Evaluation of the Clinical Efficacy of Pradofloxacin Tablets for the Treatment of Canine Pyoderma
A third-generation fluoroquinolone, pradofloxacin (PRA), is currently being developed to treat bacterial infections in dogs. The purpose of this study was to assess the clinical efficacy in 20 dogs affected with superficial and deep pyoderma. An initial aerobic skin culture was performed in dogs with superficial pyoderma; aerobic/anaerobic tissue culture was performed in dogs with deep pyoderma; and skin cytology and biopsies were obtained from all dogs. Pradofloxacin (approximately 3 mg/kg per os [PO]) was administered daily to all dogs. Clinical efficacy was recorded at 4 weeks for dogs with superficial pyoderma and at 3 and 6 weeks for dogs with deep pyoderma. At a mean dosage of 3.7 mg/kg PO once daily, PRA treatment resulted in an excellent to good clinical response within 3 to 6 weeks for all 20 dogs with superficial and deep pyoderma.
Introduction
Bacterial pyoderma is one of the most common causes of canine skin disease in small animal practice.1,2 Classification of pyoderma is traditionally based on depth of infection (superficial versus deep), and it commonly develops secondary to underlying causes.2 Secondary infections may result from hypersensitivities, endocrinopathies, ectoparasite infestation, or immunological conditions.2–8 Lesions typical of superficial pyoderma include follicular pustules, papules, epidermal collarettes, and alopecia.2,9 Clinical signs of deep pyoderma such as furunculosis and cellulitis are clinically evident as nodules, fistulae, scarring, and/or hemorrhagic bullae.2,9,10
Staphylococcus (S.) intermedius is the most common pathogen isolated in canine pyoderma.1,11,12 In addition to S. intermedius, chronic deep pyoderma may be associated with other pathogens such as Pseudomonas aeruginosa, Escherichia coli, Proteus spp., Bacteroides spp., Peptostreptococcus spp., and Fusobacterium spp.1,2,13,14
Treatment of both generalized superficial and deep pyoderma involves administration of systemic antibiotics for 2 to 3 weeks beyond clinical resolution of lesions, which may require 4 to 12 weeks of initial therapy.1,2,15–17 However, treatment failures are reported due to canine skin pathogens increasingly exhibiting resistance to many classes of antibiotics, including fluoroquinolones, beta-lactams, macrolides, and sulfas.16–21
Pradofloxacin (PRA) is a novel third-generation fluoroquinolone specifically developed for veterinary medicine with enhanced in vitro activity against a wide range of gram-positive, gram-negative, and anaerobic veterinary pathogens.22–24 Pradofloxacin has already demonstrated efficacy in the treatment of canine pyoderma and wound infections in clinical trials in Europe.10,25 For example, one study reported that clinical remission was obtained in 86% of dogs with deep pyoderma, with a mean treatment duration of 34 days.10 The dose used in this study was 3 mg/kg.
The bioavailability of orally administered PRA is approximately 100% in dogs.26 Following an oral dose of 3 mg/kg, a maximum serum concentration (Cmax) of 1.26 μg/mL is reached in 2.1 hours (Tmax), with an elimination half-life (T1/2) of 6.9 hours.27 The minimum inhibitory concentration (MIC) of PRA for 90% of S. intermedius isolates (MIC90) is 0.06 μg/mL, which is at least two to four times more active than other veterinary fluoroquinolones.22 At the time of writing this manuscript, the Clinical and Laboratory Standards Institute (CLSI) in the United States does not have approved breakpoint values for PRA. Breakpoints, also known as interpretive criteria, are discriminatory antimicrobial concentrations (in mg/L or μg/mL or a zone diameter in mm) used in the interpretation of results of susceptibility testing to define isolates as susceptible, intermediate, or resistant. In dogs, PRA has low in vitro plasma protein binding (29% to 37%),28 similar to enrofloxacin; this is important, because free drug concentrations often correlate well with antibacterial activity.29 Furthermore, PRA has been shown to have higher concentrations in the skin than in serum.27 Tissue concentration (compared to serum concentrations alone) is likely a better indicator for predicting potential efficacy of a drug.
Pradofloxacin is well tolerated and possesses a wide margin of safety when used according to dosage recommendations. The occurrence of adverse reactions was low in all of the clinical trials performed to date.10,25–27,30 The safety of PRA was demonstrated in dogs when it was administered at up to three times the recommended dosage (i.e., 9 mg/kg).27 The side effects observed in dogs treated with PRA included diarrhea (1/66 dogs) and polydipsia/polyuria (1/66 dogs).25 Previously reported adverse effects for other veterinary fluoroquinolones include gastrointestinal disturbances (e.g., nausea, transient vomiting, diarrhea, or mild changes in feces); however, these adverse effects usually only occur at higher doses, are not serious, and do not require discontinuation of therapy.25,31,32
In Europe and North America, PRA is under development for the treatment of canine pyoderma at the dosage of 3 mg/kg per os (PO) q 24 hours. The drug is indicated for the treatment of skin, soft tissue, respiratory, and urinary tract infections associated with susceptible gram-positive, anaerobic, and gram-negative bacterial organisms.10,17,21,23,25,30,33 The purpose of this study was to determine, in an open study design, the efficacy of PRA tablets in the treatment of naturally occurring superficial and deep pyoderma in dogs.
Materials and Methods
Dog Selection
Between June 1, 2004, and November 1, 2006, 20 privately owned adult dogs of any breed, weight, or sex that were referred to the dermatology service at the Veterinary Medical Teaching Hospital, University of California-Davis, for clinical signs consistent with superficial and/or deep pyoderma were enrolled in the study. The diagnosis was based on clinical, cytological, and histopathological features of pyoderma as described.2,9 The study was approved by the university’s Animal Care and Use Committee, and all owners signed a consent agreement.
Inclusion and Exclusion Criteria
Dogs were excluded from the study if, within 14 days prior to enrollment in the study, they had received systemic or topical antibiotic therapy, systemic or topical antifungal therapy, oral antihistamines, or killed bacterial products used to stimulate the immune system (e.g., Staphage Lysatea). Oral and/or topical glucocorticoid therapy was not allowed for at least 4 weeks prior to the study and 6 weeks prior to the study if long-acting injectable glucocorticoids (e.g., methylprednisolone acetate, triamcinolone acetonide) were administered. Dogs younger than 12 months (small/medium breeds) or 18 months of age (large/giant breeds), breeding animals, and pregnant/lactating females were excluded from the study because of the risk of fluoroquinolones causing an arthropathy in young, rapidly growing animals.34
Concurrent medications allowed during the study included heartworm and flea preventive products, topical flea treatments, prescription diets, nonsteroidal antiinflammatory drugs, vitamin/mineral or fatty acid supplements, and vaccines. Medications (e.g., thyroid supplements, cardiovascular medications, etc.) to control underlying medical conditions were permitted. Shampoos with coat-conditioning or hypoallergenic properties were allowed during the study. German shepherd dogs with pyoderma and dogs with demodicosis and any other underlying etiology of pyoderma were included in the study.
Experimental Protocol
On initial examination (day 0), the dogs’ general and dermatological histories were obtained; these included previous episodes of pyoderma, preexisting conditions, concurrent therapies, and antimicrobial treatment within the previous year. A complete dermatological examination was performed, and specific clinical signs or lesions indicative of pyoderma were documented. The affected body sites were recorded on the dorsal and ventral views of a schematic dog. All dogs had the following diagnostics performed: (1) acetate tape cytology of lesional skin (e.g., pustules, papules, crusts, epidermal collarettes, nodules, fistulae); (2) one to three 6-mm skin punch biopsies of lesional skin submitted to the histopathology service; (3) aerobic culture and antimicrobial sensitivity collected using a sterile, dry swab rolled over one epidermal collarette12 (in cases presenting with superficial pyoderma); and (4) a 6-mm skin biopsy obtained using aseptic technique and submitted for aerobic and anaerobic tissue culture (if the dog had clinical signs of deep pyoderma). Susceptibility was measured by MIC determination at the microbiology laboratory at the University of California-Davis Veterinary Medical Teaching Hospital by use of broth microdilution techniques in accordance with the CLSI. Additionally, S. intermedius isolates were sent to Microbial Research Incorporatedb for PRA MIC testing (6 months after completion of the study protocol for retrospective analysis, as PRA MIC testing was not available during the study) by use of broth microdilution techniques in accordance with the CLSI document M31-A2. The bacterial isolates were frozen and stored at −80°C in a sterile vial containing porous beads.c
All dogs had histopathological evidence of pyoderma on the basis of established criteria.9 Disease was further classified as deep pyoderma if evidence of deep folliculitis (at the level of isthmus and below), furunculosis, or cellulitis was present on skin biopsy specimens.9 Underlying etiology of the pyoderma was diagnosed by use of the above-listed standard diagnostic methods. Atopic dermatitis was diagnosed by criteria established by Willemse35 and Prelaud et al.36 A hypoallergenic diet trial was used to diagnose food hypersensitivity.5 Identification of the organisms and response to ectoparasite treatment were used to diagnose sarcoptic mange, demodicosis, and flea allergy dermatitis.37–39 Hypothyroidism was diagnosed via detection of low total thyroxine (T4), free thyroxine, and a high concentration of thyroxine-stimulating hormone,40 compared to reference ranges (1.0 to 3.6 μg/dL, 1.0 to 3.5 ng/mL, and 0 to 0.6 ng/mL, respectively) established at the veterinary microbiology laboratory at the University of California-Davis Veterinary Medical Teaching Hospital.
Adequate control of hypothyroidism was defined as having a T4 (4 to 6 hours postpill) at the upper end of the reference range40,41 determined within the 3-month period prior to presentation. Underlying diseases were defined as diseases that, when treated successfully in combination with oral antibiotics, resulted in resolution of the pyoderma or a decrease in the relapse rate of the pyoderma.
Included dogs were prescribed an approximate dose of 3 mg/kg PRA PO q 24 hours for 28 days in cases of superficial pyoderma, and for 42 days in cases of deep pyoderma. Dogs were returned for follow-up assessment of clinical progress at 28 days for superficial pyoderma and at 21 and 42 days for deep pyoderma.
Treatment Evaluation
At each follow-up visit, adverse effects associated with PRA administration were recorded, a complete dermatological examination was performed, acetate tape cytology of lesional skin was performed, and clinical efficacy was assessed based on appearance of lesions. Clinical resolution of lesions was defined as the total disappearance of all lesions, including erythema and scaling. Efficacy was considered excellent if >75% of lesions were resolving, good if 50% to 75% of lesions were resolved, and a failure if <50% of lesions had resolved. An additional 14 to 28 days of PRA were prescribed accordingly. If a dog was considered a treatment failure, PRA was discontinued, and the bacterial skin culture and skin biopsy were repeated.
Results
Study Participants
Twenty dogs (eight dogs with a deep pyoderma and 12 dogs with a superficial pyoderma) completed the study. Population characteristics are shown in Tables 1 and 2. Thirteen dogs were male (eight castrated), and seven were female (five spayed). Ages ranged from 2 to 11 years (mean 5.9 years). Body weights on day 0 of treatment ranged from 8.2 to 51 kg (mean 33.1 kg). All dogs were presented for chronic, recurrent pyoderma that was previously unresponsive to treatment with various systemic antibiotics. Seventeen dogs (i.e., all dogs except case nos. 2, 12, and 17) had received oral antibiotics within the 12-month period prior to enrollment in this study. Case no. 17 had intermittently received a topical otic medication containing gentamicin sulfate, betamethasone valerate, and clotrimazole,d in both ears during the 2-year period prior to participating in this study. Three dogs (case nos. 9, 10, and 19) had preexisting hypothyroidism that was adequately controlled on thyroid supplementation. Case no. 4 was diagnosed with hypothyroidism and started on thyroid supplementation after completion of the PRA study protocol (i.e., 42 days after initiation of PRA).
Underlying diseases diagnosed in included dogs have also been summarized in Tables 1 and 2. Allergic hypersensitivity dermatitis (i.e., flea allergy, food allergy, and/or atopic dermatitis) was diagnosed in 10 (50%) of the 20 dogs. An additional six of the 20 dogs had allergic dermatitis with a concurrent underlying disease (e.g., hypothyroidism, demodicosis, actinic keratosis); thus, 16 dogs had underlying allergic dermatitis with or without concurrent conditions.
Histopathology
According to histological criteria, eight dogs were categorized as having deep pyoderma. Case no. 1 had demodicosis and atopic dermatitis. Case no. 6 had severe, chronic, pyogranulomatous deep dermatitis and panniculitis (acid-fast stain was negative; the broad-spectrum immunohistochemical marker for bacterial or fungal organisms [bacillus Calmette–Guerin] was negative; Brown and Brenn [B&B] gram stain and Periodic acid-Schiff [PAS] stain for bacterial and fungal organisms, respectively, were also negative). Case no. 7 had intralesional dermatophyte endospores and hyphae. Case no. 8 had allergic dermatitis, mild actinic keratosis, and severe, nodular, superficial and deep dermatitis with furunculosis (special stains including B&B and PAS were negative for bacterial and fungal organisms, respectively).
Twelve dogs met the histological criteria for superficial pyoderma and included case no. 14 with demodicosis; case no. 10 with actinic keratosis, hypothyroidism, and focal furunculosis; and case no. 11 with allergic dermatitis, folliculitis, focal furunculosis, and calcinosis cutis. The latter two dogs were initially enrolled in the protocol for dogs with superficial pyoderma according to clinical lesions (despite the later discovery of focal lesions of deep pyoderma on histopathological examination).
Skin Cytology
All 20 dogs had initial skin cytology results consistent with bacterial pyoderma. All dogs had reduction or complete resolution of bacterial counts. Skin cytology results are shown in Table 3.
Bacterial Culture and Antimicrobial Susceptibility
Deep Pyoderma.
Anaerobic tissue cultures were negative for all dogs except case no. 3, in which small numbers of Peptostreptococcus anaerobius were cultured (susceptibility panel not performed). Staphylococcus intermedius was isolated in pure culture from case nos. 1, 2, 4, and 5. In addition to S. intermedius, case nos. 3 and 6 had very small numbers of Streptococcus canis growth (sensitivity panel not performed). Case nos. 7 and 8 had no aerobic bacterial growth from tissue culture. Thus, S. intermedius was isolated from six of eight dogs. Details of the culture results (including PRA MIC results) obtained on day 0 are shown in Table 4.
Superficial Pyoderma.
Staphylococcus intermedius was isolated in pure culture from nine of 12 dogs (case nos. 9 to 17). Methicillin-resistant, coagulase-negative Staphylococcus spp. was isolated in small numbers from case no. 18. Pseudomonas aeruginosa was cultured from an intact pustule in case no. 19. Case no. 20 had a negative bacterial culture. Details of the culture results (including PRA MIC results) obtained on day 0 of the study are shown in Table 5.
Clinical Results
Pradofloxacin dosages were calculated and rounded up to the nearest tablet size; this resulted in a PRA dosage range of 3.0 to 4.6 mg/kg, with a mean dosage of 3.7 mg/kg. Of the eight dogs diagnosed with deep pyoderma, one had an excellent response, and seven had a good response after 21 days of PRA administration. The dog with the excellent response at 21 days went on to have complete resolution by 42 days. Of the other seven dogs, three had excellent responses, and three had good responses after 42 days of PRA administration. The last dog had a flare of allergic dermatitis and subsequent pyoderma diagnosed 42 days after initiating PRA administration. Treatment outcomes for dogs with deep pyoderma are listed in Table 1.
Of the 12 dogs diagnosed with superficial pyoderma, two had complete resolution, nine had excellent response, and one had a good response after 28 days of PRA administration. An excellent clinical response was obtained within 28 days of treatment for case no. 17 despite the reported in vitro resistance to enrofloxacin, marbofloxacin, ciprofloxacin, and orbifloxacin. An excellent clinical response was also achieved in case no. 18 with methicillin-resistant, coagulase-negative Staphylococcus sp. (enrofloxacin and orbifloxacin susceptible) and in case no. 19 with multiantibiotic-resistant P. aeruginosa (enrofloxacin susceptible; orbifloxacin intermediate susceptibility) within 28 days of treatment. Treatment outcomes for dogs with superficial pyoderma are listed in Table 2.
Adverse Effects
All adverse events reported by the owners were mild. One dog had one episode of vomiting on the second day of PRA administration along with 2 days of diarrhea. Treatment was not required, and these signs resolved completely by the fourth day of PRA administration. On the second day of PRA administration, one dog started having soft stools at the end of an initially normal bowel movement. This condition resolved upon completion of the PRA trial. A third dog reportedly had feces that were occasionally lighter in color during PRA treatment.
Discussion
In this study, all 20 dogs diagnosed with either superficial or deep pyoderma had good, excellent, or complete responses from 21 to 42 days after initiating PRA. Within 28 days of treatment, nine of 12 dogs diagnosed with superficial pyoderma had excellent responses, and two of the 12 dogs had complete resolution. Recurrence of the superficial pyoderma within 14 days after cessation of oral antibiotic therapy with PRA was not seen in any of the 12 dogs.
While 12 dogs included in this study had superficial pyoderma based on histopathological evaluation, two of these dogs also had focal evidence of moderate (case no. 10) to severe (case no. 11) furunculosis. Both of these dogs had an excellent clinical response within 28 days of PRA treatment but required an additional 14 to 28 days of treatment with PRA for complete resolution of the deep pyoderma.
Case no. 13, a geriatric dog referred to the dermatology service, had a 2-year history of severe pruritus and recurrent pyoderma that had not previously responded to cephalexin antibiotic therapy. These confounding factors likely produced multiple changes in the skin of this dog that rendered it less able to resolve the pyoderma following antibiotic therapy. It is well documented in humans that geriatric patients have delayed healing of the skin and compromised macrophage function that lead to a decreased inflammatory response, a limited mitogenic response of keratinocytes and fibroblasts, reduced rates of collagen synthesis, and slower epithelialization.42–46 Case no. 13 had complete resolution of truncal lesions at 28 days of PRA treatment; however, the dog was only classified as having a good response because of a persistent abdominal fold intertrigo. Abdominal fold intertrigo (also known as skin fold dermatitis) cannot be resolved with systemic antibiotics. Correction of the anatomical defect (e.g., excess abdominal fold from obesity, as in case no. 13) is necessary for a cure. If correction of the anatomical defect is unattainable, long-term topical treatments will be required to remove surface organisms and the entrapped debris.2
Case no. 14 had a 2-year history of unresolved generalized demodicosis with a concomitant superficial pyoderma. Secondary pyoderma is common in dogs with demodicosis because of the underlying immunosuppression and mite infestation.2,38,47 This dog had an excellent response to PRA within 28 days of treatment.
Of the eight dogs with deep pyoderma, only one dog (case no. 2) had an underlying disease (flea allergy dermatitis) that was fully controlled and therefore allowed complete resolution of the deep pyoderma within 42 days of initiating PRA therapy. Case no. 6 had deep pyoderma secondary to a primary immune-mediated inflammatory process (nodular dermatitis and panniculitis). After 42 days of PRA treatment alone, the pyoderma substantially improved as evidenced by marked reduction in lesion size and lesional exudate. A complete resolution of the skin disease was only achieved in case no. 6 when immunosuppressive therapy was initiated. Case no. 7 had intralesional dermatophyte endospores and hyphae, demodicosis, and atopic dermatitis leading to deep pyoderma. During the study protocol, this dog was treated with PRA and systemic antimite treatment but no antifungal therapy. After 42 days of PRA treatment, clinical efficacy was noted as excellent. Case no. 8 had a 3-year history of allergic pruritus in addition to large cutaneous furuncles that would rupture periodically. Histopathological examination in this case revealed deep dermal furunculosis, chronic granulation tissue and scarring, and evidence of actinic keratosis. These confounding factors impeded complete resolution of skin disease with oral antibiotics alone. Nonetheless, while the underlying skin disease was unable to be resolved, the secondary pyoderma showed a good response to PRA therapy. Lastly, case no. 4 did not begin treatment (thyroid supplementation) for profound hypothyroidism until 42 days after initiation of PRA therapy. This dog had an excellent response to PRA treatment alone within the 42 days.
Bacterial culture-negative dogs (case nos. 7, 8, and 20) were not excluded, as the diagnosis of superficial and/or deep pyoderma routinely is made by board-certified dermatologists and primary care clinicians via clinical lesions, skin cytology, and/or histopathological features of pyoderma.1,48 In the authors’ experience, negative bacterial cultures occur regularly in the sampling of superficial and deep pyoderma. Skin scrapings, cytological examination, and skin biopsy are the most consistently useful diagnostic procedures in the evaluation of suspected canine pyoderma.1,49,50 Bacterial culture traditionally is used primarily to guide antimicrobial therapy, and a negative culture does not preclude the diagnosis of pyoderma. False-negative bacterial cultures can result from numerous factors, including improper culture technique (e.g., inappropriate lesion selection; surgically scrubbing the site [for superficial lesion] prior to obtaining superficial culture), clinical sampling error (e.g., the age of the individual lesion sampled; sampling of a secondary versus primary lesion), and/or microbiology lab error (e.g., inappropriate growth conditions/growth media; transport error). Histopathological examination of skin biopsy samples may be useful in understanding the mechanism, etiology, and state of development of the folliculitis and furunculosis syndrome.48 Clinical response to PRA in all three bacterial culture-negative dogs lends additional credence to the cytological, clinical, and histopathological diagnosis of pyoderma.
Furthermore, per this study protocol, bacterial culture was not used to guide therapy selection, and instead results were recorded and correlated with final treatment outcome. In clinical practice, it is important to remember that the use of fluoroquinolones only be considered in cases where canine pyoderma has been refractory to appropriate “first line” antibiotics. Fluoroquinolones are most useful in the management of recurrent pyoderma and in chronic, deep pyoderma cases with extensive scar tissue.32
The clinical outcomes of these cases highlight the importance of determining and treating underlying causes of superficial and deep pyoderma in order to achieve complete resolution of pyoderma.1,2 Various underlying diseases predisposing dogs to the development of pyoderma have been described.3,5–8 Pruritic skin diseases, such as allergic hypersensitivities that were present in 16 of the 20 dogs in this study, result in self-trauma and damage to the barrier function of the epidermis.51 Furthermore, S. intermedius has been shown to adhere to corneocytes preferentially in dogs with atopic dermatitis.52
In the study reported here, culture of S. intermedius from 15 (75%) of 20 dogs with superficial and deep pyoderma is in agreement with results of previously reported studies,10–12 indicating that this organism is the most common canine skin pathogen. Interestingly, case no. 17 cultured positive for a methicillin-sensitive, fluoroquinolone-resistant S. intermedius. This dog had an excellent clinical response to PRA at 28 days. It is well known that in vitro susceptibility may not always correlate with in vivo clinical response. This may be due to individual variation regarding drug concentration, metabolism, absorption, and in vivo variance of the bacterium itself and its response to the drug. Case nos. 18 and 19 had methicillin-resistant, coagulase-negative Staphylococcus sp. and P. aeruginosa, respectively. Both of these dogs also had excellent clinical response to PRA within 28 days of treatment.
In this study, management of underlying conditions may have contributed to the beneficial response of the clinical efficacy of PRA. In order to limit the variables influencing assessment of PRA clinical efficacy, only three of the 20 dogs received a concurrent medication (e.g., ivermectin for demodicosis) not used prior to inclusion in the study. The other 17 dogs had either 1) a preexisting condition that was being treated long-term (with thyroid supplementation) and a chronic recurrent pyoderma despite these therapies, or 2) additional therapies were not initiated until after the PRA trial concluded (i.e., thyroid supplementation was administered for newly diagnosed hypothyroidism, or immunosuppressive therapy was given for nodular dermatitis and panniculitis). Coexistent diseases and lack of culture positivity do not detract from the usefulness of including these cases for the evaluation of PRA clinical efficacy. In clinical practice, treatment of pyoderma is commonly initiated while managing underlying disease processes. This study highlights the benefit of antibiotic therapy in such cases.
Conclusion
Based on results of this study, at a dosage of 3 mg/kg PO q 24 hours, PRA is a safe and highly efficacious treatment for canine superficial and deep pyoderma, regardless of underlying skin condition. This is in agreement with the results of a recently published study.10
Third-generation fluoroquinolones such as PRA have enhanced activity against gram-positive bacteria relative to first- and second-generation compounds, which differentiates PRA from earlier-generation fluoroquinolone compounds used in veterinary medicine. The efficacy of many antimicrobial agents is being threatened by a global increase in the number of resistant bacterial pathogens. Therefore, third-generation fluoroquinolones, such as PRA, clearly may have important utility in veterinary medicine as single-drug therapy for conditions caused by mixed aerobic/anaerobic infections.
Staphage Lysate; Delmont Laboratories, Inc., Swarthmore, PA 19081
Microbial Research Incorporated, Fort Collins, CO 80524
Prolab Diagnostics, Austin, TX 78754
Otomax; Schering-Plough Animal Health Corp, Union, NJ 07083
Acknowledgments
The authors thank the Department of Dermatology service technicians, Nina Toebe and Judy Schettler, for their invaluable help in organizing and aiding in collection of samples.
