Mechanical deformation of elastomer medical devices can enable microbial surface colonization.

Elastomers such as silicone are common in medical devices (catheters, prosthetic implants, endoscopes), but they remain prone to microbial colonization and biofilm infections. For the first time, our work shows that rates of microbial surface attachment to polydimethylsiloxane (PDMS) silicone can be significantly affected by mechanical deformation. For a section of bent commercial catheter tubing, bacteria (P. aeruginosa) show a strong preference for the 'convex' side compared to the 'concave' side, by a factor of 4.2. Further testing of cast PDMS materials in bending only showed a significant difference for samples that were manually wiped (damaged) beforehand (1.75 × 104 and 6.02 × 103 cells/mm2 on the convex and concave sides, respectively). We demonstrate that surface microcracks in elastomers are opened under tensile stress (convex bending) to become 'activated' as sites for microbial colonization. This work demonstrates that the high elastic limit of elastomers enables these microcracks to reversibly open and close, as 'dynamic defects'. Commercial catheters have relatively high surface roughness inherent to manufacturing, but we show that even manual wiping of newly-cast PDMS is sufficient to generate surface microcracks. We consider the implication for medical devices that feature sustained, surgical, or cyclic deformation, in which localized tensile conditions may expose these surface defects to opportunistic microbes. As a result, our work showcases serious potential problems in the widespread usage and development of elastomers in medical devices.

Glycoside hydrolase processing of the Pel polysaccharide alters biofilm biomechanics and Pseudomonas aeruginosa virulence.

Pel exopolysaccharide biosynthetic loci are phylogenetically widespread biofilm matrix determinants in bacteria. In Pseudomonas aeruginosa, Pel is crucial for cell-to-cell interactions and reducing susceptibility to antibiotic and mucolytic treatments. While genes encoding glycoside hydrolases have long been linked to biofilm exopolysaccharide biosynthesis, their physiological role in biofilm development is unclear. Here we demonstrate that the glycoside hydrolase activity of P. aeruginosa PelA decreases adherent biofilm biomass and is responsible for generating the low molecular weight secreted form of the Pel exopolysaccharide. We show that the generation of secreted Pel contributes to the biomechanical properties of the biofilm and decreases the virulence of P. aeruginosa in Caenorhabditis elegans and Drosophila melanogaster. Our results reveal that glycoside hydrolases found in exopolysaccharide biosynthetic systems can help shape the soft matter attributes of a biofilm and propose that secreted matrix components be referred to as matrix associated to better reflect their influence.

Antioxidant and antimicrobial activities and UPLC-ESI-MS/MS polyphenolic profile of sweet orange peel extracts.

With growing consumer awareness, exploitation of renewable resources is cost-effective and environment friendly. This work examines the potential of citrus peels as natural antioxidants and antimicrobials for food preservation. Extraction yield, total soluble phenols and flavonoids of various citrus peels (sweet orange, lemon, tangerine and grapefruit) were optimized by varying the solvent type. While the highest extract yield (~16 â€‹g/100g) was obtained from the sweet orange peels in methanol, extraction with ethanol maximized the concentration of total phenols and flavonoids (~80 â€‹mg catechol equivalents/100 â€‹g dry weight). In addition, sweet orange peel extract showed the highest DPPH, ABTS and hydroxyl radical scavenging values. UPLC-ESI-MS/MS analysis of aqueous and ethanolic extracts of sweet orange peels revealed more than 40 polyphenolic compounds including phenolic acids and flavonoids, some of which have not been previously reported. The predominant polyphenols were narirutin, naringin, hesperetin-7-O-rutinoside naringenin, quinic acid, hesperetin, datiscetin-3-O-rutinoside and sakuranetin. The incorporation of sweet orange peel extract into two vegetable oils enhanced their oxidative stability. In addition, all citrus peel extracts possessed high antimicrobial activity against several food-borne pathogens, and the activity was highest for the sweet orange peel extract. Overall results suggested the great potential of sweet orange peels as natural antioxidant and antimicrobials, which can be efficiently extracted using a simple and low-cost method, for enhancing the storage stability and safety of vegetable oils.

Preventing Pseudomonas aeruginosa Biofilms on Indwelling Catheters by Surface-Bound Enzymes.

Implanted medical devices such as central venous catheters are highly susceptible to microbial colonization and biofilm formation and are a major risk factor for nosocomial infections. The opportunistic pathogen Pseudomonas aeruginosa uses exopolysaccharides, such as Psl, for both initial surface attachment and biofilm formation. We have previously shown that chemically immobilizing the Psl-specific glycoside hydrolase, PslGh, to a material surface can inhibit P. aeruginosa biofilm formation. Herein, we show that PslGh can be uniformly immobilized on the lumen surface of medical-grade, commercial polyethylene, polyurethane, and polydimethylsiloxane (silicone) catheter tubing. We confirmed that the surface-bound PslGh was uniformly distributed along the catheter length and remained active even after storage for 30 days at 4 °C. P. aeruginosa colonization and biofilm formation under dynamic flow culture conditions in vitro showed a 3-log reduction in the number of bacteria during the first 11 days, and a 2-log reduction by day 14 for PslGh-modified PE-100 catheters, compared to untreated catheter controls. In an in vivo rat infection model, PslGh-modified PE-100 catheters showed a ∼1.5-log reduction in the colonization of the clinical P. aeruginosa ATCC 27853 strain after 24 h. These results demonstrate the robust ability of surface-bound glycoside hydrolase enzymes to inhibit biofilm formation and their potential to reduce rates of device-associated infections.

Isolation and characterization of a novel lutein-producing marine microalga using high throughput screening.

A novel highly selective lutein-producing marine microalga was isolated and identified using high throughput screening (HTS). Phylogenetic analysis based on 18S rRNA gene sequence revealed it belongs to a microalgae class (Trebouxiophyceae), and is closely related to Auxenochlorella spp. The novel strain was designated as Auxenochlorella sp. strain LEU27. Based on HPLC-DAD and HPLC-MS analyses, strain LEU27 produced a total of 1203.9 ±â€¯98 µg g-1 dry cells of carotenoids, including a remarkable amount (996.6 ±â€¯98 µg g-1 dry cells) of pure lutein under heterotrophic growth conditions. This is the first report that describes a natural, highly selective lutein-producing marine microalga isolated by HTS approach.

Rapid and Selective Screening Method for Isolation and Identification of Carotenoid-Producing Bacteria.

Carotenoids are naturally occurring yellow to red pigments with many biological activities including antioxidant, anticancer, anti-inflammatory, membrane stabilizers, and precursors for vitamin A. These biological activities are linked with many health benefits (e.g., anticarcinogenic activity, prevention of chronic diseases, etc.), which grew the interest of several industrial sectors especially in food, feed, nutraceuticals, cosmetics, and pharmaceutical industries. The production of natural carotenoids from microbial sources such as bacteria can help meet the growing global market of carotenoids estimated at $1.5 billion in 2014 and is expected to reach 1.8 billion in 2019. This chapter demonstrates, step-by-step, the development of a rapid and selective screening method for isolation and identification of carotenoid-producing microorganisms and their carotenoid analysis. This method involves three main procedures: UV treatment, sequencing analysis of 16S rRNA genes, and carotenoids analysis using rapid and effective HPLC-diode array-MS methods.

Purification and Identification of Astaxanthin and Its Novel Derivative Produced by Radio-tolerant Sphingomonas astaxanthinifaciens.

The red diketocarotenoid, astaxanthin, exhibits extraordinary health-promoting activities such as antioxidant, anti-inflammatory, antitumor, and immune booster, which may potentially protect against many degenerative diseases such as cancers, heart diseases, and exercise-induced fatigue. These numerous health benefits and consumer interest in natural products have therefore increased the market demand of astaxanthin as a nutraceutical and medicinal ingredient in food, aquaculture feed, and pharmaceutical industries. Consequently, many research efforts have been made to discover novel microbial sources with effective biotechnological production of astaxanthin. Using a rapid screening method based on 16S rRNA gene, and effective HPLC-Diode array-MS methods for carotenoids analysis, we isolated a novel astaxanthin-producing bacterium (strain TDMA-17T) that belongs to the family Sphingomonadaceae (Asker et al., FEMS Microbiol Lett 273:140-148, 2007).In this chapter, we provide a comprehensive description of the methods used for the analysis and identification of carotenoids produced by strain TDMA-17T. We will also describe the methods of isolation and identification for a novel bacterial carotenoid (an astaxanthin derivative), a major carotenoid that is produced by the novel strain. Finally, the identification methods of the novel strain will be summarized.

Screening, Isolation, and Identification of Zeaxanthin-Producing Bacteria.

Zeaxanthin is a yellow xanthophyll, dihydroxy-carotenoid, that is naturally found in some of the green, orange, and yellow vegetables and fruits and has a powerful antioxidant activity. Epidemiological evidences suggest that increasing the consumption of zeaxanthin in the diet is associated with a lower risk of age-related macular degeneration (ARMD) and cataracts, two of the leading causes of blindness in the world. Zeaxanthin is a promising nutraceutical/colorant with many applications in feed, food, and pharmaceutical industries. Currently, the commercial production of zeaxanthin is dependent on synthetic routes with limitation in production from biological sources. However, the biotechnological production of natural zeaxanthin is favored due to its safety, potential large-scale production and consumers' preference for natural additives. In this chapter, we describe a rapid screening method based on 16S rRNA gene sequencing and effective HPLC with diode array detector/MS methods for the isolation and identification of zeaxanthin-producing bacteria and their carotenoid analysis.

Food-Safe Modification of Stainless Steel Food-Processing Surfaces to Reduce Bacterial Biofilms.

Biofilm formation on stainless steel (SS) surfaces of food-processing plants, leading to food-borne illness outbreaks, is enabled by the attachment and confinement of pathogens within microscale cavities of surface roughness (grooves, scratches). We report foodsafe oil-based slippery coatings (FOSCs) for food-processing surfaces that suppress bacterial adherence and biofilm formation by trapping residual oil lubricant within these surface cavities to block microbial growth. SS surfaces were chemically functionalized with alkylphosphonic acid to preferentially wet a layer of food-grade oil. FOSCs reduced the effective surface roughness, the adhesion of organic food residue, and bacteria. FOSCs significantly reduced Pseudomonas aeruginosa biofilm formation on standard roughness SS-316 by 5 log CFU cm-2, and by 3 log CFU cm-2 for mirror-finished SS. FOSCs also enhanced surface cleanability, which we measured by bacterial counts after conventional detergent cleaning. Importantly, both SS grades maintained their antibiofilm activity after the erosion of the oil layer by surface wear with glass beads, which suggests that there is a residual volume of oil that remains to block surface cavity defects. These results indicate the potential of such low-cost, scalable approaches to enhance the cleanability of SS food-processing surfaces and improve food safety by reducing biofilm growth.

Non-eluting, surface-bound enzymes disrupt surface attachment of bacteria by continuous biofilm polysaccharide degradation.

Bacterial colonization and biofilm formation on surfaces are typically mediated by the deposition of exopolysaccharides and conditioning protein layers. Pseudomonas aeruginosa is a nosocomial opportunistic pathogen that utilizes strain-specific exopolysaccharides such as Psl, Pel or alginate for both initial surface attachment and biofilm formation. To generate surfaces that resist P. aeruginosa colonization, we covalently bound a Psl-specific glycoside hydrolase (PslGh) to several, chemically-distinct surfaces using amine functionalization (APTMS) and glutaraldehyde (GDA) linking. In situ quartz crystal microbalance (QCM) experiments and fluorescence microscopy demonstrated a complete lack of Psl adsorption on the PslGh-bound surfaces. Covalently-bound PslGh was also found to significantly reduce P. aeruginosa surface attachment and biofilm formation over extended growth periods (8 days). The PslGh surfaces showed a ∼99.9% (∼3-log) reduction in surface associated bacteria compared to control (untreated) surfaces, or those treated with inactive enzyme. This work demonstrates a non-eluting 'bioactive' surface that specifically targets a mechanism of cell adhesion, and that surface-bound glycoside hydrolase can significantly reduce surface colonization of bacteria through local, continuous enzymatic degradation of exopolysaccharide (Psl). These results have significant implications for the surface design of medical devices to keep bacteria in a planktonic state, and therefore susceptible to antibiotics and antimicrobials.

Screening and profiling of natural ketocarotenoids from environmental aquatic bacterial isolates.

Ketocarotenoids are high-value natural pigments. The red diketocarotenoid astaxanthin particularly exhibits an extraordinary antioxidant activity, which raises its market demand for foods and nutraceuticals. We screened for ketocarotenoid-producing bacteria from both marine and freshwater environments. Phylogenetic analysis, based on 16S rRNA gene sequence, revealed 37 potential producers of ketocarotenoids that are related to α-proteobacteria, comprising 32 strains of Brevundimonas and 5 strains of Erythrobacter. Carotenoids analysis by HPLC-DAD and HPLC-MS revealed two groups; astaxanthin-producers (28 Brevundimonas strains) and adonixanthin-producers (Five Brevundimonas and 5 Erythrobacter strains). Strain FrW-Asx16 exhibited the highest carotenoid production (1060 µg g-1 dry cells with 16.6% astaxanthin). Strain FrW-Asx-5 producing 946.1 µg g-1 dry cells carotenoid exhibited the highest astaxanthin content (∼46%). The most intriguing result is the potential of producing natural colorants from freshwater bacterial isolates, and with high productivity and selectivity, suggesting a great promise for their application in food.

Evidence of coexisting microemulsion droplets in oil-in-water emulsions revealed by 2D DOSY 1H NMR.

Optimizing the macroscopic properties, shelf-life and stability of emulsion products requires a better understanding of the microstructural characteristics such as the type (nano, micro and macro) and the relative distribution of components (i.e., oil and surfactant) within the emulsion droplets. We used Diffusion-Ordered NMR Spectroscopy (DOSY NMR) to evaluate these characteristics in model oil-in-water emulsion containing Tween 80 and medium chain triglycerides (MCT). At low MCT concentrations, the solutions were transparent but from 1 to 5 wt% MCT, they became translucent then opaque. 1 wt% MCT was the upper boundary for the appearance of nanoemulsion phase. From the decays of the chemical shift signals of MCT and Tween 80, the DOSY results clearly demonstrate that the self-diffusion coefficients (D) are dependent on oil concentration. Small microemulsion droplets of almost uniform size (d = 12-22 nm) coexist with two sets of large nanoemulsion (d < 200 nm) and emulsion (d > 200 nm) droplets. The large droplets increase significantly in size with increasing MCT. The most striking result is the clear evidence for the presence of microemulsion droplets of nearly uniform size in the aqueous phase from below to above the nanoemulsion transition concentration at 1 wt% MCT.

A novel radio-tolerant astaxanthin-producing bacterium reveals a new astaxanthin derivative: astaxanthin dirhamnoside.

Astaxanthin is a red ketocarotenoid that exhibits extraordinary health-promoting activities such as antioxidant, anti-inflammatory, antitumor, and immune booster. The recent discovery of the beneficial roles of astaxanthin against many degenerative diseases such as cancers, heart diseases, and exercise-induced fatigue has raised its market demand as a nutraceutical and medicinal ingredient in aquaculture, food, and pharmaceutical industries. To satisfy the growing demand for this high-value nutraceuticals ingredient and consumer interest in natural products, many research efforts are being made to discover novel microbial producers with effective biotechnological production of astaxanthin. Using a rapid screening method based on 16S rRNA gene, and effective HPLC-Diodearray-MS methods for carotenoids analysis, we succeeded to isolate a unique astaxanthin-producing bacterium (strain TDMA-17(T)) that belongs to the family Sphingomonadaceae (Asker et al., Appl Microbiol Biotechnol 77: 383-392, 2007). In this chapter, we provide a detailed description of effective HPLC-Diodearray-MS methods for rapid analysis and identification of the carotenoids produced by strain TDMA-17(T). We also describe the methods of isolation and identification for a novel bacterial carotenoid (astaxanthin derivative), a major carotenoid that is produced by strain TDMA-17(T). Finally, we describe the polyphasic taxonomic analysis of strain TDMA-17(T) and the description of a novel species belonging to genus Sphingomonas.

Isolation, characterization, and diversity of novel radiotolerant carotenoid-producing bacteria.

Carotenoids are natural pigments that exhibit many biological functions, such as antioxidants (i.e., promote oxidative stress resistance), membrane stabilizers, and precursors for vitamin A. The link between these biological activities and many health benefits (e.g., anticarcinogenic activity, prevention of chronic diseases, etc.) has raised the interest of several industrial sectors, especially in the cosmetics and pharmaceutical industries. The use of microorganisms in biotechnology to produce carotenoids is favorable by consumer and can help meet the growing demand for these bioactive compounds in the food, feed, and pharmaceutical industries. This methodological chapter details the development of a rapid and selective screening method for isolation and identification of carotenoid-producing microorganisms based on UV treatment, sequencing analysis of 16S rRNA genes, and carotenoids' analysis using rapid and effective High-Performance Liquid Chromatography-Diodearray-MS methods. The results of a comprehensive 16S rRNA gene-based phylogenetic analysis revealed a diversity of carotenoid-producing microorganisms (104 isolates) that were isolated at a high frequency from water samples collected at Misasa (Tottori, Japan), a region known for its high natural radioactivity content. These carotenoid-producing isolates were classified into 38 different species belonging to 7 bacterial classes (Flavobacteria, Sphingobacteria, α-Proteobacteria, γ-Proteobacteria, Deinococci, Actinobacteria, and Bacilli). The carotenoids produced by the isolates were zeaxanthin (6 strains), dihydroxyastaxanthin (24 strains), astaxanthin (27 strains), canthaxanthin (10 strains), and unidentified molecular species that were produced by the isolates related to Deinococcus, Exiguobacterium, and Flectobacillus. Here, we describe the methods used to isolate and classify these microorganisms.

Novel zeaxanthin-producing bacteria isolated from a radioactive hot spring water.

Zeaxanthin is a powerful antioxidant that is widely found in vegetables and fruits. Epidemiological evidences suggest that increasing the consumption of zeaxanthin in the diet is associated with a lower risk of age-related macular degeneration, helps prevent glaucoma and cataracts, and supports normal eye health. Zeaxanthin is a promising nutraceutical with many applications in the feed, food, and pharmaceutical industries. Currently, the commercial production of zeaxanthin is still dependant on synthetic routes with limitation for the biological one. Nevertheless, the biotechnological production of zeaxanthin is emerging due to its safety, potential large-scale production, and consumers' demand and preference for natural additives. Using a rapid screening method based on 16S rRNA gene and effective high-performance liquid chromatography (HPLC)-Diodearray-MS methods for carotenoids' analysis, we isolated effective zeaxanthin-producing bacteria (strain TDMA-5(T) and -16(T)) that belong to the family Sphingobacteriaceae and Sphingomonadaceae, respectively. In this chapter, we provide a detailed description of the HPLC-Diodearray-MS methods used for rapid analysis and identification of the carotenoids produced by both strains. In addition, the polyphasic taxonomic analysis of both novel strains and the description of a novel species and genus are described.

Deinococcus depolymerans sp. nov., a gamma- and UV-radiation-resistant bacterium, isolated from a naturally radioactive site.

Four gamma- and UV-radiation-resistant bacterial strains, designated TDMA-24(T), TDMA-24-2, TDMA-24-3 and TDMA-24-4, were isolated from a fresh-water sample collected at Misasa, Tottori, Japan. Cells of these strains were Gram-reaction-positive, non-motile, non-spore-forming, rod-shaped and formed red colonies. The genomic DNA G+C contents ranged from 70.5 to 70.6 mol%. Phylogenetic analysis based on 16S rRNA gene sequences indicated that the novel isolates belong to the genus Deinococcus, the highest sequence similarities being with Deinococcus aquaticus PB314(T) (98%) and Deinococcus caeni Ho-08(T) (97 %). The polar lipid profile of strain TDMA-24(T) comprised three unidentified phosphoglycolipids, five unidentified glycolipids and seven unidentified polar lipids. MK-8 was the predominant respiratory quinone. Major fatty acids were iso-C(15 : 0), C(15 : 1)ω6c, C(15 : 0), C(16 : 0) and summed feature 3 (iso-C(15 : 0) 2-OH and/or C(16 : 1)ω7c). On the basis of their phylogenetic positions and chemotaxonomic and phenotypic characteristics, the novel isolates represent a novel species of the genus Deinococcus, for which the name Deinococcus depolymerans sp. nov. is proposed. The type strain is TDMA-24(T) ( = JCM 14369(T)  = NBRC 102115(T)  = CCUG 53609(T)).

Impact of surfactant properties on oxidative stability of beta-carotene encapsulated within solid lipid nanoparticles.

The impact of surfactant type on the physical and chemical stability of solid lipid nanoparticle (SLN) suspensions containing encapsulated beta-carotene was investigated. Oil-in-water emulsions were formed by homogenizing 10% w/w lipid phase (1 mg/g beta-carotene in carrier lipid) and 90% w/w aqueous phase (surfactant + cosurfactant) at pH 7 and 75 degrees C and then cooling to 20 degrees C. The impact of surfactant type was investigated using aqueous phases containing different water-soluble surfactants [2.4% w/w high-melting (HM) lecithin, 2.4% w/w low-melting (LM) lecithin, and 1.4% w/w Tween 60 or 1.4% w/w Tween 80] and a cosurfactant (0.6% taurodeoxycholate). The impact of the physical state of the carrier lipid was investigated by using either a high melting point lipid (tripalmitin) to form solid particles or a low melting point lipid (medium chain triglycerides, MCT) to form liquid droplets. A higher fraction of alpha-crystals was detected in solid particles prepared with high-melting surfactants (HM-lecithin and Tween 60) than with low-melting surfactants (LM-lecithin and Tween 80). With the exception of the HM-lecithin-coated solid particles, the suspensions were stable to particle aggregation during 21 days of storage. beta-Carotene degradation after 21 days of storage was 11, 97, 100, and 91% in the solid particles (tripalmitin) and 16, 21, 95, and 90% in the liquid droplets (MCT) for HM-lecithin, LM-lecithin, Tween 80, and Tween 60, respectively. These results suggest that beta-carotene may be stabilized by (1) LM- or HM-lecithin when liquid carrier lipids are used and (2) HM-lecithin when solid carrier lipids are used. The origin of this latter effect is attributed to the impact of the surfactant tails on the generation of a crystal structure better suited to maintain the chemical stability of the encapsulated bioactive.

Deinococcus aquiradiocola sp. nov., isolated from a radioactive site in Japan.

A gamma- and UV-radiation-tolerant, pale-pink strain (TDMA-uv53T) was isolated from a freshwater sample collected at Misasa (Tottori, Japan), after exposure of the water sample to UV radiation. The cells stained Gram-positive and were non-motile, rod-shaped and non-spore-forming. The DNA G+C content of the strain was 69.1 mol%. Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain TDMA-uv53T belongs to the genus Deinococcus, the highest sequence similarities being found with Deinococcus claudionis PO-04-19-125T (96 %), D. altitudinis ME-04-01-32T (96 %), D. radiomollis PO-04-20-132T (95 %), D. deserti VCD115T (91.5 %), D. hopiensis KR-140T (91.0 %) and D. sonorensis KR-87T (91.0 %). Major fatty acids were iso-15 : 0, 15 : 1omega6c, 15 : 0, 16 : 0 and summed feature 3 (iso-15 : 0 2-OH and/or 16 : 1omega7c). MK-8 was the predominant respiratory quinone. Phylogenetic distinctiveness and unique phenotypic characteristics differentiated strain TDMA-uv53T from closely related Deinococcus species. The results of our polyphasic taxonomic analyses suggested that TDMA-uv53T represents a novel Deinococcus species, for which the name Deinococcus aquiradiocola sp. nov. is proposed. The type strain is TDMA-uv53T (=JCM 14371(T) [corrected] =NBRC 102118T =CCUG 53612T).

Deinococcus misasensis and Deinococcus roseus, novel members of the genus Deinococcus, isolated from a radioactive site in Japan.

Two gamma- and UV-radiation resistant, Gram-positive, red- or pink-pigmented, rod-shaped, strictly aerobic, oxidase- and catalase-positive bacterial strains, TDMA-25T and TDMA-uv51T, were isolated from fresh water collected at Misasa, a radioactive site in Japan. Phylogenetic analysis based on 16S rRNA gene sequences placed both in a distinct lineage in the family Deinococcaceae, and the highest degrees of sequence similarity determined belonged to Deinococcus maricopensis LB-34T (88.8-89.3%), Deinococcus pimensis KR-235T (86.4-86.7%) and Deinococcus yavapaiensis KR-236T (86.1%). The DNA G+C content of the strains was 53-58 mol%. The major respiratory quinone was MK-8. The predominant fatty acids were C15:0 iso, C16:0 iso, C13:0 iso, C17:0 iso, C16:0, C13:0 anteiso, C15:0 and C12:0 iso. The strains degraded gelatin, casein, starch and Tween 80. Unique physiological characteristics, differences in their fatty acid profiles, and genotypic and phylogenetic features, differentiated strains TDMA-25T and TDMA-uv51T from closely related Deinococcus species. Hence, the two strains are described as novel species of the genus Deinococcus. The names Deinococcus misasensis sp. nov. (type strain TDMA-25T=JCM 14369=NBRC 102116=CCUG 53610) and Deinococcus roseus sp. nov. (type strain TDMA-uv51T=JCM 14370=NBRC 102117=CCUG 53611) are proposed.