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Phthiocerol dimycocerosates (DIM) and phenolglycolipids (PGL) are functionally important surface-exposed lipids of Mycobacterium tuberculosis. Their biosynthesis involves the products of several genes clustered in a 70-kb region of the M. tuberculosis chromosome. Among these products is PpsD, one of the modular type I polyketide synthases responsible for the synthesis of the lipid core common to DIM and PGL. Bioinformatic analyses have suggested that this protein lacks a functional enoyl reductase activity domain required for the synthesis of these lipids. We have identified a gene, Rv2953, that putatively encodes an enoyl reductase. Mutation in Rv2953 prevents conventional DIM formation and leads to the accumulation of a novel DIM-like product. This product is unsaturated between C-4 and C-5 of phthiocerol. Consistently, complementation of the mutant with a functional pksl5/1 gene from Mycobacterium boris BCG resulted in the accumulation of an unsaturated PGL-like substance. When an intact Rv2953 gene was reintroduced into the mutant strain, the phenotype reverted to the wild type. These findings indicate that Rv2953 encodes a trans-acting enoyl reductase that acts with PpsD in phthiocerol and phenolphthiocerol biosynthesis.

The diversity of polyketide synthases (PKSs) in sponge-associated microbial communities is analyzed to study the uncultured sponge bacteria. The results have proposed a simplified strategy for the targeted isolation of unusually small PKS genes from complex sponge-symbiont associations.

It had been assumed that production of the cytotoxie polyketide mycolactone was strictly associated with Mycobacterium ulcerans, the causative agent of Buruli ulcer. However, a recent study has uncovered a broader distribution of mycolactone-producing mycobacteria (MPM) that includes mycobacteria cultured from diseased fish and frogs in the United States and from diseased fish in the Red and Mediterranean Seas. All of these mycobacteria contain versions of the M. ulcerans pMUM plasmid, produce mycolactones, and show a high degree of genetic relatedness to both M. ulcerans and Mycobacterium marinum. Here, we show by multiple genetic methods, including multilocus sequence analysis and DNA-DNA hybridization, that all MPM have evolved from a common M. marinum progenitor to form a genetically cohesive group among a more diverse assemblage of M. marinum strains. Like M. ulcerans, the fish and frog MPM show multiple copies of the insertion sequence IS2404. Comparisons of pMUM and chromosomal gene sequences demonstrate that plasmid acquisition and the subsequent ability to produce mycolactone were probably the key drivers of speciation. Ongoing evolution among MPM has since produced at least two genetically distinct ecotypes that can be broadly divided into those typically causing disease in ectotherms (but also having a high zoonotic potential) and those causing disease in endotherms, such as humans.

Although bacterial polyketides are of considerable biomedical interest, the molecular biology of polyketide biosynthesis in Bacillus spp., one of the richest bacterial sources of bioactive natural products, remains largely unexplored. Here we assign for the first time complete polyketide synthase (PKS) gene clusters to Bacillus antibiotics. Three giant modular PKS systems of the trans-acyltransferase type were identified in Bacillus amyloliquefaciens FZB 42. One of them, pks1, is an ortholog of the pksX operon with a previously unknown function in the sequenced model strain Bacillus subtilis 168, while the pks2 and pks3 clusters are novel gene clusters. Cassette mutagenesis combined with advanced mass spectrometric techniques such as matrix-assisted laser desorption ionization--time of flight mass spectrometry and liquid chromatography-electrospray ionization mass spectrometry revealed that the pks1 (bae) and pks3 (dif) gene clusters encode the biosynthesis of the polyene antibiotics bacillaene and diflicidin or oxydifficidin, respectively. In addition, B. subtilis OKB105 (pheA [sfp.sup.0]), a transformant of the B. subtilis 168 derivative JH642, was shown to produce bacillaene, demonstrating that the pksX gene cluster directs the synthesis of that polyketide.

The use of Red/ET recombination to generate a large number of shuttle plasmids from a diverse library of chimeric domains for directed evolution of the pikromycin polyketide synthase (PKS) is reported. Red/ET recombination technology is adopted for generating new polyketide as it overcame some of the limitations of traditional approaches for PKS engineering.

Genes encoding a type I polyketide synthase (PKS) (PKS12) and a putative laccase (Gip1) that are required for aurofusarin biosynthesis by Gibberella zeae are isolated. Targeted gene deletion and complementation analyses confirmed that both Gip1 and PKS12 are required for aurofusarin production in Gibberella zeae.

The bacteria in the pallial sinuses of Bugula simplex larvae was identified and whether polyketide synthase (PKS) genes and bryostatin-like polyketides were present in the organism was determined. The reports demonstrate a symbiosis in B. simplex that is similar and evolutionarily related to that in B. neritina.

Secondary metabolite genes of the model fungus Aspergillus nidulans were screened to identify a silent gene cluster on chromosome II comprising two nonribosomal peptide synthetase (NRPS) genes, inpA and inpB, flanked by a regulatory gene scpR for secondary metabolism cross-pathway regulator. The existence of the polyketide product asperfuranone revealed that the transcription factor scpR controls the expression of the asperfuranone biosynthesis gene cluster located on different chromosomes.

The gene disruption and enzyme activity of hypC, an open reading frame in the region between the pksA (aflC) and nor-I (aflD) genes in the aflatoxin biosynthesis gene cluster is reported. The gene HypC, the anthrone oxidase involved in aflatoxin biosynthesis encodes a 17-kDa oxidase that converts norsolorinic acid anthrone to nonsolorinic acid.

The possible use of heterologous expression of bacterial biosynthetic pathways in metagenomic DNA libraries to access new natural products for drug discovery is discussed. The study demonstrated the application of Myxococcus xanthus as a potential 'universal' host for heterologous expression of polyketide biosynthetic gene clusters.

A study related to the gene fscTE, encoding a putative type II thioesterase (TEII) associated with the FR-008/candicidin gene cluster, is reported. The substrate specificity of fscTE gene is confirmed by its overexpression of fscTE under the strong constitutive ermE*p promoter in the wild-type strain, which did not suppress FR-008/candicidin formation.

The diversity of type I modular polyketide synthase (PKS) is analyzed by using PCR amplification of DNA encoding ketosynthase and acyltransferase domains in myxobacteria. The sequencing of the amplicons has shown that many PKS genes are distantly related to the sequences, indicating myxobacteria to be excellent resources for novel and diverse polyketides.

Extensive sponge-derived metagenomic libraries for diverse polyketide synthases (PKS) genes that are useful for identifying target PKS genes in other sponges are described. It indicates that the filamentous bacterial community of Discodermia dissoluta consists mainly of Entotheonella spp, an unculturable sponge-specific taxon implicated in the biosynthesis of bioactive peptides.

The cloning and characterization of a major portion of the herbimycin (hbm) biosynthetic gene cluster containing the polyketide synthase (PKS) genes and some other genes involved in post-PKS modifications are described. The results show a novel geldanamycin analog, KOS-1806 which was isolated and characterized.

The study aims to develop a culture-independent method to identify genes encoding novel and unique polyketide synthases (PKS) pathways. It also hypothesized that soil containing a large diversity of actinomycetes, as indicated by terminal restriction fragment length polymorphism (TRFLP), also contains a large diversity of PKS gene sequences.

The PCR primers are designed in the conserved regions of the ketosynthase domain (KS) to detect type I polyketide synthases (PKSI) genes in recombinant clones from a large metagenomic soil library. The phylogenetic analysis of PKSI domains led to the detection of numerous PKSI-positive clones and the selection of two promising clones for the potential production of new compounds.

A polykitide synthase (PKS) gene is identified from a strain of nextria haematococca by complete menting a mutant unable to synthesize a red perithecial pigment, pksN encodes a 2,106-amino-acid polypeptide with conserved motifs characteristic of type I PKS enzymatic domains. It is proposed that pksN encodes a novel PKS required for the perithecial red pigment biosynthesis.

Spirochetes are among the most abundant microbial groups in termite hindguts, accounting for as much as one-half of the prokaryotic community. The nutritional, physiological, and biochemical properties of Treponema primitia strains ZAS-1 and ZAS-2, H2/CO2- acetogenic spirochetes isolated from the hindgut of the California dampwood termite, is reported.