MERS-CoV detections over the past 6-weeks: 38 cases, 13 deaths.

Click to enlarge.
Laboratory confirmed MERS-CoV
cases (including deaths; green) and
deaths (red) by day (bottom, x-axis), per
week. Number of cases on
the left hand (y) axis peak at 8/week

Updating the Middle East respiratory syndrome coronavirus (MERS-CoV) graphs from just over 2-weeks ago and adding in recent weeks, we see how the cases have been accruing. 

Last week was a big week; 42% of cases from the past 6-weeks occurred then.

There are some differences in some charts when comparing to the earlier post with some of these; put that down to updated dates due to extra data being released and some cases being reported the week(s) after they occurred. I'll keep updating this figure. Those changes may keep happening.

3 new MERS-CoV cases, 2 deaths tally at 135

Three new cases were reported from the (Arabic, not English yet) Kingdom of Saudi Arabia (KSA) Ministry of Health (MOH) website and details of the entire list can be found on FluTracker's website of confirmed cases. The tally sits at 135 confirmed cases. There may, or may not, be a chunk of other cases ("probables") but they have not been confirmed.

Today's cases all had comorbidities and were (FT-FluTracker's case numbering system):

• FT#133: 75-year female (75F), contact of a case in Medina, death
• FT#134: 35M contact (Mkhalt=male contact; hat-tip Helen Branswell) in intensive care
• FT#135: 83M, Riyadh, death

Declan Butler has a nice article preempting the MERS-CoV birthday "celebrations" that is good summary, as was Helen Branswell's article earlier in the week, These show just how obvious the poor progress on some key aspects of MERS-CoV research have been. Is the slow progress due to a system that does not utilize its own Universities and research infrastructure  does the KSA not have such capacity? I don't know. But the lion's share of the MERS-CoV work does seem to await off-shore advice and direction. Principal research direction does not seem to originate from within the hot-zone of viral emergence and this may be a key difference between the rate of early progress in understanding the emergence of the MERS-CoV compared to that of SARS-CoV, H1N1pmd09, H7N9 or many other viral outbreaks.

Thanks to @HelenBranswell for input on sex of 83[M]

17 new MERS-CoV sequences bind perfectly to frontline screening PCR assay for MERS...

Click to enlarge. The primers/probe are depicted as grey boxes.
If mismatches existed they would show up as horizontal black
lines within the grey box. No mismatches are evident.
The GenBank accession numbers are
shown on the left of this alignment of 17 MERS-CoV
sequences.

Only 17 of the 45 sequences seem to include the region covered by the upE laboratory assay I just posted about in the WHO laboratory testing update but of those, the forward and reverse oligonucleotide primers and the probe all bind without any mismatch.

While that may sound like an obvious statement considering that these viruses were probably detected using that assay it isn't.

The new MERS-CoV sequences were determined using using unbiased 2nd generation high-throughput sequencing technologies that did not rely on these primers to generate them. So we are now able to check and see if there are any nucleotide changes at the target sites for the primers and probe, that would reduce the efficiency the assay.

There are no such oligonucleotide mismatches between primer and viral genes among those 17 sequences, which is good news for that assay's continued usefulness.

Built to last eh?

MERS-CoV WHO testing guidelines: September update

The World Health Organization has updates its laboratory testing guidelines. They can be accessed here.

Some key points:

• Lower respiratory tract samples are recommended since there are data to support higher viral loads (better detectability using PCR) from these samples
• The upE real-time reverse transcriptase polymerase chain reaction (RT-rtPCR) assay of Corman et al is considered highly sensitive and is recommended for screening
• The ORF1a screening assay is of equal sensitivity and the ORF1b slightly less sensitive than upE
• RdRp and N gene assay are suitable for genotyping
• Antibody testing assays are described and some seem to be very sensitive. none have been validated against a large panel of known MERS-CoV POS sera and result interpretation is still without consensus
• Laboratories should notify their local public health authorities as soon as they receive a specimen for MERS-CoV testing and all results should also be passed along to these authorities.
• Member States are asked to immediately notify WHO of initially POS laboratory results even before confirmatory testing is complete.

Pretty clear and straightforward really.

MERS-CoV genomes on GenBank...[UPDATE]

Click to enlarge. A scale schematic of the first
MERS-CoV genome, EMC/2012.

45 subgenomic (the smallest is 361 nucleotides [nt]) to full length genome (only 13; >30,000nt) sequences of the MERS-CoV have been released onto GenBank ahead of a Lancet Infectious Diseases paper arriving in days. The GenBank accession numbers range from KF600612 - KF600656 and repsenst human cases form 2012 & 2013. 

Usually (and hopefully soon), I would get the entire batch using a search of KF600612:KF600656[ACCN] at http://www.ncbi.nlm.nih.gov/nuccore. They seem to have to be downloaded manually for now.

The list of Middle East respiratory syndrome coronavirus sequences with their date of collection (DOC) includes:

• KF600612, Riyadh_1_2012 ; 30,063nt
  DOC: 23-Oct-2012 
• KF600613, Riyadh_3_2013; 30,064nt
  DOC: 05-Feb-2013
• KF600614, Al-Hasa_10e_2013; 2,151nt
  DOC:02-May-2013
• KF600615, Al-Hasa_14a_2013; 6,673nt
  DOC: 08-May-2013
• KF600616, Al-Hasa_13b_2013; 3,787nt
  DOC: 07-May-2013
• KF600617, Al-Hasa_22a_2013; 2,102nt
  DOC: 09-May-2013

Most are from cases in Al-Ahsa (adding to the previous 4), as well as from Riyadh, Buraidah and Hafr-Al-Batin

In case you read otherwise, there are no obvious signs of recombination among these viral sequences.

Thanks to @Sarah_E_Smith1 for announcing location on GenBank ahead of the Lancet paper.

The Rubik's cube of influenza A genes spins up a new lineage of H7N7

Click to enlarge. A (very) summary view of the latest
contributing influenza viruses that precedes the emergence of
human infections with influenza A(H7N9) virus
in south-east China in 2013.

Lam and a global host of collaborators, writing in Nature on the 21st of August, have identified a previously unknown influenza A(H7N7) virus line circulating in chickens. The authors indicate that more influenza viruses lurk among poultry and that active surveillance is needed. This report comes from testing 1,341 pairs of oropharyngeal and cloacal swabs and 1,006 faecal and waters samples from live bird markets (LBMs) in Wenzhou and Rizhao of Zhejiang province, as well as Shenzhen from Guangdong province.


In a complex alphabet soup of influenza A virus findings, the authors, sequenced 34 H7N7, 4 H7N9 and 19 H9N2 egg-isolated viruses but also found H7N2 and H7N3 in ducks. Animals tested were chickens, ducks, geese, pigeons  partridges and quail.

The authors note that rather than wild birds from Europe and Korea, the neuraminidase (NA or N) gene segment from H7N9 is more temporally related to those from H11N9 and H2N9 found in wild birds (wild water fowl, Northern shoveller and common teal) in Hong Kong during 2010-11 with links to domestic ducks in China prior to the H7N9 outbreak this year. Overall, domestic ducks proved to be an important mixing pot between wild birds and chickens.

And it's not just H7N9; the H7N7 found in chickens reminds us that the colours on the cubes are many and are in constant motion. These virus may become/may already be enzootic (endemic in non-humans) and so continuing exposures to live poultry in markets and backyards remains a continuing source of risk for new zoonoses.

Age and sex morbidity and mortality from avian influenza A(H7N9) virus

Click to enlarge. The majority of cases of H7N9 that occurred
worldwide earlier in 2013. Taken from Virology Down Under's
H7N9 page.

In a study co-written by yours truly using a lot of data collected for Virology Down Under, Dr Joseph Dudley and I have just described, in the Journal of Clinical Virology, the age-specific and sex-specific morbidity and mortality from the avian influenza A(H7N9) virus outbreak earlier in the year.

We sought to highlight differences between H7N9 and another zoonotic influenza A virus, H5N1. The distribution of age and sex is notably different between cases of each virus in more distant countries (Saudi Arabia vs Egypt) as it is within the same country (see Cowling et al reference in the article's discussion). Such differences and patterns may be instructive for identifying specific risk factors for an outbreak and also serve to highlight that there are differences between outbreaks which, on the surface, might be expected to have very similar courses. 

Intriguingly, there were marked similarities between H7N9 and Middle East respiratory syndrome coronavirus age and sex case distribution.

We also published the term created here on VDU, the Proportion of Fatal Cases (PFC). A percentage defined as the number of currently known fatalities divided by the number of total lab-confirmed cases including fatalities, regardless of whether they are inpatients (hospitalized) or outpatients. It was created to avoid the need for a gauge of recovered cases (released from hospital) which is linked with use of the term Case Fatality Ratio.

Most MERS may not have met a camel, but index cases may have

Donald G McNeil Jr., writing in the New York Times  a few days ago, posited the idea that camel contact, while not at all widespread among the majority of Middle East respiratory syndrome (MERS) cases, may play a role in the first cases that sparked clusters of infection; the so-called index cases.

These sorts of patterns, whether for camels, bats, baboons or cats, are key to understanding infection acquisition. 

We all look forward to reading more detailed local analyses of the sleuthing that seeks to define first contact with our MERS-CoV adversary...sometime in the future. 

Infection Scene Investigation (ISI): Kingdom of Saudi Arabia?

First noticed on Twitter from @crof.

A stroll down Polymerase Chain Reaction lane

Dr Kary Mullis, 1993 Nobel prize winner and the co-creator of the Polymerase Chain Reaction (PCR; I have aliottle on PCR over at PCR Down Under), walks down memory lane describing the process of those very early discoveries while working at Cetus Corporation.

You can check out the whole seminar here.

Some key points were:

• PCR gets 41 million hits today
• Created PCR in the Spring of 1983 as away of increasing the demand for oligonucleotides ("oligos")
• Ron Cook developed the first automated oligonucleotide synthesizer machine that could create oligonucleotides in hours instead of weeks
• Before PCR, the only way to examine a specific bit of human DNA was to clone it and such genetic modification of bacteria was a cause of concern in the early days
• Speeding up the process of identifying genetic mutations was a driving factor behind the development of PCR
• The target produced by exponential amplification resulting from binding and extension of two oligonucleotide "primers" binding to a template, would overwhelm any non-specific product
• The original manuscript was knocked back by Nature and Science and ended up in Methods in Enzymology, after other's by Saiki et al, Saiki et al, and another by Saiki et al, a Symposium by Mullis also Saiki from Cetus were published. Lesson here for us all - write faster!

The Mullis story is an colourful one - as you can get a taste of from Wikipedia, his personal website, Cracked, virusmyth (describing his prior comments on HIV), this excerpt from Making PCR: A Story of Biotechnology and his book Dancing naked in the mind field.

Thanks to John F Mackay (no relation except by All Black) of DNature, an Antipodean PCR historian (nerd), for pointing out the presentation and for discussion.

Middle East respiratory syndrome coronavirus cases amongst healthcare workers [UPDATED]

Click on image to enlarge. (a) the proportion of
MERS-CoV positives HCWs who have died (red) vs.
survived (blue), (b) the proportion of fatal cases (PFC; red) 
of MERS-CoV worldwide vs. the proportion of 
surviving cases (PSC; blue) (c) breakdown HCWs
as a proportion of all MERS-CoV cases (blue), HCW deaths 
as a proportion of all MERS cases (green) and HCW deaths
as a proportion of all MERS-CoV deaths.

With a lot of help from FluTrackers, the 2 of us have synced our lists to account for all the healthcare workers (HCWs) for which public data are available, that have been confirmed as MERS-CoV positive.

Some charts then.

We can see that HCWs make up approximately a sixth (18.2%; n=24) of all MERS-CoV cases.

Fatal infections in HCWs account for 2.3% (3/132) of all MERS-CoV cases (including living and deceased cases) and 5.4% of all MERS-CoV deaths worldwide are among HCWs (3/56). This last figure indicates that HCWs are at a relatively reduced risk of death from MERS-CoV infection when compared to other groups that have been infected.

For example:
NB: I have death data for 56 cases; age data for 125/132 cases; sex data for 120/132 cases); 27 comorbidities listed [underestimate]

• 63% of MERS-CoV deaths have occurred among those older than 55-years (50% of deaths among those >60-years; 38% among those >65-years; 59% among those <65-years)
• 46% of MERS-CoV deaths have occurred among males older than 55-years (38% among those>69-years; 30% among those >65-years; 45% among those <65-years)
• 82% of deaths )n=46) and 83% of cases have occurred in the Kingdom of Saudi Arabia
• 48% of MERS-CoV deaths occurred among those with comorbidities [this is an underestimate]So in the lower proportion of deaths represented by HCWs, while horrible in any proportion, may provide evidence to support that MERS-CoV is still not transmitting well, even in close quarters.

It may also mean that attending HCWs are adhering to good infection control and prevention practices. But it coudl just mean that we do not have data on all HCW infections/death and there are greater numbers of cases.

Finally, and perhaps most importantly, we should remember that HCWs may have some degree of resistance to disease caused by some viruses because of their constant exposure to patients with all manner of airway infections.

If HCWs may not show the same proportion of illness, but still become infected, they can act to spread cases among their contacts - patients and visitors. This was evident in the severe acute respiratory syndrome (SARS) outbreak where HCWs accounted for a fifth of all confirmed cases.¹

In other words, even a few cases in HCWs could have major implications for nosocomial outbreaks. If an emerging virus, such as the MERS-CoV, is being frequently detected in association with healthcare settings, that scenario may already be happening.

Some literature..

• Severe acute respiratory syndrome (SARS) and healthcare workers.
• Lack of SARS Transmission among Healthcare Workers, United States
• SARS in Healthcare Facilities, Toronto and Taiwan