The main aim of this research was to establish links between agro-ecological and socio-economic factors, fish intake and nutritional wellbeing in adolescent girls in Bangladesh. The omega-3 index has been proposed as a biomarker for cardiovascular (CVD) risk  and as a biomarker of fish intake—but then mainly of oily fish intake . It is not expected that changes in the omega-3 index are associated with anthropometrics and other micronutrient biomarkers and, therefore, it should not be put forward as a general marker of nutritional deficiency. However, for this specific population, where fish intake is relatively high and important, the omega-3 index could present a different but also sensitive marker of nutritional status in relation to fish intake (where micronutrients may be slightly less sensitive).
Evidence from previous studies indicates that the omega-3 index has clinical relevance, being not only a biomarker of fish intake but now also emerging as a risk factor for fatal and non-fatal cardiovascular events in high-income countries  and depression and pre-term birth in a wide range of countries, including LMIC . The observational data enabled us to develop and validate a metric, a user-friendly tool that offers a way of predicting omega-3 status by deriving an immediate risk score from responses to a few short questions on religion, salinity zone, female autonomy, diet diversity and tilapia consumption. As stated above, a composite score is calculated from the response of each girl to these questions multiplied by the logistic regression coefficients for the relevant season in which the questions are asked. This algorithm can be set up online to enable fieldworkers to simply enter the response data. The use of a metric using a few short questions is cheaper, can be done online, and avoids the complexity and cost of finger prick blood sampling and biomarker measurement based on field samples. It is ideal in contexts where the relationship between these variables has already been established such as large areas of coastal Bangladesh but would need to be reworked based on the same process for different sociological and geographical contexts.
Also, the study provides a more detailed insight into the factors of importance in health and wellbeing of adolescent girls, including modifiable risk factors, and thus ways to potentially intervene. For example, low omega-3 index levels could be an indication of general low levels of fish being accessible in the food environment or, of a lack of female autonomy, which is, in itself, challenging to measure as surveys may have to be redesigned to each specific setting and/or be controversial in some settings.
We found that the metric discriminates better in the wet season compared with the dry season, reflecting that during the dry (or lean) season, nutritional deficiency is more widespread. During this season, any interventions to address nutritional deficiencies would more likely be at a population level, rather than identifying individuals at risk. Given the seasonal impact on food intake [10, 11], this is a significant finding as the metric would enable interventions to be targeted during the wet season to those girls who are most likely to remain chronically ‘at risk’ of nutritional deficiency. The influence of seasonality on food consumption is widely acknowledged . In rural Timor Leste, fish consumption was higher in coastal than inland locations during the wet rather than dry season . Karim and Little  found household consumption and income from freshwater fish culture was strongly impacted by season and household wellbeing status in Northern Bangladesh. In relation to seafood consumption specifically, Karageorgou et al.  found differences in intake by religion (Muslims versus Christians, Hindus) which was not observed for other food groups . However, whilst differences in seasonality and religion exist, there is still much unexplained variability in the model. Other factors such as genetics or other phenotypic variables that were not measured may also play a role.
Similar to previous research [10, 11], we found that anthropometry outcomes did not show consistent correlations with the omega-3 index, highlighting the challenges of measuring nutritional wellbeing in adolescents . It is generally accepted that a limitation of anthropometric measurements is that they are unable to identify protein and micronutrient deficiencies, detect small disturbances in nutritional status, nor identify small changes in the proportions of body fat to lean body mass [42, 43]. There have been calls for the use of alternative methods to measure dietary quality among adolescents, complementing anthropometry with the systematic collection of other information, particularly dietary intake . We found that the intake of the main fish fatty acids eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), correctly reclassifies adolescent girls from intermediate to high or low risk in our specific setting in Bangladesh which is an important criterion for a novel biomarker .
This research is important for aquaculture-ecozones such as coastal Bangladesh. The use of different aquaculture-ecozones defined by salinity in our study setting has enabled us to establish relationships between seasonal fish intake, the role of women’s autonomy and health outcomes. This study has improved our understanding of factors including salinity, seasonality and within-household/community social contexts and health outcomes. We have established that higher female (ie adolescent girls’) autonomy, religion (being Hindu rather than Muslim), geographical location (i.e. living in a high or mid-saline area), and a higher dietary diversity were the strongest predictors of whole blood omega-3 index, an established marker of fish intake but also a risk marker of cardiovascular disease , in adolescent girls in the Khulna area of Bangladesh. Whilst being a major contributor to the intake of dietary energy, fat and protein, fish consumption also lowers the risk of coronary heart disease and stroke [47, 48], which may become increasingly relevant for the treatment and prevention of non-communicable diseases in LMICs. Fish production and availability in agro-ecological coastal zones is dependent on seasonal and annual fluctuations in freshwater supply creating a variable salinity gradient, which impacts on aquatic food production, and on food production more generally. The local communities living in these dynamic aquatic eco-zones are vulnerable to poverty, poor diet and health, and whilst these ecosystems produce highly valuable and nutritious aquatic foods, changes to the supply and accessibility of aquatic foods may impact on the dietary quality of the population, as well as on population health outcomes .
The main purpose of developing a metric, a statistical model used to predict the likelihood that an individual with a given set of risk factors will experience a health outcome , is to provide an effective tool to reveal these specific linkages in practice. Its application would enable the development and implementation of better informed and more integrated policies and practices in relation to aquatic food production systems and inform the design of interventions that might be appropriately targeted to promote healthy fish-containing diets and aquatic agro-ecosystems. It is anticipated that these findings will be potentially relevant for other coastal LMIC that are dependent on fish, particularly freshwater fish, to increase dietary diversity and ensure sufficient intake of energy, macro- and micronutrients. As such, the metric is intended for use in local communities living in such dynamic aquatic eco-zones or geographic areas with similar coastal saline deltas with a high concentration of people and extensive aquaculture.
The importance of social and cultural norms on intra-household access to nutrients and micronutrients in South Asia generally has long been understood . Including the omega-3 index and women’s autonomy score, a key indicator of resource allocation within the household [8, 51,52,53], in this metric are novel additions to complement existing indicators such as anthropometry. Adolescent girls from Hindu communities enjoyed more autonomy in terms of mobility, ambition and food choice compared to individuals from Muslim communities. The inclusion in the metric of one type of fish, tilapia, that are strongly correlated with omega-3 index demands further analysis. Tilapia was introduced in Bangladesh but is now occurring as natural breeding populations in ponds. Tilapia freely breeds in saline ponds for frequent harvest supporting local consumption whereas it is excluded from freshwater ponds where it is believed to compete with the key economic species, Macrobrachium. In other regions of Bangladesh, tilapia have been promoted in fresh water and the species has risen to the top three most produced and consumed fish . It is also among the most affordable of farmed fish and consumed by the poorest consumers. In Barisal, Bangladesh, consumption is particularly high in the wet season reflecting greater availability and, if not self-sourced, their affordability .
In a wider global context, this research is important for documenting the wellbeing of adolescent girls. The promotion of fish consumption as an important dietary source of protein, micronutrients and vitamins could play a key role in nutritional security in this vulnerable group . Here, we developed a risk-based prediction algorithm to identify adolescent girls at increased risk of nutritional deficiency. By combining readily measured risk factors, as informed by our previous studies [10, 11], we could produce an overall risk score which can inform more targeted and appropriate dietary and health interventions. Currently, there is no metric available that takes account of environmental, cultural and economic contexts when considering dietary health from a health policy perspective. Indeed, policies addressing the specific challenges of nutritional needs of these communities are limited by the sectoral separation of aquatic food production, the broader food sector and public health institutions [1, 2]. There is an urgent need for metrics to incorporate all these aspects. Bennett et al.  called for new metrics to better understand current and potential nutrient production, and highlight specifically the need to understand “pathways linking fish to food and nutrition security’’ (direct consumption, income, women’s empowerment).
The strength of the study is the use of primary data from a field study designed to develop the specific metric. Also, the validation of the metric by external data collected with the specific purpose is a major strength of the validity of the metric.
The study is limited by a number of assumptions for the predictors. For women’s autonomy, we assumed an equal value for each dimension in our multidimensional measure. This is the most common approach for setting weights. This approach has been used in many examples, including the United Nation’s Human Development Index but, whilst convenient, this approach has been criticised  on the basis of assuming that all dimensions are equally valuable to people. For example, in the women’s autonomy score, responses linked to food choice and nutritional food access were as important as opportunities for recreational activities and freedoms in being able to make personal decisions affecting daily life activities. Other data-driven methods to adjust the weights could be employed in future iterations .
Micronutrients are not part of the metric as either a predictor or outcome. A manuscript is currently under review which assesses the micronutrient status of the same adolescent girls . In that manuscript, we investigate the status and risk of deficiencies for iron, iodine, vitamin D and vitamin A separately. For the analysis in this study, the status of deficiencies of the four micronutrients assessed are aggregated into a score, rather than including each separately. In this way, we capture a score for generalised micronutrient riskwhich is clearly a cruder analysis. Using a combined count (score) would hide how any deficiencies may potentially differ by salinity site and season for each micronutrient. Similarly, socio-economic status could have been split into better off/worse off categories rather than just splitting by quintiles.
Yet, despite these limitations, the development of this metric, with the participation of potential users in Bangladesh, would have value to development agencies assessing strategies for other coastal or saline affected areas in terms of maintaining agricultural yield and the quality of food systems and nutritional outcomes for vulnerable groups. Its development is an iterative process as we continue to interrogate the data. These data were collected in the relatively small agro-ecological area of Khulna in Bangladesh. In order to enhance the external validity and generalisability of our findings, it is imperative that these data are validated in a larger variety of different agro-ecosystems. Bangladesh has an estimated 30 million people living in agro-ecosystems, including coastal wetlands that are vulnerable to climate change and salinisation, but such transformations are also occurring in other densely populated LMICs in Asia and further afield where we intend to validate and refine the metric further. There is also the potential for linking this metric to a future economic or policy model with costs and resource use included to evaluate the cost-effectiveness of interventions.