Limitations of the widely used GAM42a and BET42a probes targeting bacteria in the Gammaproteobacteria radiation

The 23S rRNA-targeted probes GAM42a and BET42a provided equivocal results with the uncultured gammaproteobacterium 'Candidatus Competibacter phosphatis' where some cells bound GAM42a and other cells bound BET42a in fluorescence in situ hybridization (FISH) experiments. Probes GAM42a and BET42a span positions 1027-1043 in the 23S rRNAand differ from each other by one nucleotide at position 1033. Clone libraries were prepared from PCR products spanning the 16S rRNA genes, intergenic spacer region and 23S rRNA genes from two mixed cultures enriched in 'Candidatus C. phosphatis'. With individual clone inserts, the 16S rDNA portion was used to confirm the source organism as 'Candidatus C. phosphatis' and the 23S rDNA portion was used to determine the sequence of the GAM42a/BET42a probe target region. Of the 19 clones sequenced, 8 had the GAM42a probe target (T at position 1033) and 11 had G at position 1033, the only mismatch with GAM42a. However, none of the clones had the BET42a probe target (A at 1033). Non-canonical base-pairing between the 23S rRNA of 'Candidatus C. phosphatis' with G at position 1033 and GAM42a (G-A) or BET42a (G-T) is likely to explain the probing anomalies. A probe (GAM42_C1033) was optimized for use in FISH, targeting cells with G at position 1033, and was found to highlight not only some 'Candidatus C. phosphatis' cells, but also other bacteria. This demonstrates that there are bacteria in addition to 'Candidatus C. phosphatis' with the GAM42_C1033 probe target and not the BET42a or GAM42a probe target.

Limitations to global disease control

In October 2000, Australia was declared poliomyelitis-free” by the World Health Organisation. This declaration followed some extensive six years of surveillance of all cases of acute flaccid paralysis, by the Poliomyelitis Expert Surveillance Committee (Centre for Disease Control, Commonwealth Department of Health and Aged Care, Canberra), chaired by the author. There have been seven attempts in the history of the world to eliminate a disease from the earth’s surface. The dramatic failure of five of these, the success of global smallpox eradication, and current successes and difficulties in the case of attempts to eliminate poliomyelitis worldwide, lead one to an analysis of the factors which led both to success and to failure. The global eradication of a specific disease is one of the most important endeavours which the international community can undertake. This audit reviews the details of such approaches; that such might be used as tools for the future.

The utility and limitations of chloroplast DNA analysis for identifying native British oak stands and for guiding replanting strategy

We report a method using variation in the chloroplast genome (cpDNA) to test whether oak stands of unknown provenance are of native and/or local origin. As an example, a sample of test oaks, of mostly unknown status in relation to nativeness and localness, were surveyed for cpDNA type. The sample comprised 126 selected trees, derived from 16 British seed stands, and 75 trees, selected for their superior phenotype (201 tree samples in total). To establish whether these two test groups are native and local, their cpDNA type was compared with that of material from known autochthonous origin (results of a previous study which examined variation in 1076 trees from 224 populations distributed across Great Britain). In the previous survey of autochthonous material, four cpDNA types were identified as native; thus if a test sample possessed a new haplotype then it could be classed as non-native. Every one of the 201 test samples possessed one of the four cpDNA types found within the autochthonous sample. Therefore none could be proven to be introduced and, on this basis, was considered likely to be native. The previous study of autochthonous material also found that cpDNA variation was highly structured geographically and, therefore, if the cpDNA type of the test sample did not match that of neighbouring autochthonous trees then it could be considered to be non-local. A high proportion of the seed stand group (44.2 per cent) and the phenotypically superior trees (58.7 per cent) possessed a cpDNA haplotype which matched that of the neighbouring autochthonous trees and, therefore, can be considered as local, or at least cannot be proven to be introduced. The remainder of the test sample could be divided into those which did not grow in an area of overall dominance (18.7 per cent of seed stand trees and 28 per cent of phenotypically superior) and those which failed to match the neighbouring autochthonous haplotype (37.1 per cent and 13.3 per cent, respectively). Most of the non-matching test samples were located within 50 km of an area dominated by a matching autochthonous haplotype (96.0 per cent and 93.5 per cent, respectively), and potentially indicates only local transfer. Whilst such genetic fingerprinting tests have proven useful for assessing the origin of stands of unknown provenance, there are potential limitations to using a marker from the chloroplast genome (mostly adaptively neutral) for classifying seed material into categories which have adaptive implications. These limitations are discussed, particularly within the context of selecting adaptively superior material for restocking native forests.

Limitations of the ultracentrifugal approach for measuring the effective net charge of a macroion

Limitations have been detected in a recently published method for macroion valence determination by an ultracentrifugal procedure for quantifying the Dorman distribution of small ions in macroion solutions dialyzed against buffer supplemented with chromate as an indicator ion. The limitation reflects an implicit assumption that sedimentation velocity affords an unequivocal means of separating effects of chromate binding from those reflecting the Dorman redistribution of small ions. Although the assumed absence of significant Dorman redistribution of small ions across the sedimenting macroion boundary seemingly holds for some systems, this approximation is demonstrably invalid for others. Despite preliminary signs of promise, the ultracentrifugal procedure does not afford a simple, readily applied solution to the problem of unequivocal macroion valence determination. (C) 2004 Elsevier Inc. All rights reserved.

Limitations of asymmetric parallel-beam geometry

Bragg diffraction peak profiles and intensities in asymmetric (Omega-2theta) diffraction using a mirror-based parallel-beam geometry were compared with symmetric parallel-beam (theta-2theta) and conventional Bragg - Brentano (theta-2theta) diffraction for a powdered quartz sample and the NIST standard reference material (SRM) 660a (LaB6, lanthanum hexaboride). A comparison of the intensities and line widths (full width at half-maximum, FWHM) of these techniques demonstrated that low incident angles (Omega < 5&DEG;) are preferable for the parallel-beam setup. For higher &UOmega; values, if 2θ < 2Omega, mass absorption reduces the intensities significantly compared with the Bragg - Brentano setup. The diffraction peak shapes for the mirror geometry are more asymmetric and have larger FWHM values than corresponding peaks recorded with a Bragg - Brentano geometry. An asymmetric mirror-based parallel-beam geometry offers some advantages in respect of intensity when compared with symmetric geometries, and hence may be well suited to quantitative studies, such as those involving Rietveld analysis. A trial Rietveld refinement of a 50% quartz - 50% corundum mixture was performed and produced adequate results.

Plant-based immunocontraceptive control of wildlife - "potentials, limitations, and possums"

Possums (Trichosurus vulpecula), originally introduced from Australia, are spread over 90% of New Zealand and cause major economic and environmental damage. Immunocontraception has been suggested as a humane means to control them. Marsupial-specific reproductive antigens expressed at high levels in edible transgenic plant tissue might provide a safe, effective, and cheap oral delivery bait for immuno-contraceptive control. As proof of concept, female possums vaccinated with immunocontraceptive antigens showed reduced fertility, and possums fed with potato-expressed heat labile toxin-B (LT-B) had mucosal and systemic immune responses to the antigen. This demonstrated that immunocontraception was effective in possums and that oral delivery in edible plant material might be possible. Nuclear transformation with reporter genes showed that transgenic carrot roots accumulate high levels of foreign protein in edible tissues, indicating their potential as a delivery vector. However, prior to attempts at large scale production, more effective immunocontraceptive antigen-adjuvant formulations are probably required before plant-based immunocontraception can become a major tool for immunocontraceptive control of overabundant vertebrate pests. (c) 2004 Elsevier Ltd. All rights reserved.

Practical limitations in optical entanglement purification

Entanglement purification protocols play an important role in the distribution of entangled systems, which is necessary for various quantum information processing applications. We consider the effects of photodetector efficiency and bandwidth, channel loss and mode mismatch on the operation of an optical entanglement purification protocol. We derive necessary detector and mode-matching requirements to facilitate practical operation of such a scheme, without having to resort to destructive coincidence-type demonstrations.

The voice of a child: Archaeological limitations in interpretive stories

In the archaeological record, it seems children are rarely seen. If they are, children are referred to, to explain symbolism, rituals, past lifeways, and behavior of a society or culture rather than the past lifeways of children and their relationship to family and society. This lack of investigation suffers in all forms of archaeological research. However, this bias appears to be unconscious rather than intentionally applied. Archaeology, generally, involves the nameless and faceless rather than the individual. The archaeological signature of children appears minimal. It is adults, or more succinctly, society that generates material remains. This paper discusses interpretation of sites within the context of different archaeologies, thereby providing researchers with information that may not usually be considered when approaching interpretation of sites to visitors.

Limitations on bioassays in macronutrient deficiency determination

Short-term nutrient bioassays can be used to assess labile nutrient availability in soils. These bioassays rely on a high number of plants and small soil volumes to exploit labile soil resources rapidly and assess potential nutrient deficiency. A comparison of the Neubauer bioassay with conventional pot trial assessment of P and S availability in a Yellow Kurosol was undertaken. Changes in labile soil nutrients and enzyme activity after bioassay assessment were also measured. The Neubauer bioassay was able to detect increased labile P availability following P fertiliser application to the soil. This corresponded with response to added P in a longer-term pot trial using maize. As expected, phosphatase activity increased following the bioassay and labile P was depleted by the plants. However, although a longer-term pot trial demonstrated the Yellow Kurosol was responsive to S fertilisation, labile S pools were sufficiently large that the short-term Neubauer bioassay detected no difference in S availability to plants. Both soil sulphatase activity and labile soil S were elevated following the bioassay. The short period of contact between the roots of the bioassay and the soil may have limited S uptake and therefore the ability of the bioassay to identify a S responsive soil. When using bioassay techniques to assess labile nutrient availability, it is critical that the size of the labile nutrient pool present be considered for each element, and that the period of contact between the bioassay and soil being tested is long enough for plant uptake to lower the nutrient supply to a level that limits further uptake.