LONEPINELLA SP. ISOLATED FROM WOUND INFECTIONS OF KOALAS.

We describe two cases of wound infections of koalas (Phascolarctos cinereus), one wild and one captive, in which Lonepinella-like organisms were involved. The wild adult koala was captured with bite wound injuries, as part of a koala population management program in Queensland, Australia. In both cases, there was evidence of physical trauma causing the initial wound. The captive koala suffered injury from the cage wire, and the wild koala had injuries suggestive of intermale fighting. Gram-negative bacteria isolated from both cases proved to be challenging to identify using routine diagnostic tests. The wound in the captive koala yielded a pure culture of an organism shown by whole genome sequence (WGS) analysis to be a member of the genus Lonepinella, but not a member of the only formally described species, L. koalarum. The wound of the wild koala yielded a mixed culture of Citrobacter koseri, Enterobacter cloacae and an organism shown by WGS analysis to be Lonepinella, but again not Lonepinella koalarum. Both cases were difficult to treat; the captive koala eventually had to have the phalanges amputated, and the wild koala required removal of the affected claw. The two Lonepinella isolates from these cases have a close relationship to an isolate from a human wound caused by a koala bite and may represent a novel species within the genus Lonepinella. Wound infections in koalas linked to Lonepinella have not been reported previously. Wildlife veterinarians need to be aware of the potential presence of Lonepinella-like organisms when dealing with wound infections in koalas, and the inability of commercial kits and systems to correctly identify the isolates.

Wound swabs versus biopsies to detect methicillin resistant Staphylococcus aureus in experimental equine wounds.

OBJECTIVE: To compare: (1) the load and diversity of cultivatable bacterial species isolated from tissue biopsies with cultures from surface swabs, and (2) the ability of each technique to detect methicillin-resistant Staphylococcus aureus (MRSA) in a model of MRSA-infected equine wounds. STUDY DESIGN: Experimental in vivo study. ANIMALS: Three light-breed adult horses. METHODS: Four 2.5 × 2.5 cm full-thickness skin wounds were created on the dorsolateral aspect of each forelimb. Five days later, each wound was inoculated with a pure culture of MRSA (ATCC 43300). One hundred microlitres of 0, 5 × 108 , 5 × 109 or 5 × 1010 colony forming units (CFU)/ml was used to inoculate each wound. Surface swabs (Levine technique) and tissue biopsy samples (3 mm punch biopsy) were obtained at 2, 7, 14, and 21 days after inoculation. Quantitative aerobic culture was performed using routine clinical techniques. RESULTS: A similar bacterial profile was identified from the culture of each wound-sampling technique and there was moderate correlation (R = 0.49, P < .001) between the bacterial bioburdens. Agreement was fair (κ = 0.31; 95% CI, 0.129-0.505) between the sampling techniques in identification of MRSA. Methicillin-resistant Staphylococcus aureus was isolated more frequently (P = .016) from cultures of tissue biopsies (79%; 76/96) than from surface swabs (62%; 60/96). CONCLUSION: Bacterial load and diversity did not differ between sampling techniques but MRSA was detected more often from the cultures of tissue biopsies. CLINICAL SIGNIFICANCE: Tissue biopsy should be preferred to culture swab in wounds where MRSA is suspected.

Electroceutical treatment of infected chronic wounds in a dog and a cat.

OBJECTIVES: To describe the use of an innovative printed electroceutical dressing (PED) to treat non-healing, infected chronic wounds in one dog and one cat and report outcomes. ANIMALS: A 4-year-old female spayed Mastiff and a 1-year-old female spayed domestic shorthair cat. STUDY DESIGN: Short case series. METHODS: Both cases had chronic wounds (duration: approximately 1 year for the dog and 6 3/4 months for the cat) that remained open and infected despite various wound management strategies. Both animals were treated with the PED. Observations from the records regarding wound size, antimicrobial susceptibility, and the time to healing were recorded. RESULTS: After 10 days of PED treatment in the dog and 17 days of PED treatment in the cat, the wounds had decreased in size by approximately 4.2 times in the dog and 2.5 times in the cat. Culture of punch biopsies yielded negative results. Wounds were clinically healed at 67 days in the dog and 47 days in the cat. No further treatment of the wounds was required beyond that point. CONCLUSION: Application of a PED led to closure of two chronic wounds and resolution of their persistent infection. CLINICAL SIGNIFICANCE: PEDs may provide a new treatment modality to mitigate infection and promote healing of chronic wounds.

Comparison of bacteriologic culture results for skin wound swabs and skin wound biopsy specimens.

OBJECTIVE: To compare bacteriologic culture results for superficial swab and tissue biopsy specimens obtained from dogs with open skin wounds. ANIMALS: 52 client-owned dogs. PROCEDURES: For each dog, 1 wound underwent routine preparation prior to collection of 2 specimens, 1 by superficial swab (Levine) technique and 1 by tissue biopsy. Specimens were processed for bacteriologic culture. Two observers determined whether any detected difference in culture results for the 2 types of specimen would have resulted in differing treatment plans. RESULTS: Culture results of swab and tissue biopsy specimens were identical in 11/52 (21.2%) cases. Tissue biopsy specimen and swab cultures yielded positive results for 44 (84.6%) and 40 (76.9%) wounds, respectively. With regard to mean recovery rates of bacteria from wounds with positive culture results, both the biopsy specimens and swabs yielded 3.4 bacterial species/wound. All wounds for which swab cultures yielded no growth also had negative culture results for biopsy specimens. Biopsy specimen and swab culture results were in agreement with regard to the most common bacteria cultured. In 7/52 (13%) wounds, the observers would have treated the patient differently on the basis of the results of the 2 cultures. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicated that culture of a swab collected by the Levine technique is an appropriate noninvasive alternative to culture of a tissue biopsy specimen. A negative result obtained from culture of a swab is likely to be reliable. Disagreement between the results of swab and tissue biopsy specimen cultures is likely of low clinical importance.

Effects of various wound dressings on microbial growth in perfused equine musculocutaneous flaps.

OBJECTIVE: To compare the effect of multiple wound dressings on microbial growth in a perfused equine wound model. SAMPLE: Abdominal musculocutaneous flaps from 16 equine cadavers. PROCEDURES: 8 full-thickness skin wound covered were created in each flap. Tissues were perfused with saline (0.9% NaCl) solution. Wounds were inoculated with methicillin-resistant Staphylococcus aureus (MRSA) or Pseudomonas aeruginosa (106 CFUs), incubated, and covered with a dressing containing activated charcoal, boric acid, cadexomer iodine, calcium alginate, manuka honey, nanoparticle silver, or polyhexamethylene biguanide or with a control (nonadherent gauze) dressing. Muscle biopsy specimens were obtained at baseline (immediately prior to dressing application) and 6, 12, 18, and 24 hours later for mean bacterial load (MBL) determination. The MBLs at each subsequent time point were compared with that at baseline within dressing types, and MBLs at each time point were compared among dressing types. RESULTS: MBLs in MRSA-inoculated wounds covered with cadexomer iodine dressings were significantly decreased from baseline at the 6- and 12-hour time points. For P aeruginosa-inoculated wounds, MBLs were significantly increased from baseline in all wounds at various times except for wounds with cadexomer iodine dressings. The MBLs of wounds with cadexomer iodine dressings were lower than all others, although not always significantly different from those for wounds with boric acid, manuka honey, nanoparticle silver, and polyhexamethylene biguanide dressings. CONCLUSIONS AND CLINICAL RELEVANCE: In this nonviable perfused wound model, growth of MRSA and P aeruginosa was most effectively reduced or inhibited by cadexomer iodine dressings. These results and the effect of the dressings on wound healing should be confirmed with in vivo studies.

Prospective evaluation of the influence of sampling method on bacterial culture results and antimicrobial selection in 52 dogs with infected wounds.

OBJECTIVE: To determine the influence of sampling methods on culture results and selection of antimicrobials for treating infected wounds in dogs. DESIGN: Prospective study from January to July 2016. SETTING: Three private multispecialty referral centers. ANIMALS: Fifty-two client-owned dogs with infected wounds. INTERVENTIONS: Each wound was sampled for culture by 3 methods: swabbing prior to preparation (dirty swabs), swabbing after debridement and sterile lavage (clean swabs), and biopsy. Bacterial species and phenotypes were compared. Three clinicians unaware of patient, wound, and sampling information selected antimicrobial drugs based on culture and sensitivity reports. Antimicrobials were divided into class I, II, or III based on established guidelines. The number, highest class of antimicrobial chosen, and inter-investigator agreement were examined. MEASUREMENTS AND MAIN RESULTS: Identical populations of bacteria were isolated for all 3 sampling techniques in only 31% of wounds. Significantly fewer bacterial species were isolated from biopsy samples (1.87 bacterial species per wound ± 1.14) than from clean swab samples (2.29 ± 1.18; P = 0.009) but not dirty swab samples (2.29 ± 1.29; P = 0.06). The recovery frequency for gram-positive bacteria was lower for biopsy compared to either swabbing technique (P = 0.001 for both comparisons). No difference was observed between clean and dirty swabbing techniques for any parameter examined. Sampling technique did not affect the proportion of wounds with anaerobic, gram-negative, or multi-drug resistant bacteria. The number (P = 0.28) and highest class of antimicrobial (P = 0.9) selected per wound did not differ between the 3 sampling techniques (P = 0.28). Clinician agreement was 83-90% depending on sampling technique. CONCLUSION: Although there were some differences in bacteria isolated from biopsy samples compared to swab samples from infected wounds, technique did not influence the number and highest class of antimicrobial selected by clinicians. Wound debridement prior to sampling by swabbing did not alter the number or type of bacteria isolated, nor the number or the highest class of antimicrobial selected by clinicians.

First Report on the Use of Larvae of Cochliomyia macellaria (Diptera: Calliphoridae) for Wound Treatment in Veterinary Practice.

Factors contributing to the delay or prevention of the natural wound healing process include infection and ineffective conventional treatment. Alternative therapies, such as the maggot debridement therapy (MDT), may be helpful for successful treatment in these cases. Aiming to disseminate information about the possibility of using other species of flies for wound treatment, besides the best known Lucilia sericata Meigen, 1826 (Diptera, Calliphoridae), we report here a successful MDT case with the application of Cochliomyia macellaria (Fabricius, 1775) larvae to treat an infected wound with extensive area of necrotic tissue in a dog. Five sterile larvae were applied to each square-cm of lesion and kept on the animal for only 48 h. The healing was successful, from both qualitative and quantitative points of view. The ratio of wound healing (RWH) reached almost 50% on the 5th day and 100% on the 14th day after MDT. Although the overall animal prognosis had been unfavorable, mainly due to the sepsis, the patient began to recover and had improved clinical condition from the fifth day after MDT. This study shows the importance and effectiveness of MDT in promoting faster and more complete healing of a complex wound.

Isolation of Neisseria dumasiana from a deep bite wound infection in a dog.

BACKGROUND: Neisseria spp. is a common inhabitant of the oral flora of cats and dogs; it is a potential cause of cutaneous infections in people secondary to animal bites. Neisseria dumasiana is a new species identified in the oral cavity of dogs. It has not been linked to cutaneous infections in people. OBJECTIVE: To describe the clinical and microbiological features of Neisseria dumasiana isolated from the skin of a dog. ANIMAL: A 3-year-old male neutered mixed breed dog from Florida, USA. METHODS AND MATERIALS: The dog had no travel history within or outside the United States; the dog was presented with draining tracts on the dorsum. Five months before the initial examination the dog was involved in a fight with another dog. RESULTS: An aerobic bacterial culture was performed and isolated multiple small, grey, moist, circular, convex and nonhaemolytic colonies. A Gram stain showed Gram-negative coccobacilli present in pairs. The bacteria were oxidase- and catalase-positive and negative for indole and fermentation of fructose, xylose, sucrose, mannitol and sorbitol. Polymerase chain reaction, using 16S rDNA sequencing, was positive for Neisseria dumasiana. Clinical and microbiological resolution occurred within three months of treatment. CONCLUSIONS AND CLINICAL IMPORTANCE: To the best of the authors' knowledge, this is the first report of a deep cutaneous dermatitis associated with Neisseria dumasiana in a dog. This case should stimulate veterinary clinicians and microbiologists to consider Neisseria spp. among the differential diagnoses of nonhealing wounds in dogs, particularly those associated with bite wounds.

Medical Therapy in Equine Wound Management.

Suitable use of prophylactic antimicrobial drugs for wounds depends on the accurate selection of appropriate antibiotics, dosing regimen, and duration of use. Regional intravenous delivery and intraosseous infusion of antibiotics are pivotal to a successful outcome for deep-rooted infections, inadequately perfused tissue, and infected wounds containing biofilm. Antibiotic-impregnated polymethylmethacrylate beads are predominantly helpful for wounds that have a poor blood supply and for those containing surgical implants that must remain in place.

Development of a High-Throughput ex-Vivo Burn Wound Model Using Porcine Skin, and Its Application to Evaluate New Approaches to Control Wound Infection.

Biofilm formation in wounds is considered a major barrier to successful treatment, and has been associated with the transition of wounds to a chronic non-healing state. Here, we present a novel laboratory model of wound biofilm formation using ex-vivo porcine skin and a custom burn wound array device. The model supports high-throughput studies of biofilm formation and is compatible with a range of established methods for monitoring bacterial growth, biofilm formation, and gene expression. We demonstrate the use of this model by evaluating the potential for bacteriophage to control biofilm formation by Staphylococcus aureus, and for population density dependant expression of S. aureus virulence factors (regulated by the Accessory Gene Regulator, agr) to signal clinically relevant wound infection. Enumeration of colony forming units and metabolic activity using the XTT assay, confirmed growth of bacteria in wounds and showed a significant reduction in viable cells after phage treatment. Confocal laser scanning microscopy confirmed the growth of biofilms in wounds, and showed phage treatment could significantly reduce the formation of these communities. Evaluation of agr activity by qRT-PCR showed an increase in activity during growth in wound models for most strains. Activation of a prototype infection-responsive dressing designed to provide a visual signal of wound infection, was related to increased agr activity. In all assays, excellent reproducibility was observed between replicates using this model.

Fatal Pulmonary and Cerebellar Zygomycosis due to Rhizomucor pusillus in a Ringed Seal (Pusa hispida).

A 4-year-old captive ringed seal (Pusa hispida) was treated with subcutaneous antibacterial injections for pus exuding wounds in the skin and associated blubber following a bite attack. Three months after the incident, the animal presented nystagmus and died the following day. At necropsy, there was a 25 × 18 × 25 mm well-delineated, opaque nodular mass in the lung, besides the skin ulcers and localized areas of discoloration in the blubber correlating with the bite wound and injection sites. Histopathology of the pulmonary mass demonstrated severe eosinophilic inflammatory infiltration among numerous intralesional fungal hyphae. The hyphae were irregularly branched, broad and aseptate, consistent of zygomycosis. Magnetic resonance imaging was conducted on the head, which was initially frozen intact, revealing diffuse areas of hyperintensity in the cerebellum. Restricted histopathologic examination of the cerebellum showed severe granulomatous inflammation well spread within the neuroparenchyma, associated with abundant intralesional fungal hyphae similar to those appreciated in the pulmonary mass. Molecular analyses of the fungi in the pulmonary and cerebellar tissue identified the etiologic agent in both sites as Rhizomucor pusillus. The likely route of infection is through inhalation of R. pusillus spores or fragmented hyphae from the environment that developed into an initial pulmonary infection, becoming the source of hematogenous dissemination to the cerebellum. The skin and blubber lesions likely contributed to immunosuppression. Zygomycosis is uncommon in pinnipeds, and the present report emphasizes the importance of considering zygomycete dissemination even when the primary focus is highly confined.

[Comparison of the antibacterial efficacy of polyhexanide, cold atmospheric argon plasma and saline in the treatment of canine bite wounds]. / Vergleich der Effizienz von Polyhexanid-Biguanid, Argon-Kaltplasma und Kochsalzlavage zur Dekontamination von Bisswunden beim Hund.

OBJECTIVE: To evaluate the bacterial contamination rate and to compare the efficacy of polyhexanide, cold argon plasma and saline at reducing bacterial bio-burden in dog bite wounds. MATERIAL AND METHODS: Dogs with bite-wound injuries were included when surgical debridement was pursued with subsequent treatment using either polyhexanide-biguanide lavage (A), cold argon plasma treatment (B) or saline lavage (C). Culture swabs were taken after debridement as well as after lavageor argon treatment. Statistical analysis was performed using the chi-square test. RESULTS: A total of 40 dogs were enrolled in the study (A: n = 12; B: n = 10; C: n = 18). The majority of injuries were minor and 87.5% of patients had positive bacterial culture results pre-lavage, with 19.8% of isolates classified as multidrug resistant. A reduction in wound bioburden was achieved in 8/12 patients in group A, 5/10 patients in group B and 14/18 patients in group C. Complete decontamination was achieved in 5/12 patients in group A, 2/10 in group B and 9/18 in group C. None of these differences were statistically significant nor associated with the development of complications. CONCLUSION: No statistically significant differences were detected between the treatment groups; however, the cold argon plasma treatment provided the least effective decontamination. CLINICAL RELEVANCE: Bite wounds yield a high rate of bacterial contamination, with increasing multidrug-resistance rates. Based on these preliminary results, no superior effect was detected for lavage using polyhexanidebiguanide or cold argon plasma.

Initial Management of Traumatic Wounds.

When traumatic wounds are quickly and accurately treated, morbidity and costs can be significantly decreased. Several factors, such as time delay between injury and treatment, the degree of contamination, extension and depth of the wound, and the mechanism of injury, influence the treatment and prognosis and stress the importance of a patient-specific approach. Although all traumatic wounds are contaminated, antibiotic therapy is seldom required if correct wound management is installed.

A quantitative swab is a good non-invasive alternative to a quantitative biopsy for quantifying bacterial load in wounds healing by second intention in horses.

The aim of this study was to evaluate different techniques for diagnosing wound infection in wounds healing by second intention in horses and to assess the effect of a vortex and sonication protocol on quantitative bacteriology in specimens with a histologically confirmed biofilm. In 50 wounds healing by second intention, a clinical assessment, a quantitative swab, a semi-quantitative swab, and a swab for cytology were compared to a quantitative tissue biopsy (reference standard). Part of the biopsy specimen was examined histologically for evidence of a biofilm. There was a significant, high correlation (P<0.001; r=0.747) between the outcome of the quantitative swabs and the quantitative biopsies. The semi-quantitative swabs showed a significant, moderate correlation with the quantitative biopsies (P<0.001; ρ=0.524). Higher white blood cell counts for cytology were significantly associated with lower log10 colony-forming units (CFU) in the wounds (P=0.02). Wounds with black granulation tissue showed significantly higher log10 CFU (P=0.003). Specimens with biofilms did not yield higher bacteriological counts after a vortex and sonication protocol was performed to release bacteria from the biofilm. Based on these findings, a quantitative swab is an acceptable non-invasive alternative to a quantitative biopsy for quantifying bacterial load in equine wounds healing by second intention.

Influence of intravenous regional limb perfusion with amikacin sulfate on Staphylococcus aureus bioburden in distal limb wounds in horses.

OBJECTIVE: To quantify the influence of intravenous regional limb perfusion (IVRLP) with amikacin on bacterial bioburden in limb wounds. STUDY DESIGN: In vivo, experimental. ANIMALS: Four adult horses. METHODS: Full thickness wounds created on each dorsal metacarpus were inoculated with Staphylococcus aureus. One forelimb was randomly assigned to cephalic vein IVRLP with amikacin on days 2, 3, and 4 post-wounding. Biopsies on day 2 prior to IVRLP, and on days 5 and 8 were cultured to quantify wound bioburden (CFU/g). Two horses repeated the study in a crossover design, receiving IVRLP on the opposite limb (n = 6 experiments). IVRLP was performed on both forelimbs simultaneously in 3 horses at the end of the study: limbs were perfused with a volume of new methylene blue equal to that of the amikacin, through the cephalic vein or palmar digital vein. After euthanasia, wounds were photographed to subjectively assess dye diffusion. RESULTS: The bioburden did not differ between control and IVRLP limbs overall or at any individual time point. No difference was detected between groups in terms of frequency of positive bacterial growth at any time. Methylene blue was visible in all excised tissues after IVRLP through the palmar digital vein, but was not visible in limbs perfused through the cephalic vein. CONCLUSION: IVRLP may not effectively concentrate amikacin within a wound bed and did not influence S. aureus bioburden in an experimentally created wound infection.

Antibacterial activities of Manuka and Honeydew honey-based membranes against bacteria that cause wound infections in animals.

INTRODUCTION: In this study, membranes composed of honey (Manuka or Honeydew) and pectin were developed, and the ISO 22196 method was used to evaluate their antibacterial activities against multidrug-resistant bacteria (i.e., Staphylococcus pseudointermedius, Escherichia coli, Proteus mirabilis, Pseudomonas aeruginosa) that cause wound infection in animals. The results demonstrated that both Manuka and Honeydew honey-based membranes had strong antibacterial activities against the strain of methicillin-resistant S. pseudointermedius tested. Specifically, membranes composed of Manuka honey were effective in inhibiting the growth of Gram-negative bacteria within 3 h, whereas those composed of Honeydew honey needed 24 h to neutralise bacterial growth. The antimicrobial activities of both membranes developed in this study suggest that they can be effectively used as wound dressing in veterinary clinical medicine.

Dans le cadre de cette étude, on a fabriqué des membranes à base de miel (miel de Manuka et miel de miellat) et de pectine et on a testé, selon le processus ISO 22196, leur activité antibactérienne sur des germes multirésistants provenant de blessures d'animaux (Staphylococcus pseudointermedius, E. coli, Proteus mirabilis und Pseudomonas aeruginosa). Les résultats montrent que les deux types de membranes ont une forte activité bactéricide sur les souches de Staphylococcus pseudointermedius résistantes à la méthicilline. Les membranes à base de miel de Manuka étaient également actives contre tous les germes gram négatifs ét réduisaient leur nombre en 3 heures, alors qu'un contact de 24 heures était nécessaire pour que les membranes à base de miel de miellat réduisent la croissance bactérienne. L'activité antibactérienne des membranes utilisées dans la présente étude justifie leur emploi dans la médecine vétérinaire clinique.

Postvaccination wounds associated predominantly with Arcanobacterium phocae in mink (Neovison vison) at three mink farms.

BACKGROUND: The emerging skin disease fur animal epidemic necrotic pyoderma (FENP) has been attributed to infection with Arcanobacterium phocae (ABP). The exact pathogenesis and risk factors of FENP have yet to be elucidated. ANIMALS: Three mink from each of three different mink farms (A-C) with postvaccination skin wounds at the vaccination site and six mink from an unaffected mink farm (D) that had used the same vaccine batch and vaccination site (hind leg). METHODS AND RESULTS: All mink from farms A-C had severe necrotizing to necropurulent dermatitis where they were vaccinated intramuscularly in the hind leg. ABP was the sole bacterium cultured from six of nine wounds. Using 16S-23S rDNA intergenic spacer region and BOX-PCR, the ABP isolates from these wounds were indistinguishable from isolates originating from several cases of FENP. CONCLUSIONS AND CLINICAL IMPORTANCE: This is the first report of FENP-like lesions at the site of vaccination, in the days following the procedure, associated with ABP. At farms with FENP vaccination, procedures should be considered carefully.