Respiratory viruses: the viruses we detect in the human respiratory tract

A list of the viruses we can and do detect in the (mostly upper) human respiratory tract.

The standard testing panel/diagnostic menu comprises the influenza A and B viruses, parainfluenza viruses 1-3, respiratory syncytial virus, human metapneumovirus and the adenoviruses.

Some caveats:

• I make no claims to there being a link between cause-and-detect here. Because these viruses are mostly detected using polymerase chain reaction-based methods (PCR, RT-PCR mostly in a real-time format), virus may be detected before, during or after a symptomatic period. Virus may also be found when there are no symptoms at all; an asymptomatic infection. 
• In some instances the virus types included in this list are taken from a larger number of viruses grouped within a genus, for example the genus Enterovirus or the genus Mastadenovirus. My selection is based on our lab's multi-year, PCR-based studies since it can be hard to find decent data for a list like this using the scientific literature. Yes, I should write & publish faster, I know. 227 may well be an underestimate. For example viruses that may cause symptoms of gastroenteritis and central nervous system disease like the Saffold cardioviruses or the parechoviruses (HPeV) may begin their journey towards those diseases as a respiratory infections or involve the respiratory tract in the transmission process in some way.
• Other viruses including the newer polyomaviruses (WUPyV and KIPyV) are also detected in the airways but I have not listed them here.

In short, take this figure as a guide, but a pretty good one, of the approximately 227 viruses that comprise the pool from which we get infected each year. 

Also keep in mind that most if not all viruses seem capable of being associated with a spectrum of clinical disease; from no symptoms at all (yes, even influenza) to mild and short-lived acute respiratory diseases (common colds), severe colds, bronchiolitis, wheeze in the absence of asthma, cough, sore throat, middle ear infection, pneumonia, exacerbating asthma and chronic obstructive pulmonary disease and preceding bacterial secondary infections. These viruses often circulate in seasons some of which overlap and some of which are exclusive. They can also co-occur in the same person at the same time, regardless of season.

We can vaccinate against influenza virus; but it's worth remembering that a vaccine does not stop the virus entering our nose/throat. An effective immunization stops the severe form of the disease which can result from that infection. We may still test positive in a laboratory test after we have been vaccinated, but we are unlikely to be experiencing the full form of the disease (unless the vaccine did not "take" or there was a vaccine mismatch with the intended virus that we got infected by etc). 

Specific antiviral drugs among the respiratory viruses are rare; most infections are self-limiting (you recover from them yourself) and short-lived [1]. Influenza virus is again an exception in that we have useful drugs to apply after we get infected. A specific monoclonal antibody treatment for respiratory syncytial virus (RSV) has also been successfully used for some years [2] and the broader acting ribavirin is licensed for use in humans [3].

One question that is often posed when discussing testing for some/all of the list above is: why bother? If determining the virus won't lead to applying a treatment/cure and is of no use in prevention (if a patient is currently positive for that virus), then why expend time and money to sort it all out? Four very good reasons (from reference  #4) plus some of mine (with some overlap), are listed below. 

We test to...

• Provide information about disease occurrence [4]
• Monitor trends in disease prevalence [4]
• Guide public health efforts to prevent and control diseases [4]
• Allow timely detection of outbreaks to facilitate their management and control [4]
• Inform parents of ill children (clients in some areas of health-speak) who deserve to know what their children are infected with because from that information Doctors can provide a measure of the prognosis (how long they will be like this, how bad it might get etc)
• Inform the drug researchers about the importance of each virus
• Cohort patients with certain viral infections when they are in hospitals, to keep the same viruses together and away from other susceptible but uninfected patients
• Know when one seasonal virus is on the rise (RSV) or decline (rhinoviruses) because that may inform us about when another (influenza virus) is about to take off
• Know which virus is causing disease currently as that may affect hospital resource allocations; being able to identify a bigger influenza season may trigger increased purchasing of antivirals, masks, certain supportive care equipment or consumables or drive the updating of procedures and plans
• Look for sign that an outbreaks has occurred because of a change in the virus or its "behaviour". These changes can also trigger the State to make available new funding for targeted research

These are the viruses that we interact with all the time; some more often than others at certain times of the year. Over a few years, we're likely to have fended them all off, or succumbed to the signs and symptoms of an infection by them, at one time or another. 

Great to have an immune system huh? 

Reference..

1. Antiviral Drugs for Viruses Other Than Human Immunodeficiency Virus
   http://www.ncbi.nlm.nih.gov/pubmed/21964179
2. Monoclonal antibody for reducing the risk of respiratory syncytial virus infection in children.
   http://www.ncbi.nlm.nih.gov/pubmed/23633336
3. Respiratory syncytial virus infection in adult populations.http://www.ncbi.nlm.nih.gov/pubmed/22335500
4. Queensland Government's Queensland Health Notifiable Conditions Page
   http://www.health.qld.gov.au/ph/cdb/sru_data.asp

Middle East respiratory syndrome coronavirus (MERS-CoV): summing up 100 weeks

We stand at 182 cases with 78 deaths. The proportion of fatal cases (PFC) stands at 43%.

• Median age of all cases, including deaths, sits at 53-years (missing data on 13 cases); median age of fatal cases is 60-years
• 47% of all MERS cases with data are >55-years of age; 36% are >60-years
• 65% of cases are male (missing data on 18 cases)
• Underlying comorbidities feature in most severe disease MERS cases
• Approximately 18% of MERS-CoV cases are in healthcare workers; 2.7% of all fatal MERS cases are HCWs
• 81% of case are from the Kingdom of Saudi Arabia (KSA); the Arabian peninsula is the zone of case origin
• Reliable real-time reverse transcription polymerase chain reaction (RT-rtPCR) assays exist for detection, confirmation and genotyping
• Camels have been found on multiple occasions at multiple sites in the region to have antibodies to an antigenically similar virus to the MERS-CoV and nasal swabs have been found to be MERS-CoV RNA positive, as have humans in contact with the same camels (infection direction unknown). 
• Camel, goat, monkey, alpaca and human cells lines efficiently replicate MERS-CoV (multiple intermediate sources?)
• 1 diagnostic sequence of MERS-CoV RNA has been identified in a Taphozus perforatus bat (origin of animal other infections?)
• MERS-CoV uses DPP4 (CD26) as its receptor on host cells, a molecule found on some cell lines and epithelial cells of kidney, small intestine, liver and prostate. DPP4 has a standard role in hormone and chemokine activation
• No viable antiviral therapy or cocktail exists to treat infection. No vaccine exists.
• MERS-CoV replicates well in the lower respiratory tract of lab-infected macaques
• Person-to-person (p2p) transmission of MERS-CoV is sporadic
• Genetic variation among MERS-CoV genomes suggests multiple insertions into humans from the source(s)
• Fever, cough and shortness of breath in >70% of 47 cases in KSA; runny nose in 4%; abnormal chest X-Ray in 100%
• Sample often, sample lower respiratory tract to increase chance of successful RT-PCR result 
• Testing 5,065 hospitalized patients, healthcare worker contacts and family contacts found 2% (n=106) positivity over 12-months, in Saudi Arabia 
• MERS-CoV has circulated in KSA during several mass gatherings (2x Hajj pilgrimages and Umrah) providing ample opportunity for p2p transmission. There has been no evidence for an uptick in p2p transmission. We are nowhere near the verge of a pandemic.

New coronavirus genomes....not MERS yet

Unfortunately they aren't MERS-CoV genomes.

Nonetheless, a whole lot of new feline, porcine, murine, SARS, 229E and HKU1, genomes have been directly released from the J. Craig Venter Institute.

These now appear on GenBank with non-sequential accession numbers around the KF272920-KF530271. The sequences were produced using next generation sequencing technology.

Looks like the virome is in the sights of the big data guys.

Coronavirus family tree....

The phylogenetic tree below shows the relationships among the four genera of coronaviruses (CoVs); Genus Alphacoronavirus, Betacoronavirus, Deltacoronavirus, Gammacoronavirus. 

This tree is based on full length genomes (nucleotides; aligned using Geneious Pro; Neighbor-Joining tree built using Mega with 500 bootstraps). 

The MERS-CoV clade of betacoronaviruses is marked on the left and the endemic human CoVs are indicated with yellow triangles.

Infection Prevention and Control measures for MERS..mostly as per other ARIs

Thanks to Mike Coston for help and tips.

Cases are few and details are incomplete but the authors of an article in the recent MERS-centric issue of the EMRO Journal, recommend following the basic protocols you would to suppress spread of any virus capable of causing an acute respiratory infection (ARI) with a leaning towards those that worked well to interrupt hospital-based spread of severe acute respiratory syndrome (SARS) coronavirus.

Some key points from the paper, of highest relevance to our current knowledge of the  MERS-CoV are  listed include (not in specific order or priority):

1. Identify patients with ARIs and prevent them from transmisttign the agent to helathcare worklers and patients
2. Droplet and contact precautions for people with ARIs
3. Separate ARI patients by ≥1m from other patients and from HCWs
4. Use personal protective equipment (PPE) including eye protection, gloves, long-sleeved gowns and surgical mask/procedure mask/particulate respirator if aerosol-generating procedures are to be performed (tracheal intubation alone or with cardiopulmonary resuscitation or bronchoscopy being notable risks)

Mike Coston's description of the mask debate is very helpful for #4 above.

If a particular infectious diagnosis can be made, then patients with that diagnosis, say MERS-CoV,  can be cohorted - co-located to minimize spread to uninfected patients and maximise specialised care and efficient use of available resources.

Specifically, the article includes a list of SARS-like IPC precautions listed include which may be useful for known MERS-CoV infections. Many of these apply to ARIs due to endemic respiratory viruses and novel influenza viruses in general though:

• Good hand hygiene
• Use of PPE (gloves, gown, eye protection and medical masks for HCWs, caregivers and the patient if oputside their room
• Particulate respirator for aerosol generating procedures
• Separate, adequately ventilated room

While the above is written for dealing with infection in a healthcare setting, the WHO have also just released a rapid advice document for those caring for mildly ill MERS-CoV-infected people without underlying conditions, or those recently discharged from hospital. A mashup of 16 distinct points (read the document to see the full language and exceptions) home IPC are:

• Limit contact with the ill person - maintain distance (perhaps limit exposure time?). 
• Do not allow people at increased risk to care for the ill person
• Hand hygiene and respiratory hygiene are important as are appropriate (soap and water, bl;each as recommended) cleaning of all surfaces in contact with the person or their secretions - kitchen, bathroom, toilet, bedframe, bedside tables, furniture etc
• Discard contaminated tissues, masks etc
• Clean clothes
• Do not share eating utensils food or drionk, towels or bed linen
• Caregiver to wear a mask - discard after use and do not handle while in use
• Ventilate shared spaces

Close medical supervision is recommended for symptomatic or probable MERS cases and their contacts.

The WHO home care advice also notes lack of evidence for transmission of MERS (the disease) from asymptomatic, pre-symptomatic or early-symptomatic people. Thus quarantine or isolation of asymptomatic cases is currently unnecessary but possibly exposed people should monitor their health for 14-days.

Key documents and official websites to be familiar with:

• Infection prevention and control of epidemic- and pandemic-prone acute respiratory diseases in health care 
• http://www.who.int/csr/resources/publications/swineflu/WHO_CD_EPR_2007_6/en/index.html
• Coronavirus infections, World Health Organisation.
• http://www.who.int/csr/disease/coronavirus_infections/en/index.html
• Rapid advice note on home care for patients with Middle East respiratory syndrome coronavirus (MERS-CoV) infection presenting with mild symptoms and management of contacts (080813)
• http://www.who.int/csr/disease/coronavirus_infections/MERS_home_care.pdf

Tracking MERS-CoV through time: a spikey problem

This morning on Twitter, Helen Branswell (@HelenBranswell) asked this question, with a comment...

How is this so? 16 months after the 1st known #MERS cases, there are only 9 viral sequences in the public domain. So much for collaboration.
— Helen Branswell (@HelenBranswell) August 6, 2013

So I thought a little perspective might be nice. 

The SARS epidemic had its origins around Nov 16th 2002, although the major activity started in Feb of 2003. 

• 64 human SARS-CoV genomes had been produced by September 2003 ([UPDATED:] see Science paper). That is by 317-days later, or 10-months, 13-days (perhaps less given that the genome sequences were possibly sequenced well before the paper was submitted e.g. late phase genome s seem to have been submitted to GenBank by July 2003). 
• For MERS-CoV we currently have 9 genomes at 505-days (give or take), or 1-year, 4-months.

Not that anyone needs to be reminded, but 80% of MERS-CoV cases come from the Kingdom of Saudi Arabia. The world is relying on them, or their collaborators, to turn the nucleic acid extracts used to define these cases (PCR-POSs hopefully kept in a -80'C freezer), into templates for gene or genome sequencing.

I personally don't believe we need to have complete genomes right now in order to fulfil the fairly urgent public health need to monitor the virus and notice if it changes, or is changing, or is not changing. These changes tell us whether the virus is still adapting or has settled in - perhaps having done so prior to this outbreak's indicator, severe disease. 

What else to use to track adaptation?

Perhaps the 4,000nt Spike (S) gene, or some smaller but suitably variable portion of it, could be a target for sequencing? 

Zhang and colleagues have data showing it could be used to track an animal coronavirus's adaptation to humans, through its 3 pandemic phases. This was done using phylogeny (a way to show how one sequence relates to another through time and space) of nucleic acid sequences and alignments of the translated version of these sequences. All we need is primer sequences that could be used to reliably amplify the S gene of the MERS-CoV. If anyone has those already perhaps they could publish them...if they haven't already. A very brief look at the 9 MERS-CoV genomes already shows some variety. Perhaps unsurprisingly, there is very little change among the 4 Al-Ahsa genomes; their collection dates are separated in time by 17-days.

This shows a schematic of the aligned Spike genes. The black lines within the grey boxes represent nucleotides that differ from the consensus. More differences are obvious in the earlier sequences. The oldest MERS-CoV isolate is at the bottom, the most recent, at the top (detailed below). See the full version here at VDU.

Interestingly, the phylogeny of the complete Spike genes looks  similar to that of the complete MERS-CoV genomes. However  its doe snot place the isolates in order of increasing time to the extent that the full genomes do. I also looked at a 900bp fragment of the 3' of the Spike gene - easier to amplify but a very similar tree to that of the complete Spike.

All 9 complete MERS-CoV spike protein genes (nt). Alignment in Geneious Pro, tree in MEGA 5.10.
Full version will be here at VDU.

All 9 complete MERS-CoV genomes (nt). The arrow indicates moving forward in time; the oldest MERS-CoV isolates at the bottom, the most recent at the top. Alignment in Geneious Pro, tree in MEGA 5.10.
Full version will be here at VDU.

So where does that leave us?

Adaptive pressures on the SARS-CoV drove its genome towards settling down in the late stage of the 3-phase outbreak (defined by the Chinese SARS Molecular Epidemiology Consortium), with changes in the Spike gene occurring before that. Complete genomes are clearly the gold standard - so I dial down that personal belief from earlier.

The Spike gene still seems a useful target for MERS-CoV too, although not as accurate at plotting the time of virus isolation as complete MERS-CoV genomes were in my example above. Still, it, or some part of it, is still of use as an early-warning system to alert us to viral change and it will prove easier to amplify by smaller or less genomics-focussed laboratories. Something we need to consider in order to get some information, which is far better than none.

While we've seen predictive modelling for the age of MERS-CoV, we don't actually know when the virus came to be or when it started spilling over to humans. More full genome sequences would certainly help address that question. And finding its origin.

However, perhaps we should make the trade off and use the 3' end of the Spike gene now, in an effort to keep some sort of eye on how the MERS-CoV is travelling? Anyone else have a good region that fits the bill?

Time for the bat signal? The need for an animal model for Middle East respiratory syndrome coronavirus.

Elizabeth Devitt notes in Nature Medicine, that unlike its cousin, the severe acute respiratory syndrome coronavirus (SARS-CoV), some important features of MERS-CoV including its transmission, incubation period, and ability to spread systemically within the host, have not been able to be defined for the MERS-CoV using non-human models, because the virus does not like to infect the same animals. 

When the MERS-CoV infects a larger non-human animal, the rhesus macaque monkey, the disease it produces, while still defined as pneumonia and proving the casual link between MERS-CoV infection and disease, resolved faster and was not as severe as that in humans. These animals are also not easy to work with. I wonder if older monkeys with comorbidities have been looked at in particular? [UPDATE: The macaques above live to about 25-years]. It is this population in which MERS is most severe. Nonetheless, the monkey studies provide an excellent vehicle on which to test the usefulness of 2-drug an antiviral approach (Falzano et al, described earlier) that can clear MERS-CoV infections in vitro.

While cell/tissue culture methods using primary human airway cells have proven extremely useful for looking at cellular biology, antiviral effects, and immunobiology related to MERS-CoV infection, something with legs will be needed for future vaccines and to address the list above. We've seen many examples of how animal models massively improve our understanding of influenza virus pathogenesis, if an example is needed.

Also according to Devitt, Ian Lipkin is still wading through the data from samples collected from a range of animals that may be the natural hosts for the MERS-CoV in Saudi Arabia. Meanwhile, we recently learned of another CoV (PML/2011) found in the fecal pellets from a South African Neoromicia cf. zuluensis bat in 2011. PML/2011's nearest CoV relative was the MERS-CoV - its closest viral relative found to date (at least in the conserved RdRp region used by the authors).

This all begs the question, is there a bat animal model? CoVs, but also studies of other viruses like Hendra and Nipah, would benefit from a well-defined model based on these critters. That is, if they can be worked with and if they show any signs of these diseases - which they may not. My very quick skim of the literature found that bats used for neurological studies and for Hendra virus.