Adaptation and growth of Serratia marcescens in contact lens disinfectant solutions containing chlorhexidine gluconate.

Serratia marcescens (11 of 12 strains) demonstrated an ability to grow in certain chlorhexidine-based disinfecting solutions recommended for rigid gas-permeable contact lenses. For a representative strain, cells that were grown in nutrient-rich medium, washed, and inoculated into disinfecting solution went into a nonrecoverable phase within 24 h. However, after 4 days, cells that had the ability to grow in the disinfectant (doubling time, g = 5.7 h) emerged. Solutions supporting growth of S. marcescens were filter sterilized. These solutions, even after removal of the cells, showed bactericidal activity against Pseudomonas aeruginosa and a biphasic survival curve when rechallenged with S. marcescens. Adaptation to chlorhexidine by S. marcescens was not observed in solutions formulated with borate ions. For chlorhexidine-adapted cells, the MIC of chlorhexidine in saline was eightfold higher than that for unadapted cells. Cells adapted to chlorhexidine showed alterations in the proteins of the outer membrane and increased adherence to polyethylene. Cells adapted to chlorhexidine persisted or grew in several other contact lens solutions with different antimicrobial agents, including benzalkonium chloride.     

Development of reproducible test inocula for disinfectant testing

For the development and approval of disinfectants, laboratory tests which combine repeatability and reproducibility with relevance to practical conditions are required to ensure optimum standards of efficacy under use conditions. Over the years, although an increasingly rigorous approach has been adopted in devising European and US Standard Test Methods for disinfectants, which includes specifying all aspects of test methodology, the precision of these methods remains a matter for concern. Studies of proposed European test methods indicate that, although some of the variability is methodological in origin, or is derived from errors in preparing test solutions and performing the tests, one of the major sources of error is lack of reproducibility in the performance of the test inoculum. Results indicate that day to day variability in biocide sensitivity of sequential subcultures arises not only from variations in the phenotype generated from the laboratory stock culture, but also from lack of standardization of conditions used to harvest and prepare test inocula. Further reductions in reproducibility between test periods both within or between laboratories probably arise from alterations in the genotype of the laboratory stock culture or source culture during storage under conditions of refrigeration, freezing or freeze-drying. Methods for production of reproducible inocula are considered and some studies aimed at development of test inocula with more reproducible biocide resistance are described.     

An investigation into the differences between the Bioscreen and the traditional plate count disinfectant test methods

Investigations of biocide efficacy by automated methods involving optical density measurements, e.g. using the recently published 'Bioscreen' method, are complicated by the fact that a poor correlation often exists between the log reductions obtained using the automated method vs those obtained by the traditional plate count methods. It was hypothesized that the differences observed between the two methods were due to the level of cell injury, which was masked by the optical density methods but which was recognized by the plate counts. Comparisons of log reductions following a disinfection test always showed the Bioscreen method to be overestimating the log reductions with respect to the plate counts. A correlation between colony size on the plates and the 'Bioscreen' results for a fixed disinfectant concentration and contact time was found using Global Imaging software. The results obtained also suggested that the observed colony area was dependent on the amount of injury incurred by a microbe during the disinfection process. A mathematical model of injury was developed which predicted the observed differences between the Bioscreen and the traditional plate method. The model further suggested a possible reason for biocidal lags.     

Manganese toxicity in standard culture solutions

Summary Atlas barley plants grown in standard Hoagland culture solutions developed dark brown necrotic spots on the older leaves. The symptoms varied from small freckle-like spots to large blotchy areas and were found to be associated with the concentrations of Mn and B in the culture solutions. An increase in the concentration of Mn or B in the solutions increased the intensity of the spotting. A decrease in the Mn- and B-concentrations to 0.025 ppm, or one-twentieth of the normal Hoagland value, caused the spots to almost completely disappear. Mn- and B-concentrations of 0.025 ppm are optimum only under a particular set of conditions. In deciding what concentration of Mn and B to use the number of plants, volume of solution, macro-salt concentration, and season must be taken into consideration. The recommended Hoagland concentrations of Mn and B were only slightly toxic to lettuce and non-toxic to tomato plants. Barley plants grown in the winter were able to tolerate much higher concentrations of Mn and B before showing toxicity symptoms. Mn- and B-toxicity symptoms on lettuce and barley are compared in photographs.     

Influence of possible disinfectant transfer on Staphylococcus aureus plate counts after agar contact sampling.

Asbestos-vinyl tile floor panels were mopped with three types of chemical disinfectant product, as well as after contact, with the untreated control panel were prepared according to the manufacturer's label instructions. Similar floor panels were inoculated artificially with Staphylococcus aureus. RODAC (replicate organism detection and counting) surface sampling plates were pressed to the disinfectant-treated panels or to the untreated control panel and then immediately pressed to sampling sites on the artificially inoculated floor panels. Plate counts were determined after contact with panels treated with each type of disinfectant, product, as well as after contact with the untreated control panel. Results indicate that disinfectant residues on environmental surfaces do not alter the average plate counts obtained by RODAC samplings.     

Revised methods for the Salmonella mutagenicity test

The methods for detecting carcinogens and mutagens with the Salmonella mutagenicity test were described previously (Ames et al., 1975b). The present paper is a revision of the methods. Two new tester strains, a frameshift strain (TA97) and a strain carrying an ochre mutation on a multicopy plasmid (TA102), are added to the standard tester set. TA97 replaces TA1537. TA1535 and TA1538 are removed from the recommended set but can be retained at the option of the investigator. TA98 and TA100 are retained. We discuss other special purpose strains and present some minor changes in procedure, principally in the growth, storage, and preservation of the tester strains. Two substitutions are made in diagnostic mutagens to eliminate MNNG and 9-aminoacridine. Some test modifications are discussed.     

Comparative testing of disinfectant and antiseptic products using proposed European suspension testing methods

The development of standard suspension test methods for disinfectants and antiseptics for adoption in Europe is described. Evaluation of a range of disinfectant agents and products currently used in the UK under conditions as proposed for these tests indicates that the majority of products diluted in water of standard hardness showed satisfactory activity producing a 4·5–5 log reduction in viable count within 5 min against Staphylococcus aureus, Pseudomonas aeruginosa, Enterococcus faecium, Proteus mirabilis and Candida albicans in the absence and presence of 1% albumin. All the products, when diluted with distilled water, produced greater than 5 log reduction in 60 min.     

Laboratory methods for assessing the toxicity of contact poisons to slugs

Two techniques for comparing the activity of different contact poisons to slugs in controlled conditions were used to measure the relative toxicities of five substances. A laboratory immersion test rated their median lethal concentrations as follows: ioxynil 8·3 ppm, sodium pentachlorophenate 22·0 ppm, copper sulphate 68·1–75·3 ppm, acetaldehyde 4822·0 ppm. Metaldehyde gave inconsistent results with this method but, using a dry-contact method, metaldehyde (42370 ppm) was much less toxic than copper sulphate (2027 ppm). The materials giving practical control in the field were not the most toxic of those tested.     

A comprehensive assessment of sequence-based and template-based methods for protein contact prediction

Motivation: Pair-wise residue-residue contacts in proteins canbe predicted from both threading templates and sequence-basedmachine learning. However, most structure modeling approachesonly use the template-based contact predictions in guiding thesimulations; this is partly because the sequence-based contactpredictions are usually considered to be less accurate thanthat by threading. With the rapid progress in sequence databasesand machine-learning techniques, it is necessary to have a detailedand comprehensive assessment of the contact-prediction methodsin different template conditions. Results: We develop two methods for protein-contact predictions:SVM-SEQ is a sequence-based machine learning approach whichtrains a variety of sequence-derived features on contact maps;SVM-LOMETS collects consensus contact predictions from multiplethreading templates. We test both methods on the same set of554 proteins which are categorized into ‘Easy’,‘Medium’, ‘Hard’ and ‘Very Hard’targets based on the evolutionary and structural distance betweentemplates and targets. For the Easy and Medium targets, SVM-LOMETSobviously outperforms SVM-SEQ; but for the Hard and Very Hardtargets, the accuracy of the SVM-SEQ predictions is higher thanthat of SVM-LOMETS by 12–25%. If we combine the SVM-SEQand SVM-LOMETS predictions together, the total number of correctlypredicted contacts in the Hard proteins will increase by morethan 60% (or 70% for the long-range contact with a sequenceseparation 24), compared with SVM-LOMETS alone. The advantageof SVM-SEQ is also shown in the CASP7 free modeling targetswhere the SVM-SEQ is around four times more accurate than SVM-LOMETSin the long-range contact prediction. These data demonstratethat the state-of-the-art sequence-based contact predictionhas reached a level which may be helpful in assisting tertiarystructure modeling for the targets which do not have close structuretemplates. The maximum yield should be obtained by the combinationof both sequence- and template-based predictions. Contact: yzhang{at}ku.edu Supplementary information: Supplementary data are availableat Bioinformatics online. Associate Editor: Anna Tramontano     

Efficacy of sticky and standard ovitraps for Aedes aegypti in Trinidad,West Indies

The double sticky trap (DST) is described for the first time and is evaluated along with standard ovitraps and sticky traps (STs) to determine population densities of Ae. aegypti in the urban township of St. Augustine and the rural community of Tamana, Trinidad, West Indies. Ten houses were selected at each study site. At each of the ten houses, one ovitrap, one ST, and one DST were placed using the criteria established for placement of ovitraps. The results showed the three trapping methods successfully collected Ae. aegypti mosquitoes. All three traps collected significantly more adults or eggs in St. Augustine than in Tamana. DSTs collected 2,286 adults from St. Augustine vs 316 adults from Tamana (p<0.002), STs collected 1,480 and 220 adults, respectively (p<0.01), and the ovitraps collected 2,735 and 517 eggs, respectively from St. Augustine and Tamana (p<0.002). Based on these results, the DSTs collected significantly (P<0.02) more adults than the STs. The DSTs and STs collected both adult and immature stages which can be used for toxicology, virology, and PCR studies and are suitable alternative Ae. aegypti surveillance tools for the Caribbean and Latin American region.     

The HKA test revisited: a maximum-likelihood-ratio test of the standard neutral model

We present a maximum-likelihood-ratio test of the standard neutral model, using multilocus data on polymorphism within species and divergence between species. The model is based on the Hudson-Kreitman-Aguade (HKA) test, but allows for an explicit test of selection at individual loci in a multilocus framework. We use coalescent simulations to show that the likelihood-ratio test statistic is conservative, particularly when the assumption of no recombination is violated. Application of the method to polymorphism data from 18 loci from a population of Arabidopsis lyrata provides significant evidence for a balanced polymorphism at a candidate locus thought to be linked to the centromere. The method is also applied to polymorphism data in maize, providing support for the hypothesis of directional selection on genes in the starch pathway.     

A critical assessment of standard processing methods for the preparation of palynological samples

Standard processing techniques for the isolation of organic walled dinoflagellate cysts from geological samples are examined, with particular attention to the size and type of sieve mesh used. Variations within the ‘standard’ processing techniques used by different laboratories are identified, and an assessment of the retention capacities of meshes of different sizes and different materials is carried out. Some dinoflagellate cysts and large numbers of Lycopodium spores, used for the calculations of absolute abundance data, were found to pass through 20 μm meshes. This is due to a combination of factors including: the diagonal aperture diameter of a 20 μm mesh measuring over 28 μm; the three-dimensional properties of different mesh weaves (nylon and polyester); and the non-spherical shape of the particles. Experiments demonstrate that the maximum mesh size that should be used in palynological processing is 15 μm. Nylon mesh is more practical to use than polyester as processing time is reduced, but nylon is degraded by contact with acid solutions. Meshes with apertures < 15 μm may be used, though this may be impractical for large samples containing significant quantities of fine siliciclastic or organic material.     

Efficacy of a pentaiodide resin disinfectant on Cryptosporidium parvum (Apicomplexa: Cryptosporidiidae) oocysts in vitro

The resin-I5 column developed at Kansas State University was tested for efficacy against oocysts of Cryptosporidium parvum (Apicomplexa: Cryptosporidiidae). Cesium chloride gradient-purified oocysts were passed through 1.0-cm-diameter columns with lengths of 2.5, 5.0, and 10.0 cm at 23 C. Following column passage, oocyst viability was determined both in vitro by excystation and in vivo by the ability to establish infections in suckling mice. Oocysts were found to be retained by the pentaiodide resin in a linear fashion, probably by electrostatic interactions. Linear regression analysis revealed 100% of the oocysts should be removed in such a manner using a column length of greater than or equal to 25.7 cm. When compared to untreated control oocysts, less than 12% of the oocysts that passed through the columns appeared to be affected by the resin, as assessed by excystation. Inoculation of suckling mice with these column-treated oocysts supported the excystation data and revealed the coccidian to be viable. These results indicate that oocysts of C. parvum are retained on the pentaiodide column in a 1-hit manner and that, although killing of parasites may occur within the column, the greatest effect that the column may have on the parasite is as an electrostatic retention device.     

《中国药典》中标准菌种质控新方法的研究

目的 研究《中华人民共和国药典》(简称《中国药典》)纳入的标准菌种质控新方法,并评价不同批号标准菌种的质量稳定性。方法 对脉冲场凝胶电泳(PFGE)技术进行比较研究,同时整合16SrRNA基因序列分析、多位点序列分型和基质辅助激光解吸电离飞行时间质谱等质控新方法,进行标准菌种质控新方法的建立,并对标准菌种的质量进行评价。结果 形成了适用于《中国药典》中标准菌种的方法,并通过整合的质控新方法对不同批号的标准菌种进行评价,结果显示,菌种质量稳定,遗传信息无改变。同时,建立了标准菌种16SrRNA基因标准序列、PFGE标准指纹图谱和标准基因型。结论 标准菌种质控新方法的研究,为更加全面、深入地评价标准菌种的质量提供了依据;建立的标准菌种质量控制体系及标准菌种质控鉴定信息,为标准菌种持续的质量控制奠定了重要的参比信息基础。