For cultivation-independent detection of sulfate-reducing prokaryotes (SRPs) an oligonucleotide microarray consisting of 132 16S rRNA gene-targeted oligonucleotide probes (18-mers) having hierarchical and parallel (identical) specificity for the detection of all known lineages of sulfate-reducing prokaryotes (SRP-PhyloChip) was designed and subsequently evaluated with 41 suitable pure cultures of SRPs. The applicability of SRP-PhyloChip for diversity screening of SRPs in environmental and clinical samples was tested by using samples from periodontal tooth pockets and from the chemocline of a hypersaline cyanobacterial mat from Solar Lake (Sinai, Egypt). Consistent with previous studies, SRP-PhyloChip indicated the occurrence of Desulfomicrobium spp. in the tooth pockets and the presence of Desulfonema - and Desulfomonile -like SRPs (together with other SRPs) in the chemocline of the mat. The SRP-PhyloChip results were confirmed by several DNA microarray-independent techniques, including specific PCR amplification, cloning, and sequencing of SRP 16S rRNA genes and the genes encoding the dissimilatory (bi)sulfite reductase ( dsrAB ).

[Enterobacter] sakazakii is an opportunistic pathogen that can cause infections in neonates. This study further clarifies the taxonomy of isolates described as [E.] sakazakii and completes the formal description of the proposed reclassification of these organisms as novel species and subspecies within a proposed novel genus, Cronobacter gen. nov. [E.] sakazakii was first defined in 1980, however recent polyphasic taxonomic analysis has determined that this group of organisms consists of several genomospecies. In this study, the phenotypic descriptions of the proposed novel species are expanded using Biotype 100 and Biolog Phenotype MicroArray data. Further DNA–DNA hybridization experiments showed that malonate-positive strains within the [E.] sakazakii genomospecies represent a distinct species, not a subspecies. DNA–DNA hybridizations also determined that phenotypically different strains within the proposed species, Cronobacter dublinensis sp. nov., belong to the same species and can be considered as novel subspecies. Based on these analyses, the following alternative classifications are proposed: Cronobacter sakazakii gen. nov., comb. nov. [type strain ATCC 29544T (=NCTC 11467T)]; Cronobacter malonaticus sp. nov. [type strain CDC 1058-77T (=LMG 23826T=DSM 18702T)]; Cronobacter turicensis sp. nov. [type strain z3032T (=LMG 23827T=DSM 18703T)]; Cronobacter muytjensii sp. nov. [type strain ATCC 51329T (=CIP 103581T)]; Cronobacter dublinensis sp. nov. [type strain DES187T (=LMG 23823T=DSM 18705T)]; Cronobacter dublinensis subsp. dublinensis subsp. nov. [type strain DES187T (=LMG 23823T=DSM 18705T)]; Cronobacter dublinensis subsp. lausannensis subsp. nov. [type strain E515T (=LMG 23824=DSM 18706T)], and Cronobacter dublinensis subsp. lactaridi subsp. nov. [type strain E464T (=LMG 23825T=DSM 18707T)].

Background: Enterobacter sakazakii is an opportunistic pathogen that can cause infections such as necrotizing enterocolitis, bacteraemia, meningitis and brain abscess/lesions. When the species was defined in 1980, 15 biogroups were described and it was suggested that these could represent multiple species. In this study the taxonomic relationship of strains described as E. sakazakii was further investigated. Results: Strains identified as E. sakazakii were divided into separate groups on the basis of f-AFLP fingerprints, ribopatterns and full-length 16S rRNA gene sequences. DNA-DNA hybridizations revealed five genomospecies. The phenotypic profiles of the genomospecies were determined and biochemical markers identified. Conclusion: This study clarifies the taxonomy of E. sakazakii and proposes a reclassification of these organisms.

For simultaneous identification of members of the betaproteobacterial order “Rhodocyclales” in environmental samples, a 16S rRNA gene-targeted oligonucleotide microarray (RHC-PhyloChip) consisting of 79 probes was developed. Probe design was based on phylogenetic analysis of available 16S rRNA sequences from all cultured and as yet uncultured members of the “Rhodocyclales.” The multiple nested probe set was evaluated for microarray hybridization with 16S rRNA gene PCR amplicons from 29 reference organisms. Subsequently, the RHC-PhyloChip was successfully used for cultivation-independent “Rhodocyclales” diversity analysis in activated sludge from an industrial wastewater treatment plant. The implementation of a newly designed “Rhodocyclales”-selective PCR amplification system prior to microarray hybridization greatly enhanced the sensitivity of the RHC-PhyloChip and thus enabled the detection of “Rhodocyclales” populations with relative abundances of less than 1% of all bacteria (as determined by fluorescence in situ hybridization) in the activated sludge. The presence of as yet unculturedZoogloea-,Ferribacterium/Dechloromonas-, andSterolibacterium-related bacteria in the industrial activated sludge, as indicated by the RHC-PhyloChip analysis, was confirmed by retrieval of their 16S rRNA gene sequences and subsequent phylogenetic analysis, demonstrating the suitability of the RHC-PhyloChip as a novel monitoring tool for environmental microbiology.