Literature Review of Pathogen Agnostic Molecular Testing of Clinical Specimens From Difficult-to-Diagnose Patients: Implications for Public Health.

To better identify emerging or reemerging pathogens in patients with difficult-to-diagnose infections, it is important to improve access to advanced molecular testing methods. This is particularly relevant for cases where conventional microbiologic testing has been unable to detect the pathogen and the patient's specimens test negative. To assess the availability and utility of such testing for human clinical specimens, a literature review of published biomedical literature was conducted.

Surveillance for Emerging and Reemerging Pathogens Using Pathogen Agnostic Metagenomic Sequencing in the United States: A Critical Role for Federal Government Agencies.

The surveillance and identification of emerging, reemerging, and unknown infectious disease pathogens is essential to national public health preparedness and relies on fluidity, coordination, and interconnectivity between public and private pathogen surveillance systems and networks. Developing a national sentinel surveillance network with existing resources and infrastructure could increase efficiency, accelerate the identification of emerging public health threats, and support coordinated intervention strategies that reduce morbidity and mortality. However, implementing and sustaining programs to detect emerging and reemerging pathogens in humans using advanced molecular methods, such as metagenomic sequencing, requires making large investments in testing equipment and developing networks of clinicians, laboratory scientists, and bioinformaticians.

Building a Public-Private Partnership to Enhance Laboratory Preparedness and Response in the United States.

The public health community has recognized that it cannot handle responses to all possible public health emergencies on its own. The public health sector has deep scientific expertise and excels at initial identification, complex characterization, and test development. The private sector has many resources and capabilities that can complement and augment the public health response.

Establishing a Hospital Response Network Among Children's Hospitals.

A timely and effective response to public health threats requires a broad-reaching infrastructure. Children's hospitals are focused on evaluating and managing some of the most vulnerable patients and thus have unique preparedness and response planning needs. A virtual forum was established specifically for children's hospitals during the 2014-15 Ebola outbreak, and it demonstrated the importance and utility of connecting these specialty hospitals to discuss their shared concerns.

Genetic homogeneity of Clostridium botulinum type A1 strains with unique toxin gene clusters.

A group of five clonally related Clostridium botulinum type A strains isolated from different sources over a period of nearly 40 years harbored several conserved genetic properties. These strains contained a variant bont/A1 with five nucleotide polymorphisms compared to the gene in C. botulinum strain ATCC 3502.

Analysis of neurotoxin cluster genes in Clostridium botulinum strains producing botulinum neurotoxin serotype A subtypes.

Neurotoxin cluster gene sequences and arrangements were elucidated for strains of Clostridium botulinum encoding botulinum neurotoxin (BoNT) subtypes A3, A4, and a unique A1-producing strain (HA(-) Orfx(+) A1). These sequences were compared to the known neurotoxin cluster sequences of C. botulinum strains that produce BoNT/A1 and BoNT/A2 and possess either a hemagglutinin (HA) or an Orfx cluster, respectively.

Real-time PCR detection of the nontoxic nonhemagglutinin gene as a rapid screening method for bacterial isolates harboring the botulinum neurotoxin (A-G) gene complex.

Botulinum neurotoxin (BoNT) producing clostridia contain genes encoding a specific neurotoxin serotype (A-G) and nontoxic associated proteins that form the toxin complex. The nontoxic nonhemagglutinin (NTNH) is a conserved component of the toxin complex in all seven toxin types. A real-time PCR assay that utilizes a locked nucleic acid hydrolysis probe to target the NTNH gene was developed to detect bacterial strains harboring the botulinum neurotoxin gene cluster.

Microarray-based detection of genetic heterogeneity, antimicrobial resistance, and the viable but nonculturable state in human pathogenic Vibrio spp.

The morbidity and mortality associated with Vibrio-mediated waterborne diseases necessitates the development of sensitive detection technologies that are able to elucidate the identity, potential pathogenicity, susceptibility, and viability of contaminating bacteria in a timely manner. For this purpose, we have designed a single multiplex PCR assay to simultaneously amplify 95 diagnostic regions (encompassing species/serogroup-specific, antimicrobial resistance, and known toxin markers) and combined it with a long oligonucleotide microarray to create a platform capable of rapidly detecting and discriminating the major human pathogenic species from the genus Vibrio: V. cholerae, V.

Nucleic acid amplification strategies for DNA microarray-based pathogen detection.

DNA microarray-based screening and diagnostic technologies have long promised comprehensive testing capabilities. However, the potential of these powerful tools has been limited by front-end target-specific nucleic acid amplification. Despite the sensitivity and specificity associated with PCR amplification, the inherent bias and limited throughput of this approach constrain the principal benefits of downstream microarray-based applications, especially for pathogen detection.

Potential applications of DNA microarrays in biodefense-related diagnostics.

Recent years have witnessed a logarithmic growth in the number of applications involving DNA microarrays. Extrapolation of their use for infectious diagnostics and biodefense-related diagnostics seems obvious. Nevertheless, the application of DNA microarrays to biodefense-related diagnostics will depend on solving a set of substantial, yet approachable, technical and logistical problems that encompass diverse topics from amplification efficiency to bioinformatics.