Rhizobacter gummiphilus NS21 has two rubber oxygenases (RoxA and RoxB) acting synergistically in rubber utilisation

Biodegradation of poly(cis-1,4-isoprene) (rubber) by Gram-negative bacteria has been investigated on the enzymatic level only in Steroidobacter cummioxidans 35Y (previously Xanthomonas sp. 35Y). This species produces two kinds of rubber oxygenases, RoxA_35Y and RoxB_35Y, one of which (RoxB_35Y) cleaves polyisoprene to a mixture of C₂₀- and higher oligoisoprenoids while the other (RoxA_35Y) cleaves polyisoprene and RoxB_35Y-derived oligoisoprenoids to the C₁₅-oligoisoprenoid 12-oxo-4,8-dimethyltrideca-4,8-diene-1-al (ODTD). ODTD can be taken up by S. cummioxidans and used as a carbon source. Gram-positive rubber-degrading bacteria employ another type of rubber oxygenase, latex clearing protein (Lcp), for the initial oxidative attack of the polyisoprene molecule. In this contribution, we examined which type of rubber oxygenase is present in the only other well-documented Gram-negative rubber-degrading species, Rhizobacter gummiphilus NS21. No homologue for an Lcp protein but homologues for a putative RoxA and a RoxB protein (the latter identical to a previously postulated LatA-denominated rubber cleaving enzyme) were identified in the genome of strain NS21. The roxA_NS21 and roxB_NS21 genes were separately expressed in a ∆roxA_35Y/∆roxB_35Y background of S. cummioxidans 35Y and restored the ability of the mutant to produce oligoisoprenoids. The RoxA_NS21 and RoxB_NS21 proteins were each purified and biochemically characterised. The results—in combination with in silico analysis of databases—indicate that Gram-negative rubber-degrading bacteria generally utilise two synergistically acting rubber oxygenases (RoxA/RoxB) for efficient cleavage of polyisoprene to ODTD.

     

Isolation and characterization of Streptomyces,Actinoplanes, and Methylibium strains that are involved in degradation of natural rubber and synthetic poly(cis-1,4-isoprene)

Rubber-degrading bacteria were screened for the production of clearing zones around their colonies on latex overlay agar plates. Novel three bacteria, Streptomyces sp. strain LCIC4, Actinoplanes sp. strain OR16, and Methylibium sp. strain NS21, were isolated. To the best of our knowledge, this is the first report on the isolation of a Gram-negative rubber-degrading bacterium other than γ-proteobacteria. Gel permeation chromatography analysis revealed that these strains degraded poly(cis-1,4-isoprene) to low-molecular-weight products. The occurrence of aldehyde groups in the degradation products by NS21 was suggested by staining with Schiff's reagent and ¹H-nuclear magnetic resonance spectroscopy. The lcp gene of LCIC4, which showed 99% amino acid sequence identity with that of Streptomyces sp. strain K30, was cloned, and contained a putative twin-arginine motif at its N terminus. It is located next to oxiB, which is estimated to be responsible for oxidation of degradation intermediate of rubber in K30. Southern hybridization analysis using LCIC4 lcp probe revealed the presence of a lcp-homolog in OR16. These results suggest that the lcp-homologs are involved in rubber degradation in LCIC4 and OR16.     

Development of a homologous expression system for rubber oxygenase RoxA from Xanthomonas sp.

Aims: Natural rubber (poly‐[cis‐1,4‐isoprene]) can be cleaved into 12‐oxo‐4,8‐dimethyltrideca‐4,8‐diene‐1‐al by rubber oxygenase A (RoxA) isolated from Xanthomonas sp. RoxA is a novel type of dihaem dioxygenase with unknown cleavage mechanism of the rubber carbon backbone. Analysis of mutant RoxA after mutagenesis could be a way to investigate the function of selected amino acids of RoxA during catalysis. Unfortunately, expression of functional RoxA in recombinant Escherichia coli or in recombinant γ‐Proteobacteria such as Pseudomonas putida was not possible in our hands. Therefore, expression of recombinant RoxA in the homologous host, Xanthomonas, was performed. Methods and Results: A transformation system via electroporation was established, and a conjugation system was optimized for Xanthomonas sp. Inactivation of the chromosomal roxA gene by insertional mutagenesis resulted in inability of Xanthomonas sp. to produce active RoxA and to utilize rubber as a sole source of carbon and energy. When an intact copy of roxA was cloned under control of a rhamnose‐inducible promoter in a broad host range vector and was transferred to Xanthomonas sp., high expression levels of functional RoxA in the presence of rhamnose were obtained. Conclusions and Significance and Impact of the Study: Purification of recombinantly expressed RoxA was simplified because of drastically shortened fermentation times and because separation of RoxA from remaining rubber latex particles was not necessary with rhamnose‐induced cultures. About 6 mg purified RoxA were obtained from 1 l of cell‐free culture fluid. Purified recombinant RoxA was highly active and revealed comparable spectral properties as RoxA purified from the wild type. The results of our study are the methodical basis for molecular biological manipulation in Xanthomonas sp. and will simplify investigation into the biochemical mechanisms by which rubber can be biodegraded in the environment by this novel extracellular dihaem dioxygenase RoxA.     

Identification of a Taraxacum brevicorniculatum rubber elongation factor protein that is localized on rubber particles and promotes rubber biosynthesis

Two protein families required for rubber biosynthesis in Taraxacum brevicorniculatum have recently been characterized, namely the cis‐prenyltransferases (TbCPTs) and the small rubber particle proteins (TbSRPPs). The latter were shown to be the most abundant proteins on rubber particles, where rubber biosynthesis takes place. Here we identified a protein designated T. brevicorniculatum rubber elongation factor (TbREF) by using mass spectrometry to analyze rubber particle proteins. TbREF is homologous to the TbSRPPs but has a molecular mass that is atypical for the family. The promoter was shown to be active in laticifers, and the protein itself was localized on the rubber particle surface. In TbREF‐silenced plants generated by RNA interference, the rubber content was significantly reduced, correlating with lower TbCPT protein levels and less TbCPT activity in the latex. However, the molecular mass of the rubber was not affected by TbREF silencing. The colloidal stability of rubber particles isolated from TbREF‐silenced plants was also unchanged. This was not surprising because TbREF depletion did not affect the abundance of TbSRPPs, which are required for rubber particle stability. Our findings suggest that TbREF is an important component of the rubber biosynthesis machinery in T. brevicorniculatum, and may play a role in rubber particle biogenesis and influence rubber production.     

Biodegradation of natural and synthetic rubbers: A review

Since polymeric materials do not decompose easily, disposal of waste polymers is a serious environmental concern. Widespread studies on the biodegradation of rubbers have been carried out in order to overcome the environmental problems associated with rubber waste. This report provides an overview on the microbial degradation of natural and synthetic rubbers. Rubber degrading microbes, bacteria and fungi, are ubiquitous in the environment especially soil. The qualitative data like plate assay, scanning electron microscopy (SEM), attenuated total reflectance-Fourier transform infrared spectroscopy (ATR-FTIR) and Sturm test indicated that both natural and synthetic rubbers can be degraded by microorganisms. It has confirmed that the enzymes latex clearing protein (Lcp) and rubber oxygenase A (RoxA) are responsible for the degradation of natural and synthetic rubbers. Lcp was obtained from Gram-positive bacterium Streptomyces sp. strain K30 and RoxA from Gram-negative bacterium Xanthomonas sp. strain 35Y. Analysis of degradation products of natural and synthetic rubbers indicated the oxidative cleavage of double bonds in polymer backbone. Aldehydes, ketones and other carbonyl groups were detected as degradation products from cultures of various rubber degrading strains. This review emphasizes the importance of biodegradation in environmental biotechnology for waste rubber disposal.     

Colonization and degradation of rubber pieces by Nocardia sp.

The growth of a Nocardia sp. occurs essentially on the insoluble rubber substrate and the cells are tightly bound to the rubber in the initial stage of the growth in spite of vigorous stirring of the cultures. The colonization of rubber pieces was followed by staining with Schiff reagent, and it was revealed that not only the thickness of rubber pieces, but also their length and width greatly influenced microbial colonization and degradation of natural rubber products. Among rubber pieces of various shapes, long strips were most rapidly covered by many microbial colonies and experienced the highest rate of rubber degradation. The rate of degradation (expressed by % weight loss) of the long strips of rubber was a linear function of surface area per unit weight of rubber. Thin and wide films of rubber were also rapidly colonized and degraded, while the colonization and degradation of short and narrow pieces were substantially slower and less extensive.     

Isolation and characterization of Streptomyces, Actinoplanes, and Methylibium strains that are involved in degradation of natural rubber and synthetic poly(cis-1,4-isoprene)

Rubber-degrading bacteria were screened for the production of clearing zones around their colonies on latex overlay agar plates. Novel three bacteria, Streptomyces sp. strain LCIC4, Actinoplanes sp. strain OR16, and Methylibium sp. strain NS21, were isolated. To the best of our knowledge, this is the first report on the isolation of a Gram-negative rubber-degrading bacterium other than γ-proteobacteria. Gel permeation chromatography analysis revealed that these strains degraded poly(cis-1,4-isoprene) to low-molecular-weight products. The occurrence of aldehyde groups in the degradation products by NS21 was suggested by staining with Schiff's reagent and 1H-nuclear magnetic resonance spectroscopy. The lcp gene of LCIC4, which showed 99% amino acid sequence identity with that of Streptomyces sp. strain K30, was cloned, and contained a putative twin-arginine motif at its N terminus. It is located next to oxiB, which is estimated to be responsible for oxidation of degradation intermediate of rubber in K30. Southern hybridization analysis using LCIC4 lcp probe revealed the presence of a lcp-homolog in OR16. These results suggest that the lcp-homologs are involved in rubber degradation in LCIC4 and OR16.     

Development of rubber‐enriched dandelion varieties by metabolic engineering of the inulin pathway

Natural rubber (NR) is an important raw material for a large number of industrial products. The primary source of NR is the rubber tree Hevea brasiliensis, but increased worldwide demand means that alternative sustainable sources are urgently required. The Russian dandelion (Taraxacum koksaghyz Rodin) is such an alternative because large amounts of NR are produced in its root system. However, rubber biosynthesis must be improved to develop T. koksaghyz into a commercially feasible crop. In addition to NR, T. koksaghyz also produces large amounts of the reserve carbohydrate inulin, which is stored in parenchymal root cell vacuoles near the phloem, adjacent to apoplastically separated laticifers. In contrast to NR, which accumulates throughout the year even during dormancy, inulin is synthesized during the summer and is degraded from the autumn onwards when root tissues undergo a sink‐to‐source transition. We carried out a comprehensive analysis of inulin and NR metabolism in T. koksaghyz and its close relative T. brevicorniculatum and functionally characterized the key enzyme fructan 1‐exohydrolase (1‐FEH), which catalyses the degradation of inulin to fructose and sucrose. The constitutive overexpression of Tk1‐FEH almost doubled the rubber content in the roots of two dandelion species without any trade‐offs in terms of plant fitness. To our knowledge, this is the first study showing that energy supplied by the reserve carbohydrate inulin can be used to promote the synthesis of NR in dandelions, providing a basis for the breeding of rubber‐enriched varieties for industrial rubber production.     

Extraction and characterization of a natural rubber from Euphorbia characias latex

A natural rubber was identified and characterized for the first time in the latex of the perennial Mediterranean shrub Euphorbia characias. Four different methods, i.e., acetone, acetic acid, trichloroacetic acid, and Triton® X‐100, followed by successive treatments with cyclohexane/ethanol, were employed to extract the natural rubber. The rubber content was shown to be 14% (w/v) of the E. characias latex, a low content compared with that of Hevea brasiliensis (30–35%) but a similar content to other rubber producing plants. E. characias rubber showed a molecular weight of 93,000 with a M_w/M_n of 2.9. ¹H NMR, ¹³C NMR, and FTIR analysis revealed the characteristic of the cis‐1,4‐polyisoprene typical of natural rubber. These results provided novel insight into latex components and will ultimately benefit the broader understanding of E. characias latex composition. © 2012 Wiley Periodicals, Inc. Biopolymers 97: 589–594, 2012.     

Structural characterization of alpha-terminal group of natural rubber. 2. Decomposition of branch-points by phospholipase and chemical treatments

The treatment of deproteinized natural rubber (DPNR) latex with phospholipases A(2), B, C, and D decreased significantly the long-chain fatty acid ester contents in DPNR and also the molecular weight and Higgins' k' constant, except for phospholipase D treatment. This indicates the presence of phospholipid molecules in NR, which combine rubber molecules together. Transesterification of DPNR resulted in the decomposition of the functional group at the terminal chain-end (alpha-terminal), including phospholipids and formed linear rubber molecules. The addition of small amounts of ethanol into the DPNR solution reduced the molecular weight and shifted the molecular weight distribution (MWD) comparable to that of transesterified DPNR (TE-DPNR). The addition of diammonium hydrogen phosphate into DPNR-latex in order to remove Mg2+ ions yielded a slight decrease in molecular weight and a slight shift in MWD. The phospholipids are expected to link with mono- and diphosphate groups at the alpha-terminal by hydrogen bonding and/or ionic linkages. The decrease in the molecular weight and Huggins' k' constant of DPNR demonstrates the formation of linear molecules after decomposition of branch-points by this treatment, showing that phospholipids participate in the branching formation of NR. The branch-points formed at the alpha-terminus are postulated to originate predominantly by the association of phospholipids via micelle formation of long-chain fatty acid esters and hydrogen bonding between polar headgroups of phospholipids.     

Oligo(cis-1,4-isoprene) aldehyde-oxidizing dehydrogenases of the rubber-degrading bacterium Gordonia polyisoprenivorans VH2

The actinomycete Gordonia polyisoprenivorans strain VH2 is well-known for its ability to efficiently degrade and catabolize natural rubber [poly(cis-1,4-isoprene)]. Recently, a pathway for the catabolism of rubber by strain VH2 was postulated based on genomic data and the analysis of mutants (Hiessl et al. in Appl Environ Microbiol 78:2874–2887, 2012). To further elucidate the degradation pathway of poly(cis-1,4-isoprene), 2-dimensional-polyacrylamide gel electrophoresis was performed. The analysis of the identified protein spots by matrix-assisted laser desorption/ionization-time of flight tandem mass spectrometry confirmed the postulated intracellular pathway suggesting a degradation of rubber via β-oxidation. In addition, other valuable information on rubber catabolism of G. polyisoprenivorans strain VH2 (e.g. oxidative stress response) was provided. Identified proteins, which were more abundant in cells grown with rubber than in cells grown with propionate, implied a putative long-chain acyl-CoA-dehydrogenase, a 3-ketoacyl-CoA-thiolase, and an aldehyde dehydrogenase. The amino acid sequence of the latter showed a high similarity towards geranial dehydrogenases. The expression of the corresponding gene was upregulated > 10-fold under poly(cis-1,4-isoprene)-degrading conditions. The putative geranial dehydrogenase and a homolog were purified and used for enzyme assays. Deletion mutants for five aldehyde dehydrogenases were generated, and growth with poly(cis-1,4-isoprene) was investigated. While none of the mutants had an altered phenotype regarding growth with poly(cis-1,4-isoprene) as sole carbon and energy source, purified aldehyde dehydrogenases were able to catalyze the oxidation of oligoisoprene aldehydes indicating an involvement in rubber degradation.

     

Uncovering mechanisms of rubber biosynthesis in Taraxacum koksaghyz – role of cis‐prenyltransferase‐like 1 protein

The Russian dandelion Taraxacum koksaghyz synthesizes considerable amounts of high‐molecular‐weight rubber in its roots. The characterization of factors that participate in natural rubber biosynthesis is fundamental for the establishment of T. koksaghyz as a rubber crop. The cis‐1,4‐isoprene polymers are stored in rubber particles. Located at the particle surface, the rubber transferase complex, member of the cis‐prenyltransferase (cisPT) enzyme family, catalyzes the elongation of the rubber chains. An active rubber transferase heteromer requires a cisPT subunit (CPT) as well as a CPT‐like subunit (CPTL), of which T. koksaghyz has two homologous forms: TkCPTL1 and TkCPTL2, which potentially associate with the rubber transferase complex. Knockdown of TkCPTL1, which is predominantly expressed in latex, led to abolished poly(cis‐1,4‐isoprene) synthesis but unaffected dolichol content, whereas levels of triterpenes and inulin were elevated in roots. Analyses of latex from these TkCPTL1‐RNAi plants revealed particles that were similar to native rubber particles regarding their particle size, phospholipid composition, and presence of small rubber particle proteins (SRPPs). We found that the particles encapsulated triterpenes in a phospholipid shell stabilized by SRPPs. Conversely, downregulating the low‐expressed TkCPTL2 showed no altered phenotype, suggesting its protein function is redundant in T. koksaghyz. MS‐based comparison of latex proteomes from TkCPTL1‐RNAi plants and T. koksaghyz wild‐types discovered putative factors that convert metabolites in biosynthetic pathways connected to isoprenoids or that synthesize components of the rubber particle shell.     

First report of Bipolaris bicolor causing a leaf spot disease on rubber tree

The rubber tree (Hevea brasiliensis) is the only resource for commercial natural rubber production and thus has economic importance in Southeast Asia. A spot disease on the leaves of a rubber tree was first discovered in 2017 in Hainan, China. In this study, the fungal isolate MA1 from the infected tissues was determined to be a pathogen of the spot disease by satisfying Koch's postulates. The isolate MA1 was identified as Bipolaris bicolor based on the morphological characteristics and multigene phylogenetic analysis. Among fungicides, prochloraz, iprodione and pyraclostrobin significantly inhibited hyphal growth of B. bicolor under in vitro conditions. This study constitutes the first report on the association of B. bicolor with leaf spot disease of rubber trees worldwide.     

In vitro studies on the degradation of poly(cis‐1,4‐isoprene)

Cleavage of the backbone of poly(cis‐1,4‐isoprene) (IR) in solid rubber material was accomplished by the addition of partially purified latex clearing protein (Lcp1_VH2) using a 200‐mL enzyme reactor. Two strategies for the addition of Lcp1_VH2 were studied revealing that the daily addition of 50 µg mL^?1 of Lcp1_VH2 for 5 days was clearly a more efficient regime in comparison to a one‐time addition of 250 µg of Lcp1_VH2 at the beginning. Soluble oligo(cis‐1,4‐isoprene) molecules occurred as degradation products and were identified by ESI‐MS and GPC. Oxygenase activity of Lcp1_VH2 with solid IR particles as substrate was shown for the first time by measuring the oxygen consumption in the reaction medium. A strong decrease of the dissolved oxygen concentration was detected at the end of the assay, which indicates an increase in the number of cleavage reactions. The oligo(cis‐1,4‐isoprene) molecules comprised 1 to 11 isoprene units and exhibited an average molecular weight (M_n) of 885 g mol^?1. Isolation of the oligo(cis‐1,4‐isoprene) molecules was achieved by using silica gel column chromatography. The relative quantification of the isolated products was performed by HPLC‐MS after derivatization with 2,4‐dinitrophenilhydrazyne yielding a concentration of total degradation products of 1.62 g L^?1. Analysis of the polymer surface in samples incubated for 3 days with Lcp1_VH2 via ATR‐FTIR indicated the presence of carbonyl groups, which occurred upon the cleavage reaction. This study presents a cell‐free bioprocess as an alternative rubber treatment that can be applied for the partial degradation of the polymer. © 2018 American Institute of Chemical Engineers Biotechnol. Prog., 34:890–899, 2018     

Characterization of natural rubber biosynthesisin Ficus benghalensis

Natural rubber was identified for the first time in the latex of Ficus benghalensis, and the rubber biosynthetic activity in latex and rubber particles was investigated. ¹³C NMR analysis of samples prepared by successive extractions with acetone and benzene confirmed that the benzene-soluble residues were natural rubber, cis-1,4-polyisoprene. The rubber content in the latex of F. benghalensis was approximately 17 %. Gel permeation chromatography revealed that the molecular mass of the natural rubber from F. benghalensis was approximately 1 500 kDa. The high rubber content and large molecular size suggest that F. benghalensis is a good candidate for an alternative rubber source. Examination of latex serum from F. benghalensis by SDS-polyacrylamide gel electrophoresis revealed a small number of proteins with major proteins of 31 and 55 kDa in size. The 31-kDa protein was predominant in catalytically-active rubber particles. Determination of metal ion concentration in latex and a comparison of the effect of ethylenediamine-tetraacetic acid on in vitro rubber biosynthesis in F. benghalensis, F. carica and Hevea brasiliensis suggest that the divalent metal ion present in latex serum is an important physiological factor controlling the rubber biosynthetic activities in these plant species. Microscopic examination revealed that the rubber in F. benghalensis occurred in a series of laticifer cells located in concentric zones in the inner bark of stems and branches.     

Gene cloning and in vivo characterization of a dibenzothiophene dioxygenase from Xanthobacter polyaromaticivorans

Xanthobacter polyaromaticivorans sp. nov. 127W is a bacterial strain that is capable of degrading a wide range of cyclic aromatic compounds such as dibenzothiophene, biphenyl, naphthalene, anthracene, and phenanthrene even under extremely low oxygen [dissolved oxygen (DO)≤0.2 ppm] conditions (Hirano et al., Biosci Biotechnol Biochem 68:557–564, 2004). A major protein fraction carrying dibenzothiophene degradation activity was purified. Based on its partial amino acid sequences, dbdCa gene encoding alpha subunit terminal oxygenase (DbdCa) and its flanking region were cloned and sequenced. A phylogenetic analysis based on the amino acid sequence demonstrates that DbdCa is a member of a terminal oxygenase component of group IV ring-hydroxylating dioxygenases for biphenyls and monocyclic aromatic hydrocarbons, rather than group III dioxygenases for polycyclic aromatic hydrocarbons. Gene disruption in dbdCa abolished almost of the degradation activity against biphenyl, dibenzothiophene, and anthracene. The gene disruption also impaired degradation activity of the strain under extremely low oxygen conditions (DO≤0.2 ppm). These results indicate that Dbd from 127W represents a group IV dioxygenase that is functional even under extremely low oxygen conditions.     

生物竞争抑制作用对渤海J油田微生物降解驱油聚合物功能的影响

【目的】揭示可降解驱油用聚合物的油藏内源微生物群落组成,分析生物竞争抑制作用(bio-competitive exclusion,BCX)对微生物聚合物降解功能的影响。【方法】通过室内培养实验,观察BCX对驱油用聚合物黏度的影响,随后借助高通量测序技术分析渤海J油田中与聚合物降解相关的微生物菌种,并探寻样本中丰度较高的聚合物降解功能基因─酰胺酶、加氧酶、硫化氢生成酶基因。之后,比对测序结果,采用实时荧光定量法验证上述功能基因在样本之间的含量差异,最后进一步注释携带上述功能基因的微生物群落组成。【结果】BCX可有效地延缓驱油聚合物黏度的损失。油田中与聚合物降解相关的微生物有Acetomicrobium、 Tepidiphilus、Thermoanaerobacter、Fervidobacterium、Ralstonia、Halomonas、Roseovarius、Deferribacteraceae和Comamonadaceae等9类菌种。高通量测序分析得到样本中BCX可显著下调丰度的聚合物降解功能基因共计有7种,其中酰胺酶基因ansB、加氧酶基因ssuD在样本之间的含量经定量验证,发现...     

Rubber particles from four different species, examined by transmission electron microscopy and electron-paramagnetic-resonance spin labeling, are found to consist of a homogeneous rubber core enclosed by a contiguous, monolayer biomembrane

The physical characteristics of rubber particles from the four rubber (cis-1,4-polyisoprene) producing species Euphorbia lactiflua Phil., Ficus elastica Roxb., Hevea brasiliensis Müll. Arg., and Parthenium argentatum Gray, were investigated using transmission electron microscopy (TEM) and electron-paramagnetic-resonance (EPR) spin labeling spectroscopy. Transmission electron microscopy showed the rubber particles to be composed of a spherical, homogeneous, core of rubber enclosed by a contiguous, electron-dense, single-track surface layer. The biochemical composition of the surface layer and its single-track TEM suggested that a monolayer biomembrane was the surface structure most compatible with the hydrophobic rubber core. The EPR spectra for a series of positional isomers of doxyl stearic acid, used to label the surface layer of the rubber particles, exhibited flexibility gradients and evidence for lipid-protein interactions for all four rubber particle types that is consistent with a biomembrane-like surface. The EPR spectra confirmed that the surface biomembrane is a monolayer. Thus, rubber particles appear similar to oil bodies in their basic architecture. The EPR spectra also provided information on protein location and degree of biomembrane penetration that correlated with the known properties of the rubber-particle-bound proteins. The monolayer biomembrane serves as an interface between the hydrophobic rubber interior and the aqueous cytosol and prevents aggregation of the particles. An unexpected observation for the probes in pure polyisoprene was evidence of an intrinsic flexibility gradient associated with the stearic acid molecule itself. Received: 22 May 1999 / Accepted: 21 June 1999     

Degradation of the rubber in truck tires by a strain of Nocardia

A strip of tread compound cut from a truck tire was degraded only slightly when it was used as the sole growth substrate for a strain of Nocardia. On the contrary, its degradation was markedly enhanced by addition of a strip cut from a latex glove which the organism readily utilized as a growth substrate. When a glove strip was added, the biomass concentration in the experimental flask became more than 10-fold higher than the control without a glove strip and the colonization of the tire strip was significantly enhanced.After 8 weeks' cultivation, about 28% of the tire strip was disintegrated into very small black particles (mostly less than 30 m in diameter) and the weight of the remaining unchanged portion of the strip was about 49% of the initial weight.Four kinds of truck tire treads were attacked in differing degrees by the organism under the same conditions. The treads containing more than 70 phr (parts per hundred of rubber) of natural rubber were considerably attacked, while those with a natural rubber content of less than 55 phr were attacked only slightly. The microbial activity against the rubber in the side wall of a truck tire was relatively high, but the inner liner was hardly attacked and the bead rubber not at all.