Confirmation of electron microscopy results by direct testing of viruses adhered to grids using nucleic acid amplification techniques

Abstract

It is possible to visualize rapidly viral particles by electron microscopy (EM) in patient samples and in cell cultures, and characterize the particles on the basis of their size and morphology. In many instances, EM has contributed to the diagnosis of specific infectious agents. Four different types of viruses with different characteristics of particle size, capsid structure, the presence or absence of an envelope, genomic content and stability outside the host were screened and diagnosed by EM at the level of family/genus. The results were confirmed at the species level by elution of the sample material from the grids used for EM examination and nucleic acid amplification. This approach could be valuable in situations where the immediate diagnosis is unclear, or when new infectious agents appear.

Detection limit of negative staining electron microscopy for the diagnosis of bioterrorism-related micro-organisms

“…With both poxviruses and spores, the application of a drop of the test suspension onto the grid was more efficient than placing the grid onto a drop of suspension (grid-on-drop procedure) indicating that particle density is high enough for a sufficient sedimentation by gravity. The detection limit determined here is in the range given by other studies using virus suspensions (Katz et al 1984;Biel and Gelderblom 1999a;Johnsen et al 2006). However, detection is not only influenced by the concentration of the relevant particles but also by the overall composition of the suspension.…”

“…The detection limit determined here is in the range given by other studies using virus suspensions (Katz et al. 1984; Biel and Gelderblom 1999a; Johnsen et al. 2006).…”

The role of electron microscopy in the rapid diagnosis of viral infections — review

“…Comparison of results obtained by EM and real-time PCR in the detection of viruses from untreated tissue culture showed the superiority of EM that found 38-times higher number of viral copies than PCR (Johnsen et al 2006). EM is occasionally used in combination with other viral methods to identify the causative viruses of zoonotic outbreaks and both seasonal and non-seasonal epidemics or pandemics.…”

Reply to: Disinfection in diagnostic electron microscopy prior to preparation?

“…Furthermore, it was demonstrated recently that FA-treated viruses adhering to EM grids can be analyzed in targeted PCR, using the information obtained by EM. 6 So there are several good reasons for disinfection and GLP in EM diagnostics.…”

“…1 Saccharomyces boulardii (Sb), a probiotic agent, has been shown to be effective in prevention and treatment of diarrhea due to Clostridium difficile, 2,3 enteral feeding 4 or antibiotics, 5 and traveler's diarrhea. 6 Sb is assumed to be a subtype of Saccharomyces cerevisiae (Sc) and is usually considered safe. 7 However, several Sc infections have been reported, including fungemia, endocarditis, pneumonia, liver abscesses, peritonitis, vaginitis, and urinary tract infections, 8 and were caused by touch contamination from patients staying in the same room or enteral translocation.…”