Comparison of four diagnostic methods for detection of Helicobacter species in laboratory mice.

Four species of Helicobacter--H. muridarum, "H. rappini," H. hepaticus, and H. bilis--have been identified in the gastrointestinal tract of rodents. The association of Helicobacter species with chronic gastrointestinal diseases in mice has raised concern about their impact on research results. In this study, different methods for detection of Helicobacter species in the mouse intestinal tract were compared: polymerase chain reaction (PCR) amplification of 16S rRNA gene sequences, bacterial culture, electron microscopy, and histologic examination (Steiner stain). The PCR method was more sensitive in detecting murine Helicobacter species than was culture, electron microscopy, or histologic examination. Of the cecal specimens identified as Helicobacter species-positive by PCR, approximately 60% were identified as positive by each of the other methods. An 87.5% concordance was obtained by PCR screening of DNA from fecal and cecal specimens. Differentiation among murine Helicobacter species by colony morphologic or histologic features was not possible. Scanning electron microscopy and histologic examination indicated greater numbers of helical microorganisms, specifically H. hepaticus, in the cecum than in the colon. These results indicate that the PCR assays used can be performed on feces as a noninvasive means for rapidly screening large numbers of colony mice for murine Helicobacter species.

The amino-terminal one-third of pseudorabies virus glycoprotein gIII contains a functional attachment domain, but this domain is not required for the efficient penetration of Vero cells.

We have examined the attachment and penetration phenotypes of several glycoprotein gIII mutants of pseudorabies virus (PRV) and have identified the first one-third of gIII as a region that mediates efficient virus attachment to PK15 and Vero cells. This portion of gIII, amino acids 25 through 157 of the wild-type sequence, appeared to support attachment by binding to heparinlike molecules on cell surfaces. Virions containing the first one-third of gIII were sensitive to heparin competition and showed greatly reduced infectivity on cells treated with heparinase. PRV virions lacking the first one-third of the mature glycoprotein exhibited only residual binding to cells if challenged by vigorous washing with phosphate-buffered saline at 2 h postinfection at 4 degrees C. This residual binding was resistant to heparin competition, and strains lacking the first one-third of gIII were able to infect cells treated with heparinase as effectively as untreated cells. When we determined the penetration phenotypes for each strain, we found that gIII-mediated virus attachment was necessary for timely penetration of PK15 cells but remarkably was not required for efficient virus penetration of Vero cells. Moreover, wild-type PRV was actually prohibited from rapid penetration of Vero cells by a gIII-heparan sulfate interaction. Our results indicate that initial virus binding to heparan sulfate via glycoprotein gIII is not required for efficient PRV infection of all cell types and may in fact be detrimental in some instances.