CROSS REFERENCE TO RELATED APPLICATION [0001] This application claims benefit of US Provisional Patent Application number 61/780,551 filed 13 March 2013 the disclosure of which is incorporated herein by reference.

INCORPORATION BY REFERENCE [0002] All publications and patent applications mentioned in this specification are herein incorporated by reference to the same extent as if each individual publication or patent application was specifically and individually indicated to be incorporated by reference.

FIELD

[0003] Described herein are infection treatment devices and methods that can be used for antimicrobial treatment of an infection site. In particular, the devices described may be configured to provide antimicrobial gas/vapor/plasma in conjunction with ultrasonic stimulation for disinfection of nail, skin, or other locations. BACKGROUND

[0004] Aspects of the present invention relate to methods and apparatus for treating microbial infections of the body where bacteria, viruses, molds, yeasts and fungi play a role, particularly in the treatment of skin and nail infections. More particularly, one aspect of the present invention provides methods and devices for treatment of microbial infection of the nail such as where pathogenic fungal colonization of the nail causes onychomycosis.

[0005] Healthy nails in visibly good condition are an important and highly prized aspect of human appearance. Nails are a hard structure made of layers of keratin, a protein, which has lipid bilayers, phospholipid cell membranes and connections between the dead cells called desmosomes. Referring to FIGS. 7A-8, the nail typically includes a nail plate or body with a free end and a nail root that is held near the epidermis by a proximal nail fold. A nail matrix is located at the base of the nail at the nail root. The nail matrix consists of, among other things, rapidly dividing cells that fill with keratin and form the structure of the nail body. As shown, the nail plate is designed to be tightly held against the epidermis by the lateral and proximal folds encircling the nail body. As such, even with a healthy nail structure, there are limited options for introducing or delivering treatments into the nail body or the areas between the nail and the skin. These challenges are further exacerbated where the nail structure has been compromised by disease, injury, or infection.

[0006] Microbial infection, especially of the toes and feet is a common problem. For example, when fungal infections occur in the nail bed, nail matrix, or nail plate of fingers and toes, it has the medical term onychomycosis. Onychomycosis refers generally to an infection of the nail plate, bed, or matrix caused by any fungus, including dermatophytes, nondermatophytes, and yeasts. Onychomycosis accounts for at least a third of integumentary fungal infections and at least half of all nail disease. Onychomycosis fungus acts as a parasite, and lives on the nail keratin, by dissolving the keratin with enzymes known as keratinases. The nail becomes discolored, turning yellow or white, and keratin debris develops under the nail causing nail thickening, nail deformity, shape distortions, and separation. As the fungus grows in the substance of the nail it becomes brittle, splits and crumbles. Some commonly contagious routes include direct contact with an infected person or secondary transfer from damp surfaces in nail salons, gyms or health spas. Nail fungus can also spread over time to other nails or the skin of the patient.

[0007] Once the fungus establishes itself under a toenail or fingernail, it is difficult to cure. Generally, target sites for treatment include the nail plate, bed, and matrix depending on the severity and type of onychomycosis (e.g. distal subungual onychomycosis, white superficial onychomycosis, etc.) The common treatments for onychomycosis generally falls into three categories: a) surgical removal of all or part of the nail followed by topical treatment of the tissue, b) systemic, usually oral, administration of antifungal drugs, or c) topical application of creams, lotions, gels or solutions on the intact infected nail.

[0008] There are a number of drawbacks with these treatments. Surgical removal can result in considerable patient discomfort, has high costs, usually renders the exposed nail bed susceptible to infections, and has only a small chance of cure. Negative aspects associated with oral systemic antifungal therapy for onychomycosis include a limited cure rate, contraindications and drug interactions, toxicity, and the high cost of the medication. While the topical administration of antifungal drugs is the preferred method of administration, they have not been effective because antifungal drugs cannot readily penetrate the nail plate to reach the infection sites both under and within the nail. Also patient compliance is low due to the time consuming and complicated application protocol of the topical drugs.

[0009] Although a number of attempts have been made to effectively treat nail fungus, there is widespread dissatisfaction with the limited clinical efficacy, high cost, long treatment duration and potential side effects of available technologies and products. Recently, laser treatment has been proposed as a possible treatment option; however, there is yet no evidence to support the effectiveness of these high cost treatments for eliminating nail fungus.

[00010] Accordingly, there remains a need for effective treatment of pathological conditions of the nail; an effective antifungal topical therapy which is safe and effective in treating onychomycosis with limited side effects and does not require the removal of the nail. Such a treatment can potentially provide significant improvements over current therapies including less toxicity, shorter treatment times, lower cost and greater efficacy.

[00011] In another aspect, the invention provides devices and methods for treating wounds and promoting wound healing. Generally, the wound healing process involves the phases of hemostasis, inflammation, proliferation, and remodeling. At any one of these stages, it is important to keep the wound clean and free from infection. Infection not only slows healing but can cause discomfort and pain for the patient. Options for disinfecting mild or moderate wounds can entail topical or surface treatments such as applying antimicrobial creams or gels. More severe cases may require systemic antibiotics to control aggressive infections.

[00012] As discussed, there are several drawbacks for existing topical and systemic treatment options. The efficacy of topical treatments is limited by the deliverability of the active agents to a treatment zone. Wound sites often include exudates and/or tissue debris that block

permeability of a liquid, gel, or cream based medicant. Similarly, systemic treatments include many side effects and are not targeted to a specific wound location. Rather, an antibiotic, for example, effective against an infecting bacterial strain may also eliminate healthy bacteria flora from the patient. As such, there remains a need for an effective topical treatment option for wound therapy.

[00013] Embodiments described herein address at least these concerns. Aspects of the invention provide devices and methods for a topical therapy which is curative for microbial infections, particularly onychomycosis. Other features and advantages of the present invention will be apparent to those skilled in the art from the following detailed description.

SUMMARY OF THE DISCLOSURE

[00014] The following summary of the invention is provided to facilitate an understanding of some of the technical features related to the invention, and is not intended to be a full description of the present invention. It is meant to be exemplary and illustrative, not limiting in scope. Disclosed are device and method embodiments of the present invention that are useful for treating or disinfecting a microbial infection, including a fungal infection of the nail on the hands or feet (onychomycosis) or topical application to a microbial contaminated area of the skin. [00015] Embodiments of the invention may have several aspects. In one embodiment, the invention includes a device to effectively destroy or inactivate pathogenic microbe species that could be fungi, viruses, bacteria or yeast. A further embodiment includes a method that comprises applying a gas or vapor with antimicrobial properties onto the infected body surface either alone or in combination with ultrasound-assisted mass transfer of the antimicrobial agent.

Antimicrobial is defined as tending to destroy microbes, prevent their development or inhibit their pathogenic action and includes but is not limited to antibacterial and antifungal properties.

[00016] The gas or vapor with antimicrobial properties could be an effluent from a nonthermal plasma source or another source of oxygen free radicals such as ozone, hydrogen peroxide vapor or any combination of these and is herein called an antimicrobial agent. A single antimicrobial agent or mixture of such agents can be used. By applying an antimicrobial agent to the infected nail surface, the gas can penetrate into the interior of the nail region via the existing porosity and kill the microorganisms that cause onychomycosis. It would be advantageous to utilize an antimicrobial agent as a gas or vapor for treatment of the nail as it results in increased permeation of the antimicrobial agent into the nail as compared to a liquid or cream.

Antimicrobial agents that are adapted to kill fungus residing on the body or within such areas as the toe nail or finger nail can treat onychomycosis.

[00017] In a second aspect, the invention relates to a device where the antimicrobial agent is administered topically in combination with the use of ultrasound stimulation to improve nail penetration of the antimicrobial agent through the nail plate and onto the nail bed surface. The ultrasound stimulation makes the antimicrobial agent even more effective by its action of creating pores within the nail structure. It also helps disrupt any fungal debris or biofilms that may occupy the nail and be difficult for the antimicrobial agent to penetrate. The ultrasound created pores will increases the transport of the antimicrobial gas or vapor into and through the nail structure. When both the ultrasound stimulation and the antimicrobial agent are

concurrently delivered, it results in effective diffusion of the antimicrobial agent along a pressure path created by the ultrasound into the inner, less accessible parts of the nail region, enabling the antimicrobial agent to penetrate deeply and allowing significant reduction or elimination of the fungal infection in all components of the nail region.

[00018] One device embodiment includes a positioning element to position the treatment elements over a nail affected by onychomycosis. The positioning element can be a clip designed to fit over a toe or finger in a manner similar to clips worn on the finger for use in pulse oximetery. The clip includes two hinging portions with a spring to keep the clip closed around the toe or finger. The clip may contain one or more ultrasound piezoelectric elements mounted in the clip and a gas source with tubes directing the antimicrobial gas onto the nail when the clip is worn. The apparatus may include an on/off switch, a power source and control circuitry for controlling the operation of the treatment elements, all or part of which may be incorporated into the clip or be located externally.

[00019] In another embodiment, the invention includes a method of treating antimicrobial infections such as onychomycosis, in an individual. The method provides a treatment element, a positioning element a control circuit and a power source; positioning the treatment element over a nail affected by onychomycosis; and operating the device to treat onychomycosis. The treatment element may comprising a nonthermal plasma source, an ultrasound piezoelectric element or both.

[00020] In accordance with one embodiment of the invention, a nonthermal plasma generator promotes delivery of an antimicrobial agent both over and through the nail and to the nail bed where the nonthermal plasma effluent or gas composition acts as an antimicrobial agent.

Another aspect may be an electrically operated ultrasound stimulation device suitable to facilitate delivery of the antimicrobial agent into and through the nail plate. In an alternate embodiment, the antimicrobial agent is preferably concurrently delivered to at least one of the cuticle, the nail folds, the lunula, the matrix, and the hyponychium utilizing ultrasound stimulation to assist the antimicrobial agent into the nail to treat onychomycosis.

[00021] In another embodiment, a method for treatment may include a first application of ultrasound stimulation to the nail for increasing porosity of the nail, then a second treatment of the antimicrobial agent to the nail surface for killing the fungus and spores both on, under and within the nail. A preferred method for treatment is when ultrasound stimulation and antimicrobial agent are concurrently delivered to at least one of the cuticle, the nail folds, the lunula, the matrix, and the hyponychium where the ultrasound-assisted plasma gas antimicrobial agent cures the treated nail fungal infection.

[00022] The methods and embodiments described above can achieve markedly better cure rates, improvement and/or prophylaxis in the treatment of onychomycosis than the current oral and topical administration of drugs. This method enables one to bypass the systemic route used by oral medication and its concurrent systemic side effects by placing the medication exactly where it is needed.

[00023] Any of the embodiments described herein may be used alone or together with one another in any combination. Inventions encompassed within this specification may also include embodiments that are only partially mentioned or alluded to. Other features, benefits, and advantages of the invention will be apparent upon a review of the detailed disclosure, including the specification, drawings, and claims. BRIEF DESCRIPTION OF THE DRAWINGS

[00024] The novel features of the invention are set forth with particularity in the claims that follow. A better understanding of the features and advantages of the present invention will be obtained by reference to the following detailed description that sets forth illustrative

embodiments, in which the principles of the invention are utilized, and the accompanying drawings of which:

[00025] The accompanying figures, in which like reference numerals refer to identical or fiinctionally-similar elements throughout the separate views and which are incorporated in and form a part of the specification, further illustrate the present invention and, together with the detailed description of the invention, serve to explain the principles of the present invention.

[00026] FIG. 1A and IB illustrate a microbial therapy method separated into two treatment steps to kill the fungus within the nail;

[00027] FIG. 2 illustrates a device configuration where ultrasound stimulation and the direct application of the antimicrobial agent are applied simultaneously;

[00028] FIG. 3 A and 3B illustrate a second constructible therapy device where the ultrasound stimulation and the indirect application of the antimicrobial agent are applied simultaneously;

[00029] FIG. 4 is a block diagram that illustrates combining a nonthermal plasma generator with a reactive oxygen species generator that can incorporate additional reactive oxygen species into the effluent output that is sent to a treatment chamber;

[00030] FIG. 5 illustrates a perspective side view showing a toe being inserted into an antimicrobial therapy toe clip device;

[00031] FIG. 6 illustrates a perspective view of a toe fully inserted into an antimicrobial therapy toe clip device;

[00032] FIG. 7A illustrates a superficial view of a healthy nail.

[00033] FIG. 7B illustrates a cross-sectional view of the nail in FIG. 7A.

[00034] FIG. 7C shows a longitudinal section of the nail in FIG. 7A

[00035] FIG. 8 schematically illustrates the proximal area of the nail structure.

DETAILED DESCRIPTION

[00036] The following description is presented to enable one of ordinary skill in the art to make and use the invention and is provided in the context of a patent application and its requirements. Various modifications to the preferred embodiment and the generic principles and features described herein will be readily apparent to those skilled in the art. Thus, the present invention is not intended to be limited to the embodiment shown but is to be accorded the widest scope consistent with the principles and features described herein. [00037] Embodiments of the present invention provide apparatus' and methods for creating an ultrasound-assisted, nonthermal plasma generated effluent that has antimicrobial properties. The embodiments described can be employed for general wound therapy and/or, specifically, for skin or nail treatments. For example, the ultrasound-assisted nonthermal plasma can penetrate the toenail or fingernail and kill the primary microorganism that causes onychomycosis.

[00038] Aspects of the present invention include a treatment device that provides nonthermal plasma gas, reactive oxygen gas or vapor, ultrasound stimulation or a combination thereof to a patient. A nonthermal plasma generator alone, a ultrasound transducer alone, a reactive oxygen generator alone or some combination of these is herein referred to as a "treatment element". There are multiple technologies that have been used for nonthermal plasma generation at atmospheric pressure to which the present invention may be applicable. Nonthermal plasma gas at atmospheric pressure have been generated by a microwave-induced plasma systems, dielectric barrier discharge, corona discharge, and atmospheric pressure plasma jet. U.S. Patent No.

7572998 describes some representative, but not exclusive, plasma generators to which the present invention may be applied. This patent is hereby expressly incorporated by reference herein in its entirety for all purposes. The nonthermal plasma generator is electrically connected to both a power supply and electrical control circuit to control both the duration and intensity of the plasma gas effluent.

[00039] The active content of the nonthermal plasma effluent can include singlet oxygen (Ό ₂ ), hydroxide (OH), hydrogen peroxide (H ₂ 0 ₂ ), ozone (0 ₃ ), nitrous oxide (N ₂ 0), nitrogen dioxide (N0 ₂ ) and other excited molecules of air constituents that includes other reactive nitrogen and reactive oxygen species. Charged particles, electric fields and UV light are also generated. The ionized gases last for very short periods of time (less than a second), but free radicals and reactive oxygen and nitrogen species that are electrically neutral last long enough to be effective many meters away from the source in destroying fungus, bacteria and their spores. These free radicals denature critical lipid, protein and nucleic acid contents of the microbes, ultimately causing cell death. Research has demonstrated the effectiveness of plasma gas as well as nitrogen and oxygen free radicals such as ozone or hydrogen peroxide vapor or a combination of these in causing retarded fungal growth and fungal death. The byproduct of the plasma process is water (H ₂ 0) and carbon dioxide (C0 ₂ ).

[00040] The nonthermal plasma gas effluent can be directed to the treatment site in two primary configurations: Direct mode or Indirect mode. The direct mode configuration puts the treatment area within the visible plasma region discharge or plume, between 0.0 mm and 5.0 mm for most apparatus'. In direct mode, UV light, charged particles and electric fields, in addition to reactive neutral species, can directly reach the treatment surface. The indirect mode configuration will have the treatment surface between 5.0 mm and 1.0 meter away so that the charged particles and electric field will have dissipated or decayed and not reach the treatment surface. The indirect mode configuration may benefit from a gas delivery system, where the nonthermal plasma gas effluent is directed to the treatment surface through tubing, inline fans, connectors and ports for input, delivery and output of the antimicrobial agent to the treatment surface.

[00041] For plasma gas application in the indirect mode configuration, an optional porous material, element, or member may be utilized for optimal distribution of the gas across the treatment area, such as a nail treatment site. It can be manufactured from any suitable material which assists the spread of the gas across the entire treatment area (e.g. nail infection site) and is inert to the reactive species so that the antifungal agent concentration remains constant. Suitable materials include, but are not limited to foam, woven material, non-woven material, sponge, polymers such as hydrogels, conducting material, non-conducting material, paper, cardboard, plastic, synthetic materials, natural materials, fabric, porous metals, porous glass, or a combination thereof. Preferably, the material of the base is made from polyester. Optionally, the base can be made up of a plurality of materials, which can be stacked or connected in a co-planar way by any suitable attachment means.

[00042] The treatment can occur for a predetermined period and the antimicrobial agent can be applied for a sufficient time to achieve an effective killing of the microbes within the target treatment zone such as fungus in the nail structure or bacteria in an open wound. For example, a sufficient time for application is a time from about 10 seconds to about 4 hours. In one embodiment, antimicrobial agent is applied for a time from about 1 minute to about 15 minutes. In yet another embodiment, antimicrobial agent is applied for a time from about 5 minutes to about 20 minutes. In a further embodiment, antimicrobial agent is applied for a time from about 30 minutes to about 1 hour. In other embodiments, the treatment application may be cyclical in nature where the electrical control circuit will cycle the device on and off for a predetermined period of time, for example a 50% duty cycle (1 minute on l\ minute off) for a 30 minute to 1 hour treatment period.

[00043] The amount of active antimicrobial agent entering or permeating the treatment site will depend on such factors as its polarity, structure, antimicrobial activity, treatment duration and penetration rate into the nail. The therapeutically effective amount may vary depending on the subject and the severity of the affliction. Generally, the amount of the active antimicrobial agent in the composition may be any amount effective for kill the infecting microorganism. In some cases, the amount of active antimicrobial agent is between about 0.1 parts per million to about 1000 parts per million.. [00044] In addition, or alternatively, a gas or vapor may be used with or independently from the nonthermal plasma gas to provide antimicrobial treatment. As described above, the efficacy of topical creams, gels, and liquids is heavily dependent on accessibility to target treatment sites.

For example, an infected nail can have an asymmetric porous structure created in the nail by the fungus as it consumes the keratin. The asymmetric structure creates many passive bottlenecks for liquid flow due to abrupt widening of the flow path. A fluid that approaches a widened flow path actively stops flow because of capillary forces. Antifungal liquids, gels and creams cannot get into nail channels, chambers, and crevices infected by the fungus. Similarly, tissue debris and other exudates at a wound site impedes the perfusion and permeation of a topical liquid/gel into the infected areas. Likewise, biofilms can form on top or within the wound, blocking access to the infection site by antimicrobial agents.

[00045] Advantageously, a gas-based topical antimicrobial treatment can permeate into each crack, crevice and chamber without overcoming capillary forces. In an optimal environment, the gas has the additional advantage of maintaining a dry breathable treatment site. Layering a liquid or cream on top of a wound hinders gas transport to the site, which slows healing and can actually provide a moist, dark environment conducive to microbial proliferation.

[00046] In additional embodiments, the devices and methods described also provide for ultrasound assisted wound and infection therapy. Although gas, plasma, and vapor therapy are more effective at reaching infection sites compared to traditional topical creams and liquids, another way to improve efficacy is by reducing pathway obstruction at the treatment site.

[00047] In another aspect, delivery of the antimicrobial agent to the treatment site is improved by using ultrasound stimulation to increase permeability of the site. For example, for nail fungal treatment, a second barrier to topical liquid antifungals and possibly topical gas-based antifungals is also created by the fungus. Sometimes there is not a crack, crevice or chamber to penetrate because the fungus has created a barrier in one of two ways. The keratin debris formed by the fungus can create blockages in any passage that stops the penetration of the antifungal treatment. Also, the fungus can form biofilms, a community of fungus encompassed by a self- created extracellular matrix. These extracellular matrix supplies an improved structure for the colony that holds some of the nutrients from the surrounding environment and offers protection from other microbial forces, any host response and efficiently resists the action of antifungals.

[00048] Ultrasound stimulation can be used to break down passageway obstructions and barriers preventing medicant penetration. The ultrasound stimulation enhances transport by cavitation, acoustic pressure, and heating; all physical phenomena that will speed up the process of mass transfer. Ultrasound waves that propagate into the treatment area result in alternating high-pressure (compression) and low-pressure (rarefaction) cycles. During the low-pressure cycle, high-intensity ultrasound waves create small vacuum bubbles or voids in the liquid. When the bubbles attain a volume at which they can no longer absorb energy, they collapse violently during a high-pressure cycle. This phenomenon is termed cavitation. The resulting shear forces of cavitation can create pores within the treatment area, disrupt cell envelopes mechanically and improve material transfer.

[00049] In the case of the infected nail structure, ultrasound induced cavitation can create pores, ingress passage, and turbulent (chaotic) air flow within the nail structures that will break up the keratin debris and biofilms and allow for adequate penetration of the antifungal gas throughout the nail structure.

[00050] Similarly, for wound treatment, the ultrasound stimulation can improve permeability of an antimicrobial agent by unblocking or creating access points in the wound site. For example, ultrasound cavitation can create pores in biofilms formed at a surface of a wound to allow a gas antimicrobial agent to penetrate through the biofilm into an infection site within the wound.

[00051] Ultrasound stimulation is generated by the phenomenon known as the piezoelectric effect, in which high frequency, alternating, electric current is applied across a quartz or silicone dioxide crystal, or across certain other polycrystalline materials such as lead- zirconate- titanate (PZT) and barium titanate to create rhythmic deformation. As the piezoelectric crystal undergoes rhythmic deformation due to electric current, it can produce ultrasonic vibrations. An apparatus that can create ultrasound stimulation is an ultrasound transducer. A generator that is connected to the ultrasound transducer generate a current that vibrates the ultrasound transducer. See, for example, U.S. Pat. No. 6,385,487, the disclosure of which is incorporated herein by reference.

[00052] Ultrasound stimulation to improve delivery of an antimicrobial agent to a treatment site would preferably be in the vibration range from 10 kHz to 15 MHz. More optimally between 40 kHz and 1.5 MHz. Previous research has shown that low-frequency transducers are better suited for drug delivery because they result in more cavitation. Ultrasound assisted mass transfer is also dependent on ultrasound intensity and the intensity would preferably be in the range from 10 mW/cm ² to 7 W/cm ² . More preferably are intensities from 0.5 W/cm ² to 3 W/cm ² to optimally improve nail permeation to the nonthermal plasma gas that functions as the antimicrobial agent.

[00053] The ultrasound stimulation can be applied for a sufficient time to achieve an effective amount of antimicrobial agent permeation. For example, a sufficient time for application is a time from about 10 seconds to about 60 minutes. In one embodiment, the ultrasound stimulation duration is from about 1 minute to about 15 minutes. In yet another embodiment, ultrasound stimulation is applied for a time from about 5 minutes to about 20 minutes. In a further embodiment, ultrasound stimulation is applied for a time from about 10 minutes to about 30 minutes.

[00054] An optional interfacing material can be used as an acoustic coupling layer to improve impedance matching between the ultrasound transducer and a treatment site such as the nail. The material can form a layer between the ultrasound transducer and the nail to efficiently couple the ultrasound stimulation energy into the nail. Without derogating from the generality of the optional interfacing materials, one example of an interfacing material may be a polyacrylamide gel. Another example would be a hydrogel material or a cellular polypropylene polymer foam material that would both be appropriate acoustic coupling layers. It is noted that nothing herein is meant to restrict an interfacing material or its placement location and placement can be on an area comprising both skin and nail, such as on a margin of the nail.

[00055] In some cases, an index matching material/gel/fluid may be used to facilitate the delivery of the ultrasound stimulation to treatment site. The index matching material may be selected based on the properties of the antimicrobial agent used.

[00056] FIG. 1 illustrates the method of applying the antimicrobial therapy separated into two treatment steps. FIG. I B illustrates the first step. Initially, the nail 100 is treated with an ultrasound stimulation apparatus 105 that includes an electrical generator 1 10 and an ultrasound transducer 1 15. Typically, but not required, is a polyacrylamide gel material 120 placed between the nail and the ultrasound transducer to act as an acoustic coupling layer. After ultrasound stimulation treatment, the ultrasound stimulation apparatus is removed from the nail.

[00057] FIG. IB illustrates the second treatment step using a nonthermal plasma generator 125 in a direct mode configuration where the plasma flow nozzle 130 is brought to the nail 100 surface and the plasma effluent output 135 coats the nail surface. For this application, the nonthermal plasma generator has an air or gas composition input 140 within the generator body 145 that also encloses an electrode 150 and quartz tube 155 for generating the plasma plume or plasma effluent output 135.

[00058] Any sequence of application of an antimicrobial agent in the form of plasma effluent output and ultrasound stimulation is possible according to the present invention, including, but not limited to the following additional options: (1) The only step needed for antimicrobial disinfection and cure is the application of an antimicrobial agent in the form of plasma effluent output onto the nail; or (2) The first treatment step involves the application of an antimicrobial agent in the form of plasma effluent output to the nail and the second treatment step is the application of ultrasound stimulation; or (3) The simultaneous treatment of the nail with plasma effluent output and ultrasound stimulation in a single treatment step and is further explained in connection with FIG. 2-FIG. 3 herein.

[00059] Although shown as a nail in FIGS. 1A- 1 B, it is to be appreciated, that the methods described herein can apply to any treatment site. The devices and methods described can be modified for other anatomical locations without detracting from the contemplated embodiments.

[00060] Additionally, the antimicrobial agent may be a plasma gas or, alternatively, a gas or vapor that is not a ionized gas species. The gas or vapor can be, for example, a reactive oxygen gas that provides radicals for interacting with microbial targets. As such, an antimicrobial gas/vapor may be applied to disinfect and treat a wound site.

[00061] The antimicrobial gas agent may be used in conjunction with UV light, ultrasound, or any of the other features described herein. For example, ultrasonic stimulation may be applied at any time during the treatment cycle. Ultrasound may be used to create pores in biofilms or tissue debris prior to, during, or after gas application to enhance penetration of the antimicrobial agent at the infection site and permeability to the antimicrobial agent. Alternatively, the gas or vapor treatment may be applied without a complementary application of ultrasound, especially in situations where enhanced penetration is not needed at an accessible treatment site.

[00062] FIG. 2 illustrates a direct mode antimicrobial device 200 that is a combination of a nonthermal plasma generator 205 and an ultrasound stimulation apparatus 210 that is cylindrical in shape and has an opening 215 at it's center that allows for passage of the plasma effluent output 1205. The ultrasound stimulator 210 includes first end 1206 and a second end 1208. The first end 1206 is adapted for delivering both an ultrasonic stimulation and a gas/plasma gas to the treatment site. As shown, the second end 1208 is configured to couple or interface with a plasma generator 205. In some cases, the plasma generator includes a lumen 1210 that is aligned with the ultrasound stimulator to deliver plasma gas to the treatment site through the ultrasound stimulator 210. In some embodiments, the plasma gas generator includes a distal port that aligns with the opening 215 on the ultrasound stimulator 210.

[00063] In further embodiments, a fan or compressed air cylinder or pump for moving air or gas composition through the plasma generator 205 and into the ultrasound stimulator are incorporated. The plasma generator 205 may include a delivery port 1204 for receiving and delivering the gas flow to the plasma generator to generate plasma effluent output 1205.

[00064] As can be appreciated, the device 200 may be dimensioned for a specific treatment site. As shown, the device 200 has a nail facing surface that has a width or diameter

dimensioned for application to the nail area. In some cases, the nail facing surface has a width between about 0.1mm to about 10mm. Additionally, the lumens or openings on device 200 may be designed to optimize the amount of plasma delivered to the treatment site. In a direct mode, the lumen 1205 of the generator may have a length between about 0.1mm to about lm to minimize dissipation of the active components in the plasma.

[00065] In some embodiments, the ultrasound stimulation 220 and plasma effluent output 135 are applied to the nail/treatment site 100 simultaneously; however, this is not required.

[00066] FIG. 3 illustrates another embodiment of a combination antimicrobial treatment element 300 for the nail 100 with the nonthermal plasma effluent delivered in an indirect mode configuration. The FIG. 3A shows a side view where the nonthermal plasma generator is separate from the treatment element 300 and the plasma effluent is delivered to a porous spreading element 305 by a delivery tube 310 connected to a gas inlet port 325. FIG. 3B shows a top view where the power source and control circuitry of the ultrasound transducer 315 is also separate from the treatment element 300 but is in electrical communication by virtue of wiring 320.

[00067] As shown treatment element 300 includes an ultrasound stimulator 315 that has a surface 1302 adapted for interfacing with a treatment site. The surface 1302 can be dimensioned for application of the ultrasonic stimulation to a particular anatomical location such as the nail. Additionally, the surface 1302 may be designed to directly engage or contact the treatment site, or, alternatively, indirectly engage the site via a coupling gel.

[00068] Additionally, treatment element 300 may include as part of its assembly a gas or plasma generator that is not directly located at the treatment site. The plasma or gas generator may be in fluid communication with the treatment element 300 through any suitable means including the tubing element 310. Moreover, the plasma or gas generator may be located at about 0.1mm to about lm from the treatment element 300.

[00069] In further embodiments, the porous element 305 may be positioned around the ultrasound transducer 305. In such embodiments, the porous element 305 spreads the antimicrobial agent around or against a treatment site. In some cases, the porous element 305 may cover some or a portion of the surface 1302. In other embodiments, the porous element 305 is a bandage or gauze material that is configured to be worn around a treatment site.

[00070] FIG. 4 illustrates a block diagram 400 combining a nonthermal plasma generator 405 with a reactive oxygen species generator 410 that will incorporate additional reactive oxygen species into the effluent output in a treatment chamber. The resulting antimicrobial agent generated by these two components is directed to a treatment chamber 415 by a main fan 420 through delivery tubing 425. An exhaust fan 430 pulls off the treatment gas and scrubs it through an air filter 435 for a final exhaust output of air and water vapor. In contrast to the devices of FIG. 1-FIG. 3, this alternative configuration for an antimicrobial device has an oxygen radical generator added into the system so that it enriches the antimicrobial effluent that treats the nail. Embodiments that are possible but not limited to are the addition of a hydrogen peroxide solution bubbler as the reactive oxygen species generator or an ozone generator as the oxygen radical addition to the effluent flow.

[00071] The treatment chamber may be a bootie-shaped liner or bag that may be worn on the foot where the entire bag may be replaced after each use. The bootie may contain an array of ultrasound piezoelectric elements across its top-front position that face towards the inserted toe nails along with an array of gas inlet ports for delivery of the antifungal agent over the nails. The bootie may also incorporate a porous member between the gas source and foot for better spreading of the gas over the entire nail area. The bootie may be made from any flexible and/or elastomeric material that is capable of conforming to a digit and positioning the treatment element over the nail area. The apparatus may also include a power source and control circuitry for controlling the operation of the device, all or part of which may be incorporated into the bootie or be located externally.

[00072] Alternatively, the device shown in FIG. 4 may only include a plasma generator or the reactive gas species. In such variations, the treatment chamber provides the antimicrobial gas agents with or without additional treatment modalities such as UV light and ultrasound.

[00073] FIG. 5 illustrates a perspective side view showing a patient's toe being inserted into an antimicrobial therapy toe clip 500. The toe clip acts as a positioning element with the first (or upper) housing 505 that contains the treatment element pivoted with respect to the second (or lower) housing 510 to enlarge the opening 515 to enable easy insertion of the toe of the patient and easy positioning of the treatment element over the nail 100. Spring 520 provides a counter force which brings the two housings to a closed position after the pivot force is released. The nonthermal plasma generator is, in this embodiment, separate from the antimicrobial therapy toe clip 500 and the plasma gas effluent is delivered to the clip by delivery tubing 525 that attaches to the toe clip through a gas inlet port 530.

[00074] It should be understood that the location of the various electronic elements in these embodiments is a matter of design choice. For example, the electrical connections could be located in the first housing or the circuit board could be located in the first housing.

Alternatively, in one embodiment, the primary control circuitry could be located externally in a control box that is connected to the antimicrobial therapy toe clip by both wires and tubes.

Power for the control box is provided by a standard plug-in cable. The important consideration for use with this embodiment is that the plasma effluent output and ultrasound stimulation be directed at the infected nail for treatment.

[00075] FIG. 6 illustrates a perspective view of a toe fully inserted into the antimicrobial therapy toe clip 600 after the user has positioned the device and released the pivot force. The spring 615 causes the first housing 605 and the second housing 610 to uniformly grip the inserted toe targeted for antimicrobial treatment.

[00076] In some of the embodiments disclosed herein, the terms "treatment" and

"antimicrobial" have sometimes been used. For purposes of this specification, the term

"treatment" encompasses any treatment of a microbial infection such as onychomycosis, and includes: preventing the disease from occurring in a subject which may be predisposed to the disease but has not yet been diagnosed as having it; inhibiting the disease, i.e., arresting its development; and/or curing or relieving the disease, i.e., causing regression of the disease with both microbiological and clinical improvement. "Antimicrobial" is defined as tending to destroy microbes, prevent their development or inhibit their pathogenic action and includes but is not limited to antibacterial, antiviral and antifungal properties.

[00077] As can be appreciated, although the devices and methods may be described using nail anatomy or disease as exemplary indication/target for treatment, the embodiments contemplated are not limited to the nail, fungal infections, or the specific anatomical structures described. Rather, the embodiments provide for at least three treatment modalities including (1) gas/vapor/plasma antimicrobial treatment for infections or wound therapy; (2) ultrasound assisted therapy for gas/vapor treatment; and (3) gas/vapor/plasma antimicrobial treatment for onychomycosis with or without ultrasound stimulation.

[00078] The apparatus' and methods above has been described in general terms as an aid to understanding details of preferred embodiments of the present invention. In the description herein, numerous specific details are provided, such as examples of components and/or methods, to provide a thorough understanding of embodiments of the present invention. Some features and benefits of the present invention are realized in such modes and are not required in every case. One skilled in the relevant art will recognize, however, that an embodiment of the invention can be practiced without one or more of the specific details, or with other apparatus, systems, assemblies, methods, components, materials, parts, and/or the like. In other instances, well- known structures, materials, or operations are not specifically shown or described in detail to avoid obscuring aspects of embodiments of the present invention. It is to be understood that other variations and modifications of the embodiments of the present invention described and illustrated herein are possible in light of the teachings herein and are to be considered as part of the spirit and scope of the present invention.