Participation of Acyl-Coenzyme A Synthetase FadD4 of Pseudomonas aeruginosa PAO1 in Acyclic Terpene/Fatty Acid Assimilation and Virulence by Lipid A Modification

The pathogenic bacterium Pseudomonas aeruginosa possesses high metabolic versatility, with its effectiveness to cause infections likely due to its well-regulated genetic content. P. aeruginosa PAO1 has at least six fadD paralogous genes, which have been implicated in fatty acid (FA) degradation and pathogenicity. In this study, we used mutagenesis and a functional approach in P. aeruginosa PAO1 to determine the roles of the fadD4 gene in acyclic terpene (AT) and FA assimilation and on pathogenicity. The results indicate that fadD4 encodes a terpenoyl-CoA synthetase utilized for AT and FA assimilation. Additionally, mutations in fadD paralogs led to the modification of the quorum-sensing las/rhl systems, as well as the content of virulence factors pyocyanin, biofilm, rhamnolipids, lipopolysaccharides (LPS), and polyhydroxyalkanoates. In a Caenorhabditis elegans in vivo pathogenicity model, culture supernatants from the 24-h-grown fadD4 single mutant increased lethality compared to the PAO1 wild-type (WT) strain; however, the double mutants fadD1/fadD2, fadD1/fadD4, and fadD2/fadD4 and single mutant fadD2 increased worm survival. A correlation analysis indicated an interaction between worm death by the PAO1 strain, the fadD4 mutation, and the virulence factor LPS. Fatty acid methyl ester (FAME) analysis of LPS revealed that a proportion of the LPS and FA on lipid A were modified by the fadD4 mutation, suggesting that FadD4 is also involved in the synthesis/degradation and modification of the lipid A component of LPS. LPS isolated from the fadD4 mutant and double mutants fadD1/fadD4 and fadD2/fadD4 showed a differential behavior to induce an increase in body temperature in rats injected with LPS compared to the WT strain or from the fadD1 and fadD2 mutants. In agreement, LPS isolated from the fadD4 mutant and double mutants fadD1/fadD2 and fadD2/fadD4 increased the induction of IL-8 in rat sera, but IL1-β cytokine levels decreased in the double mutants fadD1/fadD2 and fadD1/fadD4. The results indicate that the fadD genes are implicated in the degree of pathogenicity of P. aeruginosa PAO1 induced by LPS-lipid A, suggesting that FadD4 contributes to the removal of acyl-linked FA from LPS, rendering modification in its immunogenic response associated to Toll-like receptor TLR4. The genetic redundancy of fadD is important for bacterial adaptability and pathogenicity over the host.

Cardioprotective effects of omega 3 fatty acids from fish oil and it enhances autoimmunity in porcine cardiac myosin-induced myocarditis in the rat model

This experiment proposed to investigate the efficiency of omega 3 fatty acids from fish that improves autoimmune against myocarditis in the rat. Fish oil was extracted from fresh Tuna fish and performed FAME analysis and mice bioassay. The autoimmune myocarditis was induced by subcutaneous injection of porcine cardiac myosin (PCM) into the footpads of rats on the first and seventh day. Rats were dissected on the 21st day to analyze the histopathological, hemodynamic, echocardiographic factors, and immunohistochemistry expressions. In the study, 73.90% of total fatty acids were recorded. Histological analysis revealed that omega 3 fatty acids administrated groups showed tremendous development in the multifocal myocardia hyaline degeneration and necrosis with inflammatory changes. Moreover, omega 3 fatty acids inhabited the expressions of inflammatory cells (CD4, CD8 and CD11b) and suppressed the level of NF-κB. The echocardiographic factors such as heartbeat, SBP, DBP, levels of LVDs, LVDd, LVPW percentage of LVFS, EF, expression levels of inflammatory cytokines (TNF, IL-1β, IFN-ɤ, IL-2, and IL-6) also significantly suppressed by omega 3 fatty acids. Hence, the present study proved that consuming fatty acid-enriched fish might be a successful therapy for improving the inflammatory profile, regenerates the heart tissues, and controlled the production of inflammatory cells.

The Fatty Acid Methyl Ester (FAME) profile of Phytophthora agathidicida and its potential use as diagnostic tool.

Phytophthora diseases cause devastation to crops and native ecosystems worldwide. In New Zealand, Phytophthora agathidicida is threatening the survival of kauri, an endemic, culturally and ecologically important tree species. The current method for detecting P. agathidicida is a soil bating assay that is time-consuming and requires high levels of expertise to assess, thus limiting the analytical sample throughput. Here, we characterized the fatty acid methyl ester (FAME) profile of P. agathidicida. We also compared it with the FAME profile of P. cinnamomi and assessed the efficacy of FAME analysis as a diagnostic tool for detecting the pathogen in soil samples. In FAME analysis, the total fatty acid content is isolated from a sample and converted to FAMEs for analysis, a process that takes less than a day. Unique fatty acid acyl chains can serve as biomarkers for specific organisms. We detected 12 fatty acids in P. agathidicida, two of which (20:4ω6 and 20:5ω3) show promise as potential Phytophthora specific biomarkers. Collectively, these findings advance our fundamental understanding of P. agathidicida biology and provide a promising technique to increase the rate of sample processing and the speed of pathogen detection for P. agathidicida in soil.

Isolation and Description of Catonella massiliensis sp. nov., a Novel Catonella Species, Isolated from a Stable Periodontitis Subject

The genus Catonella currently counts a unique species, C. morbi, isolated from periodontal pockets and associated with periodontitis and endodontic infections. This study contributed to the taxonomical and clinical knowledge of this genus by describing a novel species isolated from a saliva sample from a man in clinical gingival health following successful treatment of periodontitis. Morphological and chemotaxonomic characteristics were investigated using different growth conditions, pH, and temperature. Cellular fatty acid methyl ester (FAME) analysis was conducted by gas chromatography/mass spectrometry (GC/MS). Phylogenetic analysis based on 16S rRNA, orthologous average nucleotide identity (OrthoANI), and digital DNA-DNA hybridization (dDDH) relatedness were performed. Strain Marseille-Q4567T was found to be an anaerobic and non-spore-forming rod-shaped bacterium that grew at 28–41.5 °C (optimum 37 °C), pH 6.5–8.5 (optimum pH 7.5), and 5–10 g/L of NaCl (optimum 5 g/L). The predominant cellular fatty acid was C16:0 (64.2%), followed by unsaturated structures C18:1n9 (12.5%) and C18:2n6 (7.8%). Based on 16S rRNA sequence comparison, the closest phylogenetic neighbor was C. morbi ATCC 51271T (98.23% similarity). The OrthoANI and dDDH values between strain Q4567T and C. morbi ATCC 51271T were respectively 79.43% and 23.8%. Therefore, we concluded that strain Marseille-Q4567T represents a novel species of the genus Catonella, for which the name Catonella massiliensis sp. nov. is proposed (= CSUR Q4567).

Variation in Mycobacterium tuberculosis genotype and molecular phenotype influence clinical phenotype of Pulmonary tuberculosis and Tuberculous Meningitis infection in host

In the present study, we investigated tissue specific colonization and virulence characteristics of two different Mycobacterium tuberculosis (MTB) clinical isolates derived from patients with Pulmonary TB (PTB) and Tuberculous Meningitis (TBM). We retrospectively studied a total of 1458 patients diagnosed with TB between 2003 and 2013. Of these, archived sputum and CSF samples were available for 323 TBM and 157 PTB patients. We selected a total of 10 sputum and CSF isolates from each group for further molecular characterization. Methodologies employed included, Gas chromatographic analyses of Fatty Acid Methyl Esters (GC-FAME) followed by MTB genotyping to assess strain diversity. We further assessed the molecular phenotype of each strain through in-vitro cytokine assays & murine MTB model. Our comparative genomics data illustrated two diverse genotypes belonging to H37RV (PTB) and CCDC5079 linkage (TBM), highlighting major variation in membrane protein composition and enzymes that make different mycolic acids. The differential cytokine response by both the strains & GC-FAME analysis further corroborated this variation in membrane composition. This was in agreement with KasIII enzyme, LppA and desaturase related variation in protein. Both MTB strains in mice showed diverse pathogenesis with CCDC5079 infected mice exhibiting higher dissemination to brain compared to the H37RV strain which developed progressive pulmonary disease. These observations suggest that variation in the MTB membrane composition could play an important role in differential colonization of these strains. The study warrants further investigation of membrane proteins with respect to blood brain barrier invasion and pathogenesis.

Polyextremotolerant Bacillus cereus groups isolated from Algerian arid ecosystems and bioprospecting for their bioactive compounds

In previous work, 70 Bacillus sp. isolates originating from 4 soil plots from southwestern Algeria were studied for their poten al plant growth promoting (PGP). In this study, pH, organic ma er, texture, Calcium carbonate, and Electrical conduc vity of the soil samples were determined, 08 selected isolates were idenfied based on rRNA 16S and FAME analysis using GC. Antimicrobial activity against bacteria and fungi were also investigated. All the four soils were moderately Calcium carbonate, pH of soil was alkaline. Our results showed that the electrical conductivity was high for all soils and the four soil isolates in semi-arid and arid regions were poor in organic matter. In addition, Fatty acid analysis showed the presence of 17 di erent fa y acids. Three fa y acids, 15:0 iso, 16:0, and 17:0, 17:0 were present in all isolates. Thus, the isolates were classified into two clusters which were con rmed the results of 16SrRNA sequence analysis. Preliminary screening revealed that the Bacillus thuringiensis isolates showed strong an bacterial and an fungal proper es against S. aureus (between 7mm to 11mm) and A. niger (between 7mm to 8mm). The aim of the present study is the screening of eight Bacillus cereus strains isolated from four rhizosphere soil sample of South-Western region of Algeria province for their an microbial activity.

Description and Genome Analysis of Methylotetracoccus aquaticus sp. nov. , a Novel Tropical Wetland Methanotroph, with the Amended Description of Methylotetracoccus gen. nov.

We enriched and isolated a novel gammproteobacterial methanotroph; strain FWC3, from tropical freshwater wetland, near Nagaon beach, Alibag, India. FWC3 is a coccoid, flesh pink/peach pigmented, non-motile methanotroph and the cells are present in pairs and as tetracocci. The culture can grow on methane (20%) as well as on a wide range of methanol from concentrations (0.02%-5%). Based on the comparison of genome data, FAME analysis, morphological characters and biochemical characters, FWC3 belongs to the tentatively and newly but not validly described genus ‘Methylotetracoccus’ of which only a single species strain was described, Methylotetracoccus oryzae C50C1. The ANI index between FWC3 and C50C1 strains is 94%, and the DDH value is 55.7%, less than the cut-off values 96% and 70%, respectively. The genome size of FWC3 is smaller (3.4 Mbp) compared to that of C50C1 (4.8 Mbp). Additionally, the FAME profile of FWC3 shows differences in cell wall fatty acid profiles compared to Methylotetracoccus oryzae C50C1. Also, there are other differences on the morphological, physiological and genomic levels. We propose FWC3 to be a member of a novel species of the genus Methylotetracoccus, for which the name Methylotetracoccus aquaticus is proposed. Also, an amended description of the genus Methylotetracoccus gen. nov. is given here. FWC3 is available in two international culture collections with the accession numbers: MCC 4198 (Microbial Culture collection, India) and JCM 33786 (Japan Collection of Microorganisms, Japan).

Characterization of siderophore producing arsenic-resistantStaphylococcus sp. strain TA6 isolated from contaminated groundwater of Jorhat, Assam and its possible role in arsenic geocycle

The presence of arsenic in sediments, carbonaceous rocks are geogenic, while its entry into the aquifers is mediated by several factors including microorganisms. It is well known that the microorganisms play a crucial role in the biogeochemical cycle of different elements. However, the precise role of bacteria in regulating the concentration of arsenic in Brahmaputra valley has not been investigated in detail. In this paper, we report the isolation of arsenic resistant bacterium TA6 with active arsenate reduction efficiency. The isolate was able to grow in arsenate concentration (250 mM) and arsenite (30 mM). Along with resistance to inorganic arsenic, it showed cross-tolerance to other heavy metals like Hg+2, Cd+2, Co+2, Ni+2, Cr+2. The bacterium also had a high siderophore activity (78.7 ± 0.004 μmol), which is positively correlated with the resistance aptitude. The biochemical test showed the TA6, a gram-positive bacterium which can hydrolyze starch and casein, produce catalase enzyme and utilizes citrate as a metabolic trait. Molecular and chemotaxonomic identification of TA6 based on 16S rRNA and FMAE analysis showed similarity with members ofStaphylococcusgenus with significant difference in sequence similarity and fatty acid composition. Based on 16S rRNA and FAME analysis it was identified asStaphylococcus sp.TA6. Rate of biotransformation showed bacterium could reduce ~88.2% of initial 2mM As(V) into As(III). The characterization of arsenate reductase enzyme with NADPH coupled assay showed the highest activity at pH 5.5 and temperature 50°C.