Transcriptome analysis and cytochrome P450 monooxygenase reveal the molecular mechanism of Bisphenol A degradation by Pseudomonas putida strain YC-AE1.

BACKGROUND: Bisphenol A (BPA) is a rapid spreading organic pollutant that widely used in many industries especially as a plasticizer in polycarbonate plastic and epoxy resins. BPA reported as a prominent endocrine disruptor compound that possesses estrogenic activity and fulminant toxicity. Pseudomonas putida YC-AE1 was isolated in our previous study and exerted a strong degradation capacity toward BPA at high concentrations; however, the molecular degradation mechanism is still enigmatic. RESULTS: We employed RNA sequencing to analyze the differentially expressed genes (DEGs) in the YC-AE1 strain upon BPA induction. Out of 1229 differentially expressed genes, 725 genes were positively regulated, and 504 genes were down-regulated. The pathways of microbial metabolism in diverse environments were significantly enriched among DEGs based on KEGG enrichment analysis. qRT-PCR confirm the involvement of BPA degradation relevant genes in accordance with RNA Seq data. The degradation pathway of BPA in YC-AE1 was proposed with specific enzymes and encoded genes. The role of cytochrome P450 (CYP450) in BPA degradation was further verified. Sever decrease in BPA degradation was recorded by YC-AE1 in the presence of CYP450 inhibitor. Subsequently, CYP450bisdB deficient YC-AE1 strain △ bisdB lost its ability toward BPA transformation comparing with the wild type. Furthermore, Transformation of E. coli with pET-32a-bisdAB empowers it to degrade 66 mg l-1 of BPA after 24 h. Altogether, the results showed the role of CYP450 in biodegradation of BPA by YC-AE1. CONCLUSION: In this study we propose the molecular basis and the potential role of YC-AE1cytochrome P450 monooxygenase in BPA catabolism.

Resveratrol biosynthesis, optimization, induction, bio-transformation and bio-degradation in mycoendophytes.

Resveratrol (3,4,5-trihydroxystilbene) is a naturally occurring polyphenolic stilbene compound produced by certain plant species in response to biotic and abiotic factors. Resveratrol has sparked a lot of interest due to its unique structure and approved therapeutic properties for the prevention and treatment of many diseases such as neurological disease, cardiovascular disease, diabetes, inflammation, cancer, and Alzheimer's disease. Over the last few decades, many studies have focused on the production of resveratrol from various natural sources and the optimization of large-scale production. Endophytic fungi isolated from various types of grapevines and Polygonum cuspidatum, the primary plant sources of resveratrol, demonstrated intriguing resveratrol-producing ability. Due to the increasing demand for resveratrol, one active area of research is the use of endophytic fungi and metabolic engineering techniques for resveratrol's large-scale production. The current review addresses an overview of endophytic fungi as a source for production, as well as biosynthesis pathways and relevant genes incorporated in resveratrol biosynthesis. Various approaches for optimizing resveratrol production from endophytic fungi, as well as their bio-transformation and bio-degradation, are explained in detail.

Prevalence and genotyping distribution of Enterocytozoon bieneusi in diarrheic pigs in Chongqing and Sichuan provinces, China.

The microsporidian fungal pathogen Enterocytozoon bieneusi is a unicellular parasite that infects humans and various animals, including pigs. Currently, there are few data on E. bieneusi infection a in diarrheic pigs in Chongqing and Sichuan Provinces, China. This study aims to determine the prevalence and genotype distribution of E. bieneusi in diarrheic pigs. In total, 514 fecal samples from diarrheic pigs were obtained from 14 large-scale farms in Chongqing and Sichuan Provinces (326 suckling pigs, 17 weaned pigs, 65 fattening pigs, and 106 sows). To identify the E. bieneusi genotypes, genomic DNA was isolated from the samples and tested by nested PCR, targeting the internal transcribed spacer region of the rRNA followed by DNA sequence analysis. The overall prevalence of E. bieneusi was 79.8% (410/514), with rates of 84.9% (90/106) in sows and 64.7% (11/17) in weaned pigs. We found 61 different genotypes, including seven known genotypes (E, F, CHG1, Peru8, CAF1, B, and BEB17) and 54 novel genotypes. These 54 new genotypes are variants of eight known genotypes (SDD2, A, B, HLJD-IV, PigSpEb1, O, JLD-I, and BEB17) based on their sequence similarities. Phylogenetically, all of the identified genotypes clustered with counterparts belonging to Group 1 and Group 2 of E. bieneusi. Therefore, we found a higher prevalence of E. bieneusi in sows than in preweaned and weaned pigs. These findings indicate that diarrheic pigs could be a potential reservoir host, which can contaminate the environment and be a source of microsporidia in humans and other animals.

Mycobacterium tuberculosis PE17 (Rv1646) promotes host cell apoptosis via host chromatin remodeling mediated by reduced H3K9me3 occupancy.

Tuberculosis caused by Mycobacterium tuberculosis remains a serious global public health threat. M. tuberculosis PE and PPE proteins are closely involved in pathogen-host interaction. To explore the predicted function of the M. tuberculosis PE17 (Rv1646), we heterologously expressed PE17 in a non-pathogenic Mycobacterium smegmatis strain (Ms_PE17). PE17 can reduce the survival of M. smegmatis within macrophages associated with altering the transcription levels of inflammatory cytokines IL1ß, IL6, TNFα, and IL10 in Ms_PE17 infected macrophages through JNK signaling. Furthermore, macrophages apoptosis was increased upon Ms_PE17 infection in a caspases-dependent manner, accompanied by the activation of the Endoplasmic Reticulum stress IRE1α/ASK1/JNK signaling pathway. This can be largely interpreted by the epigenetic changes through reduced H3K9me3 chromatin occupancy post Ms_PE17 infection. To our knowledge, this is the first report that PE17 altered the macrophages apoptosis via H3K9me3 mediated chromatin remodeling.

Mycobacterium tuberculosis RKIP (Rv2140c) dephosphorylates ERK/NF-κB upstream signaling molecules to subvert macrophage innate immune response.

Mycobacterium tuberculosis (Mtb) survival and virulence largely reside on its ability to manipulate the host immune response. We have previously shown that M. tuberculosis Raf kinase inhibitor protein (RKIP) Rv2140c regulates diverse phosphorylation events in M. smegmatis. However, its role during infection is unknown. In this report, we show that Rv2140c can mimic the mammalian RKIP function. Rv2140c inhibit the activation of extracellular signal-regulated kinase (ERK) and nuclear factor κB (NF-κB) via decreasing the phosphorylation capacity of upstream mediators MEK1, ERK1/2, and IKKα/ß, thus leading to a reduction in pro-inflammatory cytokines IL-1ß, IL-6, and TNF-α. This effect can be reversed by RKIP inhibitor locostatin. Furthermore Rv2140c mediates apoptosis associated with activation of caspases cascades. This modulation enhances the intracellular survival of M. smegmatis within macrophage. We propose that Rv2140c is a multifunctional virulence factor and a promising novel anti-Tuberculosis drug target.

Mycobacterium tuberculosis PPE10 (Rv0442c) alters host cell apoptosis and cytokine profile via linear ubiquitin chain assembly complex HOIP-NF-κB signaling axis.

Tuberculosis caused by Mycobacterium tuberculosis infection remains one of the top ten causes of deaths worldwide. M. tuberculosis genome devoted 10% capacity for highly repeated PE/PPE genes family. To explore the role of PPE10 in host-pathogen interaction, PPE10 encoding gene Rv0442c was heterologously expressed in the nonpathogenic M. smegmatis strain. PPE10 altered the bacterial cell surface properties, colony morphology, and biofilm formation. Ms_PPE10 showed more resistance to stress conditions such as diamide, and low pH, as well as higher survival within the macrophage. Moreover, the host's cell apoptosis was regulated via decreased expression of caspases, IL-1, IL-6, and TNF-α through the Linear Ubiquitin Chain Assembly Complex (LUBAC) HOIP-NF-κB signaling axis. The study revealed novel insights into the mechanism of action of the PPE family.

Mycobacterium tuberculosis Rv0580c Impedes the Intracellular Survival of Recombinant Mycobacteria, Manipulates the Cytokines, and Induces ER Stress and Apoptosis in Host Macrophages via NF-κB and p38/JNK Signaling.

The Mycobacterium tuberculosis (M. tb) genome encodes a large number of hypothetical proteins, which need to investigate their role in physiology, virulence, pathogenesis, and host interaction. To explore the role of hypothetical protein Rv0580c, we constructed the recombinant Mycobacterium smegmatis (M. smegmatis) strain, which expressed the Rv0580c protein heterologously. We observed that Rv0580c expressing M. smegmatis strain (Ms_Rv0580c) altered the colony morphology and increased the cell wall permeability, leading to this recombinant strain becoming susceptible to acidic stress, oxidative stress, cell wall-perturbing stress, and multiple antibiotics. The intracellular survival of Ms_Rv0580c was reduced in THP-1 macrophages. Ms_Rv0580c up-regulated the IFN-γ expression via NF-κB and JNK signaling, and down-regulated IL-10 expression via NF-κB signaling in THP-1 macrophages as compared to control. Moreover, Ms_Rv0580c up-regulated the expression of HIF-1α and ER stress marker genes via the NF-κB/JNK axis and JNK/p38 axis, respectively, and boosted the mitochondria-independent apoptosis in macrophages, which might be lead to eliminate the intracellular bacilli. This study explores the crucial role of Rv0580c protein in the physiology and novel host-pathogen interactions of mycobacteria.

Mycobacterium tuberculosis Raf kinase inhibitor protein (RKIP) Rv2140c is involved in cell wall arabinogalactan biosynthesis via phosphorylation.

Mycobacterium tuberculosis Rv2140c is a function unknown conserved phosphatidylethanolamine-binding protein (PEBP), homologous to Raf kinase inhibitor protein (RKIP) in human beings. To delineate its function, we heterologously expressed Rv2140c in a non-pathogenic M. smegmatis. Quantitative phosphoproteomic analysis between two recombinant strains Ms_Rv2140c and Ms_vec revealed that Rv2140c differentially regulate 425 phosphorylated sites representing 282 proteins. Gene ontology GO, and a cluster of orthologous groups COG analyses showed that regulated phosphoproteins by Rv2140c were mainly associated with metabolism and cellular processes. Rv2140c significantly repressed phosphoproteins involved in signaling, including serine/threonine-protein kinases and two-component system, and the arabinogalactan biosynthesis pathway phosphoproteins were markedly up-regulated, suggesting a role of Rv2140c in modulating cell wall. Consistent with phosphoproteomic data, Rv2140c altered some phenotypic properties of M. smegmatis such as colony morphology, cell wall permeability, survival in acidic conditions, and active lactose transport. In summary, we firstly demonstrated the role of PEBP protein Rv2140c, especially in phosphorylation of mycobacterial arabinogalactan biosynthesis proteins.

Differential Isoniazid Response Pattern Between Active and Dormant Mycobacterium tuberculosis.

Isoniazid (isonicotinic acid hydrazide, INH) is an effective frontline antituberculosis drug. INH targets several Mycobacterium tuberculosis processes, including mycolic acid biosynthesis, DNA synthesis, and redox potential. M. tuberculosis responds to INH stress by altering the expression level of crucial genes involved in various pathways. In this study, we summarize the induced gene expression pattern of active M. tuberculosis upon INH exposure. Most genes triggered by INH are involved in processes such as mycolic acid biosynthesis, a compensatory response, stress response, and drug efflux. These patterns are absent in dormant M. tuberculosis. The differential INH response pattern can inform future novel measures against M. tuberculosis.

Methylation in Mycobacterium-host interaction and implications for novel control measures.

Methylation epigenetically regulates many pivotal biological processes. Mycobacterium tuberculosis, the pathogen of tuberculosis, can modulate host methylome. The methylated genes, sites, signaling pathway, chromatin remodeling, especially the immune related genes such as cytokines and chemokines, drug resistance and vaccines efficacy relevant genes were summarized in this paper. The results showed that methylation plays important roles in immune evasion, pathogenesis, persistence, disease progression, active, drug responder and non-responder. This will inform better practice for the development of new drugs and vaccines to eradicate tuberculosis.

Biodegradation of endocrine disruptor Bisphenol A by Pseudomonas putida strain YC-AE1 isolated from polluted soil, Guangdong, China.

BACKGROUND: Bisphenol A is an important organic chemical as an intermediate, final and inert ingredient in manufacturing of many important products like polycarbonate plastics, epoxy resins, flame retardants, food-drink packaging coating, and other. BPA is an endocrine disruptor compound that mimics the function of estrogen causing damage to reproductive organs. Bacterial degradation has been consider as a cost effective and eco-friendly method for BPA degradation compared with physical and chemical methods. This study aimed to isolate and identify bacterial strain capable to degrade and tolerate high concentrations of this pollutant, studying the factors affecting the degradation process and study the degradation mechanism of this strain. RESULTS: YC-AE1 is a Gram negative bacterial strain isolated from soil and identified as Pseudomonas putida by 16S rRNA gene sequence and BIOLOG identification system. This strain found to have a high capacity to degrade the endocrine disruptor Bisphenol A (BPA). Response surface methodology using central composite design was used to statistically optimize the environmental factors during BPA degradation and the results obtained by significant model were 7.2, 30 °C and 2.5% for optimum initial pH, temperature and inoculum size, respectively. Prolonged incubation period with low NaCl concentration improve the biodegradation of BPA. Analysis of variance (ANOVA) showed high coefficient of determination, R2 and Adj-R2 which were 0.9979 and 0.9935, respectively. Substrate analysis found that, strain YC-AE1 could degrade a wide variety of bisphenol A-related pollutants such as bisphenol B, bisphenol F, bisphenol S, Dibutyl phthalate, Diethylhexyl phthalate and Diethyl phthalate in varying proportion. Pseudomonas putida YC-AE1 showed high ability to degrade a wide range of BPA concentrations (0.5-1000 mg l- 1) with completely degradation for 500 mg l- 1 within 72 h. Metabolic intermediates detected in this study by HPLC-MS were identified as 4,4-dihydroxy-alpha-methylstilbene, p-hydroxybenzaldeyde, p-hydroxyacetophenone, 4-hydroxyphenylacetate, 4-hydroxyphenacyl alcohol, 2,2-bis(4-hydroxyphenyl)-1-propanol, 1,2-bis(4-hydroxyphenyl)-2-propanol and 2,2-bis(4-hydroxyphenyl) propanoate. CONCLUSIONS: This study reports Pseudomonas putida YC-AE1 as BPA biodegrader with high performance in degradation and tolerance to high BPA concentration. It exhibited strong degradation capacity and prominent adaptability towards a wide range of environmental conditions. Moreover, it degrades BPA in a short time via two different degradation pathways.