Zika virus infection of the nervous system

FlavivirusEvidence is mounting that Zika virus is neurotropic (able to infect cells of the nervous system) and neurovirulent (causes disease of the nervous system) in humans.

The most recent evidence comes from a case report of an 81 year old French man who developed meninogoencephalitis 10 days after returning from a 4 week cruise to New Caledonia, Vanuatu, Solomon Islands, and New Zealand (meningoencephalitis is infection of the meninges – the membranes that cover the brain – and the brain). His symptoms included fever, coma, paralysis, and a transient rash. A PCR test revealed Zika virus genomes in the cerebrospinal fluid, and infectious virus was recovered after applying the CSF to Vero cells in culture.

A second case report concerns a 15 year old girl in Guadeloupe who developed left hemiparesis (weakness of one side of the body), left arm pain, frontal headache, and acute lower back pain. After admission she developed dysuria (difficulty urinating) that required catheterization. PCR revealed the presence of Zika virus genomes in her serum, urine, and cerebrospinal fluid; other bacterial and viral infections were ruled out.

Until very recently Zika virus was believed to cause a benign infection comprising rash, fever, joint pain, red eyes, and headache. There is now strong evidence that the virus can cause congential birth defects, and growing evidence that the virus is neurotropic and neurovirulent. Previously the entire Zika virus genome was recovered from brain tissue of an aborted fetus.

Zika virus is classified in the family Flaviviridae, and other members are known to be neurotropic, including West Nile virus, Japanese encephalitis virus, and tick-borne encephalitis virus. West Nile virus infection may lead to acute flaccid paralysis, meningitis, encephalitis, and ocular manifestations. Examination of additional cases of Zika virus infection will be needed to document the full spectrum of illness caused by this virus.

Update: Neurotropism of Zika virus is also indicated by the findings that the virus infects human cortical neural progenitors.

TWiV 331: Why is this outbreak different from all other outbreaks?

On episode #331 of the science show This Week in Virology, the TWiV team discusses the possible association of the respiratory pathogen enterovirus D68 with neurological disease.

You can find TWiV #331 at www.microbe.tv/twiv.

Enterovirus D68 infections in North America

EV-D68

Enterovirus D68 by Jason Roberts

An outbreak of respiratory disease caused by enterovirus D68 began in August of this year with clusters of cases in Missouri and Illinois. Since then 691 infections have been confirmed in 46 states in the US.

The number of confirmed infections is likely to increase in the coming weeks, as CDC has developed a more rapid diagnostic test. Previously it was necessary to amplify the viral genome by polymerase chain reaction, followed by nucleotide sequencing to determine the identity of the agent. The new test utilizes real time, reverse transcription PCR which is specific for the EV-D68 strains that have been circulating this summer.

Since its discovery in California in 1962, EV-D68 has been rarely reported in the United States (there were 26 isolations from 1970-2005). Beginning in 2009 it was more frequently linked to respiratory disease outbreaks in North America, Europe, Asia, and Africa. It seems likely that the virus was always circulating, but we never specifically looked for it.

The current EV-D68 outbreak is the largest ever reported in North America. Enterovirus infections are not rare – there are millions every year in the US – but why EV-D68 has been so frequently isolated this year is unknown. One possibility is that the CDC, after the initial outbreak in August 2014, began looking specifically for the virus.

Sequence analysis of the EV-D68 viral genomes indicate that 3 different strains are involved in the US outbreak. These viruses are related to EV-D68 strains that have previously circulated in the US, Europe, and Asia. The sequences are available at GenBank as follows: US-IL-14/18952, US-KY-14/18951, US-MO-14/18950, US-MO-14/18949, US-MO-14/18948, US-MO-14/18947, and US-MO-14/18946.

Most of the illness caused by EV-D68 in the US has been respiratory disease, mainly in children. Five of the 691 confirmed EV-D68 cases were fatal, but whether the virus was responsible is not known.

There have also been some cases of polio-like illness in children in several states associated with EV-D68. In Colorado the virus was isolated from four of 10 children with partial paralysis and limb weakness. Previously there had been one report of an association of EV-D68 with central nervous system disease. In this case viral nucleic acids were detected in cerebrospinal fluid. EV-D68 probably does not replicate in the human intestinal tract because the virus is inactivated by low pH. If the virus does enter the central nervous system, it may do so after first replicating in the respiratory tract, and then entering the bloodstream.

There are no vaccines or antivirals to prevent or treat EV-D68 infection. Most infections will resolve without intervention save for assistance with breathing. As the fall ends in North America, so will infections with this seasonal virus.

Oral polio vaccine-associated paralysis in a child despite previous immunization with inactivated virus

Poliovirus by Jason Roberts

Poliovirus by Jason Roberts

Vaccine-associated poliomyelitis caused by the oral poliovirus vaccine is rare, but its occurrence in a healthy, immunocompetent 6-month old child was highly unusual because the child had been previously immunized with two doses of the injected, inactivated poliovirus vaccine (IPV).

The three poliovirus vaccine strains developed by Albert Sabin (OPV, oral poliovirus vaccine) contain mutations which prevent them from causing paralytic disease. When the vaccine is ingested, the viruses replicate in the intestine, and immunity to infection develops. While replicating in the intestinal tract, the vaccine viruses undergo mutation, and OPV recipients excrete neurovirulent polioviruses. These so-called vaccine-derived polioviruses (VDPV) can cause poliomyelitis in the recipient of the vaccine or in a contact. During the years that the Sabin poliovirus vaccines were used in the US, cases of poliomyelitis caused by VDPV occurred at a rate of about 1 per 1.4 million vaccine doses, or 7-8 per year. Once the disease was eradicated from the US in 1979, the only cases of polio were caused by the Sabin vaccine.

To prevent vaccine-associated poliomyelitis, in 1997 the US switched to an immunization schedule consisting of two doses of IPV followed by one dose of OPV. The US then switched to using IPV exclusively in 2000. The child in this case essentially had a polio immunization course similar to that utilized in the US from 1997-2000: two doses of IPV, one dose of OPV. Why did the child develop poliomyelitis?

One clue comes from the fact that after the switch to an IPV-OPV schedule in 1997, there were still three cases of VAPP in 1998 and three in 1999. Another hint comes from a study of immune responses in children given multiple doses of IPV. Most of the children receiving two doses of IPV produced antibodies against types 1 and 2 poliovirus (92 and 94%), but only 74% of children produced antibodies against type 3 poliovirus.

The final piece of information needed to solve this puzzle is that the child in this case had vaccine-associated poliovirus caused by the type 3 strain, which was isolated from his feces.

Therefore, the child in this case most likely did not produce sufficient antibodies to type 3 poliovirus after receiving the two doses of IPV. As a consequence, when he was given OPV, he developed type 3 vaccine-associated poliomyelitis.

This case of VAPP could have been prevented: the child was born in Canada, and as customary in that country since 1995, he received two doses of IPV. At 5 months of age the child and his family visited China, where his parents decided to continue his immunizations according to the local schedule. China still uses OPV, so that is what the child received.

Polio-like paralysis in California

Enterovirus

Image credit: Jason Roberts

Recently a number of children in California have developed a poliomyelitis-like paralysis. The cause of this paralysis is not yet known, and information about the outbreak is scarce. Here is what we know so far:

  • At least 5, and perhaps as many as 20 children have suffered weakness or paralysis in one or more limbs. The median age of the patients is 12 years and the cases have been reported since 2012.
  • One group of 5 patients recently presented at the American Academy of Neurology Annual meeting developed full paralysis within 2 days, and have not recovered limb function in 6 months.
  • The cases are all located within a 100-mile radius.
  • A mild respiratory illness preceded paralysis in some of the children.
  • Enterovirus type 68 has been recovered from the stool of some of the patients.

I do not have any more information on this outbreak other than what I’ve obtained from ProMedMail. I have worked on enteroviruses, including poliovirus, for over 30 years, so I thought I might speculate on what might be transpiring.

What is a polio-like illness? Acute flaccid paralysis (AFP) is the term used to describe the sudden onset of weakness in limbs. AFP can have many etiologies, including viruses, bacteria, toxins, and systemic disease. It is used by the World Health Organization to maximize the ability to detect all cases of poliovirus. Confirmation that AFP is caused by poliovirus requires demonstration that the virus is present in the infected individual.

Is poliovirus the cause? I do not believe that poliovirus is causing the paralysis of children in California. I understand that they have all been immunized against poliovirus. In addition, should immunization have failed in any of these children, it seems unlikely that wild type polioviruses would be circulating in this area. Vaccine-derived polioviruses can cause paralysis but the US has not used this type of vaccine since 2000.

What might be causing the paralysis? AFP has both infectious and non-infectious etiologies. One possibility is that  a non-polio enterovirus is involved. Poliovirus is classified within the genus Enterovirus in the family Picornaviridae. Other enteroviruses besides poliovirus are known to cause paralytic disease, such as Coxsackieviruses, echoviruses, and many enteroviruses including types 70, 71, 89, 90, 91,96, 99, 102, and 114.

Most enterovirus infections can be associated with different clinical syndromes besides paralysis (such as respiratory disease), and therefore diagnosis is difficult. Stool is generally the most sensitive specimen for establishing an enterovirus infection. However, the virus may no longer be present at onset of symptoms. Polio is much easier to diagnose in individuals with AFP from whom virus can be identified: paralysis is the main serious symptom caused by infection. However note that 99 out of 100 poliovirus infections are asymptomatic or present with undifferentiated viral illness. The incidence of paralytic disease caused by other enteroviruses is even lower – for example 1 in 10,000 EV71 infections are paralytic. If all of the 20 California cases are caused by enteroviruses, this means that there have been many more infections without symptoms.

In one study of non-polio AFP in India, no virus could be isolated in 70% of the cases. Enterovirus 71 was the single most prevalent serotype associated with non-polio AFP. This virus currently causes large outbreaks of hand, foot, and mouth disease throughout Asia, with many fatalities and cases of acute flaccid paralysis. EV71 is known to circulate within the United States.

What about enterovirus 68? It has been reported that EV68 has been isolated from some of the paralyzed children. This isolation does not mean that the virus has caused the paralysis. Enterovirus infections of the respiratory and gastrointestinal tracts are very common and often do not result in any signs of disease. Random samplings of healthy individuals frequently demonstrate substantial rates of enterovirus infections.

Enterovirus type 68 was first isolated in California from an individual with respiratory illness. The virus is known to cause clusters of acute respiratory disease, and there is at least one report of its association with central nervous system disease. I believe it is an unlikely cause of the paralytic cases in California based solely on the past history of the virus and the fact that other enteroviruses are more likely to cause paralysis. It is not clear to me why enterovirus 68 would evolve to become substantially more neurotropic: entering the central nervous system is a dead end because the infection cannot be transmitted to a new host.

All of the above is pure speculation based on very little data. The paralysis might not even be caused by an infection. At this point a great deal of basic epidemiology needs to be done to solve the problem – if indeed it can be solved at all. Based on its population, California would be expected to have about 75 cases of acute flaccid paralysis each year of various etiologies, suggesting that the current number of cases is not unusual or unexpected.

Update: N. Gopal Raj wrote a story last year about acute flaccid paralysis in India, which has the highest rate of non-polio AFP in the world, with 60,000 cases reported in 2011.

Poliovirus silently (and not so silently) spreads

Poliovirus by Jason RobertsPoliovirus has been found in sewage in Israel. The virus detected is not vaccine-derived poliovirus; it is wild-type 1 poliovirus, the strain that occurs naturally in the wild and which the World Health Organization is trying very hard to eradicate from the planet.

As part of the global effort to eradicate poliovirus, environmental samples from many countries are routinely examined for the presence of the virus. Wild type poliovirus was detected in 30 sewage sample from 10 different sites, collected from 3 February to 30 June 2013 in Israel. No cases of paralytic disease have been detected in that country. This is not a surprising finding because only roughly one in 100 individuals infected with poliovirus develop paralysis.

During poliovirus infection, the virus replicates in the gastrointestinal tract and is shed in the feces. Most of the poliovirus-positive sewage samples were from southern Israel. Finding the virus at multiple sites suggests that the virus has been in the population for an extended period of time and that multiple individuals are shedding virus.

Why is wild type virus circulating in Israel? Poliovirus vaccine coverage in Israel is high but it is never 100%, and non-immunized individuals are hosts for the virus. Another explanation is related the the use of inactivated poliovirus vaccine (IPV) in Israel. Immunization with IPV, which is administered intramuscularly, does not protect the alimentary tract from infection. Therefore poliovirus can replicate in the intestine of immunized individuals, but once the virus reaches the blood its spread is blocked by anti-viral antibodies.

An interesting question is whether there is wild type poliovirus in the United States. I suspect that if we looked for poliovirus in our sewage, we would find it. However we no longer carry out surveillance of sewage for poliovirus and therefore we do not know if it is present.

Genetic analysis of wild type poliovirus from Israel suggests that it is related to the strain found in December 2012 in sewers in Cairo, Egypt. That virus in turn is closely related to virus from Pakistan, one of three countries from which wild type poliovirus has not been eradicated (the others are Nigeria and Afghanistan).

There is an ongoing outbreak of poliomyelitis in the Horn of Africa, with 65 cases in Somalia and 8 cases in Kenya. The virus causing that outbreak came from Nigeria. Somalia and Kenya had been free of polio since 2007 and 2011, respectively.

WHO has concluded that the risk of further international spread of wild type poliovirus from Israel as moderate to high. As long as there are individuals who are not immune, there will be a risk of poliovirus infection, in part due to the silent infections caused by the virus.

Microbes After Hours: West Nile virus

This discussion of West Nile virus was recorded at the headquarters of the American Society for Microbiology during a “Microbes After Hours” event on May 6, 2013. The speakers are Dr. Lyle Petersen Lyle R. Petersen, M.D., M.P.H., director of the Division of Vector-Borne Diseases at CDC, and Dr. Roberta DeBiasi, MD, FIDSA, Associate Professor of Pediatrics at George Washington University School of Medicine, Acting Chief and Attending Physician in the Division of Pediatric Infectious Diseases at Children’s National Medical Center, and investigator at Children’s Research Institute in the Center for Translational Science in Washington, D.C.

MWV Episode 70 – Microbes After Hours – West Nile Virus from microbeworld on Vimeo.

Coxsackie NY and the virus named after it

coxsackie-nyRecently while driving north on the New York State Thruway I passed the exit for the town of Coxsackie, NY (population 8,884). I grabbed my camera and photographed the exit sign, and reminded myself to write about the virus named after this small town.

In the summer of 1947 there were several small outbreaks of poliomyelitis in upstate New York. Gilbert Dalldorf, the director of the Wadsworth Laboratory in Albany, NY, and his associate Grace M. Sickles investigated this outbreak. In particular they sought polioviruses that could replicate in mice. This search was motivated by the fact that research on poliovirus required the use of monkeys which were extremely expensive. Dalldorf had attended the Fourth International Congress for Microbiology in 1947 where he heard that very young mice – suckling mice – could readily be infected with Theiler’s virus.

Dalldorf and Sickles made fecal suspensions from two children suspected of having poliomyelitis, and inoculated these into adult and suckling mice. Only the suckling mice (1 – 7 days old) developed paralysis; animals more than one week old were resistant to infection. The damage responsible for limb paralysis was widespread lesions in skeletal muscles, not in the central nervous system as occurs with poliovirus. Further study revealed that the viruses could be distinguished serologically from poliovirus.

Not only had Dalldorf and Sickles identified the first members of a very large group of human viruses, but they also introduced and popularized a new and inexpensive animal into the virology laboratory – the suckling mouse. In 1949 Dalldorf suggested that the new viruses be called Coxsackie viruses, because the first recognized human cases were residents of that New York village. This unique name is of native North American origin.

Over ten years later the importance of this work was recognized by Dr. Max Finland of Boston City Hospital:

The isolation by Dalldorf and Sickles of viruses which produced paralysis with destructive lesions of muscle in sucking mice and hamsters, from the stools of two children with signs of paralytic poliomyelitis was an achievement that may rank in importance with Landsteiner and Popper’s production of human poliomyelitis in monkeys.

In subsequent years many different Coxsackieviruses were isolated that cause a variety of clinical syndromes. Today at least 30 serotypes of Coxsackieviruses are classified in the enterovirus genus of the Picornaviridae. The viruses are classified into groups A or B depending upon the pathological effect in suckling mice.

Not every locale is pleased to have a virus named after it. In May 1993, an outbreak of an unexplained pulmonary illness occurred in the southwestern United States, in an area shared by Arizona, New Mexico, Colorado and Utah called “The Four Corners.” Muerto Canyon was proposed as the name for the etiologic agent of the disease, because the virus was first isolated from a rodent near the canyon. However after residents objected, the name Sin nombre virus was given to the agent of hantavirus pulmonary syndrome.

Dalldorf G, & Sickles GM (1948). An Unidentified, Filtrable Agent Isolated From the Feces of Children With Paralysis. Science (New York, N.Y.), 108 (2794), 61-62 PMID: 17777513

Chronology of an acute infection

poliomyelitisThe clinical signs of an acute viral infection – characterized by rapid onset of disease with a short but sometimes severe course – are often obvious. These include coughing, sneezing, rashes and poxes, or headache. What takes place within the body while these symptoms develop?

Examination of the time course of an acute infection such as poliomyelitis reveals significant differences in the onset of clinical symptoms and appearance of virus or antiviral antibodies. The data in the figure (click for a larger view) come from studies of the disease in humans, which was common in the US until the early 1960s and therefore was extensively described in many medical texts. Within 2 days of exposure, virus levels begin to rise in feces and throat secretions, but no signs of disease are evident until day 7, when headache, sore throat, and nausea develop. The appearance of these non-specific symptoms (e.g. not unique to infection with any particular viral infection) correspond with peak levels of virus in the blood. Both subside by day 10. Antiviral antibodies begin to accumulate in the blood at the end of the viremic stage, around days 9-10.

Ninety-nine percent of all poliovirus infections end at this stage. In the remainder, the virus enters the central nervous system where it multiplies rapidly and reaches peak levels at day 13-14. At this time, headache and nausea return, accompanied by stiffness, pain, and in some cases, paralysis. During this central nervous system phase of the infection, levels of virus in throat secretions decline but remain high in the feces.

This typical pattern of acute infection has several practical consequences. Shedding of virus soon after exposure, in the pre-symptomatic period, facilitates the spread of infection. These individuals do not develop serious illness and remain in contact with others. By the time symptoms appear – either the mild symptoms or paralysis – virus in the blood is already declining. An antiviral compound given at this time would have no effect on the outcome of disease. Antibodies peak late in infection – too late to have a significant impact on the course of disease. This finding provided the early clues that lymphocytes, not antibodies, play a major role in clearing many acute viral infections.

If this hand-drawn schematic looks dated, it is because it was taken from a 1959 book entitled “Viral and Rickettsial Infections of Man”, edited by Rivers and Horsfall. It contains chapters written by the virology luminaries of the time, including George Hirst, Igor Tamm, A.D. Hershey, John Enders, Edwin Lennette, Albert Sabin, Jonas Salk, and David Bodian. Much of the information is dated, but some of it – including the course of poliomyelitis – established the basic principles of viral pathogenesis that will never be obsolete.

Poliovirus

Poliovirus by Jason Roberts

Poliovirus is the etiologic agent of the paralytic disease known as poliomyelitis. It’s also the virus I’ve worked on for most of my career. The World Health Organization is in the midst of a massive effort to eradicate the disease, an undertaking that has encountered a number of obstacles. In coming posts I’d like to discuss the polio eradication effort, but first we need some background on poliovirus.

Poliovirus is a member of a family of viruses called the Picornaviridae. Viruses are classified into families, and then genera, based on many criteria, including physical and biological properties. The Picornaviridae includes many other human pathogens, such as Coxsackieviruses, echoviruses, enteroviruses, hepatitis A virus, and rhinoviruses (which cause the common cold). The International Committee for the Taxonomy of Viruses is responsible for virus classification and maintains the Universal Virus Database, where you can find information about most known viruses.

Poliovirus is a rather small and simple virus. It is composed of a shell, or capsid, made of protein, as shown.

The poliovirus capsid is about 30 nanometers in diameter. Within the capsid is the information to make new virus particles – a single molecule of ribonucleic acid, or RNA. In the image, part of the capsid has been cut away to reveal the viral RNA. When the virus infects a cell, the RNA genome enters the cell and programs it to make new virus particles. These virus particles are released from the cell and go on to infect new cells.

In humans, poliovirus is ingested, and replicates in cells of the gastrointestinal tract. Newly synthesized virus particles are released into the intestine and shed in the feces. Transmission of poliovirus to another human occurs through contact with virus-containing feces or contaminated water. After multiplying in the gastrointestinal tract, poliovirus may enter the spinal cord and brain. Destruction of motor neurons by the virus leads to limb paralysis.

Poliomyelitis became a common disease at the turn of the 20th century. Two vaccines were developed in the 1950s that can effectively prevent the disease – inactivated poliovaccine (IPV, developed by Jonas Salk) and live, oral poliovaccine (OPV, developed by Albert Sabin). In 1988 the World Health Organization announced that it would eradicate poliomyelitis from the globe by the year 2000. We’ll discuss that goal, and obstacles that might prevent it from being attained, in subsequent posts.