A model of base-call resolution on broad-spectrum pathogen detection resequencing DNA microarrays

Oligonucleotide microarrays offer the potential to efficiently test for multiple organisms, an excellent feature for surveillance applications. Among these, resequencing microarrays are of particular interest, as they possess additional unique capabilities to track pathogens’ genetic variations and perform detailed discrimination of closely related organisms. However, this potential can only be realized if the costs of developing the detection microarray are kept at a manageable level. Selection and verification of the probes are key factors affecting microarray design costs that can be reduced through the development and use of in silico modeling. Models created for other types of microarrays do not meet all the required criteria for this type of microarray. We describe here in silico methods for designing resequencing microarrays targeted for multiple organism detection. The model development presented here has focused on accurate base-call prediction in regions that are applicable to resequencing microarrays designed for multiple organism detection, a variation from other uses of a predictive model in which perfect prediction of all hybridization events is necessary. The model will assist in simplifying the design of resequencing microarrays and in reduction of the time and costs required for their development for new applications

Porphyrin-modified antimicrobial peptide indicators for detection of bacteria

This study demonstrates the potential of porphyrin modified antimicrobial peptides for indication of bacterial targets on the basis of changes in the spectrophotometric characteristics of the construct. Detection is a result of changes in the structure of the antimicrobial peptide upon target binding. Those constructs comprised of peptides that offer little or no change in conformation upon interaction with bacterial cells demonstrated negligible changes in absorbance and fluorescence when challenged using Escherichia coli or Bacillus cereus. CD analysis confirms the presence/absence of conformational changes in the porphyrin-peptide constructs. Differing spectrophotometric responses were observed for constructs utilizing different peptides. The incorporation of metals into the porphyrin component of the constructs was shown to alter their spectrophotometric characteristics as well as the resulting absorbance and fluorescence changes noted upon interaction with a target. The described constructs offer the potential to enable a new type of biosensing approach in which the porphyrin-peptide indicators offer both target recognition and optical transduction, requiring no additional reagents

Resequencing microarray probe design for typing genetically diverse viruses: human rhinoviruses and enteroviruses

<p>Abstract</p> <p>Background</p> <p>Febrile respiratory illness (FRI) has a high impact on public health and global economics and poses a difficult challenge for differential diagnosis. A particular issue is the detection of genetically diverse pathogens, i.e. human rhinoviruses (HRV) and enteroviruses (HEV) which are frequent causes of FRI. Resequencing Pathogen Microarray technology has demonstrated potential for differential diagnosis of several respiratory pathogens simultaneously, but a high confidence design method to select probes for genetically diverse viruses is lacking.</p> <p>Results</p> <p>Using HRV and HEV as test cases, we assess a general design strategy for detecting and serotyping genetically diverse viruses. A minimal number of probe sequences (26 for HRV and 13 for HEV), which were potentially capable of detecting all serotypes of HRV and HEV, were determined and implemented on the Resequencing Pathogen Microarray RPM-Flu v.30/31 (<it>Tessarae RPM-Flu</it>). The specificities of designed probes were validated using 34 HRV and 28 HEV strains. All strains were successfully detected and identified at least to species level. 33 HRV strains and 16 HEV strains could be further differentiated to serotype level.</p> <p>Conclusion</p> <p>This study provides a fundamental evaluation of simultaneous detection and differential identification of genetically diverse RNA viruses with a minimal number of prototype sequences. The results demonstrated that the newly designed RPM-Flu v.30/31 can provide comprehensive and specific analysis of HRV and HEV samples which implicates that this design strategy will be applicable for other genetically diverse viruses.</p

Enabling methods for community health mapping in developing countries

<p>Abstract</p> <p>Background</p> <p>Spatial epidemiology is useful but difficult to apply in developing countries due to the low availability of digitized maps and address systems, accurate population distributions, and computational tools. A community-based mapping approach was used to demonstrate that participatory geographic information system (PGIS) techniques can provide information helpful for health and community development.</p> <p>Results</p> <p>The PGIS process allowed for the rapid determination of sectional (neighborhood) boundaries within the city of Bo, Sierra Leone. When combined with data about hospital laboratory visits, a catchment area for one hospital in Bo could be established. A survey of households from within the catchment area determined that the average population per household (about 6 individuals) was similar to that found in the 2004 census. However, we also found that the average house was inhabited by more than one household, for an average of 17.5 inhabitants per residential building, which is critical information to know when estimating population size using remote imagery that can detect and enumerate buildings.</p> <p>Conclusions</p> <p>The methods developed in this paper serve as a model for the involvement of communities in the generation of municipal maps and their application to community and health concerns.</p

Short Communication Reflectance-based detection of oxidizers in ambient air

This study used two types of paper supported materials with a prototype, reflectance-based detector for indication of hydrogen peroxide vapor under ambient laboratory conditions. Titanyl based indicators provide detection through reaction of the indicator resulting in a dosimeter type sensor, while porphyrin based indicators provide a reversible interaction more suitable to continuous monitoring applications. These indicators provide the basis for discussion of characteristics important to design of a sensor system including the application environment and duration, desired reporting frequency, and target specificity

Short Communication Reflectance-based detection of oxidizers in ambient air

This study used two types of paper supported materials with a prototype, reflectance-based detector for indication of hydrogen peroxide vapor under ambient laboratory conditions. Titanyl based indicators provide detection through reaction of the indicator resulting in a dosimeter type sensor, while porphyrin based indicators provide a reversible interaction more suitable to continuous monitoring applications. These indicators provide the basis for discussion of characteristics important to design of a sensor system including the application environment and duration, desired reporting frequency, and target specificity

Metaproteomic evidence of changes in protein expression following a change in electrode potential in a robust biocathode microbiome

Microorganisms that respire electrodes may be exploited for biotechnology applications if key pathways for extracellular electron transfer (EET) can be identified and manipulated through bioengineering. To determine whether expression of proposed Biocathode-MCL EET proteins are changed by modulating electrode potential without disrupting the relative distribution of microbial constituents, metaproteomic and 16S rRNA gene expression analyses were performed after switching from an optimal to suboptimal potential based on an expected decrease in electrode respiration. Five hundred and seventy-nine unique proteins were identified across both potentials, the majority of which were assigned to three previously defined Biocathode-MCL metagenomic clusters: a Marinobacter sp., a member of the family Chromatiaceae, and a Labrenzia sp. Statistical analysis of spectral counts using the Fisher's exact test identified 16 proteins associated with the optimal potential, five of which are predicted electron transfer proteins. The majority of proteins associated with the suboptimal potential were involved in protein turnover/turnover, motility, and membrane transport. Unipept and 16S rRNA gene expression analyses indicated that the taxonomic profile of the microbiome did not change after 52 hours at the suboptimal potential. These findings show that protein expression is sensitive to the electrode potential without inducing shifts in community composition, a feature that may be exploited for engineering Biocathode-MCL

A Previously Uncharacterized, Nonphotosynthetic Member of the Chromatiaceae Is the Primary CO_2-Fixing Constituent in a Self-Regenerating Biocathode

Biocathode extracellular electron transfer (EET) may be exploited for biotechnology applications, including microbially mediated O_2 reduction in microbial fuel cells and microbial electrosynthesis. However, biocathode mechanistic studies needed to improve or engineer functionality have been limited to a few select species that form sparse, homogeneous biofilms characterized by little or no growth. Attempts to cultivate isolates from biocathode environmental enrichments often fail due to a lack of some advantage provided by life in a consortium, highlighting the need to study and understand biocathode consortia in situ. Here, we present metagenomic and metaproteomic characterization of a previously described biocathode biofilm (+310 mV versus a standard hydrogen electrode [SHE]) enriched from seawater, reducing O_2, and presumably fixing CO_2 for biomass generation. Metagenomics identified 16 distinct cluster genomes, 15 of which could be assigned at the family or genus level and whose abundance was roughly divided between Alpha- and Gammaproteobacteria. A total of 644 proteins were identified from shotgun metaproteomics and have been deposited in the the ProteomeXchange with identifier PXD001045. Cluster genomes were used to assign the taxonomic identities of 599 proteins, with Marinobacter, Chromatiaceae, and Labrenzia the most represented. RubisCO and phosphoribulokinase, along with 9 other Calvin-Benson-Bassham cycle proteins, were identified from Chromatiaceae. In addition, proteins similar to those predicted for iron oxidation pathways of known iron-oxidizing bacteria were observed for Chromatiaceae. These findings represent the first description of putative EET and CO_2 fixation mechanisms for a self-regenerating, self-sustaining multispecies biocathode, providing potential targets for functional engineering, as well as new insights into biocathode EET pathways using proteomics

Automated identification of multiple micro-organisms from resequencing DNA microarrays

There is an increasing recognition that detailed nucleic acid sequence information will be useful and even required in the diagnosis, treatment and surveillance of many significant pathogens. Because generating detailed information about pathogens leads to significantly larger amounts of data, it is necessary to develop automated analysis methods to reduce analysis time and to standardize identification criteria. This is especially important for multiple pathogen assays designed to reduce assay time and costs. In this paper, we present a successful algorithm for detecting pathogens and reporting the maximum level of detail possible using multi-pathogen resequencing microarrays. The algorithm filters the sequence of base calls from the microarray and finds entries in genetic databases that most closely match. Taxonomic databases are then used to relate these entries to each other so that the microorganism can be identified. Although developed using a resequencing microarray, the approach is applicable to any assay method that produces base call sequence information. The success and continued development of this approach means that a non-expert can now perform unassisted analysis of the results obtained from partial sequence data