# Development and Control of Biofilms in Diabetic Foot Infections: A Narrative Review
## 8 Biofilm Formation  
The primary criterion for biofilm formation in the foot of a patient is the presence of exogenous glucose, which is commonly observed in individuals with diabetes mellitus. Elevated glucose levels in the blood upregulate genes responsible for producingextracellular polymeric substances, which are the building blocks found in biofilms [1]. The most common microorganisms found in diabetic foot ulcers are Methicillin-resistant Staphylococcus aureus (MRSA) and _Pseudomonas aeruginosa_[54]; other organisms include _E. coli_ and _K. pneumoniae_. Hyperglycaemia increases glucose availability in tissues and wound fluids, an essential carbon source for bacterial growth and biofilm formation. Elevated glucose enhances the expression of virulence and biofilm-associated genes. For example, _Staphylococcus aureus_ biofilm formation is promoted by advanced glycation end products (AGEs), which induce extracellular DNA (eDNA) release via the sigB regulatory pathway. AGEs, a hallmark of hyperglycamic conditions, stimulate biofilm formation and enhance bacterial adhesion and immune evasion. AGEs have been shown to upregulate GlnS, a regulator of sigB, in _S. aureus_, leading to increased biofilm-associated virulence factor expression. In _Pseudomonas aeruginosa_, hyperglycaemia increases intracellular cyclic di-GMP levels, which directly promotes biofilm maturation and delays wound healing in diabetic models. This demonstrates that hyperglycaemia exacerbates biofilm development through intracellular signalling pathways that regulate EPS production. Hyperglycaemia impairs neutrophil and macrophage function, reducing the ability to clear biofilm infections. Additionally, insulin treatment in diabetic wounds has been linked to increased biofilm formation via changes in bacterial signalling pathways, including cyclic di-GMP signalling, which contributes to delayed healing. _P. aeruginosa_ produces excessive alginate, which protects _S. aureus_ biofilms from the effects of antibiotics [9]. This overproduction is primarily due to mutations in the mucA gene, which upregulate the alginate-synthesizing operon [10]. Additionally, there is a cooperative interaction between _S. aureus_ and _P. aeruginosa_, as the production of virulence factors by _P. aeruginosa_, such as hydrogen cyanide and pyocyanin, along with factors from _S. aureus_, including alpha-hemolysin and Panton-Valentine leukocidin, enhances the antibiotic resistance of biofilms formed by both organisms [14]. The antibiotic resistance associated with these biofilms are summarized in Table 3.  
_E. coli_ and _K. pneumoniae_ biofilms share many genes necessary for biofilm formation. The incidence of glycosuria in diabetes mellitus further facilitates the biofilm formation of _E. coli_[55]. Moreover, it has been observed that diabetes mellitus often lead to urinary tract infections caused by _E. coli_[56]. Additionally, the prevailing conditions in diabetes mellitus can reduce the antimicrobial activity of certain molecules like that of the antimicrobial peptide psoriasis [57]. Therefore, diabetes mellitus significantly contributes to biofilm formation which progressively leads to various complications, including diabetic foot ulcers. \begin{table}
\begin{tabular}{c c c c} \hline \hline
**Bacteria** & **Multi Drug Resistance** & **Biofilm Formation** & **References** \\  & **Status** & & \\ \hline _Escherichia coli_ & Yes & ++ & [8; 12; 58; 59; 60] \\ _Pseudomonas aeruginosa_ & Yes & ++++ & [8; 12; 58] \\ _Proteus_ sp. & Yes & + & [8; 12; 58] \\ _Klebsiella pneumoniae_ & Yes & + & [8; 12; 58; 60] \\ _Staphylococcus aureus_ & Yes & ++++ & [8; 12; 58] \\ _Citrobacter_ sp. & Yes & ++ & [8; 12; 58; 60] \\ _Acinetobacter baumannii_ & Yes & ++++ & [8; 12; 58; 61] \\ \hline \hline \end{tabular}  
* Number of + signs indicate the extent of biofilm forming ability of pathogen; For example, ++++ indicates very high biofilm forming ability while + indicates low biofilm forming ability.  
\end{table} 
Table 3: Presence Of Antibiotic resistance and Biofilm forming ability.