16 S rDNA primers and the unbiased assessment of thermophile diversity.

Our understanding of thermophile diversity is based predominantly on PCR studies of community DNA. "Universal" and domain-specific rRNA gene PCR primers have historically been used for the assessment of microbial diversity without adequate regard to the degree of specificity of primer pairs to different prokaryotic groups. In a reassessment of the published primers commonly used for "universal" and archaeal 16 S rDNA sequence amplification we note that substantial variations in specificity exist. An unconsidered choice of primers may therefore lead to significant bias in determination of microbial community composition. In particular, Archaea-specific primer sequences typically lack specificity for the Korarchaeota and Nanoarchaea and are often biased towards certain clades. New primer pairs specifically designed for "universal" archaeal 16 S rDNA sequence amplification, with homology to all four archaeal groups, have been designed. Here we present the application of these new primers for preparation of 16 S libraries from thermophile communities.

Metagenomics, gene discovery and the ideal biocatalyst.

With the rapid development of powerful protein evolution and enzyme-screening technologies, there is a growing belief that optimum conditions for biotransformation processes can be established without the constraints of the properties of the biocatalyst. These technologies can then be applied to find the "ideal biocatalyst" for the process. In identifying the ideal biocatalyst, the processes of gene discovery and enzyme evolution play major roles. However, in order to expand the pool genes for in vitro evolution, new technologies, which circumvent the limitations of microbial culturability, must be applied. These technologies, which currently include metagenomic library screening, gene-specific amplification methods and even full metagenomic sequencing, provide access to a volume of "sequence space" that is not addressed by traditional screening.

Review and re-analysis of domain-specific 16S primers.

The Polymerase Chain Reaction (PCR) has facilitated the detection of unculturable microorganisms in virtually any environmental source and has thus been used extensively in the assessment of environmental microbial diversity. This technique relies on the assumption that the gene sequences present in the environment are complementary to the "universal" primers used in their amplification. The recent discovery of new taxa with 16S rDNA sequences not complementary to standard universal primers suggests that current 16S rDNA libraries are not representative of true prokaryotic biodiversity. Here we re-assess the specificity of commonly used 16S rRNA gene primers and present these data in tabular form designed as a tool to aid simple analysis, selection and implementation. In addition, we present two new primer pairs specifically designed for effective "universal" Archaeal 16S rDNA sequence amplification. These primers are found to amplify sequences from Crenarchaeote and Euryarchaeote type strains and environmental DNA.

Bacterial community analysis of Indonesian hot springs.

We report the first attempts to describe thermophilic bacterial communities in Indonesia's thermal springs using molecular phylogenetic analyses. 16S rRNA genes from laboratory cultures and DNA directly amplified from three hot springs in West Java were sequenced. The 22 sequences obtained were assignable to the taxa Proteobacteria, Bacillus and Flavobacterium, including a number of clades not normally associated with thermophily.

Writing easily read patient education handouts: a computerized approach.

A patient education handout is a potentially powerful tool for communicating information to patients. Patients will derive the greatest benefit from what they can easily read and understand. This article describes the use of computerized readability formulas in writing patient education handouts. Suggestions are offered for writing such handouts are illustrated.

Increased readability improves the comprehension of written information for patients with skin disease.

Many standardized methods are available with which to evaluate the reading ease and comprehensibility of written material. Techniques depend on mechanical analysis of sentence length, multiple prepositional phrases, direct phraseology, and arrangement of printed materials on the page. Those techniques were used to analyze a pamphlet designed for patient education by the American Academy of Dermatology. The pamphlet scored a reading ease grade of 45, corresponding to what is considered difficult reading and at a level commonly found in academic journals. We rewrote the pamphlet and increased its reading ease score to 62.4, corresponding to material that appears in standard digest-type magazines. We gave both versions of the pamphlet to a group of first-year medical students and to a group of middle-class patients from a dermatology practice. In both groups the modified version led to greater understanding of the written material. We conclude that the usefulness of patient education materials now being distributed by dermatologists could be significantly improved by being rewritten according to well-recognized formulas.

Circumferential periosteal release in the treatment of children with leg-length inequality.

We present the results of 38 children with leg inequality treated by circumferential periosteal release. Leg-length discrepancy was expressed as a percentage of the longer limb. All patients showed a decrease in percentage difference at one year after operation, the mean difference dropping from 7.24% to 5.45%. The size of the response was directly related to the age of the patient at operation, being more pronounced in the younger patients. The response was not related to sex, diagnosis, or rate of growth of the patient immediately preceding operation.