Recent Advances in Aptamer-Based Biosensors for Bacterial Detection Aptamer-based biosensors have gained significant attention for their versatility, cost-efficiency, and high affinity and specificity in detecting bacterial pathogens. These biosensors integrate aptamers with optical, electrochemical, and mass-sensitive techniques to enable rapid and sensitive detection of bacteria, toxins, and biomarkers. This review highlights the development of aptamers, their structural characterization, and chemical modifications that enhance their recognition properties and stability in complex biological matrices. It also discusses recent examples of aptasensors for bacterial detection and explores the challenges and future perspectives of aptamer-based bacterial detection. Aptamers are small, single-stranded DNA or RNA molecules that can bind to targets with high specificity and sensitivity. They are selected using the SELEX technique, which involves multiple rounds of selection and amplification to enrich sequences that bind to the target. The optimal random region length for aptamers is estimated to be 20–50 nucleotides, while the primers are typically 20 nucleotides long. The SELEX process has been modified to improve efficiency and reliability, and the use of complex initial nucleic acid libraries has increased. Aptamers can be selected to detect specific bacteria directly by using whole cells or spores as targets. This approach, known as cell-SELEX, allows for the generation of specific aptamers against a particular bacterium or even a specific clone. However, cell-SELEX can be challenging due to the negatively charged bacterial cell surface, which can repel DNA. Additionally, the presence of dead cells in a suspension can lead to nonspecific binding, affecting the selection process. Aptamers can also be selected to target bacterial toxins or surface molecules, providing indirect detection of bacteria. This is particularly important for food, medical, and environmental safety. The detection of these toxins relies on labor-intensive processes and expensive laboratory equipment, but aptamers offer a promising alternative. Aptasensors for bacterial detection have been developed using various techniques, including electrochemical, optical, and colorimetric methods. Electrochemical aptasensors use aptamers immobilized on the working electrode to convert the recognition event into a measurable electrical signal. These sensors can detect bacteria in very small volumes with high sensitivity. Optical aptasensors, such as those based on plasmon surface plasmon resonance (SPR), localized SPR (LSPR), and surface-enhanced Raman scattering (SERS), offer high sensitivity and robustness. Colorimetric biosensors provide the possibility for naked-eye detection of bacterial spores, which are highly resistant to environmental conditions. The review also discusses the challenges and future perspectives of aptamer-based bacterial detection, emphasizing the need for further research to improve the sensitivity, specificity, and practicality of these biosensors. The development of user-friendly, low-cost, and point-of-care diagnostic devices remains a key goal in the fieldRecent Advances in Aptamer-Based Biosensors for Bacterial Detection Aptamer-based biosensors have gained significant attention for their versatility, cost-efficiency, and high affinity and specificity in detecting bacterial pathogens. These biosensors integrate aptamers with optical, electrochemical, and mass-sensitive techniques to enable rapid and sensitive detection of bacteria, toxins, and biomarkers. This review highlights the development of aptamers, their structural characterization, and chemical modifications that enhance their recognition properties and stability in complex biological matrices. It also discusses recent examples of aptasensors for bacterial detection and explores the challenges and future perspectives of aptamer-based bacterial detection. Aptamers are small, single-stranded DNA or RNA molecules that can bind to targets with high specificity and sensitivity. They are selected using the SELEX technique, which involves multiple rounds of selection and amplification to enrich sequences that bind to the target. The optimal random region length for aptamers is estimated to be 20–50 nucleotides, while the primers are typically 20 nucleotides long. The SELEX process has been modified to improve efficiency and reliability, and the use of complex initial nucleic acid libraries has increased. Aptamers can be selected to detect specific bacteria directly by using whole cells or spores as targets. This approach, known as cell-SELEX, allows for the generation of specific aptamers against a particular bacterium or even a specific clone. However, cell-SELEX can be challenging due to the negatively charged bacterial cell surface, which can repel DNA. Additionally, the presence of dead cells in a suspension can lead to nonspecific binding, affecting the selection process. Aptamers can also be selected to target bacterial toxins or surface molecules, providing indirect detection of bacteria. This is particularly important for food, medical, and environmental safety. The detection of these toxins relies on labor-intensive processes and expensive laboratory equipment, but aptamers offer a promising alternative. Aptasensors for bacterial detection have been developed using various techniques, including electrochemical, optical, and colorimetric methods. Electrochemical aptasensors use aptamers immobilized on the working electrode to convert the recognition event into a measurable electrical signal. These sensors can detect bacteria in very small volumes with high sensitivity. Optical aptasensors, such as those based on plasmon surface plasmon resonance (SPR), localized SPR (LSPR), and surface-enhanced Raman scattering (SERS), offer high sensitivity and robustness. Colorimetric biosensors provide the possibility for naked-eye detection of bacterial spores, which are highly resistant to environmental conditions. The review also discusses the challenges and future perspectives of aptamer-based bacterial detection, emphasizing the need for further research to improve the sensitivity, specificity, and practicality of these biosensors. The development of user-friendly, low-cost, and point-of-care diagnostic devices remains a key goal in the field