Background and Objectives: Chronic non-specific low back pain (CNLBP) persists beyond 12 weeks. Manual therapy recommended for CNLBP demonstrates short-term efficacy. Pain Neuroscience Education (PNE) teaches patients to modify pain perception through explanations, metaphors, and examples, targeting brain re-education. Motivational Interviewing (MI) enhances motivation for behavioral change, steering patients away from ambivalence and uncertainty. These approaches collectively address the multifaceted nature of CNLBP for effective management. The aim of this study was to investigate a manual therapy intervention combined with PNE with MI on pain, pressure pain threshold (PPT), disability, kinesiophobia, catastrophizing, and low back functional ability in individuals experiencing CNLBP. Materials and Methods: Sixty adults with CNLBP were randomly divided into three equal groups (each n = 20). The first group received manual therapy and PNE with integrated MI (combined therapy group), the second group underwent only manual therapy (manual therapy group), and the third group followed a general exercise program at home (control group). Pain in the last 24 h was assessed using the Numeric Pain Rating Scale (NPRS), functional ability with the Roland-Morris Disability Questionnaire (RMDQ), PPT in the lumbar region through pressure algometry, kinesiophobia with the Tampa Scale for Kinesiophobia (TSK), catastrophizing with the Pain Catastrophizing Scale (PCS), and performance using the Back Performance Scale (BPS) at baseline, in the fourth week, and six months post-intervention. Results: Statistically significant differences between the intervention groups and the control group were found in both the fourth-week measurement and the six-month follow-up, as evident in the NPRS and RMDQ scores, as well as in the total values of tested PPTs (p < 0.05). Differences were also observed between the two intervention groups, with a statistically greater improvement in the combined therapy group at both time points (fourth week and six-month follow-up) (p < 0.05). Regarding the TSK and PCS scores in the fourth week, statistically significant differences were observed between the two intervention groups compared to the control group, as well as between the two intervention groups (p < 0.05). However, in the six-month follow-up, statistically significant differences were found only between the combined therapy group and the other two groups, with the combined therapy group showing significant improvements (p < 0.05). In relation to BPS, both intervention groups exhibited statistically significant differences compared to the control group in the fourth week, without any significant differences between the two intervention groups. However, in the six-month follow-up, significant differences were noted between the combined therapy group and the other two groups (p < 0.05), with combined therapy demonstrating greater improvement. Conclusions: The addition of PNE with integrated MI enhanced the positive effects of a manual therapy intervention in all outcome measures. The combination of manual therapy plus PNE with integrated MI appeared to provide greater improvements compared to the isolated application of manual therapy, and these improvements also lasted longer. These short- and long-term positive effects are likely attributed to the combination of PNE with integrated MI, which contributed to increasing the effectiveness of the treatment. Further studies are required to investigate the optimum dosage of manual therapy and PNE with integrated MI in individuals with CNLBP.