Transcription factor shapes chromosomal conformation and regulates gene expression in bacterial adaptation.

Genomic mutations allow bacteria to adapt rapidly to adverse stress environments. The three-dimensional conformation of the genome may also play an important role in transcriptional regulation and environmental adaptation. Here, using chromosome conformation capture, we investigate the high-order architecture of the Zymomonas mobilis chromosome in response to genomic mutation and ambient stimuli (acetic acid and furfural, derived from lignocellulosic hydrolysate). We find that genomic mutation only influences the local chromosome contacts, whereas stress of acetic acid and furfural restrict the long-range contacts and significantly change the chromosome organization at domain scales. Further deciphering the domain feature unveils the important transcription factors, Ferric uptake regulator (Fur) proteins, which act as nucleoid-associated proteins to promote long-range (>200 kb) chromosomal communications and regulate the expression of genes involved in stress response. Our work suggests that ubiquitous transcription factors in prokaryotes mediate chromosome organization and regulate stress-resistance genes in bacterial adaptation.

Gene identification and semisynthesis of the anti-inflammatory oleanane-type triterpenoid wilforlide A.

Wilforlide A is one of the main active constituents produced in trace amounts in Tripterygium wilfordii Hook F, which has excellent anti-inflammatory and immune suppressive effects. Despite the seeming structural simplicity of the compound, the biosynthetic pathway of wilforlide A remains unknown. Gene-specific expression analysis and genome mining were used to identify the gene candidates, and their functions were studied in vitro and in vivo. A modularized two-step (M2S) technique and CRISPR-Cas9 methods were used to construct engineering yeast. Here, we identified a cytochrome P450, TwCYP82AS1, that catalyses C-22 hydroxylation during wilforlide A biosynthesis. We also found that TwCYP712K1 to K3 can further oxidize the C-29 carboxylation of oleanane-type triterpenes in addition to friedelane-type triterpenes. Reconstitution of the biosynthetic pathway in engineered yeast increased the precursor supply, and combining TwCYP82AS1 and TwCYP712Ks produced abrusgenic acid, which was briefly acidified to achieve the semisynthesis of wilforlide A. Our work presents an alternative metabolic engineering approach for obtaining wilforlide A without relying on extraction from plants.

[Association between auditory processing and problem behaviors in preschool children: the mediating role of executive function]. / å¬å¤„ç†ä¸Žå­¦é¾„å‰å„¿ç«¥é—®é¢˜è¡Œä¸ºçš„å ³ç³»ï¼šæ‰§è¡ŒåŠŸèƒ½çš„ä¸­ä»‹ä½œç”¨.

OBJECTIVES: To investigate the association between auditory processing and problem behaviors in preschool children, as well as the mediating role of executive function. METHODS: A total of 2 342 preschool children were selected from 7 kindergartens in Nanjing, China from June to August 2021. They were evaluated using Preschool Auditory Processing Assessment Scale, Conners Parent Symptom Questionnaire, and Behavior Rating Inventory of Executive Functioning-Preschool version. Children with different demographic features were compared in the scores and the abnormality rates of auditory processing, problem behaviors, and executive function. The influencing factors of the total scores of auditory processing, problem behaviors, and executive function were evaluated using multiple linear regression analysis. Whether executive function was a mediating factor between auditory processing and executive function was examined. RESULTS: Sex and grade were the main influencing factors for the total score of auditory processing (P<0.05), and sex, grade, parental education level, and family economic status were the main influencing factors for the total scores of problem behaviors and executive function (P<0.05). The auditory processing score (rs=0.458, P<0.05) and problem behavior score (rs=0.185, P<0.05) were significantly positively correlated with the executive function score, and the auditory processing score was significantly positively correlated with the problem behavior score (rs=0.423, P<0.05). Executive function played a partial mediating role between auditory processing and problem behaviors, and the mediating effect accounted for 33.44% of the total effect. CONCLUSIONS: Auditory processing can directly affect the problem behaviors of preschool children and indirectly affect problem behaviors through executive function.

Molecular mechanism of engineered Zymomonas mobilis to furfural and acetic acid stress.

Acetic acid and furfural (AF) are two major inhibitors of microorganisms during lignocellulosic ethanol production. In our previous study, we successfully engineered Zymomonas mobilis 532 (ZM532) strain by genome shuffling, but the molecular mechanisms of tolerance to inhibitors were still unknown. Therefore, this study investigated the responses of ZM532 and its wild-type Z. mobilis (ZM4) to AF using multi-omics approaches (transcriptomics, genomics, and label free quantitative proteomics). Based on RNA-Seq data, two differentially expressed genes, ZMO_RS02740 (up-regulated) and ZMO_RS06525 (down-regulated) were knocked out and over-expressed through CRISPR-Cas technology to investigate their roles in AF tolerance. Overall, we identified 1865 and 14 novel DEGs in ZM532 and wild-type ZM4. In contrast, 1532 proteins were identified in ZM532 and wild-type ZM4. Among these, we found 96 important genes in ZM532 involving acid resistance mechanisms and survival rates against stressors. Furthermore, our knockout results demonstrated that growth activity and glucose consumption of mutant strains ZM532∆ZMO_RS02740 and ZM4∆ZMO_RS02740 decreased with increased fermentation time from 42 to 55 h and ethanol production up to 58% in ZM532 than that in ZM532∆ZMO_RS02740. Hence, these findings suggest ZMO_RS02740 as a protective strategy for ZM ethanol production under stressful conditions.

[Application of Preschool Auditory Processing Assessment Scale in children with attention deficit hyperactivity disorder]. / å­¦é¾„å‰å„¿ç«¥å¬å¤„ç†è¯„ä¼°é‡è¡¨åœ¨æ³¨æ„ç¼ºé™·å¤šåŠ¨éšœç¢å„¿ç«¥ä¸­çš„åº”ç”¨ç ”ç©¶.

OBJECTIVES: To investigate the characteristics of auditory processing (AP) in preschool children with attention deficit hyperactivity disorder (ADHD) using Preschool Auditory Processing Assessment Scale (hereafter referred to as "auditory processing scale"). METHODS: A total of 41 children with ADHD and 41 typically developing (TD) children were assessed using the auditory processing scale, SNAP-IV rating scale, and Conners' Kiddie Continuous Performance Test (K-CPT). The auditory processing scale score was compared between the TD and ADHD groups. The correlations of the score with SNAP-IV and K-CPT scores were assessed. RESULTS: Compared with the TD group, the ADHD group had significantly higher total score of the auditory processing scale and scores of all dimensions except visual attention (P<0.05). In the children with ADHD, the attention deficit dimension score of the SNAP-IV rating scale was positively correlated with the total score of the auditory processing scale (rs30=0.531, P<0.05; rs27=0.627, P<0.05) as well as the scores of its subdimensions, including auditory decoding (rs=0.628, P<0.05), auditory attention (rs=0.492, P<0.05), and communication (rs=0.399, P<0.05). The hyperactivity-impulsivity dimension score of the SNAP-IV rating scale was positively correlated with the hyperactivity-impulsivity dimension score of the auditory processing scale (rs=0.429, P<0.05). In the children with ADHD, the attention deficit dimension score of the K-CPT was positively correlated with the total score (rs30=0.574, P<0.05; rs27=0.485, P<0.05) and the hyperactivity-impulsivity dimension score (rs=0.602, P<0.05) of the auditory processing scale. CONCLUSIONS: Preschool children with ADHD have the risk of AP abnormalities, and the auditory processing scale should be used early for the screening and evaluation of AP abnormalities in children.

Enhancing Secretion of Endoglucanase in Zymomonas mobilis by Disturbing Peptidoglycan Synthesis.

Zymomonas mobilis (Z. mobilis) is a potential candidate strain for consolidated bioprocessing (CBP) in lignocellulosic biorefinery. However, the low-level secretion of cellulases limits this CBP process, and the mechanism of protein secretion that is affected by cell wall peptidoglycan is also not well understood. Here, we constructed several penicillin-binding protein (PBP)-deficient strains derived from Z. mobilis S192 to perturb the cell wall peptidoglycan network and then investigated the effects of peptidoglycan on the endoglucanase secretion. The results showed that extracellular recombinant endoglucanase production was significantly enhanced in PBP mutant strains, notably, Δ1089/0959 (4.09-fold) and Δ0959 (5.76-fold) in comparison to parent strains. For PBP-deficient strains, the growth performance was not significantly inhibited, but cell morphology was altered. In addition, enhanced antibiotic sensitivity and reduced inhibitor tolerance were also detected in our study. The concentration of intracellular soluble peptidoglycan was increased, especially for single-gene deletion. Outer membrane permeability of PBP-deficient strains was also improved, notably, Δ1089/0959 (1.14-fold) and Δ0959 (1.07-fold), which might explain the increased endoglucanase extracellular secretion. Our findings indicated that PBP-deficient Z. mobilis was capable of increasing endoglucanase extracellular secretion via cell wall peptidoglycan disturbance, and it will provide a foundation for the development of CBP technology in Z. mobilis in the future. IMPORTANCE Cell wall peptidoglycan has the function to maintain cell robustness and acts as the barrier to secret recombinant proteins from the cytoplasm to extracellular space in Z. mobilis and other bacteria. Herein, we perturbed the peptidoglycan synthesis network via knocking out PBPs (ZMO0197, ZMO0959, ZMO1089) to enhance recombinant endoglycanase extracellular secretion in Z. mobilis S912. This study could lay the foundation for understanding the regulatory network of cell wall synthesis and guide the construction of CBP strains in Z. mobilis.

Metabolic engineering of Zymomonas moblis for ethylene production from straw hydrolysate.

Biological ethylene production is a promising sustainable alternative approach for fossil-based ethylene production. The high glucose utilization of Z. mobilis makes it as a promising bioethylene producer. In this study, Zymomonas mobilis has been engineered to produce ethylene through the introduction of the synthetic ethylene-forming enzyme (EFE). We also investigated the effect of systematically knocking out the competitive metabolic pathway of pyruvate in an effort to improve the availability of pyruvate for ethylene production in Z. mobilis expressing EFE. Guided by these results, we tested a number of conjectures that could improve the α-ketoglutarate supply. Optimization of these pathways and different substrate supplies resulted in a greater production of ethylene (from 1.36 to 12.83 nmol/OD600/mL), which may guide future engineering work on ethylene production using other organisms. Meanwhile, we achieved an ethylene production of 5.8 nmol/OD600/mL in the ZM532-efe strain using enzymatic straw hydrolysate of corn straw as the sole carbon source. As a preferred host in biorefinery technologies using lignocellulosic biomass as feedstock, heterologous expression of EFE in Z. mobilis converts the non-ethylene producing strain into an ethylene-producing one using a metabolic engineering approach, which is of great significance for the utilization of cellulosic biomass in the future. KEY POINTS: • Heterologous expression of EFE in Z. mobilis successfully converted the non-ethylene producing strain into an ethylene producer (1.36 nmol/OD600/mL). Targeted modifications of the central carbon metabolism can effectively improve ethylene production (peak production: 8.3 nmol/OD600/mL). • The addition of nutrients to the medium can further increase the production of ethylene (peak production: 12.8 nmol/OD600/mL). • The ZM532-efe strain achieved an ethylene production of 5.8 nmol/OD600/mL when enzymatic hydrolysate of corn straw was used as the sole carbon source.

Self-construal modulates context-based social comparison preferences in outcome evaluations.

Self-construal (SC) describes how people perceive the relationship between themselves and others and is usually divided into interdependent and independent types. Several studies have been conducted on how people with independent and interdependent SC process their own and others' outcomes. However, few studies have investigated the influence of SC on outcome evaluation in a social comparison context. To explore this, we randomly assigned participants to interdependent and independent SC priming groups and analyzed the affects and electrophysiological responses generated when they played gambling games with two pseudo-players. The results showed that self-gambling state, SC, and social comparison interacted to influence feedback-related negativity (FRN). In the self-win condition, performances that differed from others elicited more negative FRN than evenness for both the interdependent and independent groups. In the self-loss condition, this effect was only found in the independent group. These results suggest that the outcome evaluation patterns in social contexts are not fixed but vary according to self-gambling state and SC.

Features of Cognitive Ability and Central Auditory Processing of Preschool Children With Minimal and Mild Hearing Loss.

OBJECTIVE: This study aimed to investigate the current status of cognitive development and central auditory processing development of preschool children with minimal and mild hearing loss (MMHL) in Nanjing, China. METHOD: We recruited 34 children with MMHL and 45 children with normal hearing (NH). They completed a series of tests, including cognitive tests (i.e., Wechsler Preschool and Primary Scale of Intelligence and Continuous Performance Test), behavioral auditory tests (speech-in-noise [SIN] test and frequency pattern test), and objective electrophysiological audiometry (speech-evoked auditory brainstem response and cortical auditory evoked potential). In addition, teacher evaluations and demographic information and questionnaires completed by parents were collected. RESULTS: Regarding cognitive ability, statistical differences in the verbal comprehensive index, full-scale intelligence quotient, and abnormal rate of attention test score were found between the MMHL group and the NH group. The children with MMHL performed poorer on the SIN test than the children with NH. As for the auditory electrophysiology of the two groups, the latency and amplitude of some waves of the speech-evoked auditory brainstem response and cortical auditory evoked potential were statistically different between the two groups. We attempted to explore the relationship between some key indicators of auditory processing and some key indicators of cognitive development. CONCLUSIONS: Children with MMHL are already at increased developmental risk as early as preschool. They are more likely to have problems with attention and verbal comprehension than children with NH. This condition is not compensated with increasing age during the preschool years. The results suggest a possible relationship between the risk of cognitive deficit and divergence of auditory processing. SUPPLEMENTAL MATERIAL: https://doi.org/10.23641/asha.22670473.

Development and Evaluation of Orthographic Knowledge Awareness Scale for Children Aged 6-12 Years.

Objective: This study primarily aimed to develop an orthographic knowledge awareness scale in Mandarin for children aged 6-12 years. Related factors affecting orthographic knowledge awareness in children were analyzed, and a basis for individualized intervention was provided to improve reading and writing. Methods: A conceptual framework for orthographic knowledge awareness in children aged 6-12 years was determined through a detailed reading of the literature on Chinese character orthography, combined with qualitative interviews of the target population and consultation with experts. The orthographic knowledge awareness scale initially consisted of three versions: for grades 1-2 (210 items), grades 3-4 (207 items), and grades 5-6 (220 items), accumulating a total of 637 items. The initial scale was then used for the study involving children aged 6-12 years in Maanshan City, Jiangsu Province. Various approaches to screening items were comprehensively used to determine the formal version of the orthographic knowledge awareness scale. The official scale was ultimately used to conduct the third round of surveys among 1,354 children aged 6-12 years in ordinary primary schools located in 5 cities in Jiangsu Province, namely, Changzhou, Lianyungang, Nantong, Xuzhou, and Yangzhou. The reliability, validity, and discriminating power of the formal scale were evaluated. Results: A total of 360 items were included in the formal version of the orthographic knowledge awareness scale. The formal scale was divided into three versions for grades 1-2, 3-4, and 5-6. Each grade version consisted of 120 items. The scale was composed of the stroke awareness test, radical awareness test, and left-right reversal test. The cumulative variance contribution rates of grades 1-2, 3-4, and 5-6 were 82.47, 61.71, and 64.19%, respectively. The Cronbach's α coefficients of the three-grade version of the scale were 0.989, 0.946, and 0.938; the split-half reliability coefficients were 0.925, 0.766, and 0.847; and the test-retest reliability coefficients were 0.847, 0.895, and 0.8928, respectively. Conclusion: The proposed orthographic knowledge awareness scale for children aged 6-12 years exhibits good reliability and validity. The formal scale consisted of two dimensions: identification of left-right reversal at the stroke and radical levels and the left-right reversal at the whole character level. The two dimensions can more comprehensively reflect the ability of children to discriminate orthographic structures.

The development, validity, reliability, and norm of a preschool auditory processing assessment scale in China.

BACKGROUND: Children with auditory processing deficits may face problems with language, learning, and social communication. AIMS: To develop a Chinese auditory processing assessment scale for preschool children and establish the norms of the scale. METHODS AND PROCEDURES: The predictive version of the scale was formed by a literature review, qualitative interviews, expert consultation, and a pre-test with a small sample. Nine kindergartens in Nanjing were selected by a stratified cluster sampling plan. First, 734 children from two kindergartens were selected for the large sample pre-test of the scale. Then, 1526 children from four kindergartens and 1151 children from three kindergartens were selected for the reliability and validity analysis and confirmatory factor analysis, respectively. The standardized norm data of the scale were established based on the 3411 points of scale data of the nine kindergartens. Finally, the clinical usefulness of the scale was analyzed by comparing the results of objective auditory processing tests in children with normal and abnormal auditory processing prompted by the score on the scale. OUTCOMES AND RESULTS: The preschool auditory processing assessment scale includes 5 dimensions and 30 items. The Cronbach's alpha value of the scale is greater than 0.9. The confirmatory factor analysis results verify that the scale structure is reasonable. The percentile norm of the scale was established. The results of electrophysiological tests of the normal and abnormal auditory processing groups were statistically different (P < 0.05). CONCLUSIONS AND IMPLICATIONS: The developed preschool auditory processing assessment scale has good reliability and validity. The scale is suitable for clinical application.

Establishing Normative Data for the Number Cancelation Test Among Children in Kindergartens and Primary Schools in China.

This study aimed to suggest an attention assessment tool using a Digital Pen for measuring the temporal-spatial parameters during the Number Cancelation Test (NCT), and then to establish the normative data for the NCT among children in kindergartens and primary schools in China by recruiting a total of 989 children (496 males). Four measures, i.e., selective attention (SA), speed of cognitive processing (SpC), averaged time of circlings (ATC), and averaged circumference of circled curves (ACCC), were proposed to evaluate the NCT performance. They basically have a development trend with fast speed in the beginning before Grade 1 or 2 of primary schools, and then enter an extremely slow development period (with ceiling or floor effect). SA and SpC have gender and grade main effects, while ATC and ACCC have the grade main effect, only. In particular, females have higher SA scores than males in middle class of kindergarten, and Grade 2-Grade 5 of primary school, but no gender differences in other grades; females have higher SpC scores than males in middle class of kindergarten, and Grade 3-4 of primary school, but no gender differences in other grades. More importantly, in clinical practice, if SA or SpC measure of a child is below than the 5th centile (i.e., p5 level) of his/her grade-specific normative data, then this child may be predicted to have a high-risk of learning disabilities. Findings suggest that the proposed method can be used for early screening of learning disabilities by setting appropriate cut-off values.

Development, Reliability, and Validity of the Preschool Learning Skills Scale: A Tool for Early Identification of Preschoolers at Risk of Learning Disorder in Mainland China.

Background: Early identification of children at risk of learning disorders (LD) may mitigate the adverse effects of delayed intervention by guiding children to receive preventive services at an earlier age. However, there is no assessment tool for the early identification of children at risk of LD in Mainland China. Therefore, this study aimed to create a Chinese version of the Preschool Learning Skills Scale and investigate its validity and reliability. Methods: Firstly, a pilot scale was designed based on literature review and expert review. Secondly, a pre-survey of the pilot scale was conducted. In phase 3, a formal survey was carried out to test the reliability and validity of the scale by involving 2,677 preschool children from 7 kindergartens. Data were collected using a checklist for demographic characteristics, the preschool learning skills scale, the Behavior Rating Inventory of Executive Function-Preschool Version (BRIEF-P), and Conners' Rating Scales. Results: The final scale included 38 items under seven factors. The reliability and validity tests confirmed that the Cronbach's alpha, split-half reliability, and test-retest reliability coefficients of the scale were 0.946, 0.888, and 0.941, respectively. The Spearman correlations of factor-total score ranged from 0.685 to 0.876. The results of criterion-related validity showed a direct and significant association between the preschool learning skills scale with the BRIEF-P (r = 0.641, P < 0.001) and the cognitive problems factor of Conners' Rating Scales (r = 0.564, P < 0.001). The model had a good fit (χ 2 /df = 3.489, RMSEA = 0.047, RMR = 0.024, CFI = 0.912, TLI = 0.900, and IFI = 0.912). Multigroup confirmatory factor analysis supported the structural and measurement invariance on the preschool learning skills scale across gender and grade. Conclusions: The developed preschool learning skills scale has good reliability and validity, indicating that the scale can be used to identify preschool children at risk of LD and can be recommended for use in clinical research and practice.

Key Glycosyltransferase Genes of Panax notoginseng: Identification and Engineering Yeast Construction of Rare Ginsenosides.

Panax notoginseng is one of the most famous valuable medical plants in China, and its broad application in clinical treatment has an inseparable relationship with the active molecules, ginsenosides. Ginsenosides are glycoside compounds that have varied structures for the diverse sugar chain. Although extensive work has been done, there are still unknown steps in the biosynthetic pathway of ginsenosides. Here, we screened candidate glycosyltransferase genes based on the previous genome and transcriptome data of P. notoginseng and cloned the full length of 27 UGT genes successfully. Among them, we found that PnUGT33 could catalyze different ginsenoside substrates to produce higher polarity rare ginsenosides by extending the sugar chain. We further analyzed the enzymatic kinetics and predicted the catalytic mechanism of PnUGT33 by simulating molecular docking. After that, we reconstructed the biosynthetic pathway of rare ginsenoside Rg3 and gypenoside LXXV in yeast. By combining the Golden Gate method and overexpressing the UDPG biosynthetic genes, we further improved the yield of engineering yeast strain. Finally, the shake-flask culture yield of Rg3 reached 51 mg/L and the fed-batch fermentation yield of gypenoside LXXV reached 94.5 mg/L, which was the first and highest record.

Electrophysiological Screening for Children With Suspected Auditory Processing Disorder: A Systematic Review.

Objective: This research aimed to provide evidence for the early identification and intervention of children at risk for auditory processing disorder (APD). Electrophysiological studies on children with suspected APDs were systematically reviewed to understand the different electrophysiological characteristics of children with suspected APDs. Methods: Computerized databases such as PubMed, Cochrane, MEDLINE, Web of Science, and EMBASE were searched for retrieval of articles since the establishment of the database through May 18, 2020. Cohort, case-control, and cross-sectional studies that evaluated the literature for the electrophysiological assessment of children with suspected APD were independently reviewed by two researchers for literature screening, literature quality assessment, and data extraction. The Newcastle-Ottawa Scale and 11 entries recommended by the Agency for Healthcare Research and Quality were used to evaluate the quality of the literature. Results: In accordance with the inclusion criteria, 14 articles were included. These articles involved 7 electrophysiological testing techniques: click-evoked auditory brainstem responses, frequency-following responses, the binaural interaction component of the auditory brainstem responses, the middle-latency response, cortical auditory evoked potential, mismatch negativity, and P300. The literature quality was considered moderate. Conclusions: Auditory electrophysiological testing can be used for the characteristic identification of children with suspected APD; however, the value of various electrophysiological testing methods for screening children with suspected APD requires further study.

Cellulosic ethanol production by consortia of Scheffersomyces stipitis and engineered Zymomonas mobilis.

BACKGROUND: As one of the clean and sustainable energies, lignocellulosic ethanol has achieved much attention around the world. The production of lignocellulosic ethanol does not compete with people for food, while the consumption of ethanol could contribute to the carbon dioxide emission reduction. However, the simultaneous transformation of glucose and xylose to ethanol is one of the key technologies for attaining cost-efficient lignocellulosic ethanol production at an industrial scale. Genetic modification of strains and constructing consortia were two approaches to resolve this issue. Compared with strain improvement, the synergistic interaction of consortia in metabolic pathways should be more useful than using each one separately. RESULTS: In this study, the consortia consisting of suspended Scheffersomyces stipitis CICC1960 and Zymomonas mobilis 8b were cultivated to successfully depress carbon catabolite repression (CCR) in artificially simulated 80G40XRM. With this strategy, a 5.52% more xylose consumption and a 6.52% higher ethanol titer were achieved by the consortium, in which the inoculation ratio between S. stipitis and Z. mobilis was 1:3, compared with the Z. mobilis 8b mono-fermentation. Subsequently, one copy of the xylose metabolic genes was inserted into the Z. mobilis 8b genome to construct Z. mobilis FR2, leading to the xylose final-consumption amount and ethanol titer improvement by 15.36% and 6.81%, respectively. Finally, various corn stover hydrolysates with different sugar concentrations (glucose and xylose 60, 90, 120 g/L), were used to evaluate the fermentation performance of the consortium consisting of S. stipitis CICC1960 and Z. mobilis FR2. Fermentation results showed that a 1.56-4.59% higher ethanol titer was achieved by the consortium compared with the Z. mobilis FR2 mono-fermentation, and a 46.12-102.14% higher ethanol titer was observed in the consortium fermentation when compared with the S. stipitis CICC1960 mono-fermentation. Furthermore, qRT-PCR analysis of xylose/glucose transporter and other genes responsible for CCR explained the reason why the initial ratio inoculation of 1:3 in artificially simulated 80G40XRM had the best fermentation performance in the consortium. CONCLUSIONS: The fermentation strategy used in this study, i.e., using a genetically modified consortium, had a superior performance in ethanol production, as compared with the S. stipitis CICC1960 mono-fermentation and the Z. mobilis FR2 mono-fermentation alone. This result showed that this strategy has potential for future lignocellulosic ethanol production.

Genome shuffling enhances stress tolerance of Zymomonas mobilis to two inhibitors.

BACKGROUND: Furfural and acetic acid are the two major inhibitors generated during lignocellulose pretreatment and hydrolysis, would severely inhibit the cell growth, metabolism, and ethanol fermentation efficiency of Zymomonas mobilis. Effective genome shuffling mediated by protoplast electrofusion was developed and then applied to Z. mobilis. RESULTS: After two rounds of genome shuffling, 10 different mutants with improved cell growth and ethanol yield in the presence of 5.0 g/L acetic acid and 3.0 g/L furfural were obtained. The two most prominent genome-shuffled strains, 532 and 533, were further investigated along with parental strains in the presence of 7.0 g/L acetic acid and 3.0 g/L furfural. The results showed that mutants 532 and 533 were superior to the parental strain AQ8-1 in the presence of 7.0 g/L acetic acid, with a shorter fermentation time (30 h) and higher productivity than AQ8-1. Mutant 533 exhibited subtle differences from parental strain F34 in the presence of 3.0 g/L furfural. Mutations present in 10 genome-shuffled strains were identified via whole-genome resequencing, and the source of each mutation was identified as either de novo mutation or recombination of the parent genes. CONCLUSIONS: These results indicate that genome shuffling is an efficient method for enhancing stress tolerance in Z. mobilis. The engineered strains generated in this study could be potential cellulosic ethanol producers in the future.