Antibacterial and anti-biofilm effects of fatty acids extract of dried Lucilia sericata larvae against Staphylococcus aureus and Streptococcus pneumoniae in vitro

Abstract

Development of new effective antimicrobial drugs is still a big challenge to date due to microbial infection remains an inevitable problem against human health. In this study, fatty acids extract of dried Lucilia sericata larvae (L. sericata larvae) was obtained and evaluated by gas chromatograph-mass spectrometry (GC-MS), and its antibacterial activity against Staphylococcus aureus (S. aureus) and Streptococcus pneumoniae (S. pneumoniae) was investigated. We found that LFAs (fatty acids extract of dried Lucilia sericata larvae) exhibited effective antibacterial activity against S. aureus and S. pneumoniae with minimal inhibitory concentrations (MICs) of 125 μg/mL and 100 μg/mL, respectively. The bacterial wall and membrane were the main targets, which was confirmed by fluorescence microscopy, scanning electron microscopy and transmission electron microscopy. Furthermore, a notable anti-biofilm activity against S. aureus and S. pneumoniae was also observed, which was able to both prevent biofilm formation and eradicate mature biofilms of these bacteria. As a promising antibacterial agent, LFAs showed good application prospects in clinical practice.

Larval Therapy and Larval Excretions/Secretions: A Potential Treatment for Biofilm in Chronic Wounds? A Systematic Review

“…Eighty-three percent (10/12) of the studies analyzed were in vitro assessments of the effects of larvae/larval secretions on biofilm [ 84 , 85 , 86 , 87 , 88 , 89 , 90 , 91 , 93 , 94 ], with 17% (2/12) of studies being ex vivo in nature, one using a porcine-skin-based model [ 92 ] and the other a human-skin-based model [ 95 ]. In 67% (8/12) of studies, a passive secretion collection strategy was utilized, whereby live larvae were immersed in a suitable diluent, such as sterile H 2 O or 0.9% phosphate-buffered saline (PBS) solution [ 84 , 85 , 86 , 87 , 88 , 89 , 90 , 93 ], whereas 17% (2/12) utilized an extraction process that involved the milling and further processing of dried L. sericata larvae [ 94 , 95 ], 8% (1/12) utilized a recombinant enzyme derived from L. sericata secretions [ 91 ], and 8% (1/12) applied live larvae directly to their biofilm model [ 92 ]. For those studies that did not use a larval-derived recombinant (11/12), 25% (3/12) did not state the instar of larvae used for their extraction or treatment protocol [ 89 , 92 , 94 ], 25% (3/12) employed 3rd-instar larvae [ 85 , 93 , 95 ], 17% (2/12) used instar 1 and 3 larvae [ 86 , 88 ], and 25% (3/12) collected secretions from instar 2 and 3 larvae [ 84 , 87 , 90 ].…”

“…In 67% (8/12) of studies, a passive secretion collection strategy was utilized, whereby live larvae were immersed in a suitable diluent, such as sterile H 2 O or 0.9% phosphate-buffered saline (PBS) solution [ 84 , 85 , 86 , 87 , 88 , 89 , 90 , 93 ], whereas 17% (2/12) utilized an extraction process that involved the milling and further processing of dried L. sericata larvae [ 94 , 95 ], 8% (1/12) utilized a recombinant enzyme derived from L. sericata secretions [ 91 ], and 8% (1/12) applied live larvae directly to their biofilm model [ 92 ]. For those studies that did not use a larval-derived recombinant (11/12), 25% (3/12) did not state the instar of larvae used for their extraction or treatment protocol [ 89 , 92 , 94 ], 25% (3/12) employed 3rd-instar larvae [ 85 , 93 , 95 ], 17% (2/12) used instar 1 and 3 larvae [ 86 , 88 ], and 25% (3/12) collected secretions from instar 2 and 3 larvae [ 84 , 87 , 90 ]. Excluding the study that applied live larvae (1/12) [ 92 ], 73% of the remaining studies (8/11) used total protein concentrations to quantify their extracts [ 84 , 86 , 87 , 88 , 89 , 90 , 91 , 93 ], with 18% (2/11) quantifying their extracts in mg/mL (mg of milled larvae per mL of water) [ 94 , 95 ] and 9% (1/11) not quantifying their extracts [ 85 ].…”

“…For those studies that did not use a larval-derived recombinant (11/12), 25% (3/12) did not state the instar of larvae used for their extraction or treatment protocol [ 89 , 92 , 94 ], 25% (3/12) employed 3rd-instar larvae [ 85 , 93 , 95 ], 17% (2/12) used instar 1 and 3 larvae [ 86 , 88 ], and 25% (3/12) collected secretions from instar 2 and 3 larvae [ 84 , 87 , 90 ]. Excluding the study that applied live larvae (1/12) [ 92 ], 73% of the remaining studies (8/11) used total protein concentrations to quantify their extracts [ 84 , 86 , 87 , 88 , 89 , 90 , 91 , 93 ], with 18% (2/11) quantifying their extracts in mg/mL (mg of milled larvae per mL of water) [ 94 , 95 ] and 9% (1/11) not quantifying their extracts [ 85 ]. Seventy-five percent (9/12) of the studies employed a 96-well-microtiter-plate-based biofilm assay [ 84 , 85 , 86 , 87 , 88 , 90 , 91 , 93 , 94 ], 8% (1/12) used a modified Lubbock chronic-wound pathogenic-biofilm model [ 89 ], 8% (1/12) used an ex vivo human-dermal-skin model [ 95 ], and 8% (1/12) utilized an ex vivo pig explant model [ 92 ].…”

“…Excluding the study that applied live larvae (1/12) [ 92 ], 73% of the remaining studies (8/11) used total protein concentrations to quantify their extracts [ 84 , 86 , 87 , 88 , 89 , 90 , 91 , 93 ], with 18% (2/11) quantifying their extracts in mg/mL (mg of milled larvae per mL of water) [ 94 , 95 ] and 9% (1/11) not quantifying their extracts [ 85 ]. Seventy-five percent (9/12) of the studies employed a 96-well-microtiter-plate-based biofilm assay [ 84 , 85 , 86 , 87 , 88 , 90 , 91 , 93 , 94 ], 8% (1/12) used a modified Lubbock chronic-wound pathogenic-biofilm model [ 89 ], 8% (1/12) used an ex vivo human-dermal-skin model [ 95 ], and 8% (1/12) utilized an ex vivo pig explant model [ 92 ]. The organisms used to generate biofilms in these studies were as follows: 67% (8/12) of studies used Staphylococcus aureus [ 84 , 87 , 88 , 91 , 92 , 93 , 94 , 95 ], 50% (6/12) used Pseudomonas aeruginosa [ 84 , 86 , 89 , 90 , 92 , 95 ], 25% (3/12) used Staphylococcus epidermidis [ 85 , 88 , …”

“…Seventy-five percent (9/12) of the studies employed a 96-well-microtiter-plate-based biofilm assay [ 84 , 85 , 86 , 87 , 88 , 90 , 91 , 93 , 94 ], 8% (1/12) used a modified Lubbock chronic-wound pathogenic-biofilm model [ 89 ], 8% (1/12) used an ex vivo human-dermal-skin model [ 95 ], and 8% (1/12) utilized an ex vivo pig explant model [ 92 ]. The organisms used to generate biofilms in these studies were as follows: 67% (8/12) of studies used Staphylococcus aureus [ 84 , 87 , 88 , 91 , 92 , 93 , 94 , 95 ], 50% (6/12) used Pseudomonas aeruginosa [ 84 , 86 , 89 , 90 , 92 , 95 ], 25% (3/12) used Staphylococcus epidermidis [ 85 , 88 , 91 ], 17% (2/12) used Enterobacter cloacae [ 88 , 93 ], and Klebsiella oxytoca [ 88 ], Enterococcus faecalis [ 88 ], and Proteus mirabilis [ 93 ] were used in 8% (1/12) studies. Fifty-eight percent (7/12) of studies used wound- or contaminated-medical-device-derived isolates [ 85 , 88 , 89 , 90 , 91 , 93 , …”