Unrecognized diabetes in parturients can cause poor obstetric outcomes. Several professional associations recommend screening for diabetes in pregnancy at the first antenatal visit, although the most appropriate test and threshold remain unclear. Measurement of hemoglobin A1c (HbA1c) can be added to the routine early pregnancy tests, with a level of 6.5% or greater (48 mmol/mol) recommended as the diagnostic cutoff for diabetes. However, the optimal threshold in pregnancy is likely lower. This prospective cohort study was performed to determine the optimal early pregnancy HbA1c threshold to detect diabetes and whether this threshold identifies women at increased risk of adverse pregnancy outcomes. The association of the HbA1c levels was compared with diagnoses of preexisting diabetes or gestational diabetes mellitus (GDM) as measured by the standard oral glucose tolerance test (OGTT) performed at less than 20 weeks’ gestation. Diabetes was defined as fasting ≥126 mg/dL (7.0 mmol/L) or 2 hours ≥200 mg/dL (11.1 mmol/L) and GDM as fasting ≥92 mg/dL (5.1 mmol/L), 1 hour ≥180 mg/dL (10.0 mmol/L), or 2 hours ≥153 mg/dL (8.5 mmol/L). Women were referred for management of GDM only when fasting levels were ≥99 mg/dL (5.5 mmol/L) or 2 hours ≥162 mg/dL (9.0 mmol/L). HbA1c testing was performed for 16,122 women at a median of 47 days’ (interquartile range, 38–62 days’) gestation. The mean ± SD HbA1c level was 5.3% ± 0.3% (34 ± 3.3 mmol/mol); 33 (0.2%) women had HbA1c of 6.5% or greater (48 mmol/mol). The OGTTs were performed at less than 20 weeks in 974 of 4201 women at a median of 99 days (interquartile range, 84–113 days). Fifteen women met the OGTT criteria for diabetes, with a median HbA1c of 6.5% (48 mmol/mol; range, 5.9%–8.9%; 41–74 mmol/mol). The OGTT criteria for GDM were met by 170 women at less than 20 weeks, with a median HbA1c of 5.8% (40 mmol/mol; range, 4.8%–7.3%; 29–56 mmol/mol). The optimal HbA1c screening threshold for detecting diabetes was 5.9% or greater (41 mmol/mol) and was met by 2.9% of the total cohort. Sensitivity was 100% (95% confidence interval [CI], 91.8%–100%), specificity 97.4% (95% CI, 95.5%–99.2%), positive predictive value 18.8% (95% CI 15.7%–22.4%), and negative predictive value 100% (95% CI, 100%–100%). For early GDM, this HbA1c threshold was highly specific at 98.4% (95% CI, 97.0–99.9), but less sensitive at 18.8% (95% CI, 6.6%–31.1%). In total, 128 women (74.0%) with early HbA1c of 5.9% or greater who had an OGTT met GDM criteria at some point in the pregnancy; 18 (10.4%) of these women met diabetes criteria. Seventy-three of 78 patients had a postnatal OGTT; 34 (46.6%) were abnormal, 10 (13.7%) had diabetes, 24 (32.9%) had impaired glucose tolerance, and 8 (11.0%) had impaired fasting glucose (100–109 mg/dL; 5.6–6.0 mmol/L). Of the 29 women with HbA1c of 6.5% or greater at less than 20 weeks, 25 (86.2%) had a postnatal OGTT, 20 (80%) had diabetes, and 5 (20%) had impaired glucose tolerance. Compared with women with HbA1c of less than 5.9%, those with HbA1c of 5.9% or greater had more than 2-fold increased relative risk (RR) of preeclampsia, shoulder dystocia, and major congenital anomaly; more than a 3-fold increased RR of perinatal death; and more than a 1.5-fold increased RR of delivery at less than 37 weeks. Four perinatal deaths occurred for women with HbA1c of 5.9% to 6.4% who received routine pregnancy care. Hemoglobin A1c of 5.9% or greater identified all women with diabetes in those who had an OGTT at less than 20 weeks’ gestation. A threshold of 6.5% or greater would have missed almost 50% of these parturients and is thus too high for screening purposes. Future studies are needed to confirm these results and to assess the impact of early detection and treatment on maternal and fetal outcomes and the cost-effectiveness of HbA1c compared with OGTT.