Glutamine promotes antibiotic uptake to kill multidrug-resistant uropathogenic bacteria.

The prevalence of multidrug-resistant bacteria has been increasing rapidly worldwide, a trend that poses great risk to human and animal health and creates urgent need for pharmaceutical and nonpharmaceutical approaches to stop the spread of disease due to antimicrobial resistance. Here, we found that alanine, aspartate, and glutamate metabolism was inactivated, and glutamine was repressed in multidrug-resistant uropathogenic Escherichia coli using a comparative metabolomics approach. Exogenous glutamine promoted ß-lactam­, aminoglycoside-, quinolone-, and tetracycline-induced killing of uropathogenic E. coli and potentiated ampicillin to eliminate multidrug-resistant Pseudomonas aeruginosa, Acinetobacter baumannii, Klebsiella peneumoniae, Edwardsiella tarda, Vibrio alginolyticus, and Vibrio parahaemolyticus. Glutamine-potentiated ampicillin-mediated killing was effective against biofilms of these bacteria in a mouse urinary tract infection model and against systemic infection caused by E. coli, P. aeruginosa, A. baumannii, or K. peneumoniae in a mouse model. Exogenous glutamine stimulated influx of ampicillin, leading to the accumulation of intracellular antibiotic concentrations that exceeded the amount tolerated by the multidrug-resistant bacteria. Furthermore, we demonstrated that exogenous glutamine promoted the biosynthesis of nucleosides including inosine, which in turn interacted with CpxA/CpxR and up-regulated OmpF. We validated the physiological relevance of the mechanism by showing that loss of purF, purH, cpxA, or ompF elevated antibiotic resistance in antibiotic-sensitive strains. In addition, glutamine retarded the development of ampicillin resistance. These results may facilitate future development of effective approaches for preventing or managing chronic, multidrug-resistant bacterial infections, bacterial persistence, and difficult-to-treat bacterial biofilms.

Octupolar Acrylonitrile-Bridged 2D-Conjugated Polymers Enable Bright Far-Red Emission with Intense Two-Photon Absorption via Alkoxylation Chemistry.

Herein, alkoxylation chemistry is introduced as a "one-stone-three-birds" solution for exploring a new family of highly-fluorescent octupolar 2D-conjugated organic polymers/frameworks (OCOPs/OCOFs) combining far-red emission, high fluorescence quantum yield (QY), and strong two-photon absorption (TPA). Both alkoxy-substituted OCOP and OCOF comprising acrylonitrile-bridged strongly-coupled donor3-(acceptor core) chromophores densely packed in either disordered or ordered forms, exhibit significantly redshifted emission. They produce high QY of 22.2% and 27.8% in tetrahydrofuran, large TPA cross section of 600 and 1124 GM, and 2-3 folds and 15-30 folds that of non-alkoxylate amorphous counterpart respectively. Combined theoretical and experimental studies reveal unique "one-stone-three-birds" role of the alkoxylation in realizing red-shifted-emission, improved QY and TPA enabled by inducing steric hindrance effect for weakened π-π stacking, and triggering p-π conjugation effect for electronically engineering octupolar chromophores, while the crystalline engineering enables enforced coplanarity conformation and improved π-electron delocalization for further improved QY and TPA. The robust and biocompatible pentoxy-substituted polymer can be used not only as metal-free red-emissive phosphor for efficient warm white light-emitting diodes, but also as efficient two-photon fluorescence probes for bio-imaging.

The comprehensive detection of miRNA, lncRNA, and circRNA in regulation of mouse melanocyte and skin development.

BACKGROUND: Pigmentation development, is a complex process regulated by many transcription factors during development. With the development of the RNA sequencing (RNA-seq), non-coding RNAs, such as miRNAs, lncRNAs, and circRNAs, are found to play an important role in the function detection of related regulation factors. In this study, we provided the expression profiles and development of ncRNAs related to melanocyte and skin development in mice with black coat color skin and mice with white coat color skin during embryonic day 15 (E15) and postnatal day 7 (P7). The expression profiles of different ncRNAs were detected via RNA-seq and also confirmed by the quantitative real-time PCR (qRT-PCR) method. GO and KEGG used to analyze the function the related target genes. RESULTS: We identified an extensive catalogue of 206 and 183 differently expressed miRNAs, 600 and 800 differently expressed lncRNAs, and 50 and 54 differently expressed circRNAs, respectively. GO terms and pathway analysis showed the target genes of differentially expressed miRNA and lncRNA. The host genes of circRNA were mainly enriched in cellular process, single organism process. The target genes of miRNAs were mainly enriched in chromatin binding and calcium ion binding in the nucleus. The function of genes related to lncRNAs are post translation modification. The competing endogenous RNA (ceRNA) network of lncRNAs and circRNAs displays a complex interaction between ncRNA and mRNA related to skin development, such as Tcf4, Gnas, and Gpnms related to melanocyte development. CONCLUSIONS: The ceRNA network of lncRNA and circRNA displays a complex interaction between ncRNA and mRNA related to skin development and melanocyte development. The embryonic and postnatal development of skin provide a reference for further studies on the development mechanisms of ncRNA during pigmentation.

The comprehensive detection of miRNA, lncRNA, and circRNA in regulation of mouse melanocyte and skin development

BACKGROUND: Pigmentation development, is a complex process regulated by many transcription factors during development. With the development of the RNA sequencing (RNA-seq), non-coding RNAs, such as miRNAs, lncRNAs, and circRNAs, are found to play an important role in the function detection of related regulation factors. In this study, we provided the expression profiles and development of ncRNAs related to melanocyte and skin development in mice with black coat color skin and mice with white coat color skin during embryonic day 15 (E15) and postnatal day 7 (P7). The expression profiles of different ncRNAs were detected via RNA-seq and also confirmed by the quantitative real-time PCR (qRT-PCR) method. GO and KEGG used to analyze the function the related target genes. RESULTS: We identified an extensive catalogue of 206 and 183 differently expressed miRNAs, 600 and 800 differently expressed lncRNAs, and 50 and 54 differently expressed circRNAs, respectively. GO terms and pathway analysis showed the target genes of differentially expressed miRNA and lncRNA. The host genes of circRNA were mainly enriched in cellular process, single organism process. The target genes of miRNAs were mainly enriched in chromatin binding and calcium ion binding in the nucleus. The function of genes related to lncRNAs are post translation modification. The competing endogenous RNA (ceRNA) network of lncRNAs and circRNAs displays a complex interaction between ncRNA and mRNA related to skin development, such as Tcf4 , Gnas , and Gpnms related to melanocyte development. CONCLUSIONS: The ceRNA network of lncRNA and circRNA displays a complex interaction between ncRNA and mRNA related to skin development and melanocyte development. The embryonic and postnatal development of skin provide a reference for further studies on the development mechanisms of ncRNA during pigmentation.

Comparison of miRNA-101a-3p and miRNA-144a-3p regulation with the key genes of alpaca melanocyte pigmentation.

BACKGROUND: Many miRNA functions have been revealed to date. Single miRNAs can participate in life processes by regulating more than one target gene, and more than one miRNA can also simultaneously act on one target mRNA. Thus, a complex regulatory network involved in many processes can be formed. Herein, the pigmentation regulation mechanism of miR-101a-3p and miR-144a-3p was studied at the cellular level by the overexpression and equal overexpression of miR-101a-3p and miR-144a-3p. RESULTS: Results revealed that miR-101a-3p and miR-144a-3p directly regulated the expression of microphthalmia-associated transcription factor, thereby affecting melanin synthesis. CONCLUSIONS: The two miRNAs with the same binding site in the same gene independently excreted each other's function. However, the inhibitory effect of miR-144a-3p was stronger than that of miR-101-3p in alpaca melanocytes, although both decreased melanin production.

Glycine, serine and threonine metabolism confounds efficacy of complement-mediated killing.

Serum resistance is a poorly understood but common trait of some difficult-to-treat pathogenic strains of bacteria. Here, we report that glycine, serine and threonine catabolic pathway is down-regulated in serum-resistant Escherichia coli, whereas exogenous glycine reverts the serum resistance and effectively potentiates serum to eliminate clinically-relevant bacterial pathogens in vitro and in vivo. We find that exogenous glycine increases the formation of membrane attack complex on bacterial membrane through two previously unrecognized regulations: 1) glycine negatively and positively regulates metabolic flux to purine biosynthesis and Krebs cycle, respectively. 2) α-Ketoglutarate inhibits adenosine triphosphate synthase, which in together promote the formation of cAMP/CRP regulon to increase the expression of complement-binding proteins HtrE, NfrA, and YhcD. The results could lead to effective strategies for managing the infection with serum-resistant bacteria, an especially valuable approach for treating individuals with weak acquired immunity but a normal complement system.

Phenylalanine enhances innate immune response to clear ceftazidime-resistant Vibrio alginolyticus in Danio rerio.

Antibiotic-resistant bacteria becomes a major threat to the economy and food safety in aquaculture. Although the antibiotic-dependent strategy is still the mostly adopted option, the development of antibiotic-free approach is urgently needed to ameliorate the severe situation of the global antibiotic resistance. In the present study, we showed that modulating the metabolism of zebrafish, Danio reiro, would enhance D. rerio to clear ceftazidime-resistant Vibrio alginoyticus (Caz-R) in vivo. By generating Caz-R in vitro, we found Caz-R stays longer than ceftazidime-sensitive V. alginoyticus (Caz-S) in D. rerio, where Caz-R induced less potent immune response than that of Caz-S. The differential immune response was associated with different metabolism of the host. Through functional metabolomics, we identified a crucial biomarker, phenylalanine. The abundance of phenylalanine was increased in both of Caz-S and Caz-R infected hosts but the abundance was higher in Caz-S infected group. This specific difference indicated phenylalanine could be a metabolite required to clear Caz-R by the host. Exogenous phenylalanine would enhance the host's ability to remove Caz-R, which was through upregulated production of lysozyme and C3b. Thus, our study demonstrates a novel strategy to boost host's immune response to combat against antibiotic-resistant bacteria.

NaCl promotes antibiotic resistance by reducing redox states in Vibrio alginolyticus.

The development of antibiotic resistance in Vibrio alginolyticus represents a threat to human health and fish farming. Environmental NaCl regulation of bacterial physiology is well documented, but whether the regulation contributes to antibiotic resistance remains unknown. To explore this, we compared minimum inhibitory concentration (MIC) of V. alginolyticus cultured in different media with 0.5%-10% NaCl, and found that the MIC increased as the NaCl concentration increased, especially for aminoglycoside antibiotics. Consistent with this finding, internal NaCl also increased, while intracellular gentamicin level decreased. GC-MS-based metabolomics showed different distributions of pyruvate cycle intermediates among 0.5%, 4% and 10% NaCl. Differential activity of enzymes in the pyruvate cycle and altered expression of Na(+)-NQR led to a reducing redox state, characterized by decreased levels of NADH, proton motive force (PMF) and ATP. Meanwhile, NaCl negatively regulated PMF as a consequence of the reducing redox state. These together are responsible for the decreased intracellular gentamicin level with the increased external level of NaCl. Our study reveals a previously unknown redox state-dependent mechanism regulated by NaCl in V. alginolyticus that impacts antibiotic resistance.

Identification and efficacy of glycine, serine and threonine metabolism in potentiating kanamycin-mediated killing of Edwardsiella piscicida.

We previously showed that glucose potentiated kanamycin to kill multidrug-resistant Edwardsiella piscicida through activation of the TCA cycle. However, whether other regulatory mechanism is involved requires further investigation. By quantitative proteomics technology, iTRAQ, we systematically mapped the altered proteins in the presence of glucose and identified 94 differentially expressed proteins. The analysis of the altered proteins by pathways, amino acid biosynthesis and metabolism were enriched. And the most significantly altered eight amino acids tyrosine, phenylalanine, valine, leucine, isoleucine, glycine, serine and threonine were investigated for their potentiation of kanamycin to kill EIB202, where glycine, serine and threonine showed the strongest efficacy than the others. The combinations of glycine and serine or glucose with glycine, serine or threonine had the best effects. Moreover, pyruvate dehydrogenase, α-ketoglutarate dehydrogenase and succinate dehydrogenase activities were increased as well as the proton motive force (PMF) and intracellular kanamycin. Finally, inhibitors that disrupt PMF production abolished the potentiation. These results shed light on the mechanism of how glucose promoting the amino acids biosynthesis and metabolism to potentiate kanamycin to kill antibiotic-resistant bacteria. More importantly, our results suggested that adjusting amino acid biosynthesis and metabolism might be a strategy to become phenotypic resistance to antibiotics in bacteria. SIGNIFICANCE: Tackling antibiotic resistance is an emerging issue in current years. Despite the efforts made toward developing new antibiotics, the progress is still lagged behind expectation. Novel strategies are required. The use of metabolite to revert antibiotic resistant is highly appreciated in recent years due to the less toxicity, more economic and high efficacy. As a continued study of our previous report on glucose potentiating kanamycin to kill antibiotic-resistant bacteria. The current study further expands the previous discovery on the mechanism of how glucose potentiate this effect. This result provides more basis on the action of glucose in reverting antibiotic resistance. And more importantly, we may derive more metabolites other than glucose to manage antibiotic resistance.

The depressed central carbon and energy metabolisms is associated to the acquisition of levofloxacin resistance in Vibrio alginolyticus.

The overuse and misuse of antibiotics lead to bacterial antibiotic resistance, challenging human health and intensive cultivation. It is especially required to understand for the mechanism of antibiotic resistance to control antibiotic-resistant pathogens. The present study characterized the differential proteome of levofloxacin-resistant Vibrio alginolyticus with the most advanced iTRAQ quantitative proteomics technology. A total of 160 proteins of differential abundance were identified, where 70 were decreased and 90 were increased. Further analysis demonstrated that crucial metabolic pathways like TCA cycle were significantly down-regulated. qRT-PCR analysis demonstrated the decreased gene expression of glycolysis/gluconeogenesis, the TCA cycle, and fatty acid biosynthesis. Moreover, Na(+)-NQR complex gene expression, membrane potential and the adenylate energy charge ratio were decreased, indicating that the decreased central carbon metabolism is associated to the acquisition of levofloxacin resistance. Therefore, the reduced central carbon and energy metabolisms form a characteristic feature as fitness costs of V. alginolyticus in resistance to levofloxacin. BIOLOGICAL SIGNIFICANCE: The overuse and misuse of antibiotics lead to bacterial antibiotic resistance, challenging human health and intensive cultivation. Understanding for the antibiotic resistance mechanisms is especially required to control these antibiotic-resistant pathogens. The present study characterized the differential proteome of levofloxacin-resistant Vibrio alginolyticus using the most advanced iTRAQ quantitative proteomics technology. A total of 160 differential abundance of proteins were identified with 70 decreases and 90 increases by liquid chromatography matrix assisted laser desorption ionization mass spectrometry. Most interestingly, crucial metabolic pathways such as the TCA cycle sharply fluctuated. This is the first report that the reduced central carbon and energy metabolisms form a characteristic feature as a mechanism of V. alginolyticus in resistance to levofloxacin.

Six genes of ompA family shuffling for development of polyvalent vaccines against Vibrio alginolyticus and Edwardsiella tarda.

Polyvalent vaccines against more than one species of pathogens are especially important due to the complex ecosystem in aquaculture. We have previously shown that the development of polyvalent vaccines by shuffling six ompA genes from different bacteria with V. parahaemolyticus VP0764 primers. Here, we used the same 6 genes, V. alginolyticus VA0764 and VA1186, V. parahaemolyticus VP0764 and VP1186, E. tarda ompA and E. coli ompA, but with E. tarda ompA primers to develop new polyvalent vaccines. By this approach, we identified 7 potential polyvalent vaccines that were effective against both V. alginolyticus and E. tarda infections. Furthermore, the innate immunity triggered by the vaccines were also explored in three groups, no protection (group I), protection against V. alginolyticus (group II), and protection against both V. alginolyticus and E. tarda (group III). The transcription of IL-1ß, IL-6, IL-8, C3b and NF-kB were significantly increased in group II and group III but not group I, where the expression level of group III was higher than group II. In addition, differential activities of succinate dehydrogenase were detected among the three groups. These results indicate the expansion of polyvalent vaccine reservoir with the same shuffling genes but different primers, and promote the understanding of the mechanisms of polyvalent vaccines based on vaccine-induced innate immunity.

Pyruvate cycle increases aminoglycoside efficacy and provides respiratory energy in bacteria.

The emergence and ongoing spread of multidrug-resistant bacteria puts humans and other species at risk for potentially lethal infections. Thus, novel antibiotics or alternative approaches are needed to target drug-resistant bacteria, and metabolic modulation has been documented to improve antibiotic efficacy, but the relevant metabolic mechanisms require more studies. Here, we show that glutamate potentiates aminoglycoside antibiotics, resulting in improved elimination of antibiotic-resistant pathogens. When exploring the metabolic flux of glutamate, it was found that the enzymes that link the phosphoenolpyruvate (PEP)-pyruvate-AcCoA pathway to the TCA cycle were key players in this increased efficacy. Together, the PEP-pyruvate-AcCoA pathway and TCA cycle can be considered the pyruvate cycle (P cycle). Our results show that inhibition or gene depletion of the enzymes in the P cycle shut down the TCA cycle even in the presence of excess carbon sources, and that the P cycle operates routinely as a general mechanism for energy production and regulation in Escherichia coli and Edwardsiella tarda These findings address metabolic mechanisms of metabolite-induced potentiation and fundamental questions about bacterial biochemistry and energy metabolism.

Construction, immune protection and innate immune response of shuffled polyvalent ompAs vaccines.

Our previous studies demonstrated that molecular breeding via DNA shuffling directs the evolution of polyvalent vaccines with desired traits, which leads to generation of polyvalent ompA vaccines using Vibrio alginolyticus VA0764 primers. Here, we replaced VA0764 primers with Edwardsiella tarda ompA primers to generate new polyvalent ompA vaccines by DNA shuffling of the same five ompA genes from four species of bacteria E. tarda, V. parahaemolyticus, V. alginolyticus and Escherichia coli. We identified four polyvalent vaccine candidates from a eukaryotic expressing library EompAs-FE containing 82 ompAs using active immune protection against V. alginolyticus and E. tarda. Furthermore, we explored mechanisms of polyvalent vaccine candidates by investigation of the innate immune response to these ompAs, and found that expression of IL-1ß, IL-8, IL-15, COX-2, IFN-γ, TLR-1, TLR-3 and C3b genes was elevated as a characteristic feature of these polyvalent vaccine candidates. These results indicate that use of different primers to construct a DNA library selects new evolution of polyvalent vaccines with desired traits, and polyvalent ompA vaccines elicit high innate immune response.

Boosted TCA cycle enhances survival of zebrafish to Vibrio alginolyticus infection.

Vibrio alginolyticus is a waterborne pathogen that infects a wide variety of hosts including fish and human, and the outbreak of this pathogen can cause a huge economic loss in aquaculture. Thus, enhancing host's capability to survive from V. alginolyticus infection is key to fighting infection and this remains still unexplored. In the present study, we established a V. alginolyticus-zebrafish interaction model by which we explored how zebrafish survived from V. alginolyticus infection. We used GC-MS based metabolomic approaches to characterize differential metabolomes between survival and dying zebrafish upon infection. Pattern recognition analysis identified the TCA cycle as the most impacted pathway. The metabolites in the TCA cycle were decreased in the dying host, whereas the metabolites were increased in the survival host. Furthermore, the enzymatic activities of the TCA cycle including pyruvate dehydrogenase (PDH), α-ketoglutaric dehydrogenase (KGDH) and succinate dehydrogenase (SDH) also supported this conclusion. Among the increased metabolites in the TCA cycle, malic acid was the most crucial biomarker for fish survival. Indeed, exogenous malate promoted zebrafish survival in a dose-dependent manner. The corresponding activities of KGDH and SDH were also increased. These results indicate that the TCA cycle is a key pathway responsible for the survival or death in response to infection caused by V. alginolyticus, and highlight the way on development of metabolic modulation to control the infection.

Edwardsiella tarda Tunes Tricarboxylic Acid Cycle to Evade Complement-Mediated Killing.

Evasion of complement-mediated killing is a common phenotype for many different types of pathogens, but the mechanism is still poorly understood. Most of the clinic isolates of Edwardsiella tarda, an important pathogen infecting both of human and fish, are commonly found serum-resistant. To explore the potential mechanisms, we applied gas chromatography-mass spectrometry (GC-MS)-based metabolomics approaches to profile the metabolomes of E. tarda EIB202 in the presence or absence of serum stress. We found that tricarboxylic acid (TCA) cycle was greatly enhanced in the presence of serum. The quantitative real-time PCR (qRT-PCR) and enzyme activity assays validated this result. Furthermore, exogenous succinate that promotes the TCA cycle increased serum resistance, while TCA cycle inhibitors (bromopyruvate and propanedioic acid) that inhibit TCA cycle, attenuated serum resistance. Moreover, the enhanced TCA cycle increased membrane potential, thus decreased the formation of membrane attack complex at cell surface, resulting serum resistance. These evidences suggested a previously unknown membrane potential-dependent mechanism of serum resistance. Therefore, our findings reveal that pathogen mounts a metabolic trick to cope with the serum complement-mediated killing.

Expression of glucose transporter 8 (GLUT8) in spermatogenesis of adult boar testes.

The present study was designed to investigate the glucose transporter 8 (GLUT8) expression and localization in adult boar testis. Localization and expression of GLUT8 were conducted with Western Blotting, immunohistochemistry and reverse transcription-polymerase chain reaction (RT-PCR) methods GLUT8 protein and mRNA were expressed in the boar testes. The results of Western Blotting analysis showed specificity of the antibody for protein of boar testes. The immunohistochemistry results showed that GLUT8 protein mainly localized in spermatocytes, round spermatids and elongated spermatids of the seminiferous tubules in the adult boar testes. And the GLUT8 expression persists during eight stages of boar spermatogenesis. GLUT8 may mainly provide glucose for the later stage of germ cell differentiation in adluminal compartment in adult boar testes. These results suggested that GLUT8 is important for the spermatogenesis in the adult boar testes.

Exogenous l-Valine Promotes Phagocytosis to Kill Multidrug-Resistant Bacterial Pathogens.

The emergence of multidrug-resistant bacteria presents a severe threat to public health and causes extensive losses in livestock husbandry and aquaculture. Effective strategies to control such infections are in high demand. Enhancing host immunity is an ideal strategy with fewer side effects than antibiotics. To explore metabolite candidates, we applied a metabolomics approach to investigate the metabolic profiles of mice after Klebsiella pneumoniae infection. Compared with the mice that died from K. pneumoniae infection, mice that survived the infection displayed elevated levels of l-valine. Our analysis showed that l-valine increased macrophage phagocytosis, thereby reducing the load of pathogens; this effect was not only limited to K. pneumoniae but also included Escherichia coli clinical isolates in infected tissues. Two mechanisms are involved in this process: l-valine activating the PI3K/Akt1 pathway and promoting NO production through the inhibition of arginase activity. The NO precursor l-arginine is necessary for l-valine-stimulated macrophage phagocytosis. The valine-arginine combination therapy effectively killed K. pneumoniae and exerted similar effects in other Gram-negative (E. coli and Pseudomonas aeruginosa) and Gram-positive (Staphylococcus aureus) bacteria. Our study extends the role of metabolism in innate immunity and develops the possibility of employing the metabolic modulator-mediated innate immunity as a therapy for bacterial infections.

GC-MS-Based Metabolome and Metabolite Regulation in Serum-Resistant Streptococcus agalactiae.

Streptococcus agalactiae causes severe systemic infections in human and fish. In the present study, we established a pathogen-plasma interaction model by which we explored how S. agalactiae evaded serum-mediated killing. We found that S. agalactiae grew faster in the presence of yellow grouper plasma than in the absence of the plasma, indicating S. agalactiae evolved a way of evading the fish immune system. To determine the events underlying this phenotype, we applied GC-MS-based metabolomics approaches to identify differential metabolomes between S. agalactiae cultured with and without yellow grouper plasma. Through bioinformatics analysis, decreased malic acid and increased adenosine were identified as the most crucial metabolites that distinguish the two groups. Meanwhile, they presented with decreased TCA cycle and elevated purine metabolism, respectively. Finally, exogenous malic acid and adenosine were used to reprogram the plasma-resistant metabolome, leading to elevated and decreased susceptibility to the plasma, respectively. Therefore, our findings reveal for the first time that S. agalactiae utilizes a metabolic trick to respond to plasma killing as a result of serum resistance, which may be reverted or enhanced by exogenous malic acid and adenosine, respectively, suggesting that the metabolic trick can be regulated by metabolites.

N-acetylglucosamine enhances survival ability of tilapias infected by Streptococcus iniae.

Streptococcus iniae infection has emerged as a serious fish health and economic problem in the global aquaculture operations. Current antibiotic options are few and possess severe practical limitations and potential adverse environmental impacts. The major factor contributing to the large burden of S. iniae disease in aquaculture is the lack of fundamental knowledge of innate immunity against the pathogen. In the present study, we use a tilapia model to explore which metabolites are crucial for the defense against the infection caused by S. iniae. We establish GC/MS based metabolic profile of tilapia liver and then compare the metabolic difference between survivals and the dying fish post the bacterial infection. We identify elevating N-acetylglucosamine in survival group as the most crucial metabolite differentiating the survivals from the dying in these fish infected by S. iniae. Exogenous N-acetylglucosamine significantly elevates survival ability of tilapia against the infection caused by S. iniae. Our findings highlight the importance of metabolic strategy against bacterial infections.

Characterization of protein species and weighted protein co-expression network regulation of Escherichia coli in response to serum killing using a 2-DE based proteomics approach.

Posttranslational modifications, providing covalent alterations to extend their functions, show protein species on 2-DE gels, but our knowledge on protein species is still limited. In the present study, characteristics of protein species are determined in Escherichia coli using 2-DE based proteomics. In the E. coli proteome, 691 unique proteins (representing 1096 protein spots) accounting for 15.37% of gene-coding proteins of the bacterium are identified. Out of them, 191 have 596 protein species. Proteins with higher abundance, a higher proportion of Glu, Gly, Lys, and higher pI are more likely to have protein species. Further investigation on bacterial serum resistance indicates that more proteins with protein species are found in the bacterium in response to serum stress. A weighted protein co-expression network shows that protein species are related to topological connection as a result of protein regulation. The node protein IleS is demonstrated to contribute to serum resistance using a gene-deleted mutant. These results have revealed general characteristic features of bacterial species, and also provided novel insights into the biological significance of bacterial protein species, particularly the role in serum resistance.