Abstract
Aim/background: Pakistan produces over 60 billion liters of milk annually, with more than 90% handled through informal, unregulated supply chains lacking proper hygiene, veterinary oversight, and quality control. This widespread informality increases the risk of milk adulteration, antimicrobial residues, and zoonotic pathogen transmission, posing serious public health challenges.
Methods: A total of 171 raw milk samples (cow, buffalo, and mixed) were collected from March to August 2024 from farms, producers, vendors, and retail outlets in Tehsil Maidan, District Lower Dir, a key rural dairy hub in Khyber Pakhtunkhwa province. Fat, protein, Solids-Not-Fat (SNF), lactose, pH and freezing point were analyzed using Lacto-Scan S60 while β-lactam, tetracycline, chloramphenicol residues and AflatoxinM1 (AFM1) were detected via Rapid One-Step Assay (ROSA) and Brucella abortus antibodies using indirect Enzyme-Linked Immunosorbent Assay (ELISA). Data were statistically analyzed using SPSS software (version 26.0; IBM Corp., USA) applying one-way ANOVA and descriptive statistics to determine mean differences (p < 0.05).
Results: Fat content ranged from 3.56–4.69% and protein from 3.01–3.45%, with lower values in retail milk outlets indicating possible skimming or dilution for adulteration. Values of pH ranged from 7.08–7.33, exceeding normal limits and suggesting potential subclinical mastitis. SNF (6.57–7.43%) and lactose (2.91–3.29%) values were below the national standards, while freezing points (–0.33 to –0.39°C) numbers indicated possible water adulteration. Common adulterants such as starch, urea, detergent, salt, and formalin were detected at varying levels across milk sources, with the highest frequency observed in vendor and shop samples. These findings underscore intentional malpractice and reinforce the need for stricter quality control. Likewise, β-lactam and tetracycline residues were detected in 14.03% of samples, with concentrations ranging from 1.5–12.4 µg/kg, exceeding established maximum residue limits (MRLs) in several cases. Chloramphenicol, a banned antibiotic, was detected in 1.75% of samples at 1.1 µg/kg. Aflatoxin M1 (AFM1) was found in 61.4% of samples, with concentrations between 0.03–0.26 µg/L, surpassing the EU limit of 0.05 µg/L in approximately 68% of positive samples. Antibodies of Brucella abortus were detected in 7.01% of raw milk samples, indicating a potential zoonotic risk from unpasteurized milk in informal dairy supply chains.
Conclusion: These findings highlight systemic gaps in feed quality management, veterinary drug oversight, and milk handling practices. Addressing these issues requires routine residue screening, farmer education on safe practices, and the implementation of innovative rapid testing technologies such as portable, field-deployable kits. Furthermore, aligning national milk safety regulations with Codex Alimentarius standards is essential to safeguard public health and ensure compliance with the international food safety norms.

Key words: Antibiotic Residues; Dairy Supply Chain; Milk Adulteration; Aflatoxin M1; Brucella Abortus; Food Safety