Bio-Prospecting Laccases in the Bacterial Diversity of Activated Sludge From Pulp and Paper Industry

Activated sludge is an artificial ecosystem known to harbor complex microbial communities. Bacterial diversity in activated sludge from pulp and paper industry was studied to bioprospect for laccase, the multicopper oxidase applicable in a large number of industries due to its ability to utilize a wide range of substrates. Bacterial diversity using 454 pyrosequencing and laccase diversity using degenerate primers specific to conserved copper binding domain of laccase like multicopper oxidase (LMCO) genes were investigated. 1231 OTUs out of 11,425 sequence reads for bacterial diversity and 11 OTUs out of 15 reads for LMCO diversity were formed. Phylum Proteobacteria (64.95 %) with genus Thauera (13.65 %) was most abundant followed by phylum Bacteriodetes (11.46 %) that included the dominant genera Paludibacter (1.93 %) and Lacibacter (1.32 %). In case of LMCOs, 40 % sequences showed affiliation with Proteobacteria and 46.6 % with unculturable bacteria, indicating considerable novelty, and 13.3 % with Bacteroidetes. LMCOs belonged to H and J families.     

Characterization of a laccase-like multicopper oxidase from newly isolated Streptomyces sp. C1 in agricultural waste compost and enzymatic decolorization of azo dyes

Laccases or laccase-like multicopper oxidases (LMCOs) could catalyze the oxidation of various substrates coupled to the reduction of oxygen to water. In this study, eight strains with laccase activity were isolated from composting samples in different phases, among which strain C1 isolated from the thermophilic-phase sample presented the highest laccase activity. The purified LMCO of strain C1 showed a single protein band on SDS-PAGE gel with a molecular mass of about 38 kDa. The novel laccase showed alkaline resistance and moderate thermostability. The enzyme activity was activated by some metal ions such as Cu²⁺, Co²⁺ and Fe³⁺ at the concentration of 1 mM, while was strongly inhibited in the presence of Hg²⁺. The LMCO could efficiently decolorize the indigo carmine and diamond black PV with syringaldehyde as mediator, which suggested a great potential for dye decolorization in the textile industry. The novel strain was identified as Streptomyces sp. C1. The finding of new laccase-producing Streptomyces sp. C1 in this study will also contribute to the further explanation of the function of Actinomycetes in the thermophilic phase of composting.     

Solubilisation and mineralisation of [14C]lignocellulose from wheat straw by Streptomyces cyaneus CECT 3335 during growth in solid-state fermentation

Nine Streptomyces strains were screened for their ability to solubilise and mineralise ¹⁴C-labelled lignin during growth in solid-state fermentation. Streptomyces viridosporus was confirmed as an active lignin-degrading organism along with a new isolate, Streptomyces sp. UAH 15, further classified as Streptomyces cyaneus CECT 3335. This organism was able to solubilise and mineralise the [¹⁴C]lignin fraction of lignocellulose (44.96 ± 1.77% and 3.41 ± 0.48% respectively) after 21 days of incubation. Cell-free filtrates from Streptomyces sp. grown in solid-state fermentation were capable of solubilising up to 20% of the [¹⁴C]lignin after 2 days incubation, with most of the product detected in the acid-soluble rather than in the water-soluble fraction. Identification of the extracellular enzymes produced during growth of S. cyaneus CECT 3335 revealed that extracellular peroxidase and phenol oxidase activities were present, with the activity of phenol oxidase being 100 times greater than peroxidase activity. The activity of these two enzymes was found to correlate with both solubilisation and mineralisation rates. This is the first report of phenol oxidase activity produced by a Streptomyces strain during growth in solid-state fermentation. A role for the enzyme in the solubilisation and mineralisation of lignocellulose by S. cyaneus is suggested. Received: 12 May 1997 / Accepted: 19 May 1997     

Gene structure and molecular analysis of the laccase-like multicopper oxidase (LMCO) gene family in <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis>

Completed genome sequences have made it clear that multicopper oxidases related to laccase are widely distributed as multigene families in higher plants. Laccase-like multicopper oxidase (LMCO) sequences culled from GenBank and the Arabidopsis thaliana genome, as well as those from several newly cloned genes, were used to construct a gene phylogeny that clearly divided plant LMCOs into six distinct classes, at least three of which predate the evolutionary divergence of angiosperms and gymnosperms. Alignments of the predicted amino acid sequences highlighted regions of variable sequence flanked by the highly conserved copper-binding domains that characterize members of this enzyme family. All of the predicted proteins contained apparent signal sequences. The expression of 13 of the 17 LMCO genes in A. thaliana was assessed in different tissues at various stages of development using RT-PCR. A diversity of expression patterns was demonstrated with some genes being expressed in a constitutive fashion, while others were only expressed in specific tissues at a particular stage of development. Only a few of the LMCO genes were expressed in a pattern that could be considered consistent with a major role for these enzymes in lignin deposition. These results are discussed in the context of other potential physiological functions for plant LMCOs, such as iron metabolism and wound healing.     

Bio-prospecting Bacterial Diversity of Hot Springs in Northern Himalayan Region of India for Laccases

Bacterial diversity of hot springs of northern Himalayan region of India was studied and explored for laccases, the multicopper enzymes applicable in a large number of industries due to their ability to utilize a wide range of substrates. 220 operational taxonomic units (OTUs) out of 5551 sequence reads for bacterial diversity and 3 OTUs out of 19 sequence reads for Laccase like multicopper oxidases (LMCOs) diversity were generated. Bacteroidetes (74.28%) was the most abundant phylum including genus Paludibacter (66.96%), followed by phylum Proteobacteria (24.53%) including genera Chitinilyticum (7.55%) and Cellvibrio (6.14%). In case of laccase diversity, three LMCO sequences showed affiliation with proteobacteria and one with two domain laccase from uncultivable bacteroidetes. LMCO sequences belonged to H and N families.     

Ferroxidase activity in a laccase-like multicopper oxidase from Liriodendron tulipifera.

Ferroxidase activity was detected in a laccase-like multicopper oxidase (LMCO) produced in transgenic tobacco cells expressing an LMCO cDNA (Ltlacc2.2) cloned from yellow-poplar (Liriodendron tulipifera). This marks the first report of ferroxidase activity associated with a plant laccase and suggests that some members of this plant enzyme family may have physiological functions based on activities other than their more widely recognized phenoloxidase activity. Recent work with LMCOs from bacteria, yeast and mammals has shown that metal oxidase activities in these enzymes can be important for their primary physiological functions, With respect to ferroxidase activity in certain plant LMCOs, it is proposed that the high levels of LMCO expression in plant vascular tissues may reflect the need for high-efficiency iron uptake pumps in tissues that undergo lignification during normal development. Such iron uptake pumps would function to minimize levels of free iron so that reactive oxygen species do not reach toxic levels when H2O2 is generated for peroxidase-mediated monolignol coupling during lignin deposition.     

Production and Characterization of Polymeric Lignin Degradation Intermediates from Two Different Streptomyces spp.

Previous investigations have identified a quantitatively major intermediate of lignin degradation by Streptomyces viridosporus. The intermediate, a modified lignin polymer, acid-precipitable polymeric lignin (APPL), is released as a water-soluble catabolite and has been recovered in amounts equivalent to 30% of the lignin originally present in a corn stover lignocellulose substrate after degradation by this actinomycete. In the present work, APPLs were collected at various time intervals from cultures of two highly ligninolytic Streptomyces sp. strains, S. viridosporus T7A and S. badius 252, growing on corn stover lignocellulose. APPL production was measured over time, and the chemistry of APPLs produced by each organism after different time intervals was compared. Chemical characterizations included assays for lignin, carbohydrate, and ash contents, molecular weight distributions by gel permeation chromatography, and chemical degradation analyses by permanganate oxidation, acidolysis, and alkaline ester hydrolysis. Differences between the organisms were observed in the cultural conditions required for APPL production and in the time courses of APPL accumulation. S. viridosporus produced APPL in solid-state fermentation over a 6- to 8-week incubation period, whereas S. badius produced as much or more APPL, but only in liquid culture and over a 7- to 8-day incubation period. The chemistry of the APPLs produced also differed. S. viridosporus APPL was more lignin-like than that of S. badius and was slowly modified further over time, although no change in molecular weight distribution over time was observed. In contrast, S. badius APPL was less lignin-like and increased substantially in average molecular weight over time. Results indicated that differing mechanisms of lignin metabolism may exist in these two Streptomyces sp. strains. S. viridosporus APPL probably originates from the heart of the lignin and is released largely as the result of β-ether cleavage and other oxidative reactions. S. badius APPL probably originates in the same manner; however, after release as a water-soluble catabolite, lower-molecular-weight intermediates of lignin degradation are repolymerized with APPL in a reaction catalyzed by an extracellular phenol oxidase. The chemical analyses and the presence of extracellular phenol oxidase in S. badius, but not in S. viridosporus, support this conclusion.     

Isolation from haddock tissue of psychrophilic bacteria with maximum growth temperature below 20 degrees C.

Protoplast fusion was investigated as a technique for genetically manipulating two lignin-degrading Streptomyces strains, Streptomyces viridosporus T7A and Streptomyces setonii 75Vi2. Four of 19 recombinants tested showed enhanced production of acid-precipitable polymeric lignin (APPL), producing 155 to 264% more APPL from corn stover lignocellulose than was produced by the wild-type S. viridosporus T7A. APPLs are lignin degradation intermediates known to be potentially valuable chemical products produced by bioconversion of lignin with Streptomyces spp. The prospects of utilizing protoplast fusion to construct APPL-overproducing Streptomyces strains was considered especially promising.     

Atmospheric N Deposition Increases Bacterial Laccase-Like Multicopper Oxidases: Implications for Organic Matter Decay

Anthropogenic release of biologically available nitrogen (N) has increased dramatically over the last 150 years, which can alter the processes controlling carbon (C) storage in terrestrial ecosystems. In a northern hardwood forest ecosystem located in Michigan in the United States, nearly 20 years of experimentally increased atmospheric N deposition has reduced forest floor decay and increased soil C storage. This change occurred concomitantly with compositional changes in Basidiomycete fungi and in Actinobacteria, as well as the downregulation of fungal lignocelluloytic genes. Recently, laccase-like multicopper oxidases (LMCOs) have been discovered among bacteria which can oxidize β-O-4 linkages in phenolic compounds (e.g., lignin and humic compounds), resulting in the production of dissolved organic carbon (DOC). Here, we examined how nearly 2 decades of experimental N deposition has affected the abundance and composition of saprotrophic bacteria possessing LMCO genes. In our experiment, LMCO genes were more abundant in the forest floor under experimental N deposition whereas the abundances of bacteria and fungi were unchanged. Experimental N deposition also led to less-diverse, significantly altered bacterial and LMCO gene assemblages, with taxa implicated in organic matter decay (i.e., Actinobacteria, Proteobacteria) accounting for the majority of compositional changes. These results suggest that experimental N deposition favors bacteria in the forest floor that harbor the LMCO gene and represents a plausible mechanism by which anthropogenic N deposition has reduced decomposition, increased soil C storage, and accelerated phenolic DOC production in our field experiment. Our observations suggest that future rates of atmospheric N deposition could fundamentally alter the physiological potential of soil microbial communities.     

Major Streptomyces species associated with fissure scab of potato in South Africa including description of Streptomyces solaniscabiei sp. nov

Streptomyces species are the causal agents of several scab diseases on potato tubers. A new type of scab symptom, caused by Streptomyces species, was observed in South Africa from 2010 onwards. The disease was initially thought to be caused by a single Streptomyces species, however, subsequent isolations from similar symptoms on other potato tubers revealed diversity of the Streptomyces isolates. The objective of this study was to characterise these isolates in order to determine what are the major species involved in the disease. This was done by sequencing and phylogenetic analyses of the 16S rDNA as well as five housekeeping genes, investigation of growth on different culture media, standard phenotypic tests and scanning electron microscopy of culture morphology. The presence of the pathogenicity island (PAI) present in plant pathogenic Streptomyces species was also investigated. The genomes of eight isolates, selected from the three main clades identified, were sequenced and annotated to further clarify species boundaries. Three isolates of each of the three main clades were also inoculated onto susceptible potato cultivars in order to establish the pathogenicity of the species. The results of the phylogenetic and genome analyses revealed that there are three main species involved, namely, Streptomyces werraensis, Streptomyces pseudogriseolus and a novel Streptomyces species that is described here as Streptomyces solaniscabiei sp. nov., with strain FS70^T (=?PPPPB BD 2226^T?=?LMG 32103^T) as the type strain. The glasshouse trial results showed that all three of the Streptomyces species are capable of producing fissure scab symptoms. None of the Streptomyces isolates from fissure scab contained the full PAI and the mechanism of disease initiation still needs to be determined. Genomic comparisons also indicated that S. gancidicus Suzuki 1957 (Approved Lists 1980) is a later heterotypic synonym of S. pseudogriseolus Okami and Umezawa 1955 (Approved Lists 1980).

     

Diversity of laccase-like multicopper oxidase genes in Morchellaceae: identification of genes potentially involved in extracellular activities related to plant litter decay

Despite the important role played by soil-inhabiting ascomycetes in plant litter decay processes, studies on the diversity and function of their laccase-like multicopper oxidase (LMCO) genes are scarce. In the present work, the LMCO gene diversity in 15 strains representing nine Morchellaceae and one Discinaceae species was evaluated by PCR. One to six different genes were found within the species, representing 26 different sequence types. Cluster analysis revealed LMCO genes belonging to four main gene families encoding different protein classes (Class I-IV). To identify the genes related to extracellular activities and potentially involved in litter decay processes, liquid cultures were induced by different aromatic compounds. Morchella conica and Verpa conica showed the strongest LMCO activity enhancement in the presence of the naturally occurring phenolic compound guaiacol, and their expressed LMCO genes were identified by sequencing. Only genes belonging to the gene families encoding the Class II and III proteins were expressed. Both genes (Class II and III) of the mycorrhizal-like strain M. conica were exclusively expressed in the presence of guaiacol. In contrast to the saprotrophic strain V. conica, the gene encoding the Class III protein was constitutively expressed as it was also found in control cultures without guaiacol.     

Extracellular enzyme production and fungal mycelia degradation of antagonistic Streptomyces induced by fungal mycelia preparation of cucurbit plant pathogens

Streptomyces are beneficial soil microorganisms and potential candidates for biocontrol agents against soilborne pathogenic fungi of cucurbit plants. Extracellular enzymes such as cellulase, chitinase and glucanase produced by Streptomyces are important components of actinomycete-fungus antagonism. This study aimed to investigate the influence on extracellular enzymes production and fungal mycelia degradation by antagonistic Streptomyces of mycelia preparation of pathogenic fungi (MPPF) of cucurbit plants. The results showed that the antagonistic Streptomyces displayed high extracellular enzyme activities to varying degrees when MPPF was used as the sole carbon source. The MPPF from Fusarium proliferatum, Fusarium oxysporum f. sp. niveum and Alternaria tenuissima were the most effective carbon sources in enhancing the cellulase activity of Streptomyces globisporus C7, Streptomyces globisporus subsp. globisporus C28 and Streptomyces kanamyceticus C49, respectively. S. globisporus subsp. globisporus C28, Streptomyces pactum A12 and S. kanamyceticus C49 cultured in the medium containing MPPF from Fusarium equiseti showed the highest chitinase activity (12.35, 12.50 and 15.06 U, respectively) of all the MPPF treatments. Glucanase activity of Streptomyces carnosus A11 was enhanced greatly (9.26 U) when MPPF from A. tenuissima was used as the sole carbon source. A hyphal intertwining and degradation phenomenon was observed when the antagonistic Streptomyces came across the pathogenic fungal mycelia, which was due to a synergistic effect of the extracellular enzymes produced by the antagonistic Streptomyces.     

Nitrite reductase genes as functional markers to investigate diversity of denitrifying bacteria during agricultural waste composting

The purpose of this study was to investigate the diversity of denitrifier community during agricultural waste composting. The diversity and dynamics of the denitrifying genes (nirK and nirS) were determined using polymerase chain reaction–denaturing gradient gel electrophoresis (PCR-DGGE). Relationships between physico-chemical parameters and denitrifying genes structures were simultaneously evaluated by redundancy analysis (RDA). Phylogenetic analysis indicated that nirK clones grouped into six clusters and nirS clones into two major clusters, respectively. The results showed a very high diversity of nir gene sequences within composting samples. RDA showed that the nirK and nirS gene structures were significantly related to pH and pile temperature (P?<?0.05). Significant amounts of the variation (49.2 and 38.3 % for nirK and nirS genes, respectively) were explained by pH and pile temperature, suggesting that those two parameters were the most likely ones to influence, or be influenced by the denitrifiers harboring nirK and nirS genes.     

Respiratory chain composition and activity in some thermotolerant streptomycetes

The cytochrome composition of membrane preparations from 3 thermotolerant species of Streptomyces was determined from difference spectra. All contained b, c and a-type haemoproteins and in addition, 2 species (Streptomyces thermoviolaceus and Streptomyces thermoflavus) possessed cytochrome oxidase d. A CO-reacting b cytochrome similar to cytochrome oxidase o could also be detected in all 3 organisms. Thermotolerance of the respiratory chains showed no correlation with cytochrome composition. In general, Streptomyces thermonitrificans had the most thermostable activities, whilst those of S. thermoviolaceus were the most thermolabile. Growth temperature exerted no qualitative effects on respiratory chain composition but did affect the thermostability of some substrate-supported oxidase activities.     

Diversity of Telomere Palindromic Sequences and Replication Genes among Streptomyces Linear Plasmids

Streptomyces sp. linear plasmids and linear chromosomes usually contain conserved terminal palindromic sequences bound by the conserved telomeric proteins Tap and Tp, encoded by the tap and tpg genes, respectively, as well as plasmid loci required for DNA replication in circular mode when the telomeres are deleted. These consist of iterons and an adjacent rep gene. By using PCR, we found that 8 of 17 newly detected linear plasmids in Streptomyces strains lack typical telomeric tap and tpg sequences. Instead, two novel telomeres in plasmids pRL1 and pRL2 from the eight strains and one conserved telomere in pFRL1 from the other strains were identified, while multiple short palindromes were also found in the plasmids. The complete nucleotide sequence of pRL2 revealed a gene encoding a protein containing two domains, resembling Tap of Streptomyces and a helicase of Thiobacillus, and an adjacent gene encoding a protein similar to Tpg of Streptomyces and a portion of the telomere terminal protein pTP of adenoviruses. No typical iterons-rep loci were found in the three plasmids. These results indicate an unexpected diversity of telomere palindromic sequences and replication genes among Streptomyces linear plasmids.     

Enhancing macrolide production in Streptomyces by coexpressing three heterologous genes

Antibiotic production in Streptomyces can often be increased by introducing heterologous genes into strains that contain an antibiotic biosynthesis gene cluster. A number of genes are known to be useful for this purpose. We chose three such genes and cloned them singly or in combination under the control of the strong constitutive ermE* promoter into a ?C31-derived integrating vector that can be transferred efficiently by conjugation from Escherichia coli to Streptomyces. The three genes are adpA, a global regulator from Streptomyces coelicolor, metK, encoding S-adenosylmethionine synthetase from S. coelicolor, and, VHbS, hemoglobin from Vitreoscilla. The substitutions with GC in VHbS was intended to convert codons from lower usage to higher, yet causing no change to the encoded amino acid. Plasmids containing either one of these genes or genes in various combinations were introduced into Streptomyces sp. FR-008, which produces the macrolide antibiotic FR-008-III (also known as candicidin D). The largest increase in FR-008-III production was achieved by the plasmid containing all three genes. This plasmid also increased avermectin production in Streptomyces avermitilis, and is likely to be generally useful for improving antibiotic production in Streptomyces.     

Microbial community succession and lignocellulose degradation during agricultural waste composting

The changes of microbial community during agricultural waste composting were successfully studied by quinone profiles. Mesophilic bacteria indicated by MK-7 and mesophilic fungi containing Q-9 as major quinone were predominant and seemed to be important during the initial stage of composting. Actinobacteria indicated by a series of partially saturated and long-chain menaquinones were preponderant during the thermophilic period. While Actinobacteria, fungi and some bacteria, especially those microbes containing MK-7(H4) found in Gram-positive bacteria with a low G+C content or Actinobacteria were found cooperate during the latter maturating period. Since lignocellulsoe is abundant in the agricultural wastes and its degradation is essential for the operation of composting, it’s important to establish the correlation between the quinone profiles changes and lignocellulose degradation. The microbes containing Q-9 or Q-10(H2) as major quinone were found to be the most important hemicellulose and cellulose degrading microorganisms during composting. While the microorganisms containing Q-9(H2) as major quinone and many thermophilic Actinobacteria were believed to be responsible for lignin degradation during agricultural waste composting.     

Rapid and Specific Method for Evaluating Streptomyces Competitive Dynamics in Complex Soil Communities

Quantifying target microbial populations in complex communities remains a barrier to studying species interactions in soil environments. Quantitative PCR (qPCR) assays were developed for quantifying pathogenic Streptomyces scabiei and antibiotic-producing Streptomyces lavendulae strains in complex soil communities. This assay will be useful for evaluating the competitive dynamics of streptomycetes in soil.Streptomyces spp. are ubiquitous soil bacteria that are noted for their capacity to produce a vast array of bioactive compounds, including antibiotics (10). Antibiotic-mediated species interactions are believed to be important to Streptomyces fitness and plant disease biocontrol in soil, and yet quantitative data on Streptomyces interactions in soil are limited. Moreover, because the impacts of one species on another can be mediated through interactions with other microbes in the community, detecting these impacts requires a sensitive and accurate method for quantifying the target populations within a complex community. Here, we describe a sensitive and specific assay that targets a short hypervariable region of the 16S rRNA gene to distinguish among Streptomyces organisms in complex soil communities. Streptomyces strains DL93 (Streptomyces lavendulae, an antibiotic producer that is effective in plant disease biocontrol [9]) and DL87 (Streptomyces scabiei, a plant pathogen) were studied in the present work. This approach has significant potential to shed light on the diversity and complexity of Streptomyces species interactions in soil.     

Chitinase Genes in Lake Sediments of Ardley Island, Antarctica

A sediment core spanning approximately 1,600 years was collected from a lake on Ardley Island, Antarctica. The sediment core had been greatly influenced by penguin guano. Using molecular methods, the chitinolytic bacterial community along the sediment core was studied over its entire length. Primers targeting conserved sequences of the catalytic domains of family 18 subgroup A chitinases detected group A chitinases from a wide taxonomic range of bacteria. Using quantitative competitive PCR (QC-PCR), chitinase gene copies in each 1-cm section of the whole sediment column were quantified. QC-PCR determination of the chitinase gene copies indicated significant correlation with phosphorus and total organic carbon concentration, suggesting a historical connection between chitinase gene copies and the amount of penguin guano input into the lake sediment. Most of the chitinase genes cloned from the historic sediment core were novel. Analysis of the chitinase gene diversity in selected sediment layers and in the fresh penguin deposits indicated frequent shifts in the chitinolytic bacterial community over time. Sequence analysis of the 16S rRNA genes of chitinolytic bacteria isolated from the lake sediment revealed that the isolates belonged to Janthinobacterium species, Stenotrophomonas species of γ-Proteobacteria, Cytophaga species of the Cytophaga-Flexibacter-Bacteroides group, and Streptomyces and Norcardiopsis species of Actinobacteria. Chitinase gene fragments were cloned and sequenced from these cultivated chitinolytic bacteria. The phylogeny of the chitinase genes obtained from the isolates did not correspond well to that of the isolates, suggesting acquisition via horizontal gene transfer.     

Succession of Microbial Communities during Hot Composting as Detected by PCR–Single-Strand-Conformation Polymorphism-Based Genetic Profiles of Small-Subunit rRNA Genes

A cultivation-independent technique for genetic profiling of PCR-amplified small-subunit rRNA genes (SSU rDNA) was chosen to characterize the diversity and succession of microbial communities during composting of an organic agricultural substrate. PCR amplifications were performed with DNA directly extracted from compost samples and with primers targeting either (i) the V4–V5 region of eubacterial 16S rRNA genes, (ii) the V3 region in the 16S rRNA genes of actinomycetes, or (iii) the V8–V9 region of fungal 18S rRNA genes. Homologous PCR products were converted to single-stranded DNA molecules by exonuclease digestion and were subsequently electrophoretically separated by their single-strand-conformation polymorphism (SSCP). Genetic profiles obtained by this technique showed a succession and increasing diversity of microbial populations with all primers. A total of 19 single products were isolated from the profiles by PCR reamplification and cloning. DNA sequencing of these molecular isolates showed similarities in the range of 92.3 to 100% to known gram-positive bacteria with a low or high G+C DNA content and to the SSU rDNA of γ-Proteobacteria. The amplified 18S rRNA gene sequences were related to the respective gene regions of Candida krusei and Candida tropicalis. Specific molecular isolates could be attributed to different composting stages. The diversity of cultivated bacteria isolated from samples taken at the end of the composting process was low. A total of 290 isolates were related to only 6 different species. Two or three of these species were also detectable in the SSCP community profiles. Our study indicates that community SSCP profiles can be highly useful for the monitoring of bacterial diversity and community successions in a biotechnologically relevant process.