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Military service and related risk factors for amyotrophic lateral sclerosis

Background The cause of amyotrophic lateral sclerosis (ALS) is unknown, but occupations have been explored as a potential proxy measure of risk. There is a substantial body of literature connecting military service to ALS. We aimed to summarize and assess the quality of this evidence. Methods Systematic review of the literature, including observational studies which explored one of the following exposures: general military service (army, air force, marines, or navy); or specific exposures associated with military service measured among military personnel. The outcome of interest was ALS incidence, which could include onset, diagnosis, or death from ALS. Results A total of 2642 articles were screened. Following exclusion, 19 articles remained for inclusion in the systematic review, including 1 meta‐analysis and 18 original observational studies. Most studies were of moderate quality. In general, the relationship between military service was suggestive of an increased risk, particularly among Gulf War and WWII veterans. Exposure to pesticides (including Agent Orange) certain chemicals (exhaust, burning agents), heavy metals, and head trauma appeared to increase the risk of ALS among military personnel. Conclusions There is a possible association between military service and the subsequent development of ALS; however, the evidence was limited. Studies were generally hindered by small sample sizes and inadequate follow‐up time. Future studies should endeavor to objectively measure specific exposures, or combinations thereof, associated with military service, as this will be of vital importance in implementing preventative strategies into military organizations.

The cause of ALS is unknown; however, men, people of European descent, and those a family history are more likely to develop the disease. 4 In recent years, occupations have been explored as a potential proxy measure for ALS risk. In two reports from the United States, it was found that a disproportionately large fraction of men affected by ALS have a history of military service. 4,5 Military personnel are subjected to intense physical exertion and are more likely than their civilian counterparts to be exposed to certain agents, including lead, pesticides, smoking, and trauma, thought to be linked to ALS. 6 In 2017, a meta-analysis concluded that there is indeed an increased risk of ALS associated with service in the military. 7 Since this publication, more studies have explored this relationship. As well, researchers have begun to explore the effect of specific military-related exposures which might contribute to this elevated risk of ALS. 6 We aimed to not only update the findings from the meta-analysis, but also highlight these putative military-related risk factors for ALS.
To our knowledge, this is the first study to summarize and assess the quality of the evidence through a systematic review of the literature concerning military service and military-related exposures and ALS.

| Eligibility criteria
Eligibility criteria were developed a priori with no deviations during implementation. We included observational study designs appropriate for the study of etiology: case-control, cohort, or meta-analyses.
Studies must have explored one of the following exposures: general military service (army, air force, marines, or navy); or specific exposures associated with military service measured among military personnel. The outcome of interest was ALS incidence, which could include onset, diagnosis, or death due to ALS. Typically, death is not considered a measure of incidence; however, in the context of ALS, it has been used as such due to the rapidly fatal nature of the disease. 8 Studies were included if they reported a risk estimate (odds ratio, rate ratio, risk ratio, hazard ratio, and standardized morbidity ratio); included a minimum of 5 cases of ALS; and were published in English. Case reports, intervention, descriptive studies of incidence/ prevalence and natural history studies of ALS were excluded. No restrictions were imposed in terms of age, sex, ethnicity, or publication date. If an original study was included in a meta-analysis, we reported only the results of the meta-analysis.

| Search strategy
We performed a comprehensive literature search using a strategy developed by persons with expertise in neuroepidemiology and systematic review methodology, with the aid of two research librarians at the Karolinska University Library. Medline, Embase, and Web of Science were searched for articles published up to June 25, 2019.
The detailed search strategy for all three databases is available in the Appendix. Finally, reference lists within articles were screened, and experts were consulted to identify papers which may have been missed.

| Data extraction
Articles were stored in EndNote where duplicates were deleted.
Articles were screened at three levels: title, abstract, and full paper based on the inclusion criteria. Titles and abstracts were screened by one reviewer (KAM), and full papers were reviewed by three independent reviewers (KAM, KAS, and LMS), with disagreements resolved by consensus.

| Quality assessment
The AMSTAR (A MeaSurement Tool to Assess systematic Reviews) 2 9 was employed for the quality assessment of meta-analyses, and the Newcastle-Ottawa Scale (NOS) 10 was used to evaluate the quality of observational studies. The AMSTAR 2 does not generate a quality score, but aids in the identification of high-quality meta-analyses. 9 The NOS has 3 subscales, selection, comparability, and exposure/outcome, with points awarded on each subscale for a maximum of 9 points. 10 Two reviewers (KAM and KAS) independently scored each original observational study using the NOS, with discrepancies resolved by consensus. All studies were retained, regardless of quality score.

| RE SULTS
A total of 2642 articles were screened, 2 of which were identified from reference lists of relevant articles. Of these, 102 were reviewed at the full-paper level. Following exclusion, 19 articles remained, including 1 meta-analysis and 18 original observational studies. Reasons for exclusion included, but were not limited to: lack of military personnel (n = 51), review paper (n = 11), and absence of a risk estimate (n = 10). For the full screening results, see the PRISMA flowchart in Figure 1.
Most studies were of moderate quality, with a median score of 7 of a possible 9 (range: 4 to 8). Funding for the included studies came primarily from government agencies (n = 16) and non-profit foundations (n = 3). Studies were classified into three groups: those which explored general military service; military service during specific wars; and exposures associated with military service. Details of the 18 original articles are available in Table 1.

| General military service
The relationship between general military service and ALS risk has been explored extensively. Our results included one meta-analysis from 2017, which encompassed 9 studies, 7 and 3 original studies. [11][12][13] While the meta-analysis suggested an increased risk, the three subsequent studies reported no relationship between military service and ALS risk. [11][12][13] The meta-analysis by Tai et al 7 summarized evidence between 1981 and 2016 regarding the risk of ALS associated with general military service. The analysis included 6 case-control and 3 cohort studies, with moderate heterogeneity between studies (I 2 = 55%).
Among the 9 studies, the pooled odds ratio (OR) was 1.29 (95% confidence interval [CI]: 1.08-1.54), suggesting a significant increased risk of ALS among military personnel. Sensitivity analyses revealed that studies of higher quality were more likely to yield a positive association between military service and ALS risk than studies of moderate quality. The quality of the review itself was high, based on the AMSTAR criteria; however, there were noted limitations to the included studies, such as small sample sizes, biased control selection, and potentially imprecise case selection. 7 A case-control study in New England, USA compared newly-diagnosed ALS patients to controls seen for other neurological conditions in two neurology clinics. 11  Similarly, linked registries in Sweden were accessed to explore the relationship between occupation and risk of ALS, defined using ICD codes from hospital and physician billings. 13 Among 4567 ALS cases, 31(0.68%) had stated in the 1990 census that they were employed in the armed forces, compared to 195/22,402 controls (0.87%). Following adjustment for age, sex, and education level, there was no risk of ALS among armed forces personnel (OR: 0.77; 95%CI: 0.52 to 1.14). 13

| Gulf War (1990-1991)
Of the 6 cohort studies published, each employed different methods to identify persons with ALS from the same source population of American veterans. Three reported no association, [14][15][16] while three reported a significantly increased risk of ALS among those deployed to the Gulf War. [17][18][19] The first cohort study to report on the risk of ALS associated with the Gulf War was published 9 years following the end of the war. 14 The primary outcome was hospitalization for ALS (based on ICD  were associated with risk of ALS death. There were fewer than five women who had served during any of the other war periods and died of ALS. 22 As an extension, the same research group followed this cohort for an additional 9 years. 23   Blood lead levels were compared among 184 ALS cases from the National Registry of Veterans and 194 controls from the GENEVA cohort. 27 A doubling of blood lead levels was associated with a statistically significantly increased odds of ALS (OR: 2.6; 95% CI:

| Exposures related to military service
1.9-3.7) after adjustment for age. 27 Using the same study design, researchers examined blood levels of selenium, zinc, copper, and manganese among 163 ALS cases and 229 controls. 28  In 2006, the USA Department of Veterans Affairs commissioned a report which concluded that "there is limited and suggestive evidence of an association between military service and later development of ALS". 30 Eleven years later, a high-quality meta-analysis concluded that there is indeed an increased risk; albeit a moderate one. 7 Since the publication of this meta-analysis, we found an additional three studies that found no altered risk of ALS associated with military service. [11][12][13] There were common limitations to studies which explored this relationship. First, death due to ALS was used as an outcome in one-third of studies. 7 While death has been shown to be an appropriate surrogate for ALS incidence, 8 it may be confounded by military service, which can be related to survival. 31 Further, most studies were completed on American veterans. Historically, veterans have had greater access to healthcare than the general American population, which may have also contributed to an ascertainment bias, in which military personnel were over-represented.
Generally, studies of higher quality tended to report a positive association between military service and ALS. For instance, one of the largest studies, a population-representative study from Denmark, indicated that military service was associated with a 1.3fold increased risk of ALS. 32 This study also revealed a gradient in which a ten-year increase in years employed by the military was associated with increased odds of developing ALS. 32 Five years prior to the report on military service and ALS, the USA Veterans Affairs made a policy decision to provide disability compensation to Gulf War veterans who developed ALS. 30 Subsequent to this decision, more evidence has emerged to support an association between deployment to the Gulf War and ALS. Three cohort studies reported a significantly increased risk of ALS among persons deployed to the Gulf War, [17][18][19] while three reported no association. [14][15][16] It should be noted that the number of ALS cases from the six cohort studies was small, ranging from 6 14 to 40. 18 Further, all studies drew from the same population of American veterans, but each used different methods of case ascertainment, making it unclear how much repetition in cases occurred between studies.
The highest quality study, containing the largest number of cases, found an almost two-times increased risk of ALS associated with deployment. 18 The three earlier studies that reported no altered risk (one of which was a follow-up of the same cohort) had fewer than 25 cases. It is certainly possible that they were under-powered to detect a difference. Furthermore, all studies were completed in the decade following the end of the war, which may not provide adequate follow-up time given that 85% of the persons deployed to the Gulf War were under the age of 35 in 1990, 18 and the onset of ALS peaks between 50 and 75 years of age. Again, the evidence varies, but the higher quality studies point to an elevated risk of ALS associated with Gulf War service. As the number of cases increases, future studies will be necessary to confirm or refute this relationship.
Of the other major 20th century wars, only military service during WWII was consistently associated with the subsequent development of ALS. Three cohort studies explored these relationships, one of which was a follow-up to an earlier study using the same sources. [21][22][23] All were limited to the study of men, used self-report to define the exposure and death records as the outcome. None had information on deployment status; therefore, if the risk factor for ALS was an exposure that occurred during deployment, then the true risk may be even higher. Reasons for an increased risk in WWII are not known. One specific factor to this period-ionizing radiation-was not found to increase the risk of ALS in Japan. 33 20,25 This finding is in concordance with results from a 2012 meta-analysis which reported a significantly increased risk of ALS associated with pesticide exposure. 34 However, both studies were based on self-reported exposures and when the South Korean study attempted to use a more objective measure (known battalion/company location), the association disappeared. 25 Proximity to explosions also appeared to increase ALS risk, as noted by two studies. 20,24 The first likely related to the release of nerve agents into the atmosphere following the explosion of a munitions factory. 24 The second was based on 151 ALS cases who self-reported being within one mile of an explosion. 20 It is unclear how an explosion would subsequently lead to ALS, but it may be related to traumatic brain injury or perhaps trauma, both of which have been associated with an increased risk of ALS. 35,36 A second study provides further support for a role of traumatic brain injury among military personnel and ALS risk.
Self-reported head trauma was associated with an increased risk of ALS. 26 Cigarette smoking was not found to increase ALS risk among military personnel, contrary to findings from the general population where smoking is considered an established risk factor for ALS. 37 Each of these self-reported exposures is susceptible to recall bias or reporting error. For instance, a study published after the final search for this review used a more objective measure of traumatic brain injury (US Department of Defense registry data) and found no increased risk of ALS among 139 Veterans with definite ALS. 38 Further, one study reported an association between pyridostigmine bromide (prophylactic pills used in the Gulf War) use and ALS, but found that when soldiers who had implausible values for these exposures (persons who served before the Gulf War) were removed, the association disappeared. 20 Last, heavy metal levels in blood were explored in military personnel and findings were suggestive of an increased risk of ALS associated with higher levels of lead and copper and lower levels of selenium and zinc. 27,28 The roles of these metals in mitigating or aggravating oxidative stress may explain these findings. 39 Copper and lead are the most common heavy metals in ammunition fumes. It is possible that these metals serve as a proxy for a wider range of exposures (i.e., various particulate matter and gases from ammunition fumes) and that the combined exposure is a risk factor for ALS.
However, these levels were measured on prevalent cases of ALS; therefore, it is impossible to determine whether they represent a cause or consequence of the disease.
All in all, the evidence regarding specific risk factors for ALS was scattered. Most evidence was based on self-reported exposures and is likely susceptible to reporting errors. None of the factors identified had sufficient evidence to characterize as true etiological factors.

| Strengths and limitations
Strengths of this review include its comprehensive methodology, including a thorough search of three large databases. Articles were screened by multiple trained reviewers and disagreements resolved by consensus, but it is still possible that some studies were missed.
Publication bias may have influenced results; however, 6 of the 18 original studies reported null findings, suggesting that this body of literature may be less susceptible to this form of bias. Ultimately, the quality of a review hinges on the quality of the original research.
While the average quality score of the included studies was high, there were some common limitations which should be noted. Many of the studies included a small number of ALS cases. This is an inherent challenge to studying a rare disease, but it may have resulted in a lack of statistical power to detect differences between groups. Most studies reported on both sexes; however, overall there were very few female cases of ALS included. There is a clear preponderance of men with ALS, by a factor of 1.2-1.5. 3 The military is also a heavily male-dominated field; between 1973 and 2010, women never represented more than 14% of active-duty military personnel in the United States. 40 It is possible that men bear a greater biological risk than women, or they may be more likely to be exposed to environmental risk factors, such as those common among military personnel. Most studies adequately controlled for the sex disparity through matching or adjustment of their models, but the findings of this systematic review pertain mostly to men due to the overwhelming proportion of men in each cohort. The studies were also predominantly of Americans of European descent, meaning that the generalizability of these findings to other groups is not known. None of the studies explored a cumulative risk score, and only a single study investigated the effect modification of multiple variables. 26 The case-control studies of prevalent ALS cases may have been subject to survivor bias, in that only persons who survive long enough to participate in a study would be included. Cohort studies were largely limited by a lack of adequate follow-up time given that persons typically serve in the military as young adults, while ALS affects older adults.

| Future directions
In their profession, military personnel are exposed to strenuous physical activity, trauma, and contaminants. 6 Although many questions remain unanswered, there appears to be an increased risk of ALS among military personnel, particularly among individuals who were deployed to WWII or the Gulf War. Understanding specific exposures, or combinations thereof, associated with ALS is of vital importance in order to implement preventative strategies into military organizations. Future studies should endeavor to access large source populations and report their power calculations to ensure that null results are not misinterpreted; consider duration of exposures; and include adequate follow-up time to ensure that they are capturing a true "at-risk" population. Standardization of study design and exposure definitions may be required to better understand these relationships through international efforts.

ACK N OWLED G EM ENTS
Special thank you to Sabina Gillsund and Magdalena Svanberg, librarians with the Karolinska University Library, for their fundamental work in helping to develop and implement the literature search strategy.

CO N FLI C T O F I NTE R E S T
KA McKay, KA Smith, L Smertinaite, F Fang, C Ingre, and F Taube report no disclosures.