Reciprocal natural hybridization between Lycoris aurea and Lycoris radiata (Amaryllidaceae) identified by morphological, karyotypic and chloroplast genomic data.

BACKGROUND: Hybridization is considered as an important model of speciation, but the evolutionary process of natural hybridization is still poorly characterized in Lycoris. To reveal the phylogenetic relationship of two new putative natural hybrids in Lycoris, morphological, karyotypic and chloroplast genomic data of four Lycoris species were analyzed in this study. RESULTS: Two putative natural hybrids (2n = 18 = 4 m + 5t + 6st + 3 T) possessed obvious heterozygosity features of L. radiata (2n = 22 = 10t + 12st) and L. aurea (2n = 14 = 8 m + 6 T) in morphology (e.g. leaf shape and flower color), karyotype (e.g. chromosome numbers, CPD/DAPI bands, 45S rDNA-FISH signals etc.) and chloroplast genomes. Among four Lycoris species, the composition and structure features of chloroplast genomes between L. radiata and the putative natural hybrid 1 (L. hunanensis), while L. aurea and the hybrid 2, were completely the same or highly similar, respectively. However, the features of the cp genomes between L. radiata and the hybrid 2, while L. aurea and the hybrid 1, including IR-LSC/SSC boundaries, SSRs, SNPs, and SNVs etc., were significantly different, respectively. Combining the karyotypes and cp genomes analysis, we affirmed that the natural hybrid 1 originated from the natural hybridization of L. radiata (♀) × L. aurea (♂), while the natural hybrid 2 from the hybridization of L. radiata (♂) × L. aurea (♀). CONCLUSION: The strong evidences for natural hybridization between L. radiata (2n = 22) and L. aurea (2n = 14) were found based on morphological, karyotypic and chloroplast genomic data. Their reciprocal hybridization gave rise to two new taxa (2n = 18) of Lycoris. This study revealed the origin of two new species of Lycoris and strongly supported the role of natural hybridization that facilitated lineage diversification in this genus.

The Receptor-Like Cytoplasmic Kinase STRK1 Phosphorylates and Activates CatC, Thereby Regulating H2O2 Homeostasis and Improving Salt Tolerance in Rice.

Salt stress can significantly affect plant growth and agricultural productivity. Receptor-like kinases (RLKs) are believed to play essential roles in plant growth, development, and responses to abiotic stresses. Here, we identify a receptor-like cytoplasmic kinase, salt tolerance receptor-like cytoplasmic kinase 1 (STRK1), from rice (Oryza sativa) that positively regulates salt and oxidative stress tolerance. Our results show that STRK1 anchors and interacts with CatC at the plasma membrane via palmitoylation. CatC is phosphorylated mainly at Tyr-210 and is activated by STRK1. The phosphorylation mimic form CatCY210D exhibits higher catalase activity both in vitro and in planta, and salt stress enhances STRK1-mediated tyrosine phosphorylation on CatC. Compared with wild-type plants, STRK1-overexpressing plants exhibited higher catalase activity and lower accumulation of H2O2 as well as higher tolerance to salt and oxidative stress. Our findings demonstrate that STRK1 improves salt and oxidative tolerance by phosphorylating and activating CatC and thereby regulating H2O2 homeostasis. Moreover, overexpression of STRK1 in rice not only improved growth at the seedling stage but also markedly limited the grain yield loss under salt stress conditions. Together, these results offer an opportunity to improve rice grain yield under salt stress.

Deubiquitylation and stabilization of p21 by USP11 is critical for cell-cycle progression and DNA damage responses.

p21WAF1/CIP1 is a broad-acting cyclin-dependent kinase inhibitor. Its stability is essential for proper cell-cycle progression and cell fate decision. Ubiquitylation by the multiple E3 ubiquitin ligase complexes is the major regulatory mechanism of p21, which induces p21 degradation. However, it is unclear whether ubiquitylated p21 can be recycled. In this study, we report USP11 as a deubiquitylase of p21. In the nucleus, USP11 binds to p21, catalyzes the removal of polyubiquitin chains conjugated onto p21, and stabilizes p21 protein. As a result, USP11 reverses p21 polyubiquitylation and degradation mediated by SCFSKP2, CRL4CDT2, and APC/CCDC20 in a cell-cycle-independent manner. Loss of USP11 causes the destabilization of p21 and induces the G1/S transition in unperturbed cells. Furthermore, p21 accumulation mediated by DNA damage is completely abolished in cells depleted of USP11, which results in abrogation of the G2 checkpoint and induction of apoptosis. Functionally, USP11-mediated stabilization of p21 inhibits cell proliferation and tumorigenesis in vivo. These findings reveal an important mechanism by which p21 can be stabilized by direct deubiquitylation, and they pinpoint a crucial role of the USP11-p21 axis in regulating cell-cycle progression and DNA damage responses.

Elucidation and Structural Modeling of CD71 as a Molecular Target for Cell-Specific Aptamer Binding.

Pancreatic cancer is a highly lethal malignancy associated with tissues of the pancreas. Early diagnosis and effective treatment are crucial to improving the survival rate of patients with pancreatic cancer. In a previous study, we employed the cell-SELEX strategy to obtain an ssDNA aptamer termed XQ-2d with high binding affinity for pancreatic cancer. Here, we first identify CD71 as the XQ-2d-binding target. We found that knockdown of CD71 abolished the binding of XQ-2d and that the binding affinity of XQ-2d is associated with membrane-bound CD71, rather than total CD71 levels. Competitive analysis revealed that XQ-2d shares the same binding site on CD71 with transferrin (Tf), but not anti-CD71 antibody. We then used a surface energy transfer (SET) nanoruler to measure the distance between the binding sites of XQ-2d and anti-CD71 antibody, and it was about 15 nm. Furthermore, we did molecular dynamics simulation to clarify that the spatial structure of XQ-2d and Tf competitively binding to CD71. We also engineered XQ-2d-mediated targeted therapy for pancreatic cancer, using an XQ-2d-based complex for loading doxorubicin (Dox). Because CD71 is overexpressed not only in pancreatic cancer but also in a variety of tumors, our work provides a systematic novel way of studying a potential biomarker and also promising tools for cancer diagnosis and therapy, opening new doors for effective cancer theranostics.

Identification of Vigilin as a Potential Ischemia Biomarker by Brain Slice-Based Systematic Evolution of Ligands by Exponential Enrichment.

Stroke is one of the leading causes of disability and death among adults worldwide and results in numerous biochemical alterations. However, few efficient biomarkers are clinically available to diagnose stroke because of the limitations of biomarkers and their probes. In this work, we utilized frozen brain slices of middle cerebral artery occlusion (MCAO) in a mouse model of ischemia to select a specific binding aptamer, termed LCW17, by tissue-based SELEX (systematic evolution of ligands by exponential enrichment). LCW17 was enhanced in binding in ischemic brain slices compared to sham control. We identified the binding target of LCW17 as vigilin. Vigilin is increased in ischemia brain slices and exhibits enhanced release from cultured hippocampal neurons after oxygen glucose deprivation in vitro. Taken together, ischemic brain slice-based aptamer selection will enable identification of more probes and potential target molecules for diagnosis and therapy of ischemic stroke. Aptamer LCW17 and vigilin may potentially be applied to define the molecular mechanism underlying ischemic stroke, as well as its diagnosis.

Unexpected role for p19INK4d in posttranscriptional regulation of GATA1 and modulation of human terminal erythropoiesis.

Terminal erythroid differentiation is tightly coordinated with cell-cycle exit, which is regulated by cyclins, cyclin-dependent kinases, and cyclin-dependent kinase inhibitors (CDKI), yet their roles in erythropoiesis remain to be fully defined. We show here that p19INK4d, a member of CDKI family, is abundantly expressed in erythroblasts and that p19INK4d knockdown delayed erythroid differentiation, inhibited cell growth, and led to increased apoptosis and generation of abnormally nucleated late-stage erythroblasts. Unexpectedly, p19INK4d knockdown did not affect cell cycle. Rather, it led to decreased expression of GATA1 protein. Importantly, the differentiation and nuclear defects were rescued by ectopic expression of GATA1. Because the GATA1 protein is protected by nuclear heat shock protein family (HSP) member HSP70, we examined the effects of p19INK4d knockdown on HSP70 and found that p19INK4d knockdown led to decreased expression of HSP70 and its nuclear localization. The reduced levels of HSP70 are the result of reduced extracellular signal-regulated kinase (ERK) activation. Further biochemical analysis revealed that p19INK4d directly binds to Raf kinase inhibitor PEBP1 and that p19INK4d knockdown increased the expression of PEBP1, which in turn led to reduced ERK activation. Thus we have identified an unexpected role for p19INK4d via a novel PEBP1-p-ERK-HSP70-GATA1 pathway. These findings are likely to have implications for improved understanding of disordered erythropoiesis.

Receptor kinase complex transmits RALF peptide signal to inhibit root growth in Arabidopsis.

A number of hormones work together to control plant cell growth. Rapid Alkalinization Factor 1 (RALF1), a plant-derived small regulatory peptide, inhibits cell elongation through suppression of rhizosphere acidification in plants. Although a receptor-like kinase, FERONIA (FER), has been shown to act as a receptor for RALF1, the signaling mechanism remains unknown. In this study, we identified a receptor-like cytoplasmic kinase (RPM1-induced protein kinase, RIPK), a plasma membrane-associated member of the RLCK-VII subfamily, that is recruited to the receptor complex through interacting with FER in response to RALF1. RALF1 triggers the phosphorylation of both FER and RIPK in a mutually dependent manner. Genetic analysis of the fer-4 and ripk mutants reveals RIPK, as well as FER, to be required for RALF1 response in roots. The RALF1-FER-RIPK interactions may thus represent a mechanism for peptide signaling in plants.

How many proteins can be identified in a 2DE gel spot within an analysis of a complex human cancer tissue proteome?

Two-dimensional gel electrophoresis (2DE) in proteomics is traditionally assumed to contain only one or two proteins in each 2DE spot. However, 2DE resolution is being complemented by the rapid development of high sensitivity mass spectrometers. Here we compared MALDI-MS, LC-Q-TOF MS and LC-Orbitrap Velos MS for the identification of proteins within one spot. With LC-Orbitrap Velos MS each Coomassie Blue-stained 2DE spot contained an average of at least 42 and 63 proteins/spot in an analysis of a human glioblastoma proteome and a human pituitary adenoma proteome, respectively, if a single gel spot was analyzed. If a pool of three matched gel spots was analyzed this number further increased up to an average of 230 and 118 proteins/spot for glioblastoma and pituitary adenoma proteome, respectively. Multiple proteins per spot confirm the necessity of isotopic labeling in large-scale quantification of different protein species in a proteome. Furthermore, a protein abundance analysis revealed that most of the identified proteins in each analyzed 2DE spot were low-abundance proteins. Many proteins were present in several of the analyzed spots showing the ability of 2DE-MS to separate at the protein species level. Therefore, 2DE coupled with high-sensitivity LC-MS has a clearly higher sensitivity as expected until now to detect, identify and quantify low abundance proteins in a complex human proteome with an estimated resolution of about 500 000 protein species. This clearly exceeds the resolution power of bottom-up LC-MS investigations.

Overexpression of WDR79 in non-small cell lung cancer is linked to tumour progression.

WD-repeat protein 79 (WDR79), a member of the WD-repeat protein family, acts as a scaffold protein, participating in telomerase assembly, Cajal body formation and DNA double-strand break repair. Here, we first report that WDR79 is frequently overexpressed in cell lines and tissues derived from non-small cell lung cancer (NSCLC). Knockdown of WDR79 significantly inhibited the proliferation of NSCLC cells in vitro and in vivo by inducing cell cycle arrest and apoptosis. WD-repeat protein 79 -induced cell cycle arrest at the G0/G1 phase was associated with the expression of G0/G1-related cyclins and cyclin-dependent kinase complexes. We also provide evidence that WDR79 knockdown induces apoptosis via a mitochondrial pathway. Collectively, these results suggest that WDR79 is involved in the tumorigenesis of NSCLC and is a potential novel diagnostic marker and therapeutic target for NSCLC.

Recent Advances in Nondestructive Analytical Techniques for Determining the Total Soluble Solids in Fruits: A Review.

The total soluble solids play an important role in the fruit maturity process and determine the acceptance of rich nutrients as well as economic benefits in the fruit trade. Thus, development of rapid and nondestructive techniques for evaluating soluble solids content in fruits is important. This review focuses on recent advances in nondestructive techniques for soluble solids contents of fruits including hyperspectral imaging, laser light backscattering imaging, magnetic resonance imaging, X-ray computed tomography, near-infrared spectroscopy, and Raman spectroscopy. The advantages and disadvantages of these techniques are compared and discussed, and some viewpoints about future trends are also presented.

[Research on biological and genetic characteristics of human placenta mesenchymal stem cells cultured in vitro].

OBJECTIVE: To investigate the biological characteristics and genetic features of human placenta mesenchymal stem cells (hPA-MSCs) cultured in vitro in order to assess its safety for clinical use. METHODS: The shapes of the 1st, 3rd, 5th, 7th, 10th, 13th, 17th and 20th generation hPA-MSCs cultured in vitro using serum-free culture medium were observed. Their cell cycle, cell surface markers, and karyotype were analyzed, and relevant genes and cytokines were measured. RESULTS: The shape of hPA-MSCs has remained as fusiform or short fusiform, and there was no significant change. About 93% of hPA-MSCs cells were in G0/G1 phase and remained stable. No obvious chromosomal translocation, loss or inversion was noted by karyotyping analysis. Cytokines expression level remained stable. Related gene expression level as a whole was on the decline, but the gene expression level of the first five generations showed very slight variations, with genetic characteristics remaining stable. CONCLUSION: The hPA-MSCs cultured in vitro with serum-free medium has retained stable in the first five generations.

FERONIA receptor kinase interacts with S-adenosylmethionine synthetase and suppresses S-adenosylmethionine production and ethylene biosynthesis in Arabidopsis.

Environmental inputs such as stress can modulate plant cell metabolism, but the detailed mechanism remains unclear. We report here that FERONIA (FER), a plasma membrane receptor-like kinase, may negatively regulate the S-adenosylmethionine (SAM) synthesis by interacting with two S-adenosylmethionine synthases (SAM1 and SAM2). SAM participates in ethylene, nicotianamine and polyamine biosynthetic pathways and provides the methyl group for protein and DNA methylation reactions. The Arabidopsis fer mutants contained a higher level of SAM and ethylene in plant tissues and displayed a dwarf phenotype. Such phenotype in the fer mutants was mimicked by over-expressing the S-adenosylmethionine synthetase in transgenic plants, whereas sam1/2 double mutant showed an opposite phenotype. We propose that FER receptor kinase, in response to environmental stress and plant hormones such as auxin and BR, interacts with SAM synthases and down-regulates ethylene biosynthesis.

MiR-492 contributes to cell proliferation and cell cycle of human breast cancer cells by suppressing SOX7 expression.

MicroRNAs (miRNAs) have emerged as important regulators that potentially play critical roles in cancer cell biological processes. Previous studies have shown that miR-492 plays an important role in cell tumorigenesis in multiple kinds of human cancer cells. However, the underlying mechanisms of this microRNA in breast cancer remain largely unknown. In the present study, we investigated miR-492's role in cell proliferation of breast cancer. MiR-492 expression was markedly upregulated in breast cancer tissues and breast cancer cells. Overexpression of miR-492 promoted the proliferation and anchorage-independent growth of breast cancer cells. Bioinformatics analysis further revealed sex-determining region Y-box 7 (SOX7), a putative tumor suppressor, as a potential target of miR-492. Data from luciferase reporter assays showed that miR-492 directly binds to the 3'-untranslated region (3'-UTR) of SOX7 messenger RNA (mRNA) and repressed expression at both transcriptional and translational levels. Ectopic expression of miR-492 led to downregulation of SOX7 protein, which resulted in the upregulation of cyclin D1 and c-Myc. In functional assays, SOX7 silenced in miR-492-in-transfected ZR-75-30 cells has positive effect to promote cell proliferation, suggesting that direct SOX7 downregulation is required for miR-492-induced cell proliferation and cell cycle of breast cancer. In sum, these results suggest that miR-492 represents a potential onco-miR and participates in breast cancer carcinogenesis by suppressing SOX7 expression.

Rat liver sinusoidal surface N-linked glycoproteomic analysis by affinity enrichment and mass spectrometric identification.

Glycosylation in liver is one of the most biologically important protein modifications. It plays critical roles in many physiological and pathological processes by virtue of its unique location at the blood-tissue interface, including angiogenesis, liver cancer, cirrhosis, and fibrosis. To analyze glycosylation of plasma membrane proteins in liver sinusoidal endothelial cells (LSEC), N-glycopeptides of the LSEC surface were enriched using a filter-assisted sample preparation-based lectin affinity capture method and subsequently identified with mass spectrometry. In total, 225 unique N-glycosylation sites on 152 glycoproteins were identified, of which 119 (53%) sites had not previously been determined experimentally. Among the glycoproteins, 53% were classified as plasma membrane proteins and 47 (31%) as signaling proteins and receptors. Moreover, 23 cluster of differentiation antigens with 49 glycopeptides were detected within the membrane glycoproteins of the liver sinusoidal surface. Furthermore, bioinformatics analysis revealed that the majority of identified glycoproteins have an impact on processes of LSEC. Therefore, N-glycoproteomic analysis of the liver sinusoidal surface may provide useful information on liver regeneration and facilitate liver disease diagnosis.

Rab3A is a new interacting partner of synaptotagmin I and may modulate synaptic membrane fusion through a competitive mechanism.

Rab3 and synaptotagmin have been reported to be the key proteins that have opposite actions but cooperatively play critical regulatory roles in selecting and limiting the number of vesicles released at central synapses. However, the exact mechanism has not been fully understood. In this study, Rab3A and synaptotagmin I, the most abundant isoforms of Rab3 and synaptotagmin, respectively, in brain were for the first time demonstrated to directly interact with each other in a Ca(2+)-independent manner, and the KKKK motif in the C2B domain of synaptotagmin I was a key site for the Rab3A binding, which was further confirmed by the competitive inhibition of inositol hexakisphosphate. Further studies demonstrated that Rab3A competitively affected the synaptotagmin I interaction with syntaxin 1B that was involved in membrane fusion during the synaptic vesicle exocytosis. These data indicate that Rab3A is a new synaptotagmin I interacting partner and may participate in the regulation of synaptic membrane fusion and thus the vesicle exocytosis by competitively modulating the interaction of synaptotagmin with syntaxin of the t-SNARE complex in presynaptic membranes.

Protease-activated receptor-2 modulates hepatic stellate cell collagen release and apoptotic status.

The pathogenesis of hepatic fibrosis is to be further investigated. Protease-activated receptor-2 (PAR2) plays a role in hepatic fibrosis. This study aims to elucidate the role of activation of PAR2 in the regulation of hepatic stellate cell activities. In this study, the expression of PAR2, Fas and caveolin-1 in human hepatic stellate cell line, HHStec cell (HHStecs) was assessed by real time RT-PCR and Western blot. The levels of collagen were determined by enzyme-linked immunosorbent assay. The PAR2 gene was silenced in HHStecs using RNA interference. Apoptosis of HHStecs was assessed by flow cytometry. The results showed that HHStecs expressed PAR2, which was up regulated by activation with phorbol myristate acetate (PMA). Activation of PAR2 increased the release of collagen from HHStecs. Exposure to PMA induced HHStec apoptosis, which was significantly inhibited by activation of PAR2. The PAR2 activation also suppressed the expression of caveolin-1 and Fas in HHStecs. Over expression of caveolin-1 in HHStecs blocked PAR2-reduced apoptosis. We conclude that HHStecs express PAR2. Activation of PAR2 increases HHStecs to release collagen and reduces the activation-induced HHStec apoptosis, which can be inhibited by the over expression of caveolin-1.

Activated protease receptor-2 induces GATA6 expression to promote survival in irradiated colon cancer cells.

BACKGROUND AND AIMS: The resistance to irradiation is common and a great drawback in the treatment of cancer with radiotherapy; the underlying mechanism is unclear. GATA binding protein 6 (GATA6) is associated with the pathogenesis of cancer. This study aims to investigate the role of GATA6 on compromising irradiation effect on HT55 and HT29 cells, 2 colorectal cancer cell lines. METHODS: Human colon cancer cell lines, HT55 and HT29 cells, were treated with irradiation in the culture. Apoptosis of HT55 and HT29 cells was determined by flow cytometry. The expression of PAR2 and GATA6 in HT55 and HT29 cells was analyzed by real time RT-PCR and Western blotting. The gene silence and gene over expression were employed to observe the effect of GATA6 on p53 expression in HT55 and HT29 cells. RESULTS: The results showed that HT55 and HT29 cells expressed protease-activated receptor-2 (PAR2). Irradiation induced 38.6% HT55 cell and 33.8% HT29 cell apoptosis, which reduced to 4.2% and 5.6%, respectively after activation of PAR2. Exposure to irradiation increased the expression of GATA6; the latter played a critical role in suppression of p53 expression in HT55 and HT29 cells. Inhibition of GATA6 significantly increased the effect of irradiation on HT55 and HT29 cells. CONCLUSIONS: Activation of PAR2 compromises the effect of irradiation on inducing colorectal cancer cell apoptosis, which can be prevented by inhibition of GATA6 expression.

Isolation technique and proteomic analysis of the erythrocyte ghosts of red-eared turtle (Trachemys scripta).

To proceed proteomic analysis of erythrocyte of the red-eared turtle Trachemys scripta, a method for obtaining turtle erythrocyte ghosts (TEG) was first developed. After hypotonic lysis using a special buffer, forcing the erythrocyte through the syringe with an "N"-shaped needle, applying low speed homogenizing and differential centrifugation, highly purified TEG fractions were obtained. The isolated TEG proteins were treated with in-gel digestion separated by SDS-PAGE or in-solution digestion followed by HPLC predissociation, and then identified by nano-ESI-LC MS/MS techniques. A total of 169 TEG proteins was identified, validated, and categorized. Among these proteins, tubulins, and cell-surface-located F-type ATP synthase revealed important information into the super tolerance of Trachemys scripta in anoxia and low temperature exposure. Altogether, this study not only provided a method to isolate high quality TEG and a dataset of comprehensive characterization of TEG proteins, but also provides a tool for proteomic research of all nucleated red blood cells, and thus opened a new research field for exploring the mechanisms of super tolerance of turtles in harsh environment.

Combination predicting model of traffic congestion index in weekdays based on LightGBM-GRU.

Tree-based and deep learning methods can automatically generate useful features. Not only can it enhance the original feature representation, but it can also learn to generate new features. This paper develops a strategy based on Light Gradient Boosting Machine (LightGBM or LGB) and Gated Recurrent Unit (GRU) to generate features to improve the expression ability of limited features. Moreover, a SARIMA-GRU prediction model considering the weekly periodicity is introduced. First, LightGBM is used to learn features and enhance the original features representation; secondly, GRU neural network is used to generate features; finally, the result ensemble is used as the input for prediction. Moreover, the SARIMA-GRU model is constructed for predicting. The GRU prediction consequences are revised by the SARIMA model that a better prediction can be obtained. The experiment was carried out with the data collected by Ride-hailing in Chengdu, and four predicted indicators and two performance indexes are utilized to evaluate the model. The results validate that the model proposed has significant improvements in the accuracy and performance of each component.

Co-Expression of Pig IL-2 and Fusion Bovine Cathelicidin Gene by Recombinant Plasmids in Yeast and Their Promotion of Mouse Antibacterial Defense.

In order to develop an effective and safe immunomodulator to enhance the antimicrobial bioactivity and immunity of animals against infectious bacterial diseases, a recombinant plasmid pGAPZαA-IL2-B co-expressing pig interleukin-2 (PIL-2) and fused bovine cathelicidin (FBC) genes were constructed using the 2A self-cleavage technique. After being expressed in Pichia pastoris strain SMD1168, the recombinant yeast was administered orally to 5-week-old female ICR mice. The control mice were similarly dosed with P. pastoris with a blank plasmid or FBC recombinant plasmid alone. At 28 days post-treatment, the mice were challenged intraperitoneally with virulent strains of either E. coli or S. aureus. Compared with the control groups, the mice that received recombinant yeast co-expressing PIL-2/FBC manifested significant increases in the number of leukocytes, CD4+ and CD8+ T cells, IgG, and the gene expressions of TLRs(TLR1,4,6,9), antimicrobial peptides(CRP4 and CRAMP) and cytokines (IL-2, 4, 6, 7, 12, 15, 23, IFN-γ, and TNF-α) in the blood. Furthermore, the treated mice displayed significantly higher survival than the other two control groups after the challenge. These results suggest that the antimicrobial activity and immunity of animals can be effectively enhanced by the in vivo co-expression of IL-2 and the FBS gene, which can facilitate the development of new immunopotentiation molecules to overcome the infection of antibiotic-resistant bacteria.