scholarly journals Uncovering a Microbial Enigma: Isolation and Characterization of the Streamer-Generating, Iron-Oxidizing, Acidophilic Bacterium “Ferrovum myxofaciens”

2013 ◽  
Vol 80 (2) ◽  
pp. 672-680 ◽  
Author(s):  
D. Barrie Johnson ◽  
Kevin B. Hallberg ◽  
Sabrina Hedrich
Keyword(s):  
Polymeric Materials ◽  
Molecular Techniques ◽  
Mine Waters ◽  
Content Type ◽  
Phenotypic Differences ◽  
New Family ◽  
Isolation And Characterization ◽  
Related Clone ◽  
Mine Impacted Water

ABSTRACTA betaproteobacterium, shown by molecular techniques to have widespread global distribution in extremely acidic (pH 2 to 4) ferruginous mine waters and also to be a major component of “acid streamer” growths in mine-impacted water bodies, has proven to be recalcitrant to enrichment and isolation. A modified “overlay” solid medium was devised and used to isolate this bacterium from a number of mine water samples. The physiological and phylogenetic characteristics of a pure culture of an isolate from an abandoned copper mine (“Ferrovum myxofaciens” strain P3G) have been elucidated. “F. myxofaciens” is an extremely acidophilic, psychrotolerant obligate autotroph that appears to use only ferrous iron as an electron donor and oxygen as an electron acceptor. It appears to use the Calvin-Benson-Bassham pathway to fix CO2and is diazotrophic. It also produces copious amounts of extracellular polymeric materials that cause cells to attach to each other (and to form small streamer-like growthin vitro) and to different solid surfaces. “F. myxofaciens” can catalyze the oxidative dissolution of pyrite and, like many other acidophiles, is tolerant of many (cationic) transition metals. “F. myxofaciens” and related clone sequences form a monophyletic group within theBetaproteobacteriadistantly related to classified orders, with genera of the familyNitrosomonadaceae(lithoautotrophic, ammonium-oxidizing neutrophiles) as the closest relatives. On the basis of the phylogenetic and phenotypic differences of “F. myxofaciens” and otherBetaproteobacteria, a new family, “Ferrovaceae,” and order, “Ferrovales,” within the classBetaproteobacteriaare proposed. “F. myxofaciens” is the first extreme acidophile to be described in the classBetaproteobacteria.

scholarly journals Plasmodium falciparum Line-Dependent Association ofIn VitroGrowth-Inhibitory Activity and Risk of Malaria

2012 ◽  
Vol 80 (5) ◽  
pp. 1900-1908 ◽  
Author(s):  
Josea Rono ◽  
Anna Färnert ◽  
Daniel Olsson ◽  
Faith Osier ◽  
Ingegerd Rooth ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Inhibitory Activity ◽  
Content Type ◽  
Phenotypic Differences ◽  
Erythrocyte Invasion ◽  
Growth Inhibitory ◽  
Growth Inhibitory Activity ◽  
Control Study

ABSTRACTPlasmodium falciparum's ability to invade erythrocytes is essential for its survival within the human host. Immune mechanisms that impair this ability are therefore expected to contribute to immunity against the parasite. Plasma of humans who are naturally exposed to malaria has been shown to have growth-inhibitory activity (GIA)in vitro. However, the importance of GIA in relation to protection from malaria has been unclear. In a case-control study nested within a longitudinally followed population in Tanzania, plasma samples collected at baseline from 171 individuals (55 cases and 116 age-matched controls) were assayed for GIA using threeP. falciparumlines (3D7, K1, and W2mef) chosen based on their erythrocyte invasion phenotypes. Distribution of GIA differed between the lines, with most samples inhibiting the growth of 3D7 and K1 and enhancing the growth of W2mef. GIA to 3D7 was associated with a reduced risk of malaria within 40 weeks of follow-up (odds ratio, 0.45; 95% confidence interval [CI], 0.21 to 0.96;P= 0.04), whereas GIA to K1 and W2mef was not. These results show that GIA, as well as its association with protection from malaria, is dependent on theP. falciparumline and can be explained by differences in erythrocyte invasion phenotypes between parasite lines. Our study contributes knowledge on the biological importance of growth inhibition and the potential influence ofP. falciparumerythrocyte invasion phenotypic differences on its relationship to protective immunity against malaria.

scholarly journals In Vitro Activity of Chlorhexidine Compared with Seven Antifungal Agents against 98 Fusarium Isolates Recovered from Fungal Keratitis Patients

2019 ◽  
Vol 63 (8) ◽  
Author(s):  
Claudy Oliveira dos Santos ◽  
Eva Kolwijck ◽  
Henrich A. van der Lee ◽  
Marlou C. Tehupeiory-Kooreman ◽  
Abdullah M. S. Al-Hatmi ◽  
...  
Keyword(s):  
Amphotericin B ◽  
Antifungal Agents ◽  
Culture Collection ◽  
Fungal Keratitis ◽  
Molecular Techniques ◽  
Temperate Climate ◽  
Eye Drops ◽  
Fungal Culture ◽  
Content Type

ABSTRACT Fungal keratitis is a common but severe eye infection in tropical and subtropical areas of the world. In regions with a temperate climate, the frequency of infection is rising in patients with contact lenses and following trauma. Early and adequate therapy is important to prevent disease progression and loss of vision. The management of Fusarium keratitis is complex, and the optimal treatment is not well defined. We investigated the in vitro activity of chlorhexidine and seven antifungal agents against a well-characterized collection of Fusarium isolates recovered from patients with Fusarium keratitis. The fungus culture collection of the Center of Expertise in Mycology Radboudumc/CWZ was searched for Fusarium isolates that were cultured from cornea scrapings, ocular biopsy specimens, eye swabs, and contact lens fluid containers from patients with suspected keratitis. The Fusarium isolates that were cultured from patients with confirmed keratitis were all identified using conventional and molecular techniques. Antifungal susceptibility testing was performed according to the EUCAST broth microdilution reference method. The antifungal agents tested included amphotericin B, voriconazole, posaconazole, miconazole, natamycin, 5-fluorocytosine, and caspofungin. In addition, the activity of chlorhexidine was determined. The fungal culture collection contained 98 Fusarium isolates of confirmed fungal keratitis cases from 83 Dutch patients and 15 Tanzanian patients. The isolates were collected between 2007 and 2017. Fusarium oxysporum (n = 24, 24.5%) was the most frequently isolated species followed by Fusarium solani sensu stricto (n = 18, 18.4%) and Fusarium petroliphilum (n = 11, 11.2%). Amphotericin B showed the most favorable in vitro inhibition of Fusarium species followed by natamycin, voriconazole, and chlorhexidine, while 5-fluorocytosine, posaconazole, miconazole, and caspofungin showed no relevant inhibiting effect. However, chlorhexidine showed fungicidal activity against 90% of F. oxysporum strains and 100% of the F. solani strains. Our study supports the clinical efficacy of chlorhexidine and therefore warrants its further clinical evaluation for primary therapy of fungal keratitis, particularly in low and middle income countries where fungal keratitis is much more frequent and, currently, antifungal eye drops are often unavailable.

scholarly journals In Vitro Isolation and Characterization of Oxazolidinone-Resistant Mycobacterium tuberculosis

2017 ◽  
Vol 61 (10) ◽  
Author(s):  
Matthew B. McNeil ◽  
Devon D. Dennison ◽  
Catherine D. Shelton ◽  
Tanya Parish
Keyword(s):  
Mycobacterium Tuberculosis ◽  
23S Rrna ◽  
Clinical Settings ◽  
Content Type ◽  
Isolation And Characterization ◽  
Mutant Strains ◽  
Ribosomal Protein L3 ◽  
Resistant Strains

ABSTRACT Oxazolidinones are promising candidates for the treatment of Mycobacterium tuberculosis infections. We isolated linezolid-resistant strains from H37Rv (Euro-American) and HN878 (East-Asian) strains; resistance frequencies were similar in the two strains. Mutations were identified in ribosomal protein L3 (RplC) and the 23S rRNA (rrl). All mutant strains were cross resistant to sutezolid; a subset was cross resistant to chloramphenicol. Mutations in rrl led to growth impairment and decreased fitness that may limit spread in clinical settings.

scholarly journals Cryptococcus neoformans Dual GDP-Mannose Transporters and Their Role in Biology and Virulence

2014 ◽  
Vol 13 (6) ◽  
pp. 832-842 ◽  
Author(s):  
Zhuo A. Wang ◽  
Cara L. Griffith ◽  
Michael L. Skowyra ◽  
Nichole Salinas ◽  
Matthew Williams ◽  
...  
Keyword(s):  
Cryptococcus Neoformans ◽  
Protein Glycosylation ◽  
Content Type ◽  
Phenotypic Differences ◽  
Major Virulence Factor ◽  
Transmembrane Transporters ◽  
Mutant Cells ◽  
Made In

ABSTRACTCryptococcus neoformansis an opportunistic yeast responsible for lethal meningoencephalitis in humans. This pathogen elaborates a polysaccharide capsule, which is its major virulence factor. Mannose constitutes over one-half of the capsule mass and is also extensively utilized in cell wall synthesis and in glycosylation of proteins and lipids. The activated mannose donor for most biosynthetic reactions, GDP-mannose, is made in the cytosol, although it is primarily consumed in secretory organelles. This compartmentalization necessitates specific transmembrane transporters to make the donor available for glycan synthesis. We previously identified two cryptococcal GDP-mannose transporters, Gmt1 and Gmt2. Biochemical studies of each protein expressed inSaccharomyces cerevisiaeshowed that both are functional, with similar kinetics and substrate specificitiesin vitro. We have now examined these proteinsin vivoand demonstrate that cells lacking Gmt1 show significant phenotypic differences from those lacking Gmt2 in terms of growth, colony morphology, protein glycosylation, and capsule phenotypes. Some of these observations may be explained by differential expression of the two genes, but others suggest that the two proteins play overlapping but nonidentical roles in cryptococcal biology. Furthermore,gmt1 gmt2double mutant cells, which are unexpectedly viable, exhibit severe defects in capsule synthesis and protein glycosylation and are avirulent in mouse models of cryptococcosis.

scholarly journals Shewanella algae and Microbulbifer elongatus from marine macro-algae – isolation and characterization of agar-hydrolysing bacteria

2020 ◽  
Vol 2 (11) ◽  
Author(s):  
Keerthana Ponni Kandasamy ◽  
Radhesh Krishnan Subramanian ◽  
Radhakrishnan Srinivasan ◽  
Sengali Ragunath ◽  
G. Balaji ◽  
...  
Keyword(s):  
Type Species ◽  
Degradation Products ◽  
Accession Number ◽  
Content Type ◽  
Link Type ◽  
Isolation And Characterization ◽  
Shewanella Algae ◽  
Agarase Activity ◽  
Macro Algae

Macro-algae are a good source of agar oligosaccharides, which can be obtained through bacterial enzymatic hydrolysis. The agarase enzyme secreted by the micro-organisms cleaves the cell wall of the algae and releases agar oligosaccharides as degradation products with various applications. Agarolytic bacteria were isolated from the marine algae Kappaphycus sp., and Sargassum sp., and studied for their agar-degrading properties. Among the 70 isolates, 2 isolates (A13 and Sg8) showed agarase activity in in vitro assays. The maximum agarolytic index was recorded in the isolate Sg8 (3.75 mm and 4.29 µg ml−1 agarase activity), followed by the isolate A13 (2.53 mm and 2.6 µg ml−1 agarase activity). Optimum agarase production of isolate Sg8 was observed at pH7 and at a temperature of 25 °C in 24–48 h, whereas for isolate A13 the optimum production was at pH7 and at a temperature of 37 °C in 48 h. The identities of the agarolytic isolates (Sg8 and A13) were confirmed based on microscopy, morphological, biochemical and molecular analysis as Shewanella algae [National Center for Biotechnology Information (NCBI) GenBank accession number MK121204.1] and Microbulbifer elongatus [NCBI GenBank accession number MK825484.1], respectively.

scholarly journals Immunomodulatory Metabolites Released by the Frog-Killing Fungus Batrachochytrium dendrobatidis

2015 ◽  
Vol 83 (12) ◽  
pp. 4565-4570 ◽  
Author(s):  
Louise A. Rollins-Smith ◽  
J. Scott Fites ◽  
Laura K. Reinert ◽  
Andrea R. Shiakolas ◽  
Thomas P. Umile ◽  
...  
Keyword(s):  
Batrachochytrium Dendrobatidis ◽  
Leukemia Cell ◽  
Local Environment ◽  
Amphibian Declines ◽  
Leukemia Cell Line ◽  
Content Type ◽  
Isolation And Characterization ◽  
Human T Cell

Batrachochytrium dendrobatidisis a fungal pathogen in the phylum Chytridiomycota that causes the skin disease chytridiomycosis. Chytridiomycosis is considered an emerging infectious disease linked to worldwide amphibian declines and extinctions. Although amphibians have well-developed immune defenses, clearance of this pathogen from the skin is often impaired. Previously, we showed that the adaptive immune system is involved in the control of the pathogen, butB. dendrobatidisreleases factors that inhibitin vitroandin vivolymphocyte responses and induce lymphocyte apoptosis. Little is known about the nature of the inhibitory factors released by this fungus. Here, we describe the isolation and characterization of three fungal metabolites produced byB. dendrobatidisbut not by the closely related nonpathogenic chytridHomolaphlyctis polyrhiza. These metabolites are methylthioadenosine (MTA), tryptophan, and an oxidized product of tryptophan, kynurenine (Kyn). Independently, both MTA and Kyn inhibit the survival and proliferation of amphibian lymphocytes and the Jurkat human T cell leukemia cell line. However, working together, they become effective at much lower concentrations. We hypothesize thatB. dendrobatidiscan adapt its metabolism to release products that alter the local environment in the skin to inhibit immunity and enhance the survival of the pathogen.

scholarly journals CYP101J2, CYP101J3, and CYP101J4, 1,8-Cineole-Hydroxylating Cytochrome P450 Monooxygenases from Sphingobium yanoikuyae Strain B2

2016 ◽  
Vol 82 (22) ◽  
pp. 6507-6517 ◽  
Author(s):  
Birgit Unterweger ◽  
Dieter M. Bulach ◽  
Judith Scoble ◽  
David J. Midgley ◽  
Paul Greenfield ◽  
...  
Keyword(s):  
Cytochrome P450 ◽  
Amino Acid ◽  
Amino Acid Sequences ◽  
Cytochrome P450 Monooxygenases ◽  
Content Type ◽  
E Coli ◽  
P450 Monooxygenases ◽  
Isolation And Characterization ◽  
Starting Point

ABSTRACTWe report the isolation and characterization of three new cytochrome P450 monooxygenases: CYP101J2, CYP101J3, and CYP101J4. These P450s were derived fromSphingobium yanoikuyaeB2, a strain that was isolated from activated sludge based on its ability to fully mineralize 1,8-cineole. Genome sequencing of this strain in combination with purification of native 1,8-cineole-binding proteins enabled identification of 1,8-cineole-binding P450s. The P450 enzymes were cloned, heterologously expressed (N-terminally His6tagged) inEscherichia coliBL21(DE3), purified, and spectroscopically characterized. Recombinant whole-cell biotransformation inE. colidemonstrated that all three P450s hydroxylate 1,8-cineole using electron transport partners fromE. colito yield a product putatively identified as (1S)-2α-hydroxy-1,8-cineole or (1R)-6α-hydroxy-1,8-cineole. The new P450s belong to the CYP101 family and share 47% and 44% identity with other 1,8-cineole-hydroxylating members found inNovosphingobium aromaticivoransandPseudomonas putida. Compared to P450cin(CYP176A1), a 1,8-cineole-hydroxylating P450 fromCitrobacter braakii, these enzymes share less than 30% amino acid sequence identity and hydroxylate 1,8-cineole in a different orientation. Expansion of the enzyme toolbox for modification of 1,8-cineole creates a starting point for use of hydroxylated derivatives in a range of industrial applications.IMPORTANCECYP101J2, CYP101J3, and CYP101J4 are cytochrome P450 monooxygenases fromS. yanoikuyaeB2 that hydroxylate the monoterpenoid 1,8-cineole. These enzymes not only play an important role in microbial degradation of this plant-based chemical but also provide an interesting route to synthesize oxygenated 1,8-cineole derivatives for applications as natural flavor and fragrance precursors or incorporation into polymers. The P450 cytochromes also provide an interesting basis from which to compare other enzymes with a similar function and expand the CYP101 family. This could eventually provide enough bacterial parental enzymes with similar amino acid sequences to enablein vitroevolution via DNA shuffling.

scholarly journals Characterization of Two Novel Lipopolysaccharide Phosphoethanolamine Transferases in Pasteurella multocida and Their Role in Resistance to Cathelicidin-2

2017 ◽  
Vol 85 (11) ◽  
Author(s):  
Marina Harper ◽  
Amy Wright ◽  
Frank St. Michael ◽  
Jianjun Li ◽  
Deanna Deveson Lucas ◽  
...  
Keyword(s):  
Pasteurella Multocida ◽  
Lipid A ◽  
Bacterial Pathogen ◽  
Cationic Antimicrobial Peptide ◽  
Gram Negative ◽  
Content Type ◽  
New Family ◽  
Maximum Protection ◽  
Phosphoethanolamine Transferases

ABSTRACT The lipopolysaccharide (LPS) produced by the Gram-negative bacterial pathogen Pasteurella multocida has phosphoethanolamine (PEtn) residues attached to lipid A, 3-deoxy-d-manno-octulosonic acid (Kdo), heptose, and galactose. In this report, we show that PEtn is transferred to lipid A by the P. multocida EptA homologue, PetL, and is transferred to galactose by a novel PEtn transferase that is unique to P. multocida called PetG. Transcriptomic analyses indicated that petL expression was positively regulated by the global regulator Fis and negatively regulated by an Hfq-dependent small RNA. Importantly, we have identified a novel PEtn transferase called PetK that is responsible for PEtn addition to the single Kdo molecule (Kdo1), directly linked to lipid A in the P. multocida glycoform A LPS. In vitro assays showed that the presence of a functional petL and petK, and therefore the presence of PEtn on lipid A and Kdo1, was essential for resistance to the cationic, antimicrobial peptide cathelicidin-2. The importance of PEtn on Kdo1 and the identification of the transferase responsible for this addition have not previously been shown. Phylogenetic analysis revealed that PetK is the first representative of a new family of predicted PEtn transferases. The PetK family consists of uncharacterized proteins from a range of Gram-negative bacteria that produce LPS glycoforms with only one Kdo molecule, including pathogenic species within the genera Vibrio, Bordetella, and Haemophilus. We predict that many of these bacteria will require the addition of PEtn to Kdo for maximum protection against host antimicrobial peptides.

scholarly journals Cryptosporidium Pathogenicity and Virulence

2013 ◽  
Vol 26 (1) ◽  
pp. 115-134 ◽  
Author(s):  
Maha Bouzid ◽  
Paul R. Hunter ◽  
Rachel M. Chalmers ◽  
Kevin M. Tyler
Keyword(s):  
Virulence Factors ◽  
Genetic Manipulation ◽  
Protozoan Parasite ◽  
Molecular Techniques ◽  
Host Susceptibility ◽  
Content Type ◽  
Susceptibility To Infection ◽  
Substantial Progress ◽  
Made In

SUMMARYCryptosporidiumis a protozoan parasite of medical and veterinary importance that causes gastroenteritis in a variety of vertebrate hosts. Several studies have reported different degrees of pathogenicity and virulence amongCryptosporidiumspecies and isolates of the same species as well as evidence of variation in host susceptibility to infection. The identification and validation ofCryptosporidiumvirulence factors have been hindered by the renowned difficulties pertaining to thein vitroculture and genetic manipulation of this parasite. Nevertheless, substantial progress has been made in identifying putative virulence factors forCryptosporidium. This progress has been accelerated since the publication of theCryptosporidium parvumandC. hominisgenomes, with the characterization of over 25 putative virulence factors identified by using a variety of immunological and molecular techniques and which are proposed to be involved in aspects of host-pathogen interactions from adhesion and locomotion to invasion and proliferation. Progress has also been made in the contribution of host factors that are associated with variations in both the severity and risk of infection. Here we provide a review comprised of the current state of knowledge onCryptosporidiuminfectivity, pathogenesis, and transmissibility in light of our contemporary understanding of microbial virulence.

scholarly journals In VitroandIn VivoActivities of Antibiotic PM181104

2013 ◽  
Vol 57 (11) ◽  
pp. 5315-5319 ◽  
Author(s):  
Girish Mahajan ◽  
Becky Thomas ◽  
Rajashri Parab ◽  
Zarine E. Patel ◽  
Sandip Kuldharan ◽  
...  
Keyword(s):  
Antibacterial Activity ◽  
Cyclic Peptide ◽  
Peptide Antibiotic ◽  
Active Metabolites ◽  
Gram Positive ◽  
Single Digit ◽  
Content Type ◽  
Isolation And Characterization

ABSTRACTDrug resistance has become a global threat that, if not addressed, may return us to the preantibiotic era. A way to overcome the problem of growing incidence of global antibiotic resistance is to introduce compounds belonging to classes that are new to the clinic. During a screening of the marine microbial extract library for new antibiotics, one of the extracts showed promising antibacterial activity against Gram-positive organisms. Bioactivity-guided isolation and characterization of active metabolites led to the discovery of a novel thiazolyl cyclic-peptide antibiotic, PM181104. It was isolated and characterized from a marine sponge-associated actinobacterium strain of the genusKocuria(MTCC 5269). The compound exhibited a potentin vitroantibacterial activity against a broad range of Gram-positive bacteria, including methicillin-resistantStaphylococcus aureus(MRSA) and vancomycin-resistant enterococci (VRE). The MIC values evaluated for the compound were found to be in the single-digit nanomolar range. Inin vivostudies of PM181104 in a BALB/c murine septicemia model, the compound displayed 100% effective dose (ED100) values of 2.5 and 5.0 mg/kg of body weight against MRSA and 10.0 mg/kg against VRE. In this report,in vitroandin vivostudies of PM181104 are described.

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