Detection and identification of Toscana and other phleboviruses
by RT-nested-PCR assays with degenerated primers.
Novel unclassified phleboviruses
(e.g., Fermo, Granada, Punique, and Massilia viruses), as well as recently discovered flaviviruses in sand flies, underline the importance of these neglected vectors of arboviral diseases (12-16).
For specimens yielding IgG-positive results, we performed a more specific plaque reduction neutralization test (PRNT), which can differentiate between related phleboviruses
in the United States, by using Vero E6 cells to confirm the presence of virus-specific neutralizing antibodies and a 90% plaque reduction criteria (8,9).
Two sand fly-borne phleboviruses
in the Old World were historically associated with cases of sandfly fever: Sicilian virus and Naples virus (7).
(family Bunyaviridae, genus Phlebovirus) are negative-sense tri-segmented RNA viruses for which mosquitoes, ticks, and sand flies are vectors.
The Gorgas Memorial Institute in Panama City tested dengue -negative samples from this outbreak for alphaviruses, flaviviruses, and phleboviruses
and detected Punta Toro virus species complex (PTVs) in some samples.
With the recent discovery of SFTSV, HRTV, and other novel phleboviral agents in recent years (12-14), it is apparent that tickborne phleboviruses
have been largely underrecognized as potential human disease agents.
Orthobunyaviruses and phleboviruses
are transmitted to animals and humans by blood-feeding arthropods such as mosquitoes, sandflies, and ticks (1,2).
These 2 viruses are closely related to 2 tickborne phleboviruses
: severe fever with thrombocytopenia syndrome virus, isolated in China (2), and Heartland virus, isolated in the United States (3).
Comparative neutralization tests with related viruses were not performed, because we had previously found that murine antiserum developed against the other known phleboviruses
in the United States--Sunday Canyon virus (7), Rio Grande virus (8), and Lone Star virus (9)--had no appreciable neutralizing activity against HRTV (Table 2).
Results of routine microbiologic examinations for bacteria by culture and antigen detection were negative in all cases, as were serologic and genetic tests for hantaviruses, phleboviruses
, and Rickettsiales bacteria, performed as described (10).