Purpose: To investigate whether twin pregnancies conceived by different forms of fertility treatments are associated with adverse neonatal outcomes and to examine the difference in maternal and obstetrical characteristics between patients.Methods: Our study was a retrospective analysis of twin pregnancies conceived by fertility treatments from a prospectively collected database. Treatments were stratified into two groups: group 1 (ART) consisted of in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI), and group 2 (non-ART) included intrauterine insemination (IUI) and ovulation induction (OI). Composite neonatal morbidity included respiratory distress syndrome, intraventricular hemorrhage, leukomalacia, chronic lung disease, and death prior to discharge.Results: There were 460 neonates in our study; among them, 67% (n = 310) were in group 1, and 33% (n = 150) in group 2. Group 1 patients were more likely to be older (p = 0.004), nulliparous (p = 0.01), delivered twins with lower birth weights (2278 g ± 605 vs. 2427 ± 519, p = 0.009), and had more deliveries < 32 weeks gestation (p = 0.001). In multivariable Poisson regression model, only neonatal intensive care unit admission rate was increased for group 1 twins (aRR = 1.27, 95% CI 1.003–1.60).Conclusions: After adjusting for confounders, twins conceived via ART compared to non-ART had similar neonatal outcomes. These data can help when counseling this patient population and assist in planning larger prospective cohorts.