Extracellular polymeric substances altered the physicochemical properties of molybdenum disulfide nanomaterials to mitigate its toxicity to Chlorella vulgaris.

Although the toxic effects of two-dimensional nanomaterials (2D-NMs) have been widely reported, the influence of extracellular polymeric substances (EPS) on the environmental fate and risk of 2D-NMs in aquatic environments is largely unknown, and the processes and mechanisms involved remain to be revealed. Herein, we investigated the impact of EPS secreted by microalgae (Chlorella vulgaris (C. vulgaris)) on the environmental transformation and risk of molybdenum disulfide (MoS2). We found that the attachment of EPS increased the thickness of MoS2 (from 2 nm to 5 nm), changed it from a monolayer sheet to a fuzzy multilayer structure, and promoted the formation of defects on MoS2. The blue-shift of the peak associated with the plasmon resonances in the 1 T phase and the generation of electron-hole pairs suggested that EPS altered the surface electronic structure of MoS2. EPS interacted mainly with the S atoms on the 1 T phase, and the attachment of EPS promoted the oxidation of MoS2. The reduction in hydrodynamic diameter (Dh) and the decrease in zeta potential indicated that EPS inhibited the agglomeration behavior of MoS2 and enhanced its dispersion and stability in aqueous media. Notably, EPS reduced the generation of free radicals (superoxide anion (•O2-), singlet oxygen (1O2), and hydroxyl radicals (•OH-)). Furthermore, EPS mitigated the toxicity of MoS2 to C. vulgaris, such as attenuated reduction in biomass and chlorophyll content. Compared to pristine MoS2, MoS2 + BG11 + EPS exhibited weaker oxidative stress, membrane damage and lipid peroxidation. The adsorption of EPS on MoS2 surface reduced the attachment sites of MoS2, making MoS2 less likely to be enriched on the cell surface. The findings have significant contribution for understanding the interactions between EPS and MoS2 in aquatic ecosystems, providing scientific guidance for risk assessment of 2D-NMs.

A Novel Methylation Marker NRN1 plus TERT and FGFR3 Mutation Using Urine Sediment Enables the Detection of Urothelial Bladder Carcinoma.

BACKGROUND: Aberrant DNA methylation is an early event during tumorigenesis. In the present study, we aimed to construct a methylation diagnostic tool using urine sediment for the detection of urothelial bladder carcinoma, and improved the diagnostic performance of the model by incorporating single-nucleotide polymorphism (SNP) sites. METHODS: A three-stage analysis was carried out to construct the model and evaluate the diagnostic performance. In stage I, two small cohorts from Xiangya hospital were recruited to validate and identify the detailed regions of collected methylation biomarkers. In stage II, proof-of-concept study cohorts from the Hunan multicenter were recruited to construct a diagnostic tool. In stage III, a blinded cohort comprising suspicious UBC patients was recruited from Beijing single center to further test the robustness of the model. RESULTS: In stage I, single NRN1 exhibited the highest AUC compared with six other biomarkers and the Random Forest model. At the best cutoff value of 5.16, a single NRN1 biomarker gave a diagnosis with a sensitivity of 0.93 and a specificity of 0.97. In stage II, the Random Forest algorithm was applied to construct a diagnostic tool, consisting of NRN1, TERT C228T and FGFR3 p.S249C. The tool exhibited AUC values of 0.953, 0.946 and 0.951 in training, test and all cohorts. At the best cutoff value, the model resulted in a sensitivity of 0.871 and a specificity of 0.947. In stage III, the diagnostic tool achieved a good discrimination in the external validation cohort, with an overall AUC of 0.935, sensitivity of 0.864 and specificity of 0.895. Additionally, the model exhibited a superior sensitivity and comparable specificity compared with conventional cytology and FISH. CONCLUSIONS: The diagnostic tool exhibited a highly specific and robust performance. It may be used as a replaceable approach for the detection of UBC.

Mechanisms of ZnO Nanoparticles Enhancing Phototransformation of Biologically Derived Organic Phosphorus in Aquatic Environments.

Zinc oxide nanoparticles (ZnO NPs), as the highly efficient photocatalysts, could enhance the transformation of biogenic organic phosphorus (OP) to orthophosphate (PO43-) by photodegradation, accelerating eutrophication. Conversely, orthophosphate can also transform ZnO NPs and thus potentially alter their catalytic and chemical properties. Here, we investigated the transformation mechanisms of three biogenic OP compounds and ZnO NPs under ultraviolet light (UV) illumination: inositol phosphates (IHPs), nucleic acids (DNA), and aminoethylphosphonic acid (AEP). The physicochemical characteristics of the resulting products were systematically characterized. Results show that ZnO NPs accelerated the transformation of IHPs, DNA, and AEP to inorganic phosphorus with the direct photolysis efficiencies of 98.14, 87.68, and 51.76%, respectively. The main component of the precipitates remained ZnO NPs, and Zn3(PO4)2 was identified. Zinc phytate was determined in the ZnO NP-IHP system. 31P NMR and FTIR further confirmed that the aquatic phase contained orthophosphate. Photoproduced hydroxyl (·OH) and superoxide (·O2-) were proved to play a dominant role in the OP photomineralization. Furthermore, ZnO NPs significantly enhanced the intensity of ·OH and ·O2- compared to the OP and Zn2+ solution alone. This work explored the light-induced mineralization processes of OP with ZnO NPs indicating that nanophotocatalysts may play a positive role in transformation of OP species in aquatic environments to further contribute to eutrophication.

Nanoparticles and antibiotics stress proliferated antibiotic resistance genes in microalgae-bacteria symbiotic systems.

The comprehensive effect of exogenous pollutants on the dispersal and abundance of antibiotic-resistance genes (ARGs) in the phycosphere, bacterial community and algae-bacteria interaction remains poorly understood. We investigated community structure and abundance of ARGs in free-living (FL) and particle-attached (PA) bacteria in the phycosphere under nanoparticles (silver nanoparticles (AgNPs) and hematite nanoparticles (HemNPs)) and antibiotics (tetracycline and sulfadiazine) stress using high-throughput sequencing and real-time quantitative PCR. Meanwhile, the intrinsic connection of algae-bacteria interaction was explored by transcriptome and metabolome. The results showed that the relative abundance of sulfonamide and tetracycline ARGs in PA and FL bacteria increased 103-129 % and 112-134 %, respectively, under combined stress of nanoparticles and antibiotics. Antibiotics have a greater effect on ARGs than nanoparticles at environmentally relevant concentrations. Proteobacteria, Firmicutes, and Bacteroidetes, as the primary potential hosts of ARGs, were the dominant phyla. Lifestyle, i.e., PA and FL, significantly determined the abundance of ARGs and bacterial communities. Moreover, algae can provide bacteria with nutrients (carbohydrates and amino acids), and can also produce antibacterial substances (fatty acids). This algal-bacterial interaction may indirectly affect the distribution and abundance of ARGs. These findings provide new insights into the distribution and dispersal of ARGs in microalgae-bacteria symbiotic systems.

Combined Effects of Warming and Imidacloprid on Survival, Reproduction and Population Growth of Brachionus calyciflorus (Rotifera).

Global warming and pesticide contamination are two stressors of high concern, but their combined effects on freshwater biota are controversial. This study investigated the combined effects of warming and imidacloprid (IMI) on survival (measured as life expectancy at hatching), reproduction (net reproductive rate), population growth (intrinsic rate of population increase) and sexual reproduction (proportion of sexual offspring) of Brachionus calyciflorus using a life table experiment. The results showed that compared with controls, treatments with IMI at 50-100 mg/L significantly decreased survival, reproduction and population growth of the rotifers at 20℃. The inhibiting effect at higher IMI concentrations on survival increased with increasing temperatures, but those on reproduction and population growth increased only when the temperature increased from 25℃ to 30℃. The proportion of sexual offspring decreased with increasing temperatures. When monitoring the ecological effects of pollutants, environmental temperature and the possible adaptation of rotifers to it should be taken into consideration.

Urine Cellular DNA Point Mutation and Methylation for Identifying Upper Tract Urinary Carcinoma.

BACKGROUND: To improve the selection of patients for ureteroscopy, avoid excessive testing and reduce costs, we aimed to develop and validate a diagnostic urine assay for upper tract urinary carcinoma (UTUC). METHODS: In this cohort study we recruited 402 patients from six Hunan hospitals who underwent ureteroscopy for hematuria, including 95 patients with UTUC and 307 patients with non-UTUC findings. Midstream morning urine samples were collected before ureteroscopy and surgery. DNA was extracted and qPCR was used to analyze mutations in TERT and FGFR3 and the methylation of NRN1. In the training set, the random forest algorithm was used to build an optimal panel. Lastly, the Beijing cohort (n = 76) was used to validate the panel. RESULTS: The panel combining the methylation with mutation markers led to an AUC of 0.958 (95% CI: 0.933-0.975) with a sensitivity of 91.58% and a specificity of 94.79%. The panel presented a favorable diagnostic value for UTUC vs. other malignant tumors (AUC = 0.920) and UTUC vs. benign disease (AUC = 0.975). Furthermore, combining the panel with age revealed satisfactory results, with 93.68% sensitivity, 94.44% specificity, AUC = 0.970 and NPV = 98.6%. In the external validation process, the model showed an AUC of 0.971, a sensitivity of 95.83% and a specificity of 92.31, respectively. CONCLUSIONS: A novel diagnostic model for analyzing hematuria patients for the risk of UTUC was developed, which could lead to a reduction in the need for invasive examinations. Combining NRN1 methylation and gene mutation (FGFR3 and TERT) with age resulted in a validated accurate prediction model.

Micropeptide vsp21 translated by Reovirus circular RNA 000048 attenuates viral replication.

To date, some DNA viruses and single-stranded RNA viruses have been found to generate circRNAs. However, the reports on circRNAs produced by double-stranded RNA viruses are very limited. In this study, Bombyx mori cypovirus (BmCPV), a typical double-stranded RNA virus belonging to the Reoviridae, was demonstrated to generate viral circRNAs (vcircRNAs) and a vcircRNA_000048 whose sequence corresponds with the region 164-1245 nt on the BmCPV genomic dsRNA S5 segment (GQ294468.1) was validated by PCR, Sanger sequencing, reverse transcription-rolling circle amplification, and Northern blotting. Furthermore, we verified that vcircRNA_000048 translates a micropeptide vsp21 with 21 amino acid residues in an IRES-dependent manner, and vsp21 attenuates the viral replication. These findings provided a novel clue to understanding the regulation of viral multiplication and interaction of reovirus with the host.

ß-Arrestin 2 acts an adaptor protein that facilitates viral replication in silkworm.

ß-Arrestin 2 is known to be a widely distributed adaptor protein in mammals but its function has never been reported in Lepidoptera insects. Herein, the ß-Arrestin 2 (BmArrestin 2) gene from silkworm was cloned and characterized. The spatiotemporal expression level of BmArrestin 2 was highest in the gonads at the 3rd day of 5th instar, whereas the highest and lowest abundance of BmArrestin 2 were identified in the tracheal and testis, respectively. BmArrestin 2 is mainly distributed in the cytoplasm. Furthermore, in BmN cells，overexpression of BmArrestin 2 promoted Bombyx mori nucleopolyhedrovirus (BmNPV) and B. mori cytoplasmic polyhedrosis virus (BmCPV) replication as the increment of the concentration of plasmid transfection, whereas silencing the gene with specific siRNA inhibited viral replication. Replication of BmNPV and BmCPV also was weakened using BmArrestin 2 antiserum as the increment of the concentration. Immunofluorescent staining revealed the invasion of recombinant BmNPV or BmCPV was decreased after blocking endogenous BmArrestin 2. On the other hand, BmArrestin 2 co-localizes with recombinant BmNPV and BmCPV virions in BmN cells. These results suggest that BmArrestin 2 may represent a novel target for antiviral strategies, as it is an adaptor protein that plays a key role in virus replication.

Transcriptomics and Metabolomics Revealed the Biological Response of Chlorella pyrenoidesa to Single and Repeated Exposures of AgNPs at Different Concentrations.

Increased release of engineered nanoparticles (ENPs) from widely used commercial products has threatened environmental health and safety, particularly the repeated exposures to ENPs with relatively low concentration. Herein, we studied the response of Chlorella pyrenoidesa (C. pyrenoidesa) to single and repeated exposures to silver nanoparticles (AgNPs). Repeated exposures to AgNPs promoted chlorophyll a and carotenoid production, and increased silver accumulation, thus enhancing the risk of AgNPs entering the food chain. Notably, the extracellular polymeric substances (EPS) content of the 1-AgNPs and 3-AgNPs groups were dramatically increased by 119.1% and 151.5%, respectively. We found that C. pyrenoidesa cells exposed to AgNPs had several significant alterations in metabolic process and cellular transcription. Most of the genes and metabolites are altered in a dose-dependent manner. Compared with the control group, single exposure had more differential genes and metabolites than repeated exposures. 562, 1341, 4014, 227, 483, and 2409 unigenes were differentially expressed by 1-0.5-AgNPs, 1-5-AgNPs, 1-10-AgNPs, 3-0.5-AgNPs, 3-5-AgNPs, and 3-10-AgNPs treatment groups compared with the control. Metabolomic analyses revealed that AgNPs altered the levels of sugars and amino acids, suggesting that AgNPs reprogrammed carbon/nitrogen metabolism. The changes of genes related to carbohydrate and amino acid metabolism, such as citrate synthase (CS), isocitrate dehydrogenase (IDH1), and malate dehydrogenase (MDH), further supported these results. These findings elucidated the mechanism of biological responses to repeated exposures to AgNPs, providing a new perspective on the risk assessment of nanomaterials.

[Community Distribution of the Rhizospheric and Endophytic Bacteria of Phragmites australis and Their Limiting Factors in Iron Tailings].

The composition of the rhizospheric and endophytic bacteria Phragmites australis in spring and summer around the Qiananling iron tailings reservoir in the Huairou District of Beijing was studied using Illumina high-throughput sequencing technology. Sequencing identified 40 phyla, and Proteobateria and Actinobacteria were the dominant phyla in all samples, accounting for more than 80% of the sequences in all samples. At the genus level, Pseudoarthrobacter was dominant. α-diversity analysis showed that species richness(Sobs, Chao) and species diversity indexes(Shannon, PD) of the rhizosphere soil microorganisms were significantly higher than those of endophytes. The maximum and minimum differences of the Sobs, Chao, Shannon, PD indexes were 1336, 1582.24, 6.48, and 81.18, respectively. ß-diversity analysis indicated that there were significant differences in the community compositions of rhizosphere soil microorganisms and root endophytes, while now notable differences were observed between samples in different seasons. In addition, the dominant strains in each sample(such as Pseudomonas, Arthrobacter, and Streptomyces) were highly resistant to heavy metals. Among the 774 genera, a total of 250 genera coexisted in four types of samples, indicating that the microbial community composition of different samples had some degree of similarity. Correlation analysis between soil physical and chemical properties showed that Ni, Fe, available phosphorus, effective sulphur, and organic matter were significantly related to the microbial communities. Through the prediction of COG function, it was found that the microbial metabolic functions(i.e., energy production and conversion, amino acid transport and metabolism, carbohydrate transport and metabolism, inorganic ion transport, and metabolism) were abundant.

Rh(III)-Catalyzed Regioselective Annulations of 3-Arylisoxazolones and 3-Aryl-1,4,2-dioxazol-5-ones with Propargyl Alcohols: Access to 4-Arylisoquinolines and 4-Arylisoquinolones.

The Rh(III)-catalyzed dual directing group assisted C-H activation/annulation of 3-arylisoxazolones with propargyl alcohols has been developed, which expands the application scope of isoxazolones in organic synthesis. This protocol also worked well with 3-aryl-1,4,2-dioxazol-5-ones to produce synthetically and biologically important 4-arylisoquinolones.

Involvement of Various Enzymes in the Physiology and Pathogenesis of Streptococcus suis.

Streptococcus suis causes severe infections in both swine and humans, making it a serious threat to the swine industry and public health. Insight into the physiology and pathogenesis of S. suis undoubtedly contributes to the control of its infection. During the infection process, a wide variety of virulence factors enable S. suis to colonize, invade, and spread in the host, thus causing localized infections and/or systemic diseases. Enzymes catalyze almost all aspects of metabolism in living organisms. Numerous enzymes have been characterized in extensive detail in S. suis, and have shown to be involved in the pathogenesis and/or physiology of this pathogen. In this review, we describe the progress in the study of some representative enzymes in S. suis, such as ATPases, immunoglobulin-degrading enzymes, and eukaryote-like serine/threonine kinase and phosphatase, and we highlight the important role of various enzymes in the physiology and pathogenesis of this pathogen. The controversies about the current understanding of certain enzymes are also discussed here. Additionally, we provide suggestions about future directions in the study of enzymes in S. suis.

The CpxAR Two-Component System Contributes to Growth, Stress Resistance, and Virulence of Actinobacillus pleuropneumoniae by Upregulating wecA Transcription.

Actinobacillus pleuropneumoniae is the pathogen of porcine contagious pleuropneumonia. In A. pleuropneumoniae, the CpxAR two-component system is essential for fitness and growth. The O-antigen protrudes from the outer membrane to the exterior of the cell, and the outer membrane serves as a barrier that helps the bacteria to survive in harsh environments. WecA, a undecaprenyl phosphate GlcNAc-1-phosphate transferase, is involved in O-antigen repeating unit biosynthesis. In this study, we investigated the role of CpxAR in the expression of wecA in A. pleuropneumoniae. Our results revealed that CpxR positively regulates wecA expression by directly binding to the putative promoter region of wecA. Wild-type, ΔcpxAR, ΔwecA, and complemented strains were investigated under serum, oxidative, and osmotic stresses. The ΔcpxAR and ΔwecA strains were more susceptible to these stresses than the wild-type, but the complemented strains showed phenotypes similar to those of the wild-type. Mice infected with the ΔcpxAR and ΔwecA strains exhibited lower mortality and bacterial loads in the lung than those infected with the wild-type or complemented strains. This study reveals that the CpxAR two-component system contributes to A. pleuropneumoniae growth, stress resistance, and virulence, by upregulating expression of wecA. Our findings provide new insight into the pathogenesis of A. pleuropneumoniae.

Identification and characterization of novel type of RNAs, circRNAs in crucian carp Carassius auratus gibelio.

Circular RNAs (circRNAs) with regulatory potency activity was identified from varieties of species. Crucian carp (Carassius auratus gibelio) is one of the most freshwater aquaculture species in China. Every year, huge economic damage to the farming was caused by the virus and bacterial infection. Until now, there is any information about circRNA reported from the Crucian carp. In this study, the expression pattern of circRNA in Crucian carp was investigated with transcriptomic analysis. The results showed that only 37 circRNAs were identified from the Crucian carp, and these circRNAs biogenesis was formed with canonical GU-AG splicing mechanism with unevenly distributed on the chromosomes. Wherein, most of the circRNAs were derived from the sense overlapping strategy. Reverse transcript PCR and Sanger sequencing data indicated that these circRNAs were existed authenticity in Crucian carp. The bioinformatics analysis indicated that circRNAs identified from the Crucian carp with potential miRNA sponge regulate the expression level of mRNAs. GO annotation and KEGG pathway analysis of these circRNAs showed that more than 20% circRNAs were related with catalytic activity and binding in the category of molecular function, and these circRNAs were enriched in 9 signaling pathways, such as, Wnt signaling pathway, MAPK signaling pathway, Ubiquitin mediated proteolysis et al. 220 mRNAs would be regulated by the circRNAs via miRNAs mediation. These target mRNAs were further analyzed with functional annotation and KEGG analysis. GO annotation analysis showed that several genes were related with function of nucleotide binding, transcription regulatory activity. KEGG pathway analysis showed that 5 genes were enriched in the pathway of Endocytosis. The circRNA-miRNA-mRNA regulation network indicated that one miRNA can link one or more circRNA and one or more mRNA. Overall, these results will not only help us to further understand the novel RNA transcripts in Crucian carp, but also provide the novel clue to investigate the interaction between host and pathogens from this novel circRNA molecule.

N 6-Methyladenosine Level in Silkworm Midgut/Ovary Cell Line Is Associated With Bombyx mori Nucleopolyhedrovirus Infection.

Bombyx mori nucleopolyhedrovirus (BmNPV) is one of the most serious pathogens in sericulture and causes huge economic loss annually. The roles of N6-methyladenosine (m6A) modification in silkworms following BmNPV infection are currently unclear. Here, methylated RNA immunoprecipitation with next-generation sequencing were applied to investigate the m6A profiles in silkworm midgut following BmNPV infection. A total of 9144 and 7384 m6A peaks were identified from the BmNPV-infected (TEST) and uninfected silkworm midguts (CON), respectively, which were distributed predominantly near stop codons. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis of common m6A peaks in nuclear genes revealed that these m6A-related transcripts were associated with crucial signaling pathways. Comparative transcriptome analysis showed that 1221 differential expressed m6A peaks were identified between TEST and CON, indicating that m6A modification is regulated following BmNPV infection. GO and KEGG pathway analysis of the differentially expressed m6A peaks showed their association with signal transduction, translation, and degradation. To understand further the effect of the m6A machinery on virus infection, expression levels of m6A-related genes were altered in silencing and overexpression experiments. Expression of viral structural protein VP39 was increased in BmN cells by siRNA-mediated depletion of methyltransferase-like (METTL) enzyme genes (BmMETTL3, BmMETTL14) and cytoplasmic YTH-domain family 3 (BmYTHDF3), while the reverse results were found after overexpression of the m6A-related enzymes in BmN cells. Overall, m6A modification might be a novel epigenetic mechanism that regulation BmNPV infection and interference with this mechanism may provide a novel antiviral strategy for preventing BmNPV disease.

Identification of Three Type II Toxin-Antitoxin Systems in Streptococcus suis Serotype 2.

Type II toxin-antitoxin (TA) systems are highly prevalent in bacterial genomes and have been extensively studied. These modules involve in the formation of persistence cells, the biofilm formation, and stress resistance, which might play key roles in pathogen virulence. SezAT and yefM-yoeB TA modules in Streptococcus suis serotype 2 (S. suis 2) have been studied, although the other TA systems have not been identified. In this study, we investigated nine putative type II TA systems in the genome of S. suis 2 strain SC84 by bioinformatics analysis and identified three of them (two relBE loci and one parDE locus) that function as typical type II TA systems. Interestingly, we found that the introduction of the two RelBE TA systems into Escherichia coli or the induction of the ParE toxin led to cell filamentation. Promoter activity assays indicated that RelB1, RelB2, ParD, and ParDE negatively autoregulated the transcriptions of their respective TA operons, while RelBE2 positively autoregulated its TA operon transcription. Collectively, we identified three TA systems in S. suis 2, and our findings have laid an important foundation for further functional studies on these TA systems.

The manganese efflux system MntE contributes to the virulence of Streptococcus suis serotype 2.

Manganese is an essential micronutrient to bacteria and plays an important role in bacterial physiology. However, an excess of manganese is extremely deleterious to the cell. The manganese efflux system is used to control intracellular manganese levels by some bacteria. In this study, we have identified a cation efflux family protein (MntE) that functions as a manganese export system in Streptococcus suis serotype 2. To investigate the role of mntE in S. suis 2, a mntE deletion mutant (ΔmntE) and the corresponding complementation strain (CΔmntE) were constructed. ΔmntE displayed similar growth compared to the wild-type and complementation strains under normal growth conditions, but was defective in medium supplemented with high concentrations of manganese. In addition, the mutant was more sensitive to oxidative stress conferred by diamide. Using a competitive-infection assay in the murine infection model, we demonstrated for the first time that MntE is involved in the virulence of S. suis 2. Collectively, our data indicate that manganese homeostasis controlled by the manganese efflux system MntE is important for the pathogenesis of S. suis 2.

Identification of four type II toxin-antitoxin systems in Actinobacillus pleuropneumoniae.

Toxin-antitoxin (TA) systems are small genetic elements that are widely prevalent in the genomes of bacteria and archaea. These modules have been identified in various bacteria and proposed to play an important role in bacterial physiology and virulence. However, their presence in the genomes of Actinobacillus species has received no attention. In this study, we describe the identification of four type II TA systems in Actinobacillus pleuropneumoniae, the causative agent of porcine pleuropneumonia. Reverse transcription PCR analysis revealed that the genes encoding the toxin and antitoxin are co-transcribed. Overexpression of each toxin inhibited the growth of Escherichia coli, and the toxic effect could be counteracted by its cognate antitoxin. The pull-down experiments demonstrated that each toxin interacts with its cognate antitoxin in vivo. The promoter activity assays showed that each antitoxin could autoregulate either positively or negatively the TA operon transcription. In addition, the APJL_0660/0659 TA system is present in half of the detected serovars of A. pleuropneumoniae, while the others are present in all. Collectively, we identified four type II TA systems in A. pleuropneumoniae, and this study has laid the foundation for further functional study of these TA systems.

Effects of the Qinghai-Tibetan Plateau uplift and environmental changes on phylogeographic structure of the Daurian Partridge (Perdix dauuricae) in China.

Data from eight microsatellite loci were used to assess the evolutionary demographic processes of Daurian Partridges from 285 individuals distributed among 23 populations throughout much of the species' distribution range in China. Phylogenetic analysis using microsatellite DNA data indicated that the species were geographically structured and were split into two distinctive phylogroups A and B, splitting partridges geographically from high plateaus vs. lowlands in northern China with high bootstrap support in phylogenetic trees. Each of cluster A and cluster B was comprised with two subgroups respectively. Cluster A was further subdivided into the Loess Plateau group and the Qinghai-Tibetan group. Two subgroups of cluster B corresponded to populations of Liupan Mountains and northern China, respectively. This subdivision was confirmed by Bayesian clustering and population assignment analyses of microsatellite genotypes which indicated strong isolation of these groups/populations. Our results suggested that the vicariance patterns of genetic structures of Daurian Partridge may have resulted from: (1) the uplift of the Qinghai-Tibetan Plateau and the Loess Plateau followed by the increased aridification and desertification in northern China in Middle Pleistocene, (2) the monsoon and the existence of Qinling Mountains and Liupan Mountains, and (3) the glacial cycles from the late of Middle Pleistocene to early of Late Pleistocene. In addition, BOTTLENECK analysis indicated that Daurian Partridge had been experienced recent evolutionary bottlenecks.