**Background:** Premature birth, especially before 34 weeks' gestation, is a leading cause of death and disability in children under 5 years. Omega-3 (n-3) supplementation during pregnancy has been shown to reduce prematurity risk, particularly among women with low omega-3 status. Previous analyses from the ORIP trial and the ADORE trial demonstrated that women with low n-3 levels in early pregnancy benefit most from supplementation for reducing early preterm birth (EPTB). However, it remained unclear whether n-3 status is the only important predictor for EPTB and whether it plays a role in preventing preterm birth (PTB, <37 weeks). This study aimed to identify a broad range of maternal biological, clinical, and demographic characteristics that could predict which women with singleton pregnancies benefit from n-3 supplementation to reduce their risk of both EPTB and PTB.
**Methods:** This study used data from the ORIP trial, a multicentre, double-blind RCT conducted across six tertiary care centres in four Australian states. Women with singleton pregnancies enrolled before 20 weeks' gestation (median 14 weeks) were randomised 1:1 to receive fish oil capsules (900 mg n-3 LCPUFA/day: 800 mg DHA + 100 mg EPA) or isocaloric vegetable oil control capsules until 34 weeks' gestation. The analysis included 5328 singleton pregnancies in 5305 women with known gestational length. Outcomes were EPTB (<34 weeks) and PTB (<37 weeks). Predictors included maternal age, race, socioeconomic status, education, employment, income, weight, height, smoking, alcohol intake, supplement use, diabetes, pregnancy history, and blood biomarkers of n-3 and n-6 status (ALA, EPA, DHA, EPA+DHA, total n-3, linoleic acid, arachidonic acid). DHA, EPA+DHA, and total n-3 were categorised as low, moderate, or replete based on previously identified thresholds. Logistic regression with GEE was used, with each predictor explored separately in models including treatment group, predictor, and their interaction, adjusted for stratification variables. No adjustment was made for multiple comparisons due to the exploratory nature.
**Key Results:** Overall, 91 (1.7%) pregnancies resulted in EPTB and 378 (7.1%) in PTB. For EPTB, n-3 supplementation showed no overall benefit (1.81% vs 1.61%, OR=1.13, 95% CI 0.74–1.71). However, significant effect modification by n-3 status was observed. Women with low total n-3 status (<4.2% of total fatty acids) had the greatest benefit (OR=0.30, 95% CI 0.10–0.93, interaction p=0.007), with a 70% reduction in odds of EPTB. Background risk was highest in this group (3.05% EPTB in controls). Conversely, women with replete total n-3 levels (>4.9%) had increased odds of EPTB with supplementation (OR=2.60, 95% CI 1.20–5.64). Similar patterns were seen for DHA and EPA+DHA status. No other maternal characteristics predicted EPTB benefit. For PTB, n-3 supplementation showed overall benefit (6.37% vs 7.82%, OR=0.80, 95% CI 0.65–0.99). Parity and alcohol use were significant effect modifiers. Supplementation reduced PTB odds by 35% for multiparous women (OR=0.65, 95% CI 0.49–0.87) but not for primiparous women (OR=1.03, 95% CI 0.75–1.42, interaction p=0.04). Among women who abstained from alcohol before pregnancy, supplementation reduced PTB odds by 38% (OR=0.62, 95% CI 0.45–0.86), with no effect in those who drank alcohol (OR=0.97, 95% CI 0.73–1.30, interaction p=0.04). A three-way interaction (p=0.02) suggested that n-3 supplementation was beneficial across all subgroups except primiparous women who drank alcohol (OR=1.54, 95% CI 1.01–2.34). Total n-3 status did not significantly predict PTB benefit (interaction p=0.23), though supplementing only women with low total n-3 levels would be expected to reduce PTB odds by 44% (OR=0.56, 95% CI 0.34–0.92).
**Clinical Implications:** These findings support current Australian recommendations for n-3 supplementation in women with low n-3 status during pregnancy to reduce EPTB risk. The results reinforce a 'test-and-treat' approach—measuring total n-3 status in early pregnancy (using a whole blood cut-point of <4.2% of total fatty acids) and recommending higher-dose n-3 supplements only to those with low levels. This strategy is already being evaluated in South Australia as part of routine antenatal screening. For PTB, parity and alcohol use may help identify women likely to benefit, but these exploratory findings require confirmation before clinical implementation. The study's strengths include its large sample size (5328 pregnancies), very low missing data, and biologically plausible predictor selection. Limitations include the exploratory nature with multiple comparisons (no correction), relatively few EPTB and PTB cases limiting combined analyses, and potential inaccuracies in self-reported alcohol intake. Further research is needed to understand how parity and other characteristics influence n-3 supplementation effectiveness for PTB prevention.