Sanford H. Feldman

A NOVEL MYCOPLASMA DETECTED IN ASSOCIATION WITH UPPER RESPIRATORY DISEASE SYNDROME IN FREE-RANGING EASTERN BOX TURTLES (TERRAPENE CAROLINA CAROLINA) IN VIRGINIA

Clinical signs of upper respiratory tract disease-like syndrome (URTD-LS) were observed in free-ranging eastern box turtles (Terrapene carolina carolina) from Virginia, USA (May 2001–August 2003), some of which also had aural abscesses. After a Mycoplasma sp. was detected by polymerase chain reaction (PCR), a study was undertaken to better define the range of clinical signs of disease and to distinguish mycoplasma-associated URTD-LS from other suspected causes of URTD-LS and aural abscessation in box turtles. Nasal and/or ocular swabs (from turtles possessing URTD-LS) or nasal washes (from asymptomatic turtles) were collected from turtles May 2001–August 2003; samples were assayed for Mycoplasma spp., chelonian herpesvirus, and iridoviruses by PCR testing. A partial DNA sequence (933 bases) of the small ribosomal subunit (16S rRNA) of the box turtle Mycoplasma sp. was analyzed to determine its phylogenetic relatedness to other Mycoplasma spp. of veterinary interest. Mycoplasma sp. was detected in seven (six with clinical signs of URTD-LS; one asymptomatic) of 23 fortuitously collected animals from six of 11 Virginia counties. Clinical signs in Mycoplasma sp.–infected animals included unilateral to bilateral serous to mucopurulent nasal discharge, epiphora, ocular edema, and conjunctival injection. Five Mycoplasma sp.–positive animals possessed aural abscesses; two did not. Analysis of the mycoplasma 16S rRNA gene sequence from one asymptomatic and three symptomatic animals representing four counties revealed a consensus Mycoplasma sp. sequence closely related to, but distinct from, M. agassizii. None of the samples collected contained viral DNA of chelonian herpesviruses or invertebrate and vertebrate (including FV3) iridoviruses. In conclusion, a new Mycoplasma sp. was associated with URTD-LS in native box turtles from Virginia that was not codetected with other suspected causes of chelonian upper respiratory disease; there was no proof of a direct relationship between aural abscessation and the Mycoplasma sp.

Phylogenetic Analysis of Avian Poxviruses Among Free-Ranging Birds of Virginia

Abstract Polymerase chain reaction was used to amplify a portion of the avian poxvirus core 4b gene of infected free-ranging birds that presented at the Wildlife Center of Virginia during the 2003 and early 2004 years. The species of bird infected were a great blue heron (Ardea herodias), two American crows (Corvus brachyrhyncos), two American robins (Turdus migratorius), two mourning doves (Zenaida macroura), a red-tailed hawk (Buteo jamaicensis), a blue-gray gnatcatcher (Polioptila caerulea), a northern mockingbird (Mimus polyglottos), a house finch (Carpodacus mexicanus), and a northern cardinal (Cardinalis cardinalis). Phylogenetic analysis was performed using the consensus sequences determined for each avian case in Virginia in combination with avian poxvirus core 4b gene sequence from isolates previously described in Europe and that of vaccinia virus. Alignment of DNA sequences identified areas of point mutations and, in the case of a single mourning dove, the incorporation of a triplet of nucleotides. Maximum-likelihood analysis grouped the 2003–2004 Virginia avian poxviruses into a clade distinct from those reported in European free-ranging birds, with the exception of a single case in a mourning dove that clustered within one European clade. The cladogram that resulted from our analysis of the European isolates is in agreement with those previously published. This study identified a distinct clade of avian poxvirus unique from four clades previously described and associated with epornitics in free-ranging birds, where the core 4b gene DNA sequence has been the basis of comparison.

PHYLOGENETIC CLASSIFICATION OF THE FROG PATHOGEN AMPHIBIOTHECUM (DERMOSPORIDIUM) PENNERI BASED ON SMALL RIBOSOMAL SUBUNIT SEQUENCING

We determined 1,600 base pairs of DNA sequence in the 18S small ribosomal subunit from two geographically distinct isolates of Dermosporidium penneri. Maximum likelihood and parsimony analysis of these sequences place D. penneri in the order Dermocystida of the class Mesomycetozoea. The 18S rRNA sequences from these two isolates only differ within a single region of 16 contiguous nucleotides. Based on the distant phylogenetic relationship of these organisms to Amphibiocystidium ranae and similarity to Sphaerothecum destruens we propose the organism be renamed Amphibiothecum penneri.

Molecular phylogeny of the pinworms of mice, rats and rabbits, and its use to develop molecular beacon assays for the detection of pinworms in mice

Though pinworm infestation has been prevalent since the early years of laboratory animal medicine, the genomes of these parasites have not yet been sequenced. The authors used high-fidelity polymerase chain reaction to amplify a large portion of the ribosomal gene complex of four pinworm species commonly found in lab rodents and rabbits (Aspiculuris tetraptera, Passalurus ambiguus, Syphacia muris and Syphacia obvelata). They determined DNA sequences for these complexes and carried out phylogenetic analysis. Using this information, the authors developed real-time molecular beacon assays for pinworm detection, comparing the new diagnostic approach with traditional methods such as perianal tape testing, fecal flotation and direct examination of intestinal content.

Endoluminal Negative-Pressure Therapy for Preventing Rectal Anastomotic Leaks: A Pilot Study in a Pig Model

BACKGROUND Anastomotic leak after rectal resection carries substantial morbidity and mortality. A diverting ileostomy is beneficial for high-risk anastomoses, but its creation and reversal carry a surgical risk in addition to that of resection itself. We sought an alternative method for managing complications of rectal anastomosis. METHODS We developed an endoluminal negative-pressure technology with a diverting proximal sump, and hypothesized that it would close anastomotic disruptions in pigs. We performed rectal resections on pigs, with primary anastomoses and the creation of an anastomotic defect. In animals in the treatment group we inserted an endoluminal negative-pressure device and kept it at a low level of continuous suction for 5 d. No device was inserted in a control group of animals. After the 5-d period of treatment we evaluated the anastomoses in both the treatment and control groups of animals for leakage, using contrast enemas. Specimens of anastomosed rectum were evaluated histologically for mucosal integrity and for the location and density of inflammatory responses. RESULTS Fourteen pigs were assigned to either the treatment (n=10) or control (n=4) group. Of the pigs in the treatment group, 90% had complete closure of their rectal defect, as compared with 25% of the animals in the control group (χ(2) test, p=0.04). The animals in the treatment group had only minimal mucosal and serosal inflammation, whereas those in the control group had extensive mucosal damage with associated serositis. CONCLUSIONS Endoluminal negative-pressure therapy was well-tolerated and led to successful closure of 90% of the anastomic rectal defects in the treatment group of animals in the present study. Additional evaluation of this therapy is warranted.

Association of Mycoplasma corogypsi and Polyarthritis in a Black Vulture (Coragyps atratus) in Virginia

On 10 October 2007, a Black Vulture (Coragyps atratus) was presented to the Wildlife Center of Virginia, Waynesboro, Virginia, USA, because of an inability to fly. Examination revealed multiple swollen, fluctuant joints. The bird suffered from lead toxicosis and had a prominent leukocytosis. Histopathologic evaluation revealed an acute fibrinoheterophilic polyarthritis, and results of routine aerobic and anaerobic culture of joint fluid were negative, although Mycoplasma sp. sequence-specific polymerase chain reaction was positive. Amplification of a portion of the 16S rRNA and subsequent phylogenetic analysis of the amplicon identified Mycoplasma corogypsi. This is the first report of polyarthritis being diagnosed in association with a Mycoplasma sp. in a vulture species. However, fulfilling Koch’s postulates through experimental infections is required to draw conclusions concerning an etiologic diagnosis.

Delayed Endoluminal Vacuum Therapy for Rectal Anastomotic Leaks after Rectal Resection in a Swine Model: A New Treatment Option

Anastomotic leaks are a dreaded surgical complication following colorectal operations. Creation of a temporary proximal diverting ileostomy is used in high‐risk anastomoses, however, additional surgical risk is accumulated with its creation and reversal. Endoluminal vacuum therapy has been shown to seal anastomotic defects in the prophylactic setting in a pig model and we hypothesized it could be utilized in a delayed fashion to rescue subjects with an active anastomotic leak. Yorkshire pigs underwent rectal resection, intentional leak confirmed by fluoroscopy, and endoluminal vacuum therapy device placement to low continuous suction. Following treatment, a contrast enema and necropsy was performed for gross and histopathology. Pigs underwent 2 (or 5) days of free intraperitoneal leak prior to device placement and 5 (or 7) subsequent days of endoluminal vacuum therapy. Six of seven early‐treated pigs sealed their anastomotic defect, while two of the four treated pigs in this extended group sealed the defect. Endoluminal vacuum therapy is feasible and well tolerated in a pig model, and it has been shown to seal a significant number of freely leaking anastomoses in the early period (86%). This technology warrants further study as it may provide a noninvasive means to treatment of anastomotic leaks.

Detection of pathogenic Batrachochytrium dendrobatidis using water filtration, animal and bait testing

The pathogen Batrachochytrium dendrobatidis (Bd) can be challenging to detect at endangered amphibian reintroduction sites. Pre-release Bd detection can be confounded by imperfect animal sampling and the absence of animals. In Study 1, we used historical Bd-positive sites, to concurrently evaluate water filtrates and mouth bar (tadpoles) or skin swab (caudates) samples for Bd using molecular beacon realtime PCR. In Study 2, during a natural outbreak, we used PCR to detect Bd from zoospore-attracting keratin baits (three avian, three snake species). In Study 1, no captured animals (n=116) exhibited clinical signs, although 10.6% were positive, representing three of seven species sampled. In contrast, 5.4% of water filters (n=56) were Bd-positive. In Study 2, after short incubation times, a single duck down feather tested Bd-positive. In conclusion, Bd was detected in asymptomatic amphibians and water filtrate at two sites, and from water only, at two other sites. With continued refinement, semi-quantitative Bd water filtrate screening could better define zoospore-specific disease risk, allowing better characterization of the free-living phase of the organisms life cycle. Finally, these results suggest wild aquatic birds (e.g., waterfowl) should be systematically explored as a means of Bd spread. Since large numbers of aquatic birds migrate, even low Bd transfer rates could be a significant means for disease dissemination.

Opisthotonus, torticollis and mortality in a breeding colony of Anolis sp. lizards

On clinical examination, the affected anoles presented with a reluctance to move, tremors of the distal appendages and postural distortion of the axial skeleton, including hyperextension (Fig. 1a) or twisting and serpentine distortion (Fig. 1b). Because of the acute onset of these presentations in multiple individuals, the differential diagnosis included infectious disease, metabolic disease, nutritional deficiency and toxicosis. Gross necropsy of affected lizards was unremarkable with the exception of gout affecting multiple joints. Microscopic and bacteriologic evaluations were not done. What do you think is the cause of the problems? Is it infectious, metabolic, toxic or nutritional? How would you investigate? How would you treat it?

Endoluminal Vacuum Therapy for Ivor Lewis Anastomotic Leaks: A Pilot Study in a Swine Model

Anastomotic leaks are a serious complication associated with Ivor Lewis esophagectomies. Endoluminal negative pressure vacuum devices create a possible treatment alternative to conventional surgical intervention. Ten pigs had an intrathoracic esophageal anastomosis with a 1‐cm defect. The experimental group had the device placed intraoperatively across the defect, whereas the control group did not. Once treatment was completed, a contrast fluoroscopic study and necropsy was performed. All control pigs had contrast extravasation on fluoroscopy and contamination on necropsy. The experimental group had no radiologic leak and no contamination on necropsy. The P value for leak is 0.03. This study demonstrated that endoluminal negative pressure vacuum therapy is tolerated in the swine model and is successful in facilitating the healing of anastomotic leaks. Endoluminal negative pressure vacuum therapy has potential clinical benefits, including decreased morbidity and length of hospital stay.