-/ß2-tubulin heterodimers), and they assemble together into a single microtubule. Moreover, α1- and α2-tubulins are functionally interchangeable in microtubule assembly, vegetative growth, and sexual reproduction. These results provide more insights into the functional roles of different tubulins of F. graminearum, which could be helpful for purification of tubulin heterodimers and development of new tubulin-binding agents.

Developmental validation of the Yfiler Platinum PCR Amplification Kit for forensic genetic caseworks and databases.

Y chromosome kits are successfully applied in cases where human biological material exists. With the development of genotyping ability, more Y chromosomal markers are needed for finer identification of male individuals and lineages. In this study, a developmental validation of a newly emerged Y chromosome kit that combines two different kinds of markers: 38 Y-STRs and 3 Y-indels are conducted. The results show that this kit has high sensitivity when there is a small amount of DNA (125 pg), more than one male (minor:major = 1:7), or a mixture of males and females (male:female = 125pg:1875pg), inhibited substances (800 µM hematin and more than 1600 ng/µL humic acid). The kit exhibits high precision level with a standard deviation of allele size no more than 0.14 nt. Locus DYS481 shows the largest stutter rate, with three stutters per true allele. Population samples are well identified (MP of 0.001106), and mutations can be observed in father-son pairs (46 mutations in 70 pairs, 10 in locus DYS627). Out of all the population samples, 13.2% belong to haplogroup M117-O2a2b1a1, with their ethnic group being Han Chinese. The results show that this kit can improve the performance of identifying male individuals, obtaining more unique haplotypes (increasing from 894 to 918 of 1000 male samples) and higher discrimination capacity (increasing from 0.942 to 0.955) in this study compared to previous widely used Yfiler Plus kit. Besides, it gives information about their paternal lineages in forensic genetic casework and genealogical database construction.

Genetic reconstruction and phylogenetic analysis by 193 Y-SNPs and 27 Y-STRs in a Chinese Yi ethnic group.

Yi is the seventh-largest ethnic group in China and features mountainous regional characteristics. The Liangshan Yi Autonomous Prefecture is the largest Yi agglomeration with isolated geographical conditions, profoundly impeding genetic communication. Here, we investigated 427 unrelated males of Liangshan from 193 Y-chromosome single nucleotide polymorphisms (Y-SNPs) and 27 Y-chromosome short tandem repeats (Y-STRs) to reveal the genetic structure and paternal phylogeny of the group. The haplogroup diversity reached 0.9169 with 46 different subhaplogroups by 193 Y-SNPs, and the haplotype diversity reached 0.9999 by 27 Y-STR loci. Multidimensional scaling (MDS), N-J tree, and Network were constructed to decipher and visualize the genetic relations between Yi and worldwide groups. Our results revealed: (1) the Network by Y-STRs and Y-SNPs showed the haplogroup D1a1a-M15 in the Liangshan Yi population was a ramification of Tibetan groups' expansion from west to east on the plateau; (2) the haplogroup distribution and the mismatch mutation analysis indicated the haplogroup O2a2b1a1a1a4a2-Z25929 of Liangshan Yi derived from manifold Southeast Asian immigrants; (3) a high-resolution Y-SNPs panel is vital to depict accurate paternal derivations and build an integrated and refining genetic structure of ethnic groups.

Paternal genetic structure of Kyrgyz ethnic group in China revealed by high-resolution Y-chromosome STRs and SNPs.

Kyrgyz ethnic group is one of the nomads in China, with the majority in Xinjiang and a small part of them living in Heilongjiang province. Historically, they have went through five migrations westward due to the wars. The name "Kyrgyz" means 40 tribes, originating from the primary groups of Kyrgyz. However, it is a largely understudied population, especially from the Y chromosome. In this study, we used a previously validated high-resolution Y-chromosome single nucleotide polymorphisms (Y-SNPs) and short tandem repeats (Y-STRs) system to study Kyrgyz ethnic group. A total of 314 male samples of Kyrgyz ethnic group were genotyped by 173 Y-SNPs and 27 Y-STRs. After data analysis, the results unveiled that Kyrgyz ethnic group was a population with high percentage of both haplogroup C2a1a3a1d∼-F10091 (91/134) and R1a1a1b2a2-Z2124 (109/134), which has never been reported. This implied that Kyrgyz ethnic group might have gone through bottleneck effects twice, with these two main lineages left. Mismatch analysis indicated that the biggest mismatch number in haplogroup C2a1a3a1d∼-F10091 was 10, while that of haplogroup R1a1a1b2a2-Z2124 was 20. This huge difference reflected the different substructure in two lineages, suggesting that haplogroup C2a1a3a1d∼-F10091 might have the least admixture compared to the other two lineages. After admixture modelling with other datasets, the conclusion could be drawn that Kyrgyz ethnic group had great genetic affinity with Punjabi from Lahore, Pakistan, which supported that Kyrgyz ethnic group in China was close to central Asian.

Exploitation of a novel slowly mutating Y-STRs set and evaluation of slowly mutating Y-STRs plus Y-SNPs typing strategy in forensic genetics and evolutionary research.

The Y-chromosome short tandem repeats (Y-STRs) loci with different mutation rates existing in the Y chromosome non-recombination region (NRY) allow to be applied in human forensics, genealogical researches, historical investigations and evolutionary studies. Currently, there is a high demand for pedigree search to narrow the scope of crime investigations. However, the commonly used Y-STRs kits generally contain Y-STRs with high mutation rates that could cause individuals from the same pedigree to display different haplotypes. Herein, we put forward a new strategy of Slowly Mutating (SM) Y-STRs plus Y-SNPs typing, which could not only improve the resolution and accuracy of pedigree search, but also be applicable to evolutionary research. First, we developed a nine SM Y-STRs assay by evaluating their mutation rates in 210 pedigrees. Then the gene diversity and efficiency of the SM Y-STRs and 172 Y-SNPs sets were investigated by 2304 unrelated males from 24 populations. Furthermore, network and time estimation analyses were performed to evaluate the new strategy's capability to reconstruct phylogenetic tree and reliability to infer the time to the most recent common ancestor (TMRCA). The nine SM Y-STRs assay even had a higher resolution and a comparable capacity of revealing population genetic differentiation compared to 172 Y-SNPs system. This new strategy could optimize the phylogenetic tree generated by commonly used Y-STR panels and obtain a quite consistent time estimations with the published dating.

ATP-Dependent Protease ClpP and Its Subunits ClpA, ClpB, and ClpX Involved in the Field Bismerthiazol Resistance in Xanthomonas oryzae pv. oryzae.

Resistance of Xanthomonas oryzae pv. oryzae, which causes rice bacterial leaf blight, to bismerthiazol has been detected in China since the 1990s. The strains resistant to bismerthiazol on rice plants were more sensitive to bismerthiazol than wild-type (WT) strains in vitro. Here, quantitative PCR was applied to detect the fold expression of adenosine triphosphate-dependent proteases, ClpP and its subunits, which withstand stresses including bactericides in bismerthiazol-resistant strains and their parental susceptible WT strain (ZJ173). Results showed that the expression of ClpP and its subunits was higher in bismerthiazol-resistant strains than in ZJ173. They were upregulated during the early growth phase and downregulated during the middle growth phase in ZJ173 treated with bismerthiazol but did not change in the resistant strains. ClpP and its subunits were overexpressed in X. oryzae pv. oryzae in this study; the higher expression of these genes increased sensitivity in vitro and increased resistance in vivo to bismerthiazol. Bismerthiazol inhibition of exopolysaccharide (EPS) production, biofilm production, and motility was also lower in ClpP and its subunits' overexpression mutants of X. oryzae pv. oryzae. The deletion mutants of ClpP and its subunits in ZJ173 decreased pathogenicity, biofilm production, swimming ability, EPS production, and growth in low-nutrient environments. Moreover, ClpP and its subunits may act downstream of the histidine utilization pathway, which could be inhibited by bismerthiazol in X. oryzae pv. oryzae. Taken together, our results indicated that ClpP and its subunits of X. oryzae pv. oryzae influenced resistance to bismerthiazol.

Temperature-Responded Biological Fitness of Carbendazim-Resistance Fusarium graminearum Mutants Conferring the F167Y, E198K, and E198L Substitutions.

Understanding the effects of temperature on Fusarium graminearum infection can provide theoretical guidance for chemical control of Fusarium head blight (FHB). Here, we evaluated the effects of various temperatures on biological fitness development of wild-type sensitive strain 2021 and carbendazim-resistance mutants conferring ß2-tubulin substitutions F167Y, E198K, and E198L. The results showed that mycelial growth and conidiation of four strains increased with the increase in temperature between 10 and 25°C. Conidia of F167Y displayed strong adaptability to low temperature. The virulence of the four strains was largely similar at the same temperature, showing an upward trend between 10 and 25°C. At 10°C, the hyphal growth of all strains was significantly inhibited, metabolism was slowed down, and the accumulation of secondary metabolites decreased. Subsequently, the production of deoxynivalenol (DON) and its intermediates pyruvate and aurofusarin decreased at low temperature, and the expression of DON biosynthesis-related genes Tri5, FgPK, and AUR decreased accordingly. At the same temperature, the aurofusarin production of the strains F167Y and E198K was higher than that of strains 2021 and E198L. The contents of DON and pyruvic acid in carbendazim-resistance mutants were higher than those in the wild-type strain 2021. The sensitivity of four strains to different fungicides changed at various temperatures. The sensitivity to most fungicides increased with decreasing temperature. The carbendazim-resistance mutants showed positive cross-resistance with other benzimidazoles. However, there was no cross-resistance to pyraclostrobin and azoles. These results would direct us to use fungicides preventing the infection of F. graminearum with changeable atmospheric temperature at the wheat flower stage.