Saturday, March 5, 2016

NEJM pregnancy in Rio cohort: subtleties and design issues

The most recent 'big' paper examining the Zika-microcepahly connection is just out [1]. It is by far the most comprehensive study published so far,  and shows a large range of malformations and other adverse outcomes amongst a cohort of pregnant women in Rio de Janeiro.

I want to preface this by saying the authors are clear that this is a preliminary report; and moreover they are doing very difficult research under what I can only imagine as 'war footing.' That said, there are some fairly major caveats to these findings, and it's not a smoking gun yet. First and foremost, while the title is 'Zika Virus Infection in Pregnant Women in Rio de Janeiro — Preliminary Report', a far more accurate title would be 'Risk of any adverse fetal event in pregnant women who present with rash and agree to follow up.' Why does any of this matter?

It's not clear what the measured outcome is here (perhaps in the overall cohort itself it is), but any measured malformation appears to be the closest. This is a major issue- without a well-defined outcome to measure, any abnormality becomes a study event. Clearly some are major; however one of reported cases has 'mega cisterna magna,' which a quick lit search suggests while rare, is incidental with likely very limited impacts.

Differences in follow-up and lack of blinding (?)
This is the critical issue in any cohort- differential measurement of outcomes severely weakens the entire design. In this case, all 42 of the ZIKV (+) pregnancies have had extensive (presumably research hospital level) followup, while the ZIKV (-) women 'had undergone fetal ultrasonography as part of regular prenatal care, and the results were reported as normal.' 

And then: "Of the 70 remaining women with ZIKV infection, 42 (60%) had prenatal ultrasonographic examinations, with a total of 56 studies performed." Loss of the 40% of the positives is exceedingly worrisome- especially if the women who agreed to follow-up potentially felt changes in fetal movement etc. These two issues are major red flags (threats to validity), and means this is should be considered more of a case series report than a cohort (at this point).

A secondary issue is the potential lack of blinding. The authors don't say whether the ultrasonographers were blinded to the mother's ZIKV status at the readings, but given the design this would be difficult. Lack of blinding has the potential to lead to serious biases in any study, but as
specifically stated in the bible of epi: 'The skill of the examiner and the thoroughness of the examination can have a large effect on the apparent birth prevalence of a particular defect' [2].

It's difficult to do a power analysis since the outcome is unclear, but comparing the proportions of pregnancies with any adverse outcome, it's adequately power at 0.87 (which is presumably why they published these interim results). Interestingly, the difference in stillbirths (4 in 42 pregnancies vs. 0 in 16) is not significant (p=0.20).

Other issues
'Among ZIKV-positive women, more than half reported similar illnesses in other family members, and 21% reported that their partner had been ill.' However, it's clearly apparent that the presentation between ZIKV (+) and (-) patients in Table 1 is very similar. Moreover, dengue is currently co-circulating in Brazil and differential diagnosis by clinical symptoms is very error-prone [2a]. The difference between the dengue serology and the values in Table 1 for 'history of dengue' clearly reinforce this issue.

'Information in prenatal records regarding rubella, cytomegalovirus, and Venereal Disease Research Laboratory serologic testing was reviewed.' This statement is very, very different from any quantified serological measures; and record linkages are notoriously problematic in all health systems.

How representative is this cohort?
As only women presenting with rash were enrolled, how common is rash in ZIKV infections?
There are very limited data, but one study from Micronesia found 90% of 31 confirmed patients [3]. However, the Rio cohort (is or began as?) a dengue-focused one, in which rash is less common amongst adults cases; in adults one study in Nicaragua found 40% [4]; while one in Brazil (adults and children) found 61% had rash at presentation [5]. This suggests that there are a very large number of pregnant women would not be eligible for the study.

Final thoughts
I think it's clear from the very large number of adverse events that something serious is occurring in this cohort, but I don't think these results make a water-tight case for ZIKV in isolation.

Given the high dengue seropositivity, I'd be interested to see the serotype-specific dengue serology data.  Finally, how comparable are the 'rashy' periods relative to active viremia in ZIKV and dengue infections?

[2] Rothman KJ, Greenland S, Lash TL. Modern epidemiology. Third edt. Philadelphia: Lippincott Williams & Wilkins; 2008. pg. 637.
[2a] eg,


  1. And other issues. The paper does not make clear how alternative causes for the abnormalities were ruled out. In particular, the description of ocular abnormalities and others in two babies resembles strongly descriptions of congenital toxoplasmosis. Data on complete TORCHS survey should be made available for all mothers.

    It also appears that the sonograms were not validated in several cases where abnormalities were detected but the infant was born essentially normal.

    The study does not list the individual ages of the mothers, but the range includes 17 to 46. Were the mothers at each end of the range the mothers of the low birth weight babies? At least two mothers fall into the at-risk categories defines by age.

    What about the 12 mothers that were not positive for Zika? So, yeah, despite the headlines, this study left me wanting more data before making a leap.

  2. I think this is not good for the pregnant lady what things they have to bear already not more than that either those who cannot be a mother can test this once check this