Genomic insights into Dactylosporangium sp. AC04546, a rubber degrader with three latex clearing proteins.

With more than 100 rubber-degrading strains being reported, only 9 Lcp proteins isolated from Nocardia, Gordonia, Streptomyces, Rhodococcus, Actinoplanes, and Solimonas have been purified and biochemically characterized. A new strain, Dactylosporangium sp. AC04546 (strain JCM34239), isolated from soil samples collected in Sarawak Forest, was able to grow and utilize natural or synthetic rubber as the sole carbon source. Complete genome of Strain AC04546 was obtained from the hybrid assembly of PacBio Sequel II and Illumina MiSeq. Strain AC04546 has a large circular genome of 13.08 Mb with a G+C content of 72.1%. The genome contains 11,865 protein-coding sequences with 3 latex clearing protein (lcp) genes located on its chromosome. The genetic organization of the lcp gene cluster is similar to two other reported rubber-degrading strains-Actinoplanes sp. OR16 and Streptomyces sp. CFMR 7. All 3 Lcp from strain AC04546 were expressed in Escherichia coli and exhibited degrading activity against natural rubber. The distinctiveness of strain AC04546, along with other characterized rubber-degrading strains, is reported here.

Streptomyces sp. AC04842: Genomic Insights and Functional Expression of Its Latex Clearing Protein Genes (lcp1 and lcp2) When Cultivated With Natural and Vulcanized Rubber as the Sole Carbon Source.

Rubber-degrading Actinobacteria have been discovered and investigated since 1985. Only recently, through the advancement of genomic sequencing and molecular techniques, genes and pathways involved in rubber degradation are being revealed; however, the complete degradation pathway remains unknown. Streptomyces sp. AC04842 (JCM 34241) was discovered by screening at a Culture Collection Centre in Sarawak for Actinomycetes forming a clear zone on natural rubber latex agar. Streptomyces is a dominant and well-studied soil bacterium playing an important role in soil ecology including carbon recycling and biodegradation. Streptomyces sp. AC04842 draft genome revealed the presence of 2 putative latex clearing protein (lcp) genes on its chromosome and is closely related to Streptomyces cellulosae. Under the Streptomyces genus, there are a total of 64 putative lcp genes deposited in the GenBank and UniProt database. Only 1 lcp gene from Streptomyces sp. K30 has been characterized. Unlike Streptomyces sp. K30 which contained 1 lcp gene on its chromosome, Streptomyces sp. AC04842 contained 2 lcp genes on its chromosome. Streptomyces sp. AC04842 lcp1 and lcp2 amino acid sequences showed 46.13 and 69.11%, respectively, similarity to lcp sequences of Streptomyces sp. K30. Most rubber degrading strains were known to harbor only 1 lcp gene, and only recently, 2-3 lcp homologs have been reported. Several studies have shown that lcp-homolog expression increased in the presence of rubber. To study the expression of lcp1 and lcp2 genes for Streptomyces sp. AC04842, the strain was incubated in different types of rubber as the sole carbon source. In general, the lcp1 gene was highly expressed, while the lcp2 gene expression was upregulated in the presence of vulcanized rubber. Mixtures of natural and vulcanized rubber did not further increase the expression of both lcp genes compared with the presence of a specific rubber type. In this study, we paved the way to the exploration of lcp homologs and their function in degrading different types of rubber.

Rubber Degrading Strains: Microtetraspora and Dactylosporangium.

Rubber composed of highly unsaturated hydrocarbons, modified through addition of chemicals and vulcanization are widely used to date. However, the usage of rubber, faces many obstacles. These elastomeric materials are difficult to be re-used and recovered, leading to high post-consumer waste and vast environmental problems. Tyres, the major rubber waste source can take up to 80 years to naturally degrade. Experiments show that the latex clearing proteins (Lcp) found in Actinobacteria were reportedly critical for the initial oxidative cleavage of poly(cis-1,4-isoprene), the major polymeric unit of rubber. Although, more than 100 rubber degrading strains have been reported, only 8 Lcp proteins isolated from Nocardia (3), Gordonia (2), Streptomyces (1), Rhodococcus (1), and Solimonas (1) have been purified and biochemically characterized. Previous studies on rubber degrading strains and Lcp enzymes, implied that they are distinct. Following this, we aim to discover additional rubber degrading strains by randomly screening 940 Actinobacterial strains isolated from various locations in Sarawak on natural rubber (NR) latex agar. A total of 18 strains from 5 genera produced clearing zones on NR latex agar, and genes encoding Lcp were identified. We report here lcp genes from Microtetraspora sp. AC03309 (lcp1 and lcp2) and Dactylosporangium sp. AC04546 (lcp1, lcp2, lcp3), together with the predicted genes related to rubber degradation. In silico analysis suggested that Microtetraspora sp. AC03309 is a distinct species closely related to Microtetraspora glauca while Dactylosporangium sp. AC04546 is a species closely related to Dactylosporangium sucinum. Genome-based characterization allowed the establishment of the strains taxonomic position and provided insights into their metabolic potential especially in biodegradation of rubber. Morphological changes and the spectrophotometric detection of aldehyde and keto groups indicated the degradation of the original material in rubber samples incubated with the strains. This confirms the strains' ability to utilize different rubber materials (fresh latex, NR product and vulcanized rubber) as the sole carbon source. Both strains exhibited different levels of biodegradation ability. Findings on tyre utilization capability by Dactylosporangium sp. AC04546 is of interest. The final aim is to find sustainable rubber treatment methods to treat rubber wastes.

Erratum: Basik et al. Microbial Degradation of Rubber: Actinobacteria. Polymers 2021, 13, 1989.

The authors wish to make the following changes to the published paper [...].

Microbial Degradation of Rubber: Actinobacteria.

Rubber is an essential part of our daily lives with thousands of rubber-based products being made and used. Natural rubber undergoes chemical processes and structural modifications, while synthetic rubber, mainly synthetized from petroleum by-products are difficult to degrade safely and sustainably. The most prominent group of biological rubber degraders are Actinobacteria. Rubber degrading Actinobacteria contain rubber degrading genes or rubber oxygenase known as latex clearing protein (lcp). Rubber is a polymer consisting of isoprene, each containing one double bond. The degradation of rubber first takes place when lcp enzyme cleaves the isoprene double bond, breaking them down into the sole carbon and energy source to be utilized by the bacteria. Actinobacteria grow in diverse environments, and lcp gene containing strains have been detected from various sources including soil, water, human, animal, and plant samples. This review entails the occurrence, physiology, biochemistry, and molecular characteristics of Actinobacteria with respect to its rubber degrading ability, and discusses possible technological applications based on the activity of Actinobacteria for treating rubber waste in a more environmentally responsible manner.

Luteipulveratus halotolerans sp. nov., an actinobacterium (Dermacoccaceae) from forest soil.

The taxonomic position of an actinobacterium strain, C296001T, isolated from a soil sample collected in Sarawak, Malaysia, was established using a polyphasic approach. Phylogenetically, strain C296001T was closely associated with the genus Luteipulveratus and formed a distinct monophyletic clade with the only described species, Luteipulveratus mongoliensis NBRC 105296T. The 16S rRNA gene sequence similarity between strain C296001T and L. mongoliensis was 98.7 %. DNA-DNA hybridization results showed that the relatedness of strain C296001T to L. mongoliensis was only 21.5 %. The DNA G+C content of strain C296001T was 71.7 mol%. Using a PacBio RS II system, whole genome sequences for strains C296001T and NBRC 105296T were obtained. The genome sizes of 4.5 Mbp and 5.4 Mbp determined were similar to those of other members of the family Dermacoccaceae. The cell-wall peptidoglycan contained lysine, alanine, aspartic acid, glutamic acid and serine, representing the peptidoglycan type A4α l-Lys-l-Ser-d-Asp. The major menaquinones were MK-8(H4), MK-8 and MK-8(H2). Phosphatidylglycerol, phosphatidylinositol, diphosphatidylglycerol and phosphoglycolipid were the polar lipids, while the whole-cell sugars were glucose, fucose and lesser amounts of ribose and galactose. The major fatty acids were iso-C16 : 0, anteiso-C17 : 0, iso-C16 : 1 H, anteiso-C17 : 1ω9c, iso-C18 : 0 and 10-methyl C17 : 0. Chemotaxonomic analyses showed that C296001T had typical characteristics of members of the genus Luteipulveratus, with the main differences occurring in phenotypic characteristics. On the basis of the phenotypic and chemotaxonomic evidence, it is proposed that strain C296001T be classified as a representative of a novel species in the genus Luteipulveratus, for which the name Luteipulveratus halotolerans sp. nov. is recommended. The type strain is C296001T ( = ATCC TSD-4T = JCM 30660T).