influenza virusHuman influenza viruses replicate almost exclusively in the respiratory tract, yet infected individuals may also have gastrointestinal symptoms such as vomiting and diarrhea. In mice, intestinal injury occurs in the absence of viral replication, and is a consequence of viral depletion of the gut microbiota.

Intranasal inoculation of mice with the PR8 strain of influenza virus leads to injury of both the lung and the intestinal tract, the latter accompanied by mild diarrhea. While influenza virus clearly replicates in the lung of infected mice, no replication was observed in the intestinal tract. Therefore injury of the gut takes place in the absence of viral replication.

Replication of influenza virus in the lung of mice was associated with alteration in the populations of bacteria in the intestine. The numbers of segmented filamentous bacteria (SFB) and Lactobacillus/Lactococcus decreased, while numbers of Enterobacteriaceae increased, including E. coli. Depletion of gut bacteria by antibiotic treatment had no effect on virus-induced lung injury, but protected the intestine from damage. Transferring Enterobacteriaceae from virus-infected mice to uninfected animals lead to intestinal injury, as did inoculating mice intragastrically with E. coli.

To understand why influenza virus infection in the lung can alter the gut microbiota, the authors examined immune cells in the gut. They found that Mice lacking the cytokine IL-17A, which is produced by Th17 helper T cells, did not develop intestinal injury after influenza virus infection. However these animals did develop lung injury.

Th17 cells are a type of helper T cells (others include Th1 and Th2 helper T cells) that are important for microbial defenses at epithelial barriers. They achieve this function in part by producing cytokines, including IL-17A. Th17 cells appear to play a role in intestinal injury caused by influenza virus infection of the lung. The number of Th17 cells in the intestine of mice increased after influenza virus infection, but not in the liver or kidney. In addition, giving mice antibody to IL-17A reduced intestinal injury.

There is a relationship between the intestinal microbiome and Th17 cells. In mice treated with antibiotics, there was no increase in the number of Th17 cells in the intestine following influenza virus infection. When gut bacteria from influenza virus-infected mice were transferred into uninfected animals, IL-17A levels increased. This effect was not observed if recipient animals were treated with antibiotics.

A key question is how influenza virus infection in the lung affects the gut microbiota. The chemokine CCL25, produced by intestinal epithelial cells, attracts lymphocytes from the lung to the gut. Production of CCL25 in the intestine increased in influenza virus infected mice, and treating mice with an antibody to this cytokine reduced intestinal injury and blocked the changes in the gut microbiome.

The helper T lymphocytes that are recruited to the intestine by the CCL25 chemokine produce the chemokine receptor called CCR9. These CCR9 positive Th cells increased in number in the lung and intestine of influenza virus infected mice. When helper T cells from virus infected mice were transferred into uninfected animals, they homed to the lung; after virus infection, they were also found in the intestine.

How do CCR9 positive Th cells from the lung influence the gut microbiota? The culprit appears to be interferon gamma, produced by the lung derived Th cells. In mice lacking interferon gamma, virus infection leads to reduced intestinal injury and normal levels of IL-17A. The lung derived CCR9 positive Th cells are responsible for increased numbers of Th17 cells in the gut through the cytokine IL-15.

These results show that influenza virus infection of the lung leads to production of CCR9 positive Th cells, which migrate to the gut. These cells produce interferon gamma, which alters the gut microbiome. Numbers of Th17 cells in the gut increase, leading to intestinal injury. The altered gut microbiome also stimulates IL-15 production which in turn increases Th17 cell numbers.

It has been proposed that all mucosal surfaces are linked by a common, interconnected mucosal immune system. The results presented in this study are consistent with communication between the lung and gut mucosa. Other examples of a common mucosal immune system include the prevention of asthma in mice by the bacterium Helicobacter pylori in the stomach, and vaginal protection against herpes simplex virus type 2 infection conferred by intransal immunization.

Do these results explain the gastrointestinal symptoms that may accompany influenza in humans? The answer is not clear, because influenza PR8 infection of mice is a highly artificial model of infection. It should be possible to sample human intestinal contents and determine if alterations observed in mice in the gut microbiome, Th17 cells, and interferon gamma production are also observed during influenza infection of the lung.

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CaribouRecovering viral genomes from ancient specimens can provide information about viral evolution, but not many old nucleic acids have been identified. A study of 700 year old caribou feces reveals that viruses can be protected for long periods of time – under the right conditions.

The oldest virus recovered so far is the giant Pithovirus sibericum, which was isolated from 30,000 year old Siberian permafrost. Other attempts have yielded fragments of viral genomes. It was possible to reconstruct the 1918 influenza virus from small RNAs recovered from formalin fixed and frozen human tissues. However this feat was not achieved for viral RNA in 140,000 year old Greenland ice cores, 900 year old North African barley grains, or 7,000 year old Black Sea Sediments.

Caribou feces have been frozen for the past 5,000 years in ice patches in the Selwyn Mountains of the Canadian Northwest territories. To determine if viruses could be recovered from this material, the frozen feces were thawed, resuspended in buffer, filtered, and treated with nucleases to destroy any nucleic acids not contained within a viral capsid. Sequence analysis of the remaining nucleic acids revealed two different viruses.

Ancient caribou feces associated virus (aCFV) has a single stranded, circular DNA genome distantly related to plant-infecting geminiviruses and gemycircularviruses. The entire 2.2 kb genome of aCFV was amplified from the caribou feces specimen. This reconstructed viral DNA replicated upon introduction into tobacco plant leaves.

Sequences of an RNA virus distantly related to picornaviruses of insects (such as Drosophila C virus) were also identified in the caribou feces. These viral genomes exceed 7.4 kb, but it was only possible to recover a 1.8 kb fragment of this virus, ancient Northwest Territories cripavirus (aNCV).

Neither virus was isolated from contemporary Caribou feces collected from an animal living in the same region. The authors also went to great pains to demonstrate that the two 700 year old viral genomes were not contaminants. The isolation was repeated in a different laboratory, and was not to be a consequence of contamination from any laboratory reagent or apparatus used for purification of nucleic acids.

It is not likely that aCFV or aNCV infected a caribou 700 years ago. The viruses were probably acquired when a caribou ingested plant material infected with the plant virus; perhaps insects harboring aNCV were also present on these plants. The exact hosts for both viruses are unknown.

The fact that two relatively large fragments of viral DNA and RNA were identified suggests that intact capsids were present in the caribou feces. Their preservation is probably a consequence of the low temperature of the arctic ice, and the stable icosahedral capsids characteristic of members of geminiviruses, gemycircularviruses, and cripaviruses.

We already know that viruses have been around for a long time, more than hundreds of millions of years, so what is the value of this work? Studying ancient viruses can provide insight into viral diversity and evolution. However, the value of two viral genome sequences is limited, and additional work should be done to acquire additional specimens spanning a long period of time. Similar sampling of other environments would also be desirable, but it is unlikely that large fragments of viral genomes can be recovered from specimens that are not frozen. And as the ice caps melt away, we will lose our ability to decode this important viral record.

Image credit

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On episode #314 of the science show This Week in Virology, Vincent travels to Albert Einstein College of Medicine where he speaks with Kartik, Ganjam, and Margaret about their work on Ebolavirus entry, a tumor suppressor that binds the HIV-1 integrase, and the entry of togaviruses and flaviviruses into cells.

You can find TWiV #314 at www.twiv.tv.

On episode #313 of the science show This Week in Virology, Vincent, Alan, and Rich discuss how norovirus, an enteric virus, can replace the functions of the gut microbiome.

You can find TWiV #313 at www.twiv.tv.

antibodies bound to Ebola virus GPZMapp, a mixture of three antibodies against Ebola virus, became a household name after it was used to treat two Americans who were infected while working in Liberia. The structure of these antibodies bound to the Ebola virus glycoprotein suggest how they inhibit infection and ways to improve ZMapp.

The three monoclonal antibodies that comprise ZMapp (called c13C6, c2G4, and c4G7) were produced by immunizing mice with a recombinant vesicular stomatitis virus in which the glycoprotein was replaced with that from Ebola virus. Antibodies that bound the viral glycoprotein and protected mice from infection were identified, and three were made to resemble human antibodies and produced in tobacco plants. Ecco Zmapp!

Embedded in the membrane of the filamentous Ebola virus particle are many copies of the Filovirusglycoprotein, seen as club-shaped spikes in the image to the right (image credit: ViralZone). The viral glycoprotein is essential for entry of the virus into cells. The antibodies in ZMapp are directed against the viral glycoprotein.

To determine how the antibodies bind the virus particle, they were individually mixed with purified Ebola virus glycoprotein, and the structures were determined by electron microscopy and image reconstruction. The results, shown in the illustration, indicate precisely where each antibody binds to the Ebola virus glycoprotein. The individual antibodies colored red (c2G4), yellow (c4G7), and purple (c13C6) are bound to a single Ebola virus glycoprotein in white, with the viral membrane below (Image credit).

The structures reveal that c13C6 (purple) binds at the tip of the viral glycoprotein, perpendicular to the plane of the viral membrane. The other two antibodies (red, yellow) bind at the base of the viral glycoprotein. Their binding sites overlap but are not identical (the Ebola virus glycoprotein is a trimer, and in the image, the yellow and red antibodies are shown binding to different subunits for clarity). Two other antibodies that block Ebola virus infection also bind at the base of the glycoprotein.

Antibody c13C6, which binds to the tip of the viral glycoprotein, does not neutralize viral infectivity. Nevertheless, it can protect animals from Ebola virus infection. This observation suggests that the c13C6 antibody may work in concert with complement, a collection of serum proteins, to block virus infection. It is not known why c13C6 antibody is non-neutralizing, but one possibility is that it binds to a part of the viral glycoprotein that is removed by an endosomal protease, cathepsin, before receptor binding in late endosomes.

Antibodies c2G4 and c4G7, which bind to the membrane-proximal part of the viral glycoprotein, neutralize viral infectivity. How they do so is not known, but one possibility is that they prevent structural changes of the viral protein that are essential for the fusion with the endosomal membrane, a process that delivers the viral nucleic acid into the cell cytoplasm.

These studies reveal two general areas of the Ebola virus glycoprotein that are important targets for antibodies that protect animals from Ebola virus infection. Those directed at the base of the glycoprotein neutralize infectivity while those that bind the tip do not. This information can now be used to isolate additional antibodies that bind either site. These can be used in animal protection studies to design mixtures that are even more potent than ZMapp.

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TWiV 312: She sells B cells

23 November 2014

On episode #312 of the science show This Week in Virology, the TWiVbolans discuss the finding that human noroviruses, major causes of gastroenteritis, can for the first time be propagated in B cell cultures, with the help of enteric bacteria.

You can find TWiV #312 at www.twiv.tv.

A virus that melts sea stars

17 November 2014

Sun flower sea starSea stars are lovely marine invertebrates with a round central body connected to multiple radiating legs (photo credit). In the past year millions of sea stars in the west coast waters of North America have melted into piles of slime and ossicles. Sea star associated densovirus might be the cause of this lethal disease.

Sea star wasting disease (SSWD) is characterized by lesions, limb curling and deflation, and death as the animals rapidly degrade or ‘melt’. The current outbreak began in June 2013 and has killed sea stars from Baja California, Mexico, to Southern Alaska. SSWD might be the biggest marine wildlife epizootic ever observed.

Evidence that SSWD is caused by a virus came from experiments in which extracts of diseased sea stars were passed through a filter with pore sizes small enough to allow passage of viruses but not bacteria or other microbes. When injected into healthy sea stars, these filtrates induced sea star wasting disease. Extracts of diseased sea stars collected in Vancouver, CA contained 25 nanometer virus particles, as determined by electron microscopy.

Nucleic acid sequencing was to identify the viral agent of SSWD. Virus particles were purified from diseased animals, and both DNA and RNA was extracted. Analysis of the nucleotide sequences revealed the presence of giant DNA viruses such as mimiviruses and phycodnaviruses (link to algal virus paper), and among RNA viruses, retroviruses, dicistroviruses, and parvoviruses. With few exceptions, all samples containing parvoviruses were from symptomatic asteroids, and so the authors decided to pursue the study of this virus.

Analysis of the DNA sequence data revealed the presence of a densovirus (a parvovirus) related to viruses found in Hawaiian sea urchins. The authors called this virus sea star-associated densovirus, SSaDV. Like other members of the parvovirus family, these are small (25 nm diameter), naked icosahedral viruses with a ~6 kb single stranded DNA genome. When sea stars were infected in the laboratory with filtrates from diseased animals, virus loads, determined by PCR, increased with time together with disease progression. Field surveys revealed that the virus was more abundant in diseased than in healthy sea stars. The virus was found in marine sediments, plankton, and sea urchins. Viral nucleic acid was also found in sea stars preserved in museums since 1942.

While retrovirus sequences were found in sea star tissues, the authors believe that such ‘Retroviral annotations are likely spurious because they were detected in DNA libraries (and have RNA as nucleic acids).’ The detection of retrovirus sequences in sea stars is not at all spurious! DNA copies of retrovirus genomes are produced during infection and integrated into the host genome, explaining why these sequences were detected in DNA libraries. Their absence in RNA libraries means that virus particles are not produced, as is the case in many other organisms that contain endogenous retroviruses.

The evidence that SSaDV causes sea star wasting disease is strong but not yet complete. The crucial experiment that remains to be done is to isolate infectious virus in cell culture, inoculate it into sea stars, and show that it causes wasting disease.

While the authors’ work reveals that sea star wasting disease virus has been present on the North American Pacific Coast for over 70 years, the disease is not always observed. The current outbreak has been ongoing since just June 2013. Perhaps other environmental conditions have lead to the increased susceptibility of sea stars to disease. It is important to determine if a human activity is involved in precipitating the disease, so that we can prevent future loss of sea stars. The authors suggest one possibility, that increased populations of adult sea stars in small bays and inlets have produced increased virus levels which lead to more infections. It will also be important to determine if the virus has undergone any changes in transmissibility or virulence.

Sea star associated densovirus was also found in non-asteroids, including ophiuroids (brittle stars and basket stars) and echinoids (sea urchins and sand dollars). We need to know the host range of this virus, how it is transmitted, and whether it can cause disease in other species. A troubling scenario is that the recent amplification of the virus in sea stars could lead to infection, and perhaps death of, many other marine species.

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TWiV 311: Bulldogs go viral

17 November 2014

On episode #311 of the science show This Week in Virology, Vincent visits the University of Georgia where he speaks with Zhen Fu and Biao He about their work on rabies virus and paramyxoviruses.

You can find TWiV #311, audio and video versions, at www.twiv.tv.

Phycodnaviridae virionMany well-known human viruses, including poliovirus, rabies virus, West Nile virus, can infect cells of the nervous system, leading to alterations in the function of that organ. Could a virus that infects algae also cause human neurological alterations?

Chloroviruses are large DNA-containing viruses that infect unicellular algae called zoochlorellae (pictured: image credit, ViralZone). Unexpectedly, chlorovirus DNA sequences were found in the oropharynx of 40 of 92 individuals (43.5%) who had no known physical or psychiatric illness. The clinical specimens had been obtained as part of a study of cognitive function, and it was possible to determine that presence of chlorovirus DNA was associated with a slight but statistically significant decreased performance in tests for visual motor speed, delayed memory, and attention.

When mice were fed chlorovirus-infected algae, they showed decreased performance in tests of cognitive function, such as recognition memory and sensory-motor gating. Some of these animals developed antibodies against the virus, suggesting that viral replication took place. Furthermore, feeding of chlorovirus to mice was associated with changes in gene expression in the hippocampus, the part of the brain essential for learning, memory, and behavior.

It is not known if the chlorovirus replicates in humans or in mice; only viral nucleic acids were detected. No mention is made of attempts to isolate infectious chloroviruses from humans or mice. The amount of chlorovirus in the oropharynx is not known. However the results of sequence analysis, in which low numbers of sequences were found in each person suggest very low numbers of genomes. Of course, it is possible that virus replication took place some time ago, and its effects linger after replication has subsided.

Chloroviruses are commonly found in inland waters, and the subjects could have acquired the virus via inhalation or drinking contaminated water. It is entirely possible that the virus does not replicate in humans, but is present in the oropharynx as a common environmental contaminant. Many plant and insect virus sequences can be isolated from the human intestinal tract as a consequence of the food we ingest, but there is no evidence that they can replicate at that site. Consequently, chlorovirus might not have any role in the reduced cognitive functions observed in this study. It is possible that exposure to another factor together with chloroviruses, such as heavy metals, is responsible for the observed cognitive differences.

The suggestion that a virus infection might cause subtle cognitive defects is not outlandish. For example, lymphocytic choriomeningitis virus infects rodents congenitally or immediately after birth and establishes a persistent infection of virtually all tissues. These mice show no outward signs of illness, but careful study of infected animals reveals that they are less ‘smart’ than their uninfected peers.

The results are intriguing and warrant more study, including a determination of whether an infectious chlorovirus can be isolated from humans, whether this virus can replicate in human cells in culture, and how they differ from environmental isolates. It would also be important to determine if antibodies to chloroviruses are present in humans, and if they are associated with any diseases. It is too early to conclude that a virus of algae causes altered human neurological functions.

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transmission-image-ebolaAfter the governors of New York and New Jersey decided that health workers who have returned from the Ebola virus outbreak in West Africa should be subject to a 21-day quarantine, two Nobel laureates entered the fray. Bruce Beutler feels that the quarantine is the right thing to do, while Peter Doherty says it’s wrong. Which laureate is right?

The key issue in this debate is the assumption that someone who has been infected with Ebola virus, and does not display symptoms, is not contagious. Beutler doesn’t believe that there is enough evidence for this assumption: “People may have said that without symptoms you can’t transmit Ebola. I’m not sure about that being 100 percent true. There’s a lot of variation with viruses.” (source: NJ.com). On the other hand, Doherty says “the evidence-based consensus among the professionals seems to be that this is not necessary” (source: NJ.com).

As I’ve written before, our knowledge of the non-contagious nature of Ebola virus infected patients is based on experience with previous outbreaks.  To make sure I wasn’t missing anything that Beutler had noted, I examined the published summaries of the previous outbreaks of Ebolaviruses. There have been 24 outbreaks caused by these viruses, summarized in a table by CDC.  In poring over the outbreak data, I looked for information on how the virus is transmitted. These data are typically obtained by interviewing Ebola virus patients or their families, and constructing chains of transmission – who infected who, and how the infection was transmitted. Because it is not always possible to determine exactly how transmission took place, the interviewers attempt to determine what kinds of activities are most associated with acquiring infection. These activities are called ‘risk factors’.

I was particularly interested in determining if the following was a risk factor in any of the 23 previous outbreaks: some type of contact with a healthy person who subsequently developed Ebola hemorrhagic fever (EHF). I didn’t find any. All transmissions that could be assessed involved an obviously sick individual, and never from anyone who was healthy.

The first two recognized outbreaks of infection were in 1976 in Zaire (Ebola virus) and in Sudan (Sudan virus). The former comprised 318 cases, and infection was spread by close contact with patients and by using contaminated needles. The Sudan oubreak of 284 cases started in workers of a cotton factory, and was amplified by transmission in a hospital. Transmission required close contact with an acute case, usually the act of nursing a patient.

Sudan, 1979 (Sudan virus, 34 cases) The index case worked in a textile factory. Virus was then introduced into 4 families from the local hospital. Every case, except the index patient, could be traced to a human source of infection. Twenty-nine cases occurred in chains of secondary spread in 4 families, all from direct physical contact. No illness was observed among persons who were exposed to cases in confined spaces, but without physical contact.

Gabon, 1994 (Ebola virus, 52 cases) This outbreak began in three different gold mining camps. A second wave of infections outside the camp was initiated by contact with a hospitalized patient from first wave.

DRC, 1995 (Ebola virus, 315 cases) The index case was a charcoal worker, with no known contacts with other EHF patients, probably infected in a charcoal pit or on his farm. He infected 3 members of his family and 10 secondary cases occurred among his family members. Some of these individuals introduced virus into a maternity hospital, and then a general hospital. Prior contact with a suspected patient was reported by 93.5% (159/170) of case-patients for whom data were available. Eleven patients reported receiving an injection within 3 weeks of symptom onset. No role for airborne transmission was observed in human-to-human transmission. Sleeping in the same room with patient not a risk factor.

Gabon, 1996 (Ebola virus, 37 cases) A chimpanzee found dead in the forest was eaten by people hunting for food. Eighteen people who were involved in butchering the animal became ill. Ten other cases occured in their family members. Of 190 individuals who contacted sick people, none developed EHF.

Gabon, 1996-97 (Ebola virus, 60 cases) The index case was a hunter who lived in a forest camp. A dead chimpanzee found in the forest at the time was determined to be infected with Ebola virus. Disease was spread by close contact with infected persons. This outbreak was not well documented.

South Africa, 1996 (Ebola virus, 2 cases) A nurse in South Africa developed EHF; she had been previously exposed to the blood of a sick doctor brought from Gabon. This doctor had treated EV infected patients.

Uganda, 2000-2001 (Sudan virus, 425 cases) The three most important risk factors identified: attending funerals of EHF patients; interfamilial or nosocomial transmission.  The source of infection of primary patients could not be identified. The transmission risk factors among 83 contacts were determined. The most important risk factor was direct repeated contact with a sick person’s body fluids, as occurs during care. Risk of infection was higher during late stage of disease. Simple physical contact with sick person was neither necessary nor sufficient for developing EHF. Transmission through contaminated fomites is possible – such as sleeping on the same mat, or sharing meals with a sick person. Having washed the clothing of a sick person was not a risk factor. There was no evidence for airborne transmission during this outbreak – sleeping in the same hut as an infected person was not a risk factor.

Gabon, 2001-2002 and Republic of Congo (Ebola virus, 65 and 67 cases, respectively)  All but two cases were linked to recognized chains of transmission. There were six different introductions of virus into humans during the outbreak, each related to a hunting episode. An unusually high number of animals were found dead in the rainforest at this time, mainly gorillas, chimps, and monkeys. Ebola virus was detected in the carcass of a gorilla that had been butchered by one of the index cases shortly before onset of illness. The vast majority of secondary cases were related to community based transmission, and health care workers caring for infected patients included.

Republic of Congo 2002-03 (Ebola virus, 143 cases)  Risk factors in this outbreak were determined to be participating in the funeral/burial ritual of a patient, and providing nursing care to an individual with EHF. Quote from the paper: ‘There is no contamination by air or just handshake’.

Republic of Congo 2003 (Ebola virus, 35 cases)  Risk factors in this outbreak were determined to be participating in the funeral/burial ritual of a patient, and providing nursing care to an individual with EHF.

Sudan, 2004 (Sudan virus, 17 cases)  Risk factors in this outbreak were determined to be participating in the funeral/burial ritual of a patient, and providing nursing care to an individual with EHF. There was one introduction of virus into the index case-patient, who had been hunting baboons in forest, and was in contact with fresh monkey meat 5 days before onset of symptoms. All cases were epidemiologically linked, with 4 generations of transmission observed.

Democratic Republic of Congo, 2007 (Ebola virus, 264 cases) Index case-patient bought freshly killed fruit bats from hunters for consumption. She infected 11 family members who provided care, who in turn passed infection to others in a chain of transmission.

Uganda 2007-08 (Bundibugyo virus, 149 cases) Identified a chain of transmission from a prominent individual in the community to 27 others; they had contact with him before or after his death.

I did not find risk factor analysis for outbreaks in Democratic Republic of Congo, 2008-2009 (Ebola virus, 32 cases) ; Uganda, 2012 (Sudan virus, 11 cases; and Democratic Republic of Congo, 2012 (Bundibugyo virus, 36 cases). Transmission studies on the recent Ebola virus outbreak in West Africa has not yet been published. However the recent outbreak of Ebola virus in Nigeria was traced from an index case who traveled there from Liberia, where he had cared for a sibling who died of EHD. Upon receiving medical attention in Nigeria, he infected health care workers and the disease spread further. Few of the 898 contacts linked to this index case became infected.

From this analysis it is clear that contact with an asymptomatic individual, who subsequently developed EHV, is not a risk factor for disease. All risk factors involve close contact with sick individuals, such as would occur in a health care facility, among family members caring for a sick individual, and during funeral proceedings.

I am aware that not all of the cases in these outbreaks could be traced and assigned risk factors for infection. One could argue that among these cases, some might have been acquired by contact with an individual in the incubation period. However it makes no sense that these individuals would have been missed in all of the outbreaks. The more likely conclusion is that during the incubation period, patients infected with Ebola virus are not contagious.

There is one more piece of information to support the conclusion that there is no transmission during the Ebola virus incubation period. During the Ebola virus outbreak of 2000-2001 in Uganda, blood samples were collected from health care workers who became symptomatic with EHF; of these, 27 died and 18 survived. Ebola virus RNA could be detected in the blood on the first day that symptoms were observed, at levels just above the detection threshold (illustrated; image credit). Each day thereafter, the virus load increased, much more substantially in patients with a fatal outcome. While no viral loads were determined before the onset of symptoms, it is reasonable to predict that viral loads would be even lower. This conclusion is consistent with the observation that infected patients do not transmit infection before the onset of symptoms, probably because the levels of virus that they produce are too low. The results of this study are also consistent with observations from some outbreaks that the risk of transmission is increased later in disease, when more virus is present in the blood.

These data lead to the conclusion that Ebola virus transmission does not occur during the incubation period. Therefore it is not necessary to quarantine those who might have had contact with Ebola virus infected individuals. Daily reporting of temperature and other vital signs should be sufficient to allow detection of the onset of disease.

In other words, I disagree with Dr. Beutler’s statement that there are no good data supporting lack of Ebola virus transmission during the incubation period. The fact that Dr. Doherty disagrees with Dr. Beutler just goes to show that having a Nobel Prize doesn’t mean you are always right.

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