O1010 - Active Immunization Against Tetanus

This study looked at antibody levels in subjects immunized with various kinds of tetanus vaccine. So nothing that new. But the discussion was pretty interesting:

"The first dose of alum precipitated toxoid affects the antitoxin producing cells of the body in such a manner that following a second injection there is a rather prompt increase in the antitoxin content of the blood. A certain period of time must elapse between these two injections before this release of antitoxin into the blood stream takes place...The effect of the first dose of alum toxoid is rather unique in that it cannot be replaced by the natural occurrence of tetanus and recovery from it. We have had occasion to confirm Cowles' finding that a patient who had recovered from tetanus 10 years before still required two doses of alum toxoid before any demonstrable antitoxin appeared in the blood. Following the second dose, a few days elapse before a protective titer develops. Hence, if an injury occurs during the interval between the two injections of alum toxoid and for a week or two following the second injection, passive immunization may be necessary in order to get full protection against tetanus."

"The 'repeat' or stimulating dose of plain or alum precipitated toxoid produces within one week a remarkable increase in the antitoxin content of the blood. In most cases about five days elapse after the injection of the 'repeat' dose before the antitoxin titer is brought up or above the protective level. It is stated that the period of incubation of tetanus in man is usually from six to 14 days and is directly proportional to the amount of toxin and the severity of the disease. With a short period of incubation, six days or less, the disease is almost invariably fatal. Whether the mobilization of antitoxin that occurs after the injection of the 'repeat' dose is fast enough to prevent all cases of acute tetanus is not definitely known, but more than likely it will be able to do so, since our minimum protective value of 0.1 unit is a conservative one."

"From our studies to date, it would appear that active immunization against tetanus by means of the injection of two doses of alum precipitated toxoid followed by a 'repeat' injection upon the occurrence of an injury, will prove to be of value in military practice, in civil life where frequent injuries make the repeated injection of tetanus antitoxin impossible, and in the handling of allergic individuals who are sensitive to horse serum."

So that's interesting, and explains how things pretty much still work today, except that Td is given upon injury only if one hasn't received it for a number years.

Reference:

Gold, H. Active Immunization Against Tetanus. Ann Intern Med 13, 768–782 (1939).

100 - A Comprehensive Study of Influenza in a Rural Community

Ok, here's another study that doesn't look at vaccines specifically, but rather the background of the disease they prevent, and how our bodies respond to it. In this case, influenza.

The researchers followed a small community of people in New York over the course of a flu season, looking at their antibody levels. The tests they used were neutralization (mixing serum with virus sample and seeing if the serum neutralized the virus so it didn't make mice sick) and complement fixation, though of the two tests the former seemed more useful.

In the period before the flu season, teenagers had the highest titers in the neutralization test, which were lower at younger and older ages. For complement fixation, the highest levels were in those 40 to 60 years old. Both tests observed a decrease above 60, in the elderly.

Then in the flu season, they made sure to distinguish clinical flu from the common cold or other flu-like illnesses: the flu was defined by a fever over 99 degrees F, headache, pains, general malaise, and respiratory symptoms. About 85% of the disease in the community was actually colds, which peaked in December, while the flu didn't peak until February. About 62% of the flu tested was found to be Influenza A, the rest other kinds.

So regarding the neutralization test results, having a cold didn't increase one's titers at all, but having the flu did (makes sense). Some had an increase without observing symptoms, possibly subclinical infections, too mild to be noticed. There was a definite correlation between having lower antibodies before the flu season and being likely to get the flu during the season, so titers correlated with protection. Though even those with the highest titers occasionally got sick.

One year after the season, some people's titers had returned to the same level they had been before the season (even if they got the flu), some stayed the same, the rest fell partway back. The average was 5 times more 1 year after than before.

So overall, it seems like higher titers correlate with more protection, but there's no really safe level that's 100% protective. And correlation isn't causation, so it's not clear the antibodies are doing the protecting, though neutralizing the virus seems like a good sign. Also I'm not sure how specific these tests are for specific antibodies, so they could show high levels of antibodies that aren't actually that useful, or something like that. Needs further study.

Reference:

Rickard, E. R., Lennette, E. H. & Horsfall, F. L., Jr. A Comprehensive Study of Influenza in a Rural Community. Public Health Reports (1896-1970) 55, 2146–2167 (1940).

099 - Secondary Familial Attack Rates from Pertussis in Vaccinated and Unvaccinated Children

This study is a follow-up analysis of previous results by Kendrick and Eldering of a clinical trial in Grand Rapids, Michigan, of a whole-cell pertussis vaccine.

Rather than the whole study population, this study focused only on 165 families that were definitely exposed to pertussis from one of their own members. The other members' immunity status was known from history of vaccination or the disease itself.

What they saw was that, of all 78 vaccinated subjects, only 28 got whooping cough (36%). All these were under age 7. In contrast, of those without history of vaccination or disease, 79% got sick. Of those unvaccinated under age 7, 92% were attacked (the rate decreased with increasing age of subject). 36% vs. 92% for those most vulnerable? Not bad.

Most of the primary cases, that brought the disease to a family, were in the older category, confirming that older children often bring it to their younger siblings. Interestingly, with the subjects who had the disease before, 5 of the 6 that got sick were over 7 years old, so it seems there's a period after which the immunity is not so great, even after natural infection. The study was set up to analyze that in depth though.

Also noteworthy is that, of 172 primary cases that brought the disease to the families, 157 were unvaccinated (91%), 12 were vaccinated (7%), and 3 had the disease previously. There's some herd immunity in action, perhaps, but it's hard to tell.

This wasn't a very rigorous study (lack of blinding or placebo), but considering that an intimate exposure, such as in the same household, is probably the most difficult to have immunity against, there seems to be some effect. And apparently whooping cough is so contagious that 80-90% get sick when exposed to it this way, if not immune somehow.

References:

Kendrick, P. L. Secondary Familial Attack Rates from Pertussis in Vaccinated and Unvaccinated Children. Am. J. Epidemiol. 32-SectionA, 89–91 (1940).

O984 - Tetanus Immunization

The author of this study (Philip B. Cowles) seemed skeptical of some claims made about the tetanus toxoid vaccine, so set out to test them.¹

The first was the claim that 0.01 unit of antitoxin per milliliter of serum (the level of antibodies against the tetanus toxin in the blood) was enough to protect against the symptoms of tetanus. He used guinea pigs and mice to figure out how much antitoxin was necessary to protect. He seemed to find that 0.1 unit per mL was usually enough, though not always. As another author summarizes,

"This author felt that though 0.10 unit is not sufficiently great to protect all animals against a maximum infection, it is probably much larger than is necessary to care for many infections resulting from wounds judged to be too slight for surgical attention, and probably large enough to care for the majority of injuries that receive surgical treatment."²

The second question was how long does it take for a booster shot to raise the level of antitoxin. The common practice was to give a booster when a person presented with a wound of some sort that might expose them to tetanus infection, thinking that the vaccine would raise their antibodies enough to protect. But if that doesn't happen quickly enough, it wouldn't be very useful.

Cowles tested a number of people before and after a booster. Most had about 0.02 unit or less before, and it took 4-5 days to rise up to 0.1 or higher. Is this fast enough? Tetanus seems to develop in 7-10 days, but it's possible the toxin has started working before then, before symptoms show up. So more study would be needed to say a booster is definitely helpful, but it's pretty definitely better than nothing if an antitoxin treatment is unavailable.

As a final addendum, the author mentions the question of whether having tetanus makes one immune to further tetanus. There had been reports of people have tetanus repeatedly, so obviously there isn't always natural immunity, but Cowles describes a man who had recovered from a mild tetanus attack at some point, but didn't seem to have any antitoxin in his blood. So it seems like the tetanus toxin is toxic at lower concentrations than it is antigenic (capable of producing antibodies and immunity), so it's rare to become immune naturally without dying or something. Small sample size though.

References:

1. Cowles, P. B. Tetanus Immunization. Yale J Biol Med 9, 409–416 (1937).
2. Gold, H. Active Immunization Against Tetanus. Ann Surg 114, 1060–1068 (1941).

097 - An Experiment in Immunization Against Influenza with a Formaldehyde-Inactivated Virus

There had been some good results in animals with vaccines against influenza, and possibly some good trials in humans too, though not in others; influenza is tricky though, due to antigenic drift.

So this was another study, in Hungary during an epidemic in 1937. They used formaldehyde-inactivated virus taken from infected mouse lungs, and vaccinated 306 nurses and children in Budapest institutions with a single dose. 336 were controls.

They tested some subjects before and after the vaccination, and found that antibody levels rose a decent amount after the vaccination, 24x on average. One subject got tested before, after the vaccine, and then again after getting the flu, and the disease hadn't increased the levels any higher than the vaccine had (though obviously it wasn't a protective level somehow).

The epidemic was pretty small, so there were only 34 cases total in the study: 20 in the unvaccinated, and 14 in the vaccinated. This wasn't a significant difference, so it was negative.

But they isolated virus from some of the cases and found that it was a different serotype, so the vaccine might not've been good at targeting it anyway. Oh well.

Reference:

Taylor, R. M. & Dreguss, M. An Experiment in Immunization Against Influenza with a Formaldehyde-Inactivated Virus. Am. J. Epidemiol. 31-SectionB, 31–35 (1940).

O850 - Tetanus Bacillus Recovered from Scar Ten Years after Attack

When active, the bacteria that cause tetanus can only survive in conditions completely lacking in oxygen, but they can form very tough spores that can survive oxygen, heat, drying, whatever.

In this case report, a woman had some surgery on her uterus and showed some signs of tetanus paralysis afterward; the doctors treated her with anti-tetanus serum and she recovered.

Ten years later, she needed more surgery in the same place. The doctors feared a tetanus recurrence so they gave her anti-serum prophylactically, and tried isolating bacteria from the scar tissue they removed from her body. They successfully grew some typical Clostridium tetani, which produced tetanus toxin capable of killing mice. I don't know how they tried to avoid contamination though.

They tested the patient's serum to see if there were any sign of immunity against the tetanus that seemingly had been in her body for ten years, aside from the immunity from the anti-serum they had given. But they didn't find any indication of immunity.

They cite other reports of tetanus remaining dormant in the body for long periods, even up to 14 years, and advise that surgeons treat for tetanus prophylactically when a patient has had tetanus before. Bacteria can be pretty tough.

Reference:

Bonney, V., Box, C. & MacLennan, J. Tetanus Bacillus Recovered from Scar Ten Years after Attack. BMJ 2, 10–11 (1938).

090 - Serologic Studies in Epidemic Influenza: With Particular Reference to the Persistence of Antibodies After Infection

Many virus infections seem to lead to long-term immunity, so you get it only once; at least, that's how it seemed. But one obvious exception is influenza, which you can get every year almost. Part of the reason for this is its ability to mutate frequently, becoming different enough each year that our immune system doesn't recognize it as well; but Fairbrother and Martin wondered whether this were really the whole story.

So they studied the antibody levels of people before and after an epidemic of flu and for about a year after. There were two kinds of tests they used: complement fixation, and neutralization where they mixed antibodies with virus and injected it into mice to see if they died.

As we saw before (O860), antibody levels seem to correlate with immunity, and definitely increase after infection. They saw the same thing here: after being infected in the epidemic, people's levels were much higher. For those that didn't get infected, levels were pretty variable, some high, some low.

But after about a year, antibodies in people who had been infected were much lower than they had been, nearly back to where they had been before the people were infected.

So it's possible that natural immunity to influenza wanes after a year, though that can't be concluded here because they didn't actually test immunity, only antibody levels.

Reference:

Fairbrother, R. W. & Martin, A. E. Serologic Studies in Epidemic Influenza: With Particular Reference to the Persistence of Antibodies After Infection. The Lancet 231, 718–720 (1938).

O951 - Does an attack of acute anterior poliomyelitis confer adequate immunity? Report of four second attacks in New York City in 1935

There've been other reports of reinfection with polio after an initial infection (077), so it seems immunity is not always lifelong or perfect from natural infection.

Fischer and Stillerman collected a number of reports of such second attacks, 13 from others and four of their own in New York City in 1935. These four were in children 3-9 years old, and the attacks all came 2-5 years apart.¹

The first had some paralysis in the first attack, recovered, and then had some more in the second and took years longer to recover from that. The second had paralysis the first time but none the second; the infection was nonparalytic. The third had some mild paralysis both times. The fourth had a nonparalytic infection first (it was questionable whether it was even polio), but the second attack was fatal.

So from this and other statistics, the authors calculated that second attacks happen around 2 per thousand first attacks. Considering the low attack rate in the first place, they wonder whether there's actually any immunity to polio at all. But others disagree:

"Fischer and Stillerman have raised the question as to whether the low morbidity rate in poliomyelitis would not make the incidence of second attacks rare even if no immunity occurred following the disease. In the 1935 New York City epidemic they observed four second attacks, a rate of 2 per thousand, which was within the limits of expectancy if no immunity resulted from a previous attack. However, these figures were not based on age specific rates and cannot therefore be taken as final."²

References:

1.Fischer, A. E. & Stillerman, M. Does an attack of acute anterior poliomyelitis confer adequate immunity? Report of four second attacks in new york city in 1935. JAMA 110, 569–572 (1938). 

2. Horstmann, D. M. Clinical aspects of acute poliomyelitis. The American Journal of Medicine 6, 592–605 (1949).

O860 - Investigation on Volunteers Infected with the Influenza Virus

Apparently back in the 30s, the Soviet Union was doing research on influenza, some of which used human volunteers.¹ Well, better flu research than smallpox or something, and it seems to have worked out alright.

Here, they tried infecting 72 volunteers with virus taken from infected mice, by aerosolizing it and letting them inhale it. It seemed to work in some of the volunteers, though their illness was mild; the authors speculate that the virus might have been attenuated by passage through animals.

The most interesting result was the antibody levels in the volunteers before and after inoculation. They measured antibodies by drawing blood, mixing it with live virus, and injecting it into mice. If the mice survived, the virus had been neutralized by the antibodies.

What they saw was that volunteers who did get sick had pretty low antibodies to begin with, almost none in some cases; afterward their levels were 25-100 times higher. In those that didn't get sick, the levels had started out high and risen only a little (to about the same level as the others). So antibody levels correlated well with immunity to influenza.

The authors thought this infection with somewhat-attenuated virus method might be a good strategy for immunization. The editor of this study's journal disagreed with that statement, as did later researchers:

"A paper published from the Soviet Union by Smorodintsev[sic] et al. in 1937 - frequently cited as the first paper on live virus vaccine - described the administration of a mouse-lethal strain of the...virus by protracted inhalation of atomized virus. Typical febrile influenza developed in 20% of volunteers, hardly an acceptable vaccine by present standards, and certainly not attenuated, as claimed by the authors. Remarkably, they claimed, as well, that the virus appeared not to multiply in men, but the study was a landmark in establishing unequivocally the role of the virus in the development of the disease and in demonstrating antibody response to the virus during convalescence."⁴

Others questioned whether this study could really be compared to natural infections:

"These results suggested that the level of neutralizing antibodies was of significance in determining susceptibility or immunity to influenza A in man. But the conditions under which the experimental disease was produced and the relatively large quantities of virus suspensions used seem so different from conditions encountered in the natural epidemic disease as to make comparisons between these two conditions hazardous."²

"Whether experimentally induced influenza A in human beings is entirely analogous to the naturally occurring epidemic disease may be open to some question."³

Seems like a reasonable question, but at least it's easier to control this kind of study for unwanted variables.

References:
1. Smorodintseff, A. A., Tushinsky, M. D., Drobyshevskaya, A. I., Korovin, A. A. & Osetroff, A. I. Investigation on Volunteers Infected with the Influenza Virus. The American Journal of the Medical Sciences 194, 159–170 (1937).
2. Rickard, E. R., Horsfall, F. L., Jr., Hirst, G. K. & Lennette, E. H. The Correlation between Neutralizing Antibodies in Serum against Influenza Viruses and Susceptibility to Influenza in Man. Public Health Reports (1896-1970) 56, 1819–1834 (1941).
3. Horsfall, Jr., F. L. Recent Studies in Influenza. Am J Public Health Nations Health 31, 1275–1280 (1941).
4. Kilbourne, E. D. in History of Vaccine Development (ed. Plotkin, S. A.) 137–144 (Springer New York, 2011).

088 - Studies in whooping cough: Diagnosis and immunization

First, the scary numbers: in 1936, there were more than 300,000 cases of whooping cough in the US, and 15% of infected infants died from it.

Leila Daughtry-Denmark wanted to help make this situation better, so this study was partially to figure out how to diagnose it early, partially to figure out how to determine who was susceptible, and partially to see if it was possible to make people less susceptible (by vaccination).

The cultures and vaccine used in this study were provided by Eli Lilly.

First Daughtry-Denmark tested white blood cell counts and agglutination to see if they correlated with infection, recovery, or immunity. Infection or vaccine seemed to increase the former but didn't seem to affect the latter.

Complement fixation (relating to antibody levels) seemed a much better indicator. Even young children (under 6 months) showed a good response to vaccination. Letting the vaccine age to 4 months didn't reduce its effectiveness. And the Georgia Public Health Lab repeated the test on their own and saw similar results. And they never observed fixation without either the vaccine or the actual disease.

Complement fixation also seemed to correlate with immunity, at least in a small sample (2 brothers), where when they were both exposed later, the brother with good fixation was protected and the other got sick.

Then there were some bigger tests of the vaccine, both with Sauer's version and a more concentrated kind that required fewer injections, making it more convenient. Apparently Sauer's sometimes took more than 8 injections to work, while this double-strength one only took three. They sure had some tolerance to shots back then; though I guess the current recommendation is four shots of DTaP before 1.5 years of age, so that's not so much better.

Anyway, there were 240 subjects who got the vaccine, but only 73 actually got exposed to the disease. Of these, 10 got the disease. So you could say it was 86% effective. But there was no control group, so it isn't possible to make firm conclusions. Also, only those who got Sauer's vaccine got exposed, so there wasn't really a test of the double-strength version.

So it seemed to work, but it's hard to know how effective it really was; exposure doesn't always mean disease, even in the unvaccinated.

A final comment mentioned in the article: there was a test of 50 college students who claimed to have had pertussis as children, and it found that only one had very good complement fixation, while 46 had no detectable fixation. So natural immunity doesn't seem that great for pertussis.

Reference:
Daughtry-Denmark, L. Studies in whooping cough: Diagnosis and immunization. Am J Dis Child 52, 587–598 (1936).

087 - Schick Immunity and Diphtheria Infection

Diphtheria is a bacterial infection that can be pretty serious and deadly, especially in children. Fortunately we can form very good immunity against it, either by infection or vaccination, with antibodies targeting only the toxin that the pathogen produces (diphtheria toxin). 

This means the vaccine is fairly simple to make, the only requirement being that it induces an immune response against this toxin. So people were trying different methods to accomplish this: injecting whole toxin (not a good idea), or toxin mixed with antitoxin (antibodies from someone/something else) to neutralize the toxin, or, the best, toxin inactivated just enough that it didn't cause problems but still induced immunity.

One thing I've noticed is that back when the diphtheria vaccine was being developed, some studies seemed to measure its effectiveness solely by something called the Schick test. This involved injecting a small amount of toxin into a patient's skin, then observing the spot for a reaction. Counterintuitively, a negative reaction (lack of inflammation) indicated immunity diphtheria, the idea being that the patient's immune system could neutralize the toxin before it caused problems. So they took "Schick-negative" to mean "immune."

But as with any test, it needs to be validated, to make sure that the results correlate well with the actual state of immunity being tested. Are Schick-negatives actually better protected from diphtheria? The studies I'm reviewing today look into that question.

087a - Schick Immunity and Diphtheria Infection

Not surprisingly, immunity against diphtheria is never perfect in everyone. E. Ashworth Underwood observed a number of subjects in Leeds to see if Schick-negatives ever caught the disease.¹

Underwood had been overseeing a vaccination program, using toxin-antitoxin mixtures, toxoid, or toxoid-antitoxin mixtures, that started in 1928. Between then and 1935, he observed 2197 Schick-negative patients. Some of these were naturally immune rather than vaccinated.

Of these, 20 came down with diphtheria. Two of them were just naturally immune, while the others were natural plus vaccinated. Except for 2, all were children between 3 and 10. In 4 cases, the disease was subclinical, but 13 needed antitoxin treatment. So there are two possibilities: either the Schick test can be negative in non-immune people sometimes, or the cases were infected with some type of pathogen that overcame their immunity.

Addressing the first possibility: at the time, C. diphtheriae strains were characterized as one of three types (or as atypical): mitis, intermediate, or gravis. This roughly seemed to correlate with how virulent they were, how serious a disease they could cause. Of the 20 cases, 19 were infected with gravis type; so it seemed possible that this type could overcome a higher level of immunity.

How does the case rate in Schick-negatives compare to the rate in Schick-positives? Underwood looked at about 85,000 Schick-positive individuals in Leeds, and saw a case rate of about 1.1%. At best with the Schick-negative cases, not counting the subclinical cases or those with a questionable Schick result, the case rate is at best half as much. So immunity could be said to be less than 50% effective. Not great.

Still, that doesn't mean the vaccine was useless. Leeds seemed to be experiencing an outbreak of a virulent strain that could overcome immunity, for one thing; the vaccine might be more helpful in other places with other strains. Also it's important to note that none of the 20 Schick-negative cases was fatal, whereas the case-fatality rate in Leeds in general was at least 8%, so the vaccine might still have had a benefit.

087b - The Schick Test and Active Immunisation in Relation to Epidemic Diphtheria

In this similar study, Parish and Joyce Wright observed diphtheria cases in London.² Usually immunized people didn't get it, or if they did, it was almost too mild to recognize. They did notice a ward with 27 Schick-negative children, 11 of whom seemed to be carrying gravis diphtheria without symptoms.

In a school near London, there were 248 children and 33 staff. All of these got immunized if they were Schick-positive. An outbreak occurred in 1934, started by newcomers (when vaccination was interrupted by measles, chickenpox, and scarlet fever outbreaks), resulting in 4 cases in Schick-positives and 8 in Schick-negatives (one naturally immune). Half the positives were moderate severity, and 5 of the negatives were called "mild or moderate." They also seemed to be carrying the organism for a while, even two months later.

Again, it seemed like gravis strains were the cause of the outbreak. Unfortunately, it wasn't clear how high an antibody might protect against it.

087c - Alteration in the Incidence of the Gravis, Mitis, and Intermediate Types of C. diphtheriæ in Manchester: And Their Clinical Correlation in a Further Series of 940 Cases
According to Robinson and Marshall in this study, it's difficult to distinguish the three types of diphtheria clinically.³ Though they do cause more or less severe cases on average, and have other distinguishing characteristics, such as in lab cultures. The case-fatality rates were about 15% for gravis, 7.7% for intermediate, and 0.5% for mitis, though other researchers had different results.

They report 13 cases in Schick-negative people (12 of them naturally immune, one immunized). None had a mitis strain by itself, though one had a triple-type co-infection. Three had intermediate, the rest gravis. Two of the gravis cases were fatal, but most were mild.

In people Schick-negative due to toxoid-antitoxin vaccination, 11 got diphtheria. All but 2 cases were gravis, and 5 were severe. One died. So overall, it's not possible to conclude that Schick-negatives will only have mild cases, if any. The authors recommend periodic boosters, though nothing specific.

087d - Review of the Observations which have Accumulated with regard to the Significance of Diphtheria Types in the Last Four Years (1931-1935)⁴
In this paper, K.E. Cooper and colleagues talk about the types of diphtheria again: how types can be distinguished, the severity of disease they cause, etc. Apparently virulence doesn't always correlate with toxin production, they claim.

In Cork, more than 70 people died of diphtheria for every 100,000 in the 1920s, higher than anywhere else. The problem declined after many were immunized. After that, there were 81 cases in 2 years, and 18% of them died in the general population. (22% of non-immunized cases died, while no vaccinated child died.)

In other places, there was a lot of deadly gravis, some more than in others, where other strains predominated. In Leeds, they broke down cases by age, and found that in children under 5, case-fatality was 9-13%; it was 3-11% in ages 5-10 (the high end being gravis epidemics), and 1-4% in ages 10-15. So it's pretty dangerous in young children, unfortunately.

In Cork, Leeds, Manchester, and Stafford together, 5% of all gravis cases were vaccinated, 2.5% of intermediate cases, and 1% of mitis. I'm not sure if this takes into account the proportions of each population (as in, if 95% of the population is vaccinated but only make up 5% of the cases, that's different from if 5% of the population is vaccinated but makes up 5% of the cases). But not all of the cases in vaccinated people were mild; 3 gravis cases were fatal. The conclusion was vaccination is generally helpful though.

087e - Diphtheria in Liverpool with special reference to type incidence and severity⁵
H.R. Shone and colleagues looked at the different diphtheria types again. They found that of all cases for all types, the highest rates were in ages 5-9 (about 50% of cases), another 30% in ages 0-4, 14% in 10-14, and the rest in over 14. There was maybe slightly more mitis in ages 0-4 and more gravis in 5-9 and over 14, if the numbers were accurate.

In contrast to others, they found that intermediate infections were most severe, but the difference wasn't really significant between intermediate and gravis. This was reflected in case-fatality rates: 2.4% for mitis, 10.7% for intermediate, and 6.6% for gravis. And in terms of cause of death, it seemed like mitis caused more laryngeal complications while the others were just super-toxic.

Case-fatality rates went down as age increased, so the youngest children had the highest proportion of deaths, unfortunately.

108 vaccinated patients got diphtheria, though their Schick status wasn't tested (so they might not have been Schick-negative). 50% of the cases were gravis, 34.3% intermediate. Two of the intermediate cases died, two sisters 6 and 9 years old. Another 21 were severe.

Conclusions
None of these studies really made a rigorous investigation of the connection of Schick status and immunity, so I'm not sure what can be said, except that Schick-negative status is not always indicative of total immunity. It's important to note that it didn't seem to matter whether the immunity came from a vaccine or from previous infection, especially in the face of gravis.

I don't think Schick tests are done today, though I'm not yet sure why, so maybe it was decided they weren't that useful. We'll see, hopefully.

References:

1. Underwood, E. A. Schick Immunity and Diphtheria Infection. The Lancet 225, 364–369 (1935). 

2. Parish, H. J. & Wright, J. The Schick Test and Active Immunisation in Relation to Epidemic Diphtheria. The Lancet 225, 600–604 (1935). 

3. Robinson, D. T. & Marshall, F. N. Alteration in the Incidence of the Gravis, Mitis, and Intermediate Types of C. diphtheriæ in Manchester: And Their Clinical Correlation in a Further Series of 940 Cases. The Lancet 226, 441–445 (1935). 

4. Cooper, K. E., Happold, F. C., McLeod, J. W. & Woodcock, H. E. de C. Review of the Observations which have Accumulated with regard to the Significance of Diphtheria Types in the Last Four Years (1931-1935). Proc R Soc Med 29, 1029–1054 (1936). 

5. Shone, H. R., Tucker, J. R., Glass, V. & Wright, H. D. Diphtheria in Liverpool with special reference to type incidence and severity. J. Pathol. 48, 139–154 (1939).

Measles and Disneyland: Just the Facts

This is a bit different from my usual style, but a friend of mine on Facebook posted a link to a blog called "Measles Shmeasles Goes to Disneyland" by someone named Jessica Gianelloni, and asked for my input/opinion about what it said, and I put a lot of work into putting together a response, so I figured I might as well post it here too. Note: The original blog no longer exists, as the owner shut it down for some reason, but the content can probably still be found online if you desire it.

Overall I think in this article Jessica gets a bunch of stuff wrong, and at least some of the things she gets right are badly out of context. An interesting thing to note is that I searched for the headline she cites ("Disneyland Measles Outbreak Linked To Anti-Vaccine Movement") and could only find it on The Onion. Make of that what you will.

One thing that seems correct is that the vaccine is not as effective as expected in the 1960s; one dose is not adequate. I don't know why Jessica says 3 or more doses are recommended now though; all the recommendations I could find said only two. But that's not super-important at this point.

HERD IMMUNITY
The next thing is a claim that at least 80% of people being immune is required for herd immunity, which seems accurate based on her "citation." And levels are even higher than that, around 90% vaccinated; I doubt this is incorrect. The question is that if we have such high coverage, which is predicted to prevent spread of measles, why do we have measles outbreaks? And the implication is that the vaccine must not be capable of preventing the spread.

This is a very unsophisticated analysis though. Measles is considered eradicated in the US, which doesn't mean there are no cases, but it means that any outbreaks that start are imported from other countries; once a given outbreak ends, the virus is not present in that area anymore to start any more outbreaks.

And when outbreaks do happen, it's pretty consistent that a majority of the cases are people that never had a measles vaccine; there are areas where the coverage is well below 80% of people. For examples: there was a study on outbreaks from 1989 to 1991, and the risk of catching measles was 35 times higher for unvaccinated people. In 1990 for example, unvaccinated people made up 0.5% of the population but had 17% of the cases of measles in the outbreak. That's a lot higher than would be expected if the vaccine weren't very good. This number was worse in some years, better in others.

In the current Disneyland outbreak, of 34 cases in which the vaccination status of the people was known, 28 hadn't been vaccinated (82%). Similarly, in outbreaks in the first half of 2013 and 2014, of those cases with known vaccination status, 91% and 87% (respectively) were known to be unvaccinated. Only 2% of cases in the 2013 data had received both recommended doses. So it's not really possible to say these were outbreaks in highly vaccinated populations, since it spreads mostly only between unvaccinated. This is something Jessica seems to get wrong.

One thing to note is that of those unvaccinated cases, some of them were too young to be vaccinated. This is important in light of Jessica's Palevsky quote: the reason people who vaccinate are upset with those who don't is that when outbreaks occur, it's mostly the unvaccinated that spread disease to those too young to be vaccinated.

NATURAL MEASLES IS NICE
Next is claims about how before there was a vaccine, measles was a one-time thing, a normal part of growing up, and even contributed to a person's health in many ways other than just the disease itself. But now the vaccine makes it so that instead of measles being found mostly in older children (where it is mildest), it's more common in young children (too young to be vaccinated). The numbers Jessica gives are from less than 0.5% of cases in infants before the vaccine, to 30% now. In addition, implications are that the vaccine A) does not allow mothers to pass protective antibodies to their infants, at least not as well; B) does not offer life-long protection; and C) does not provide the same alleged general health benefits as actual measles infection.

First, about epidemiology before and after the vaccine: I don't know where Jessica got these numbers, but they aren't nearly in line with what I could find. First, from a couple of studies in the US in the 1930s, that I've blogged about before: One in Detroit in 1935 (081) found that in children 0-9 years old, 6% of cases were in children under 1 year old (and 66% in 1- to 4-year-olds). The other, in 1930 in Baltimore (060) found that of cases in children 0-14 years old, 4% were in under 1 year, 8% in 1 year, and most in 1-8 years old. So that's a lot higher than under 0.5%, long before the vaccine.

In more recent outbreaks: In the Disneyland outbreak so far, 6% of the cases have been infants under 1 year old. Again in the first half of 2013, 11% of the cases were under 1 year old. So that's a bit higher than the 1930s numbers, but nowhere near the 30% that Jessica claims.

It's also worth mentioning that in the population overall, in the 1930 Baltimore study, 0.8% of children under 1 year old got sick with measles; so 8 per 1000. Compare that to today, in the 1st half of 2013, there were 18 cases in children under 1 year out of a nationwide population of about 4 million in that age range; so that's 0.0009% of infants got measles. That's 9 per million, almost 1000 times less than before the vaccine. And Jessica acknowledges that the seeming 98% decline in measles was the death rate, not the incidence rate ("Does the incidence rate when the vaccine was introduced even matter?"), so we can attribute this 1000-fold decrease to the vaccine. That seems pretty impressive. I would say, is it important that a slightly higher proportion (2x) of outbreak cases are in younger children, if younger children are much less likely (1000x) to catch it overall?

Next, implication A: the vaccine does not allow mothers to pass protective antibodies to their infants, at least not as well the actual disease. Mothers who had the natural infection pass antibodies to their infants that generally protect them for 12-15 months (as Jessica says). But studies show that mothers who only had the vaccine can also pass protective antibodies to their infants. In this 2010 study, the protection passed from vaccinated mothers to infants was similar to that from naturally immune mothers, though it faded a bit more quickly (1-3 months less time). Here is a nice graph from this study:

Leuridan 2010, Figure 2

Is this significant? Probably somewhat, but not nearly as big a difference as Jessica makes it sound, and measles is a lot more rare now too.

Next, implication B: the vaccine does not offer life-long protection like the actual disease. I didn't look too hard into the claim that the wild virus provides life-long immunity (though one of the first detailed accounts of measles did include an observation of someone who seemed to still be immune to measles after having caught it 60 years earlier. No idea if that is a common thing though). As for the vaccine, a 2012 study found that after 20 years, only 10-15% of people who had received 2 doses had no antibodies. There was also a 1998 study that found that after 12 years, about 98% of people who got the vaccine seemed to have adequate antibody levels to protect them.

At this point in my writing, Jessica shut down her blog. Not sure why. Luckily I found another copy online so I can continue to go back and see what she said.

Next, implication C: the vaccine does not provide the same alleged general health benefits as actual measles infection. Immune diseases, tumors, allergies? I think I know what she's talking about here, something I had heard of before: there was a 1985 study that compared children who got measles and either had a rash or didn't have a rash. There wasn't any comparison with vaccinated or anything, just measles infections. And it seemed to show that children who got the rash had fewer health issues later in life than those who didn't. The lack of rash was explained by children having some sort of passive immunity, either from maternal antibodies or from injections of antibodies; there was no discussion about how the vaccine might affect things. But the hypothesis was that if the body didn't completely deal with the virus all at once, the virus might lurk around and cause health problems later (the ones Jessica claims it prevents). But I don't know if there was any follow-up to this study to clarify anything.

As it stands, it seems like the vaccine might be just as helpful as full-blown measles in preventing these health issues. Other research in Africa found that "vaccine efficacy against death was much greater than the proportion of deaths attributed to acute measles disease...These observations suggest that standard titre measles vaccine may confer a beneficial effect which is unrelated to the specific protection against measles disease." And another study found maybe a slightly increased risk of allergy for those who got wild measles infections.

Overall, it doesn't seem like the data supports Jessica's claims and implications.

MEASLES IS NOT NECESSARILY DANGEROUS
Jessica then claims that measles is not something to be feared, at least not in developed countries such as the US, so a vaccine is not necessary. And even in developing countries such as Africa, the vaccine is not as helpful as sanitation and nutrition would be, especially vitamin A. The implication, I think, is that the costs and risks from the vaccine are greater than the benefits, compared to other treatments or the disease itself.

I'll address the costs and risks of measles first. The CDC in 1998 claimed that measles kills 1 or 2 people for every 1000 it infects, and this is about the same rate as for the encephalitis it causes, a serious brain inflammation. In developing countries, it can kill as many as 1 out of 4 people it infects. I don't know where these numbers come from though.

For encephalitis, I didn't find any other good numbers on that, but it seems like we haven't seen any for a while. However, you can see in the outbreaks I've cited above (Disneyland, 2013, 2014), at least 11% of those who caught it needed to be hospitalized; up to 25% sometimes. Seems pretty serious. I guess I can appreciate Jessica's faith in modern medicine, though, if she thinks being hospitalized is no big deal.

As for death rates, two people died from measles in 2003, and considering the number of cases since 2000, about 1500, that's right in the 1-2 per 1000 range. The two deaths weren't exactly in the healthiest people, but unhealthy people do exist (often through no fault of their own) and should be protected; also, it's not always possible to know who is particularly susceptible to the disease. It could be you!

Another confirmation of the death rate: in the 1999 study mentioned above, there were 26672 cases and 89 deaths. This works out to 3 per 1000 cases; right on target, unfortunately.

But risk of death is not the only factor to consider; there's also cost burden, both to individuals and to the healthcare system overall. Being hospitalized is not cheap, I'm pretty sure, especially for those who have no insurance. A study estimated that each case of measles costs about $20,000 (and that's a conservative estimate!). This is costs for treatment and also public health efforts to track and control the outbreaks. Definitely a cost worth avoiding if possible.

What about vitamin A? That's a fairly cheap and low-risk treatment, right? I don't know what African study Jessica is referring to in the blog, of course, but the World Health Organization agrees that it's worth giving vitamin A to people with measles in developing countries, and agrees with Jessica's 50% figure. A review of other studies concludes that vitamin A might help reduce severity, at least in hospitalized cases. So that's nice, though I don't know how much of an impact it could really have on the risks overall. It seems better to avoid being hospitalized and needing treatment in the first place.

So overall, a fairly high risk of hospitalization (and associated costs), a fairly low (but not negligible) risk of death, perhaps less risk with vitamin A treatment, at least in developing countries. The obvious next question: is the vaccine any better, or is it worse?

THE MEASLES VACCINE IS MORE DANGEROUS
Jessica claims that the vaccine is associated with "seizures, encephalitis, blood disorders, sensory impairments, learning disabilities, immune system suppression, inflammatory bowel disease, inflammation of the brain, and many other severe allergic reactions." Some of those are redundant or too vague for me to figure out what she's talking about, but the Institute of Medicine released a report a few years ago reviewing vaccines and the evidence for their risks, so I'll summarize some of that.

Keep in mind that, for measles itself, if we didn't have a vaccine, there would be about 500,000 to 5 million cases in the US each year, so with a 1 in 1000 to 1 in 10000 death rate (I'm being conservative, giving a range), that means 50 to 5000 deaths and 55000 to 750000 hospitalizations. Are vaccines worse than that?

Regarding encephalitis/brain inflammation, there were studies looking at more than 500,000 children. In that sample, 199 got encephalitis, which overall is about 4 in 10000, except that only 9 of those cases happened within 3 months of vaccination; 80 of them were actually before the children got vaccinated, and the rest were more than 3 months after. So hardly any were likely associated with the vaccine. Another study found no association either.

Regarding febrile seizures, the report concluded evidence was pretty good that they were associated with the vaccine, but they don't seem to cause any permanent harm or learning disability.

Regarding autism, there have been a lot of studies of that with the MMR, some better and some worse. Even of the better ones, they're consistent with their reporting of a lack of association of the vaccine with autism.

Not sure what Jessica means by "blood disorders," but studies consistently report a lack of association of the vaccine with type 1 diabetes at least.

For sensory impairments and other things, the report authors considered the link between measles itself and the vaccine as some evidence for an association, but other than that there wasn't good evidence for a link with the vaccine in particular.

For allergic reactions, there is good evidence that the vaccine is associated with anaphylaxis, but this is rare, happens right after the shot, and is treatable.

Considering all that, it seems pretty clear that we aren't aware of any reason why the vaccine should be considered more risky than measles itself for most people. And it's pretty clear from recent outbreaks that we need to choose one or the other. So those are my thoughts on the issue, based on the data I could find. Just the facts.

086 - Studies on the Relation of Tetanus Bacilli in the Digestive Tract to Tetanus Antitoxin in the Blood

One big thing about vaccines (in some circles) is the question of what the best approach is for producing the best immunity. More specifically, because many vaccines are injected into the muscles, some ask whether this might not be ideal, because the immune system naturally encounters pathogens via other routes, usually, including by swallowing, breathing, or on other mucous membranes (as with sexually transmitted diseases). Of course, some diseases do naturally transfer across the skin mainly (such as those transmitted by mosquitoes or other biting arthropods) or secondarily (HIV can pass from blood to blood if this contact is made).

Anyway, it seems intuitive that the immune system would respond better to vaccines that use the same route as the actual pathogen, rather than a different route. Intuition like this is not always accurate, though; it's important to test. There could be good reasons that using a different route could produce a better immune response for some reason, depending on the specific disease and how it interacts with the immune system. If you listened to the podcast I linked to a few weeks ago, it talks a bit about how inert antigen in the gut doesn't really do much, even if it comes from a gut organism; there needs to be some activation of the immune system, and this is easier to obtain intramuscularly (or with an attenuated live pathogen).

So specifically, the topic of this post is tetanus, and the potential for natural immunity from colonization with the tetanus bacillus, Clostridium tetani (formerly sometimes called Bacillus tetani). Tetanus usually happens when C. tetani spores get into a wound and multiply, producing a toxin called tetanospasmin, which interferes with nerves and prevents muscles from relaxing, causing paralysis. It isn't really transmitted between people, that we can tell, or from animals; the spores are pretty much just everywhere. Getting an infection with it, if survived, doesn't seem to produce useful levels of immunity against future infections, so the vaccine is useful. The modern vaccine is just tetanospasmin, deactivated so that it doesn't cause problems, but does induce an immune response. Immunity to the bacteria specifically isn't necessary.

But some might wonder, is it true that it's not possible to become immune to tetanus naturally? Wouldn't that be better than getting a vaccine (because natural, and thus presumably more effective)? These five studies looked at people and animals that seemed to carry C. tetani around in their gut without apparent symptoms, and tried to figure out if they derived any sort of immunity from their passengers.

Studies on the Relation of Tetanus Bacilli in the Digestive Tract to Tetanus Antitoxin in the Blood
In the first, TenBroeck and Bauer, living in China, had isolated C. tetani from stools of about 35% of the patients they tested. They used basic microbiological techniques; pasteurizing to kill off everything but spores, then growing the spores in specific medium. They tested the isolates for toxin production by injecting culture medium into mice to see if they got tetanus.

So then, they wanted to see if carriers of these bacteria had antitoxin in their blood; that is, antibodies that neutralized tetanospasmin.¹

Others had looked but not found any antitoxin, though some seemed to find it in cattle, but hadn't correlated it with bacteria in the gut. So what TenBroeck and Bauer did was take blood samples from people who were or were not carriers, mix it with standardized tetanus toxin in varying proportions, and inject it into mice, then observe the mice for tetanus.

Without serum, the toxin generally killed mice in 4 days. Serum from most carriers could protect mice from doses of toxin up to 25 times the minimum that would normally kill a mouse (Minimum Lethal Dose, MLD). From the others, non-carrier serum couldn't protect against 2 times the MLD, except for two that got up to 10x MLD. From carriers though, serum consistently protected against at least 10x MLD, up to almost 50x, but mostly 25x. So the presence of C. tetani in the gut seemed to correlate with the presence of antitoxin in the blood for this population.

This doesn't necessarily indicate that the carriers of C. tetani were immune to tetanus though, just that their serum could protect mice from the toxin. The authors do note that there was a low incidence of tetanus in the area though, but it's not a rigorous observation.

They also wondered if these carriers might be a source of tetanus infection for other people. They concluded it was probably a negligible effect, considering how ubiquitous the spores are anyway.

One of the authors actually swallowed a bunch of spores to see if he could become a carrier. He noticed a bit of constipation that might not've been related, and when it went away, he didn't have spores in his gut anymore. Oh well.


Others discuss the inability of others to replicate this work:

"TenBroeck and Bauer have shown that an appreciable amount of tetanus antitoxin was found in the blood serum of persons in China who carried tetanus bacilli in the digestive tract. No one else has been able to corroborate this work. They expressed the belief that this accounts for the low incidence of tetanus in China, where a large percentage of the population harbors tetanus bacilli in the digestive tract."²

"Immunization against tetanus is quite a different problem from that against diphtheria. There are no naturally immune persons. Tetanus antitoxin has never been detected in nonimmunized men, except in China, nor has it been found even after recovery from clinical tetanus."³

I will probably come back to this issue at sometime in the future with more recent studies though.

The Immunity Produced by the Growth of Tetanus Bacilli in the Digestive Tract
The second study is by the same authors (TenBroeck and Bauer), a followup to the first. In this one, they colonize guinea pigs with tetanus spores and see if that makes the animals immune to tetanus.⁴

Guinea pigs are apparently pretty easy to colonize, which is why they used them. They fed the animals one or more of five serologically distinct types of C. tetani, waited six months for antitoxin to appear in the animals' serum, and then injected them with spores or tetanus toxin to test their immunity.

They expected that having any type in the gut would produce immunity to toxin from any other type, since the toxin seems all the same. What they saw, though, is that animals fed a certain type only had immunity when injected with that same type. If they were fed multiple types, they had immunity to multiple types. When the toxin itself was injected, none seemed immune. So immunity seemed to be specific against the bacteria, not the toxin, which isn't necessarily that helpful.

They speculated that maybe immunity to other types might develop if they waited longer than six months, but that's a long time to wait for immunity. It's possible that the antitoxin might work as protection when mixed with the toxin before injecting, but not when the toxin is already in the body.

In the discussion they mentioned having seen cases of tetanus in people who were carrying spores in their gut, even of the same serological type. So it's questionable whether this route produces immunity.

Others comment on these results:

"Ten Broeck and Bauer claimed that animals fed or injected with Cl. tetani developed a type specific resistance in which antitoxin played no part. Since such immunity was specific for the serologic type it would seem that H antigen must have been involved. On the other hand Coleman (Am J Hyg 1931, 14:515) was unable to immunize guinea pigs by feeding tetanus organisms; and Coleman and Gunnison (Am J Hyg 1931, 14:526) could not demonstrate any humoral protection, other than that due to antitoxin, even against the homologous type whether H or O antigens were used for production of antiserums."⁵

So far it doesn't look that useful.

Human Intestinal Carriers of Tetanus Spores in California
The next one is also from Bauer and another author, Meyer; it's another sample of C. tetani in people's guts, this time mostly from California.⁶

Strangely, different people doing this study in different places had found very different proportions of carriers, between 0 to about 40%. California seemed to have a pretty high rate of tetanus at the time (245 cases in 3 years, 67% mortality), so Bauer and Meyer looked at people there, from San Francisco and Los Angeles hospitals.

From 487 specimens, they found spores in 120, so 24.6%. From specimens from other states, they got 26.6%. The rate in areas of CA varied from 7% to 40%. But there didn't seem to be any particular correlation with climate, geography, sex, age, or occupation (outdoors or indoors workers). Most of the spores were of one particular serological type, which they called type 1; this corresponds to the findings of others. And... that's about it.

The Distribution of B. tetani in the Intestines of Animals
The next study was by John Kerrin.⁷ He looked at stools from 100 people, presumably in the UK (since that's where he worked), and found C. tetani in none of them. So he looked in animals and other places to get a more general survey.

Half the isolates he got didn't produce tetanospasmin, but seemed to be C. tetani anyway. Dogs and rats had the highest proportion of carriers, and spores were pretty common in soil and guinea pigs too. Rabbits, horses, cows, mice, sheep, and pigs all had some (cats didn't). He also tested chickens, and some of them seemed to also.

He tried colonizing rat guts with the spores, but after he stopped feeding the spores, they all left the gut before too long. And he tested some rats that were already carriers for antitoxin, and didn't find much. It's not clear what these results mean.

Can Immunity to Tetanus be Produced by the Oral Route?
Finally, a study by Melville Manson again looked at trying to immunize guinea pigs by feeding them tetanus spores or toxin.⁸ 

The first 8 animals got fed tetanus spores. Manson confirmed them by re-isolating them from the animals' stool and injecting them into mice to make sure they produced disease. He tested the animals for immunity by injecting the toxin. Two of these animals died before the test of other stuff, but of the six that remained, none of them lasted longer than control animals (actually somewhat less time). So that didn't work. He didn't wait six months for immunity though, I notice; only 3 months, at most.

Then he fed another 8 guinea pigs the actual toxin. Two died again of other stuff, but of the six left, 2 survived and the others died. Serum from one survivor didn't seem to protect other animals against the toxin though, so there wasn't much antitoxin; maybe they just happened to be resistant.

Finally he fed toxin to another 12 animals. Three of them survived one minimum lethal dose (MLD) but 2 killed them quickly. The others died like the controls.

So overall, there might've been a little immunity after eating the toxin, but it wasn't much.

Conclusions
My take on all of this is that even if some people do have some natural immunity from colonization with C. tetani, it doesn't seem to be enough to protect against very much (at least in guinea pigs), and trying to colonize people with the organism doesn't seem reliable (and might not be safe anyway). So not a very good alternative to the modern vaccine.

There definitely could be more done to study the issue more rigorously though, and I will be on the lookout for other/better studies in the future.

References:

1. TenBroeck, C. & Bauer, J. H. Studies on the Relation of Tetanus Bacilli in the Digestive Tract to Tetanus Antitoxin in the Blood. J. Exp. Med. 37, 479–489 (1923).

2. Bigler, J. A. & Werner, M. Active immunization against tetanus and diphtheria in infants and children. J. Am. Med. Assoc. 116, 2355–2366 (1941).

3. Miller, Jr., J. J. Immunization procedures in pediatrics. J. Am. Med. Assoc. 134, 1064–1069 (1947).

4. TenBroeck, C. & Bauer, J. H. The Immunity Produced by the Growth of Tetanus Bacilli in the Digestive Tract. J. Exp. Med. 43, 361–377 (1926).

5. Gunnison, J. B. Agglutination Reactions of the Heat Stable Antigens of Clostridium tetani. J. Immunol. 32, 63–74 (1937).

6. Bauer, J. H. & Meyer, K. F. Human Intestinal Carriers of Tetanus Spores in California. J. Infect. Dis. 38, 295–305 (1926).

7. Kerrin, J. C. The Distribution of B. tetani in the Intestines of Animals. Br. J. Exp. Pathol. 10, 370–373 (1929).

8. Manson, M. H. Can Immunity to Tetanus be Produced by the Oral Route? Exp. Biol. Med. 29, 561–564 (1932).

082 - An Attempt to Increase Resistance to Pertussis in Newborn Infants by Immunizing Their Mothers During Pregnancy

With vaccine-preventable diseases, young children are often most at risk of serious health impacts or death; the younger, the higher the risk. At least, this is true of whooping cough, which had mortality rates of 26-55% in the 1930s among infants less than a year old.

However, some observed that newborns up to six months old seemed to have some resistance to some diseases—diphtheria, polio, measles, and scarlet fever, for example—especially when the mother had resistance of her own, such as immunity from having had the disease. So it seemed like the mother was transferring her immunity to the infant, probably through the placenta.

So John Lichty, Betty Slavin, and William Bradford thought it might be wise to take advantage of this transfer to give newborns more resistance until they could be vaccinated themselves around 6 months. In this study, they try immunizing mothers during pregnancy and then observing the immune response in mother and infant.¹ This was building on previous work in humans and animals with the same or other diseases, to some extent.

So they selected healthy women with normal pregnancies in obstetrics departments of Rochester hospitals, assigned them randomly to be immunized or be a control, using Sauer's whole-cell pertussis vaccine from Eli Lilly. No placebos, so no blinding of patients. They did separate observations of mothers and infants based on whether mothers had had pertussis before; i.e. history or no history. So they had four study groups: no history or vaccine, history but no vaccine, vaccine but no history, and both history and vaccine.

The way they measured immunity was a bit unusual: opsono-cytophagic index. They took blood from subjects, mixed it with dead pertussis bacteria, and observed how many dead cells the white blood cells gobbled up. They compared subjects based on the number of white cells that ate at least 20 dead bacteria; the "index" value. Presumably the immune status would affect how likely the white cells were to eat the bacteria. There was blinding in this test somewhat: the examiner counting the index didn't know the status of the subject from whom the blood was taken, so as not to be biased in counting.

Results

In total, there were 28 women immunized and 22 as controls. They observed in most groups, most infants had a lower index than their mothers; the exception was the vaccine+history group, in which a third of infants had a higher index.

I made a graph showing the values for the four groups, mothers and infants (in mothers' cases, after the vaccine, when relevant):

Opsono-cytophagic index for mothers (post-vaccine) and their infants; error bars are standard deviations reported in the study.

Overall, looking at the error bars, it doesn't seem like there's much significant difference anywhere. But looking at trends, two things stand out: infants from mothers with no history or vaccine seem lower than from mothers with either, and with both it's highest. Second, the difference between mothers and infants is largest with neither history nor vaccine, lower and similar for history or vaccine, and mothers and infants are closest with history plus vaccine.

The authors looked at a couple other things too. They observed some of the infants before they had nursed and then again after nursing for one week, to see if the colostrum affected the immunity at all. It didn't seem to make a difference.

Secondly, they looked at some clinical data for other patients; specifically, of 31 infants that died from pertussis. Eighteen of them died before 6 months of age, and 13 after. Of those that died younger, 28% of their mothers had had pertussis before giving birth; of those that died older, the number was at least 54%. Sample sizes were pretty small, but it suggests that mother's immunity does have a protective effect for the infant up to 6 months. Seems like just correlation though.

The authors concluded that vaccinating mothers seemed to help. Comparing each infant's index to mother's, the group with neither immunity had only 50% the index in infants compared to the mother; with either vaccine or history, that number went up to 75%; and with both, 100%, almost identical index. So perhaps an additive effect.

Overall, not a very rigorous study, but suggestive. Later articles were somewhat critical of the study, perhaps explaining the weak results:

"Lichty, Slavin, and Bradford attempted, as they put it, to increase resistance against pertussis in newborn infants by immunizing the mother during pregnancy. They confessed their failure. An analysis of the data revealed the following facts: The injections were given at two week intervals in the last six weeks of pregnancy. The total dose administered was 20-25 billion [cells]. Thus the dose was inadequate and too late for antibody formation which reaches its climax between one and two months after the last inoculation. The test for immunity which they employed, cytophagocytosis of the blood, has distinct limitations and has been abandoned by them in favor of mouse tests. Their figures showed no increase in cytophagocytosis of the inoculated mother's blood. Granted the validity of the test, they found no increased immunity in the mother, so that there were no antibodies transferable to the baby through the placenta."²

Even later, though, most studies citing this one seemed to focus on the safety aspect (which I forgot to mention above): of the mothers in the vaccinated group, almost the only side effect was a sore arm that wasn't bad enough to interfere with daily life. One woman had a systemic reaction with nausea and vomiting. Here's an example of a mention:

"Although phase 1 studies of maternal immunization with Tdap are in progress, studies many decades ago with whole-cell pertussis vaccine administration late in pregnancy resulted in high levels of pertussis-specific antibodies in infants and no safety concerns."³

Sometimes I wonder if people actually read old studies before citing them, but I guess usually it doesn't make much difference.

References:

1. Lichty, J. A., Slavin, B. & Bradford, W. L. An Attempt to Increase Resistance to Pertussis in Newborn Infants by Immunizing Their Mothers During Pregnancy. J Clin Invest 17, 613–621 (1938).

2. Cohen, P. & Scadron, S. J. The placental transmission of protective antibodies against whooping cough: By inoculation of the pregnant mother. JAMA 121, 656–662 (1943).

3. Healy, C. M., Rench, M. A. & Baker, C. J. Importance of Timing of Maternal Combined Tetanus, Diphtheria, and Acellular Pertussis (Tdap) Immunization and Protection of Young Infants. Clin Infect Dis. 56, 539–544 (2013).

080 - Whooping-Cough or Pertussis

As mentioned before, whooping cough can be pretty hard on children, especially young ones. In this article from 1938, Robert Cruickshank discusses whooping cough and how it compares to some other diseases in the UK at the time.

What Should We Call...

"Whooping cough" is the common term, referring to the shrill intake of air after a bout of intense coughing, but Cruickshank pointed out that even in severe cases of the infection, not all patients actually whoop. And since just "cough" or maybe "whooping and/or non-whooping cough" don't work too well, he suggests "pertussis" as a good alternative. On the other hand, as I discussed in 079, this could cause some confusion too, since not all cases of coughs with whooping are caused by B. pertussis. But obviously both names have stuck with us throughout the years.

Mortality

Different people had different estimates of how many people died from whooping cough. The case-fatality rate seemed to be between 1 and 8.5%, generally higher for younger patients. Though in Glasgow, the reported rate was 27%, and up to 44% for those less than a year old. Pretty bad.

For comparison, the rates for measles, diphtheria, and scarlet fever were 5%, 4%, and 0.4% respectively. So pertussis was the fourth leading cause of death in London ages 0-5 years, killing 434 people per year. The three leading causes were congenital causes, pneumonia, and diarrhea (presumably infections of unknown etiology). Measles was 5th.

Though despite these numbers, the death rates for these diseases had actually been decreasing over the past 70 years, at least for younger children. Cruickshank doesn't discuss why this might be. Could be better treatments, supportive care, immunization (at least for diphtheria), increasing public health in general... not clear.

Prevalence

Keeping track of cases of whooping cough wasn't mandatory throughout the UK at this time, though some areas did so. So it was only possible to estimate the prevalence. Some estimated that 44% of children in London got pertussis before age 5, and 60% by age 10. Measles was similar, diphtheria and scarlet fever less so.

In England, it seemed like pertussis came in two-year intervals, though it seemed different in other countries. This seemed to be because of the addition of susceptible people to the population (newborns), but could also because immunity after infection didn't last too long (possibly only a year; I wasn't clear on this part).

Lab Tests

It was pretty clear at this point that B. pertussis caused whooping cough (most of the time), not some virus. People infected with these bacteria developed antibodies, and antibodies produced from vaccination correlated with immunity to infection. Cruickshank discusses methods for diagnosis, those that work and those that don't.

Treatment and Prevention

Cruickshank says: "Pertussis is a disease of which it may be said that the multiplicity of remedies is an index of therapeutic failure." I think what he means is there a lot of suggestions but not many that actually seem to work. Probably like what I discussed in 078. Supportive care is good, of course, and anything that helps children breathe better. Some thought vaccine therapy or antiserum worked well, especially in the early stages of disease, but it didn't seem clear.

For controlling spread, Cruickshank mainly recommended keeping infected patients away from susceptible children, which makes sense. Pertussis isn't as contagious as measles or chickenpox, for example, so it wouldn't be too hard, even in hospitals. He thought that patients shouldn't be contagious anymore after the 4th week of disease.

If isolation of cases were impossible in any situation, he recommended vaccination as something that seemed effective. He cited Madsen's data (069) and Sauer's, showing effectiveness of their vaccines. But in the interest of more solid data, he recommends a more controlled study, and possibly a program similar to the one in place for diphtheria at the time.

Overall, not much new here, but an interesting perspective.

Reference: Cruickshank, R. Whooping-Cough or Pertussis. The Lancet 232, 33–37 (1938).

077 - Reinfection (Second Attack) in Experimental Poliomyelitis

A common thing I hear from those that don't like vaccines is that coming down with the "natural" version of the disease gives a much stronger, even life-long immunity to it, while vaccine-derived immunity only lasts a few years. The truth of this depends on the vaccine, the disease, and the person in question, of course, but it's worth asking if it's true that getting a disease makes one immune thereafter.

This study, by Dr. Simon Flexner, investigated whether monkeys that had recovered from a "natural" (though experimental) infection of polio could be reinfected with the same or related virus—a second attack. Others had previously observed second attacks in children and monkeys infected before with polio, but not in such a formal setting.

Polio tends to be more severe in monkeys, often paralyzing and killing them, but they're not as susceptible to it, so it needs to be introduced to them experimentally, in the lab, with larger doses of virus than people encounter. Because of this, it's a bit less "natural" than human infections, but easier to work with. Those that survive have high levels of anti-polio antibodies.

So Flexner took monkeys that had recovered from polio and tried to infect them again, using the same strain of virus as the first time, or a different strain. He found that it wasn't too hard to reinfect these monkeys, even with the same strain of virus. The disease sometimes was just as severe as the first time, even in those that had severe disease the first time. It was even possible in monkeys that had been hyperimmunized through a kind of vaccination.

The final question was whether monkeys could even be reinfected a third time. There had been a case report of a third attack in a child, but none in monkeys yet. Flexner took the monkeys he had left after the second attack and tried reinfecting them, but none of them got sick that time.

So "natural" immunity is not some magical shield, at least not in lab monkeys with polio.

Citation: Flexner, S. Reinfection (Second Attack) in Experimental Poliomyelitis. J Exp Med 65, 497–513 (1937).

075 - Epidemiological Studies in Influenza

People still hadn't worked out exactly what influenza was or wasn't. Was it a disease caused by a single virus, or a collection of symptoms that could be caused by multiple viruses? Part of that was an arbitrary definition. If just one virus, were there different strains? They also strongly suspected that bacterial secondary infections could play an important role sometimes.

So Thomas Francis attempted to define influenza as well as possible at the time, and give some other information about it.¹ He defined what flu is not (common colds, pneumonia, sore throat, diarrhea) and described a typical case. 

He discussed Shope's studies (049) distinguishing the swine flu virus from Haemophilus influenzae suis bacterial infection, and other animal studies in ferrets and mice (such as 074). Also important research growing flu virus on tissue culture or in eggs.

More importantly, he discussed the issue that some had suggested that people don't form immunity against the flu. It's understandable how people could get that impression, considering how the flu virus mutates enough pretty much yearly to be able to reinfect even people who had it the previous year, so we need a new flu vaccine every year. But the presence of at least some immunity is important; otherwise every flu infection could be as scary as avian flu is supposed to be.

Part of the problem was that the techniques for identifying and distinguishing strains of virus weren't very developed at the time. They could try to infect animal models with samples to isolate virus from them, but if the virus were a type that didn't infect such animals very well, it would give a false negative. This happened to Francis: he observed an epidemic of influenza in California in 1936 with all the usual clinical symptoms, but hardly any patient samples gave infectious virus. As others noted:

"Although it has been suggested by Stuart-Harris et al. that in the presence of an epidemic of respiratory disease a certain symptom complex may serve to differentiate influenza from similar but etiologically different diseases, the California epidemic studied by Francis makes this possibility seem unlikely."²

Later, people realized that this was the first identifiable observation of an outbreak of Influenza B.³ Previous studies apparently had focused mainly on Influenza A.

Lastly, Francis discussed attempts to immunize people against the flu. Mostly it was similar to the results in 074: they saw a rise in antibodies against flu for at least a few months, but didn't actually test if it were protective against infection. Interestingly, some tried inoculation with live virus, subcutaneously, and didn't see any evidence of respiratory infection or serious side effects. Looking at antibodies in people who had just recovered from flu (the way they did this was to inject mice with human serum and see if it protected them against viral infection; another technique with questionable assumptions), they found good antibodies in about 30-60%, and also found strong antibodies in about 30% of people who gave negative histories of flu. So, more work to be done.

References:

1.  Francis, T. Epidemiological Studies in Influenza. Am J Public Health Nations Health 27, 211–225 (1937).

2.  Horsfall, Jr., F. L., Hahn, R. G. & Rickard, E. R. Four Recent Influenza Epidemics: An Experimental Study. J Clin Invest 19, 379–392 (1940).

3.  Burnet, F. M., Stone, J. D. & Anderson, S. G. An Epidemic of Influenza B in Australia. The Lancet 247, 807–811 (1946).

074 - Influenza: Further Experiments on the Active Immunization of Mice

Andrewes and Smith were some of the researchers working on creating a flu vaccine, especially since people had discovered that influenza was caused by a virus, not bacteria.

The vaccine they were developing was made from infected mouse lungs, and mice were the model animal they focused on mostly, especially in this study. Mouse lungs produced a lot of virus, but it wasn't the cleanest, so what they were attempting in this study was to produce a cleaner version.

Actually there were three main goals:
1) Try to get as much virus as possible,
So less volume is needed for the same dose

2) Purify the virus as much as possible without reducing its immunizing ability,
So there aren't contaminants that could cause unnecessary reactions

3) and if possible, inactivate the virus (so it can't infect) without reducing its immunizing ability.
So that it can't possibly infect and cause disease.

For objective 1, they tried filtering the virus with membranes that the viruses were too large to pass through, but that didn't really seem to help. At some point though, their virus densities increased 10 to 100 times spontaneously, maybe through some mutation, so that worked out.

For objective 2, they wanted to remove mouse proteins from the preparation, so they tried adsorption/elution, in which they could stick the virus to something and wash everything else off, but they lost a lot of virus with this method too so it wasn't great. Filtering seemed to help though.

For objective 3, they tried inactivating the virus with formaldehyde. A solution of 0.01% could inactivate almost completely in 5 days at -2°C, and 0.02% could completely. This inactivated virus couldn't infect mice when put into their nose.

Immunization Experiments
Then they tested these preps in mice, to see which gave the best immunity against flu virus challenge. What they found was that washed live virus immunized about as well as unpurified virus (when inoculated into the skin or body cavity), and inactivated virus seemed almost as good, though it seemed like the dose they gave of this was higher than the dose of live. Even 0.1% formaldehyde-inactivated gave good immunity. Virus-free filtrate didn't help at all, so the antigen is not soluble.

They did find that virus that had been washed and then inactivated (or the reverse) didn't have much immunizing power in mice. That was unfortunate.

The immunity from each vaccine seemed to fade in mice after 6 weeks. However, this was similar to how long mice had immunity when they had gotten sick with the flu and recovered, so the vaccine was as long-lasting as natural immunity (especially considering that most of the infected mice died from the disease).

Preliminary Human Trial
Finally, they tested inactivated virus in a few human volunteers. They didn't want to use live virus, considering how others had seen what seemed like flu outbreaks from live virus vaccines (049). So 5 volunteers got washed and inactivated virus, and two more got inactivated unwashed virus. They also added 0.01% merthiolate (thimerosal) to prevent bacterial contamination just in case.

The first two had some pain, maybe from excess formaldehyde, so for the others Andrewes and Smith changed the pH to convert the formaldehyde to something else, which worked better. They didn't see any serious reactions to any version, though the ones getting unpurified virus had more tenderness (possibly sensitivity to mouse proteins).

What they saw was that in all but one volunteer, levels of antibodies against the virus rose after the first dose (not much after the second dose for some reason). This was heartening, especially with the washed+inactivated virus that hadn't worked well in mice. Even better, the levels seemed higher than in other people who had recently recovered from the flu! (Though I'm not sure the flu the people had would be the same antigenically as the virus used in this study.) And the levels still seemed high after 2.5 months. So they might be on the way to a good flu vaccine, but they weren't sure yet if antibody levels correlated well with immunity. More work to be done.

Citation: Andrewes, C. H. & Smith, W. Influenza: Further Experiments on the Active Immunization of Mice. Br J Exp Pathol 18, 43–55 (1937).