Introduction
The beneficial effects of larvae (maggots)
upon the healing of infected wounds have been recognized for hundreds of years and have an established role in necrotic or difficult to heal chronic wounds.1 These larvae found their way into wounds by accident, particularly under battlefield conditions, but it was recorded that when this occurred the wounds tended to heal more quickly and with fewer complications than comparable wounds that had not become infested.2 The infection of wound with fly larvae (maggots) is referred to as myiasis.
Zacharius was the first on record to have intentionally introduced maggots to wounds during the American Civil War in the nineteenth century.2,3 More recently, during the First World War, Baer recognized their efficacy in infection, later using maggots in peacetime to achieve a success rate of 90% in the treatment of chronic osteomyelitis.3 The advent of antibiotics during the 1940s heralded the demise of maggot therapy and this has remained the case until recently. The emergence of multi-resistant bacterial strains has brought about renewed interest in their use in certain wounds, particularly heavily contaminated/non-healing chronic wounds, or in patients unfit for anaesthesia.4
Although a number of different species, including the common housefly, have been isolated from wounds and body orifices following accidental infestation, the fly most commonly used for larval therapy is Lucilia sericata. The adults have an attractive metallic coppery green screen, hence the common name ‘green bottles’, a facultative parasite that, in humans, only attacks necrotic tissue.5 The adult fly lays large numbers of eggs in clusters or ‘rafts’ on organic matter such as suppurating wounds or carrion. The maggots hatch from eggs laid by mature adult flies. The larvae, which hatch within a day or so, produce a powerful mixture of proteolytic enzymes including collagenase, which breaks down the dead tissue to a semi-liquid form, which is then reabsorbed and digested.6,7 These initial maggots (first instars) grow from 1-2 mm to 8-10 mm, moulting twice to become third instars some five days later. They then pupate in a dry location and undergo metamorphosis to adult flies after a further week.
Maggots constitute a living biological dressing, which acts in a number of ways. Maggot secretions appear to amplify the healing effects of host epidermal growth factor and IL-6 in addition to their haemostatic properties.8 Secretions include allantoin and urea (antimicrobial activity)2,3 calcium carbonate (stimulation of phagocytosis),9 and proteolytic enzymes (slough/necrotic tissue breakdown).3 The resultant enzymatic degradation products are subsequently ingested and digested by the maggots. In vitro studies have shown maggots to inhibit and destroy a wide range of pathogenic bacteria including Methicillin Resistant Staphylococcus Aureus (MRSA), group A and B streptococci, Gram positive, aerobic and anaerobic strains.10
Micro massage of the wound by maggot movement is thought to stimulate the formation of granulation tissue and wound exudate by the host.2 The net effects of maggot therapy are accelerated debridement of slough and necrosis and a reduction in the bacterial load of the wound, leading to earlier healing, reduced wound odour and less pain. Maggots devour dead tissue and bacteria in a wound, but avoid healthy skin and muscle. Although maggots can be used successfully in a wide variety of wounds, they remain underutilized.
Aim and Objectives
Aim of the present study was to culture sterile maggots of Lucilia sericata, which could be used for treating chronic non-healing ulcers on the limbs and trunk of a patient.
Material and Methods
Preparation of Sterile Larvae
We collected the ‘Blowfly’ Lucilia sericata species from a local fish market and an entomologist did the species identification before the flies were used for the study. Lucilia sericata (green bottle fly) were introduced into sterilized meat in a bell jar to lay eggs. Creamy white eggs in clusters were retrieved from meat (Fig. 1). Few of the eggs were mounted on a glass slide and observed under light microscope.
Results
Microscopic observation revealed the eggs to be grayish oval/oblong in shape with few striations. The eggs were hatched into larvae measuring about 1 mm in length (Fig. 2). The larvae were creamy to light brown in colour and motile. The larvae were collected and transferred into sterile lukewarm sand within a protected, screened cage to pupate. The larvae developed into reddish brown coloured pupa measuring approx, 1-2 cm (Fig. 3). The adult fly develops within this puparium and emerged by rupturing the skin with the ptilinum. The newly emerged adult bears little resemblance to the familiar green bottle
little resemblance to the familiar green bottle but within a short while the wings expand and the body takes on its familiar green hue.
The adult flies were fed a mixture of honey, yeast and water insid e the cage. Temperature and humidity were kept stable well within the human comfort range. Gravid females laid eggs one week after emerging. Small cubes of meat were placed in the cage for egg laying. The meat was removed and the eggs were washed off into containers.
The external surfaces of the eggs are normally very heavily contaminated with bacteria. These must be removed or killed before hatching if the emerging larvae are to remain sterile. The eggs were disinfected using 0.5% sodium hypochlorite solution. The sterilized eggs were transferred aseptically into a sterile blood agar medium and placed in an incubator at 37oC for 24 hrs. Maggots were tested for sterility by culturing for aerobic organisms in Tryptone Soya Broth and in Thioglycollate Broth for anaerobes. Sterile maggots were kept viable for 5 days under refrigeration.
Discussion
Review of literature revealed the use of ‘Blowfly’ Lucilia sericata to be the most appropriate for maggot therapy for wounds healing.5 Maggot therapy could be used to clean festering and foul-smelling wounds. Maggots not only eat the rotten flesh, they also get rid of harmful bacteria in the wound.11 Maggots of the blowfly have been used to treat patients with infections from injuries like pressure ulcers (“bed sores”), leg and foot ulcers, stab wounds, and post-surgical wounds that don’t heal properly. Maggots can also be used successfully in a wide variety of wounds.
Using maggots heals wound faster than other non-surgical methods and does not injure healthy tissue. Maggots are implanted directly onto a wound, where they eat dead flesh, clean out dead skin, and kill harmful bacteria within 2-3 days and they’re removed from the wound, and new maggots are applied, if required. The larvae are introduced into the wound with the help of fine nylon mesh hydro colloidal dressing and absorbent pads. Apart from the above material hydrolysing agents such as varidase or scherisorb gel and semi-permeable films such as tegaderm are also used in dressing, depending on the type of wound.
A review of the literature has revealed no significant risks or adverse events causally linked with the clinical use of sterile larvae of Lucilia sericata. There is also a theoretical possibility that a patient could develop an allergic reaction to the foreign protein of the larvae but such an effect has never been reported. According to the literature, the principal disadvantage of larval therapy appears to be a tickling sensation. This sensation can be eliminated by preventing the larvae are from leaving the wound and migrating onto the surrounding skin by the use of an appropriate dressing system.12
In an age of increasing antibiotic resistance maggot therapy should be considered a viable alternative when, a wound becomes severely infected, does not heal, despite appropriate antibiotics and surgical care.
Conclusion
In our present study we successfully collected and identified the specific species of ‘greenbottle fly’ Lucilia sericata used for maggot therapy. We were also able to culture sterile larvae on the blood agar media in our first phase of study. Further, we plan to take up the maggot therapy to its clinical application in our second phase of the study to treat chronic non-healing wounds.
Acknowledgement
The authors acknowledge Dr. A Sundararaj, Veterinary Surgeon, Department of Laboratory Medicine, KJ Hospital, for his valuable help during the study and also Ms. Manju Alex, Microbiologist, Department of Laboratory Medicine, KJ Hospital for her valuable assistance in this study.
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