Three new lignan glucosides, namely, justisecundosides A (1), B (2a), and C (2b), were isolated from the whole plant of Justicia secunda together with seven known compounds (3−9). Their structures were established based on a comprehensive analysis of HR-ESI-MS, IR, UV, and CD, in conjunction with their 1D and 2D-NMR data. A putative biogenetic pathway of compounds 1−2a,b from coniferyl alcohol was proposed. In addition, the antimicrobialactivities of the extract, fractions, and some isolated compounds were assessed against multiresistant bacterial and fungal strains. Furthermore, the antiplasmodial, antileishmanial, and antitrypanosomal activities were assessed against the sensitive (3D7) and multidrug-resistant (Dd2) strains of P. falciparum, promastigote and bloodstream forms of L. donovani, and Trypanosoma brucei, respectively. Compound 4 exhibited moderate antibacterial activity against Staphylococcus aureus SA RN 46003 with a MIC value of 62.5 μg/mL. Besides, compound 6 demonstrated a very good activity against sensitive (IC₅₀Pf3D7: 0.81 μg/mL) and multidrug-resistant (IC₅₀PfDd2: 14.61 μg/mL) strains of P. falciparum while compound 4 displayed good antitrypanosomal activity (IC₅₀: 1.19 μg/mL). Also, compound 1 was the most active on the promastigote form of L. donovani with an IC₅₀ of 13.02 μg/mL.