A novel antibacterial peptide derived from Crocodylus siamensis haemoglobin hydrolysate induces membrane permeabilization causing iron dysregulation, oxidative stress and bacterial death.

AIMS: A novel antibacterial peptide from Crocodylus siamensis haemoglobin hydrolysate (CHH) was characterized for antimicrobial activity. METHODS AND RESULTS: CHHs were hydrolysed for 2 h (2 h-CHH), 4 h (4h-CHH), 6 h (6 h-CHH) and 8 h (8 h-CHH). The 8 h-CHH showed antibacterial activity against Escherichia coli, Staphylococcus aureus, Klebsiella pneumoniae and Pseudomonas aeruginosa at concentrations of 20, 20, 20 and 10 mg ml-1 (w/v) respectively. Fluorescent microscopy revealed that the 8 h-CHH had bactericidal activity against E. coli and P. aeruginosa. ß-galactosidase assay supported by RT-qPCR demonstrated that the 8 h-CHH resulted in differential expression of genes involved in iron homeostasis (ftnA and bfd) and oxidative stress (sodA, soxR and oxyR). Siderophore assay indicated that the 8 h-CHH also impaired siderophore production with diminished expression of pvdF. This pattern of gene expression suggests that the 8 h-CHH triggers the release of free ferric ions in the cytoplasm. However, decreased expression of genes associated with the SOS response (recA and lexA) in combination with neutral comet revealed that no DNA damage was caused by 8 h-CHH. Membrane permeabilization assay indicated that 8 h-CHH caused membrane leakage thought to mediate the antibacterial and iron-stress responses observed, due to loss of regulated iron transport. The novel active peptide from 8 h-CHH was determined as QAIIHNEKVQAHGKKVL (QL17), with 41% hydrophobicity and +2 net charge. CONCLUSIONS: The QAIIHNEKVQAHGKKVL fragment of C. siamensis haemoglobin is antibacterial via a mechanism that likely relies on iron dysregulation and oxidative stress which results in bacterial death. SIGNIFICANCE AND IMPACT OF THE STUDY: We have described for the first time, a novel peptide derived from C. siamensis haemoglobin hydrolysate that has the potential to be developed as a novel antimicrobial peptide.

Salivary Lysozyme in Relation to Dental Caries among Thai Preschoolers.

AIM: The objective of this study was to analyze salivary lysozyme levels and activities in Thai preschoolers with different dental caries status. STUDY DESIGN: Unstimulated saliva samples were collected from 64 preschoolers, divided into a caries free group (n = 32) and a severe early childhood caries (S-ECC) group (n = 32). RESULTS: Both groups were similar regarding gender, age, dental caries status, salivary flow rate, and salivary protein concentrations. No differences were also in the caregivers' characteristics, oral health behaviors, and feeding habits. Only professional fluoride application was less frequently found in the S-ECC group (p < 0.03). Western blotting and lysoplate assays revealed that salivary lysozyme levels and activities were significantly increased in the S-ECC group compared with the caries free group (p< 0.001; p = 0.008, respectively). CONCLUSION: The up-regulated expression of salivary lysozyme and the increased lysozyme activity in S-ECC preschoolers suggests a possible connection between salivary lysozyme and oral immunity in response to early childhood dental caries.