Oral shedding of Marburg virus in experimentally infected Egyptian fruit bats (Rousettus aegyptiacus)

Abstract

Marburg virus (Marburg marburgvirus; MARV) causes sporadic outbreaks of Marburg hemorrhagic fever (MHF) in Africa. The Egyptian fruit bat (Rousettus aegyptiacus) has been identified as a natural reservoir based most-recently on the repeated isolation of MARV directly from bats caught at two locations in southwestern Uganda where miners and tourists separately contracted MHF from 2007-08. Despite learning much about the ecology of MARV through extensive field investigations, there remained unanswered questions such as determining the primary routes of virus shedding and the severity of disease, if any, caused by MARV in infected bats. To answer these questions and others, we experimentally infected captive-bred R. aegyptiacus with MARV under high (biosafety level 4) containment. These experiments have shown infection profiles consistent with R. aegyptiacus being a bona fide natural reservoir host for MARV and demonstrated routes of viral shedding capable of infecting humans and other animals.

Keywords: Experimental infection; Marburg virus; Rousettus aegyptiacus; shedding; transmission.

Figure 1

Viral RNA in blood of Egyptian fruit bats (Rousettus aegyptiacus) inoculated in the laboratory with Marburg virus. Viral RNA levels in blood (log₁₀ tissue culture infective dose 50 [TCID₅₀]/mL equivalent) days postinfection for each bat (n=27) are shown for three groups of bats.

Figure 2

Serology results for Egyptian fruit bats (Rousettus aegyptiacus) inoculated with Marburg virus. Results for anti-Marburg virus IgG measured by enzyme-linked immunosorbent assay are shown as adjusted sum optical densities (OD) by day postinoculation for all infected bats (n=27) at the time of euthanasia for each group.

Figure 3

Marburg virus RNA in skin, liver, spleen, and kidneys of Marburg virus-infected Egyptian fruit bats (Rousettus aegyptiacus). Mean viral RNA levels measured by quantitative reverse-transcriptase–PCR in skin at inoculation site (IS), liver, spleen, and kidneys (n=3; log₁₀ TCID₅₀/g equivalent) at the time of euthanasia (days). Each symbol (point) represents the mean viral load detected in that tissue in the three bats euthanized on that day. For reference, the mean viral loads detected in blood from those same bats on that day are shown (dashed black-yellow line). TCID=tissue culture infective dose.

Figure 4

Marburg virus RNA in salivary gland, bladder, and large intestine of Egyptian fruit bats (Rousettus aegyptiacus) inoculated with Marburg virus. Mean viral RNA levels were measured by quantitative reverse transcriptase PCR (log₁₀ TCID₅₀/g equivalent) at the time of euthanasia (days). Each symbol (point) represents the mean viral load detected in that tissue in the three bats euthanized on that day. For reference, the mean viral loads detected in blood from those same bats on that day are shown (dashed black-yellow line). TCID=tissue culture infective dose.

Figure 5

Marburg virus RNA in oral and rectal swabs from Egyptian fruit bats (Rousettus aegyptiacus) inoculated with Marburg virus. Viral levels from oral (square) and rectal (circle) swabs by day postinfection. Swab viral level values (log₁₀ TCID₅₀/mL equivalent) are equivalents of a stock tested (see Materials and Methods), not the actual amount of virus in saliva or feces, because the volume of absorbed oral secretion could not be calculated. Oral swabs from which Marburg virus could be isolated are depicted by an asterisks on days 8 (n=2) and 11 (n=1). The average viremia (Ave viremia swab pos) over time for bats with positive swabs (oral or rectal) is shown. TCID=tissue culture infective dose.

Figure 6

A large, densely populated colony of Egyptian fruit bats (Rousettus aegyptiacus) at Python Cave in Queen Elizabeth National Park, Uganda is shown. The large number of bats and their affinity for close contact is thought to perpetuate the circulation of Marburg virus infection within the colony. Marburg virus has been found in this population during every sampling effort (Amman et al. 2012).