An outbreak of systemic chlamydiosis in farmed American alligators (Alligator mississippiensis).

Chlamydia spp are reported to causes systemic disease in a variety of hosts worldwide including few reports in crocodilians. Disease presentations vary from asymptomatic to fulminant disease, some of which are zoonotic. The aim of this study was to describe the pathological, immunohistochemical, and molecular findings associated with the occurrence of a previously unreported Chlamydia sp infection causing a major mortality event in farmed American alligators (Alligator mississippiensis). The outbreak presented with sudden death in juvenile alligators mainly associated with necrotizing hepatitis and myocarditis, followed by the occurrence of conjunctivitis after the initial high mortality event. The widespread inflammatory lesions in multiple organs correlated with intralesional chlamydial organisms identified via immunohistochemistry and confirmed by 23S rRNA-specific real-time quantitative polymerase chain reaction (qPCR) for Chlamydiaceae bacteria. By sequencing and phylogenetic analysis of the OmpA gene, this uncultured Chlamydia sp grouped closely with Chlamydia poikilothermis recently described in snakes. This study highlights the significance of such outbreaks in farmed populations. Enhanced epidemiological monitoring is needed to gain further insight into the biology of Chlamydia sp in alligators, disease dynamics, risk factors, and role of carrier animals.

Enhancing the one health initiative by using whole genome sequencing to monitor antimicrobial resistance of animal pathogens: Vet-LIRN collaborative project with veterinary diagnostic laboratories in United States and Canada.

BACKGROUND: Antimicrobial resistance (AMR) of bacterial pathogens is an emerging public health threat. This threat extends to pets as it also compromises our ability to treat their infections. Surveillance programs in the United States have traditionally focused on collecting data from food animals, foods, and people. The Veterinary Laboratory Investigation and Response Network (Vet-LIRN), a national network of 45 veterinary diagnostic laboratories, tested the antimicrobial susceptibility of clinically relevant bacterial isolates from animals, with companion animal species represented for the first time in a monitoring program. During 2017, we systematically collected and tested 1968 isolates. To identify genetic determinants associated with AMR and the potential genetic relatedness of animal and human strains, whole genome sequencing (WGS) was performed on 192 isolates: 69 Salmonella enterica (all animal sources), 63 Escherichia coli (dogs), and 60 Staphylococcus pseudintermedius (dogs). RESULTS: We found that most Salmonella isolates (46/69, 67%) had no known resistance genes. Several isolates from both food and companion animals, however, showed genetic relatedness to isolates from humans. For pathogenic E. coli, no resistance genes were identified in 60% (38/63) of the isolates. Diverse resistance patterns were observed, and one of the isolates had predicted resistance to fluoroquinolones and cephalosporins, important antibiotics in human and veterinary medicine. For S. pseudintermedius, we observed a bimodal distribution of resistance genes, with some isolates having a diverse array of resistance mechanisms, including the mecA gene (19/60, 32%). CONCLUSION: The findings from this study highlight the critical importance of veterinary diagnostic laboratory data as part of any national antimicrobial resistance surveillance program. The finding of some highly resistant bacteria from companion animals, and the observation of isolates related to those isolated from humans demonstrates the public health significance of incorporating companion animal data into surveillance systems. Vet-LIRN will continue to build the infrastructure to collect the data necessary to perform surveillance of resistant bacteria as part of fulfilling its mission to advance human and animal health. A One Health approach to AMR surveillance programs is crucial and must include data from humans, animals, and environmental sources to be effective.

Antimicrobial Susceptibility Patterns and Clinical Parameters in 208 Dogs with Positive Urine Cultures (2012-2014).

Urinary tract infections (UTI) occur commonly in dogs, and gram-negative enteric bacteria are the most prevalent pathogens. Clinical parameters, urinalysis, and urine culture and sensitivity results were retrieved from the medical records of 208 dogs with positive urine cultures over a 3 yr period at the Louisiana State University Veterinary Teaching Hospital. Several groups were defined including dogs presented for primary care versus referred cases; simple UTI, complicated UTI, and pyelonephritis; dogs pretreated with antimicrobials; and dogs having an indwelling catheter in place prior to sampling. Nearly 80% of dogs had complicated UTI. Of all dogs, 70% had no documented clinical signs of lower urinary tract disease (LUTD), with 68% of them showing hematuria and/or pyuria. Based on clinical signs or urinalysis, 19% of all dogs had no evidence of lower UTI. In dogs without LUTD signs the most common comorbidities were immunosuppressive treatment and severely restricted mobility (23%). Chronic recurring UTI were present in 19% of dogs with LUTD signs. Distribution of bacterial species was comparable with the existing literature and not significantly different between clinical subgroups. Isolates from dogs pretreated with antimicrobials showed decreased susceptibility to enrofloxacin. The prevalence of multidrug-resistant Escherichia coli and Staphylococcus spp. was moderate (29%).

Intramuscular Immunization of Mice with the Live-Attenuated Herpes Simplex Virus 1 Vaccine Strain VC2 Expressing Equine Herpesvirus 1 (EHV-1) Glycoprotein D Generates Anti-EHV-1 Immune Responses in Mice.

Vaccination remains the best option to combat equine herpesvirus 1 (EHV-1) infection, and several different strategies of vaccination have been investigated and developed over the past few decades. Herein, we report that the live-attenuated herpes simplex virus 1 (HSV-1) VC2 vaccine strain, which has been shown to be unable to enter into neurons and establish latency in mice, can be utilized as a vector for the heterologous expression of EHV-1 glycoprotein D (gD) and that the intramuscular immunization of mice results in strong antiviral humoral and cellular immune responses. The VC2-EHV-1-gD recombinant virus was constructed by inserting an EHV-1 gD expression cassette under the control of the cytomegalovirus immediate early promoter into the VC2 vector in place of the HSV-1 thymidine kinase (UL23) gene. The vaccines were introduced into mice through intramuscular injection. Vaccination with both the VC2-EHV-1-gD vaccine and the commercially available vaccine Vetera EHVXP 1/4 (Vetera; Boehringer Ingelheim Vetmedica) resulted in the production of neutralizing antibodies, the levels of which were significantly higher in comparison to those in VC2- and mock-vaccinated animals (P < 0.01 or P < 0.001). Analysis of EHV-1-reactive IgG subtypes demonstrated that vaccination with the VC2-EHV-1-gD vaccine stimulated robust IgG1 and IgG2a antibodies after three vaccinations (P < 0.001). Interestingly, Vetera-vaccinated mice produced significantly higher levels of IgM than mice in the other groups before and after challenge (P < 0.01 or P < 0.05). Vaccination with VC2-EHV-1-gD stimulated strong cellular immune responses, characterized by the upregulation of both interferon- and tumor necrosis factor-positive CD4+ T cells and CD8+ T cells. Overall, the data suggest that the HSV-1 VC2 vaccine strain may be used as a viral vector for the vaccination of horses as well as, potentially, for the vaccination of other economically important animals.IMPORTANCE A novel virus-vectored VC2-EHV-1-gD vaccine was constructed using the live-attenuated HSV-1 VC2 vaccine strain. This vaccine stimulated strong humoral and cellular immune responses in mice, suggesting that it could protect horses against EHV-1 infection.

Measuring the Level of Agreement Between Cloacal Gram's Stains and Bacterial Cultures in Hispaniolan Amazon Parrots ( Amazona ventralis ).

Cloacal or fecal Gram's stains and bacterial cultures are routinely performed during avian physical examinations to assess the microbial flora of the gastrointestinal tract. Although cloacal or fecal Gram's stains and bacterial cultures are considered routine diagnostic procedures, the level of agreement between the individual tests has not been determined. To investigate the level of agreement between results from Gram's stain and bacterial culture when used to assess cloacal or fecal samples from psittacine birds, samples were taken from 21 clinically healthy Hispaniolan Amazon parrots ( Amazona ventralis ) and tested by Gram's stain cytology and bacterial culture. Most bacteria (97.2%) identified by Gram's stain were gram positive. However, gram-negative organisms were identified in 7 of 21 (33.3%; 95% confidence interval: 13.3%-53.3%) birds. Escherichia coli was the only gram-negative organism identified on culture. Agreement between results of Gram's stain and culture was fair (weighted κ = 0.27). The results of this study suggest that Gram's stains and bacterial culture may need to be performed with a parallel testing strategy to limit the likelihood of misclassifying the microbial flora of psittacine patients.

Culex flavivirus and West Nile virus in Culex quinquefasciatus populations in the southeastern United States.

Little is known of the interactions between insect-only flaviviruses and other arboviruses in their mosquito hosts, or the potential public health significance of these associations. The specific aims of this study were to describe the geographic distribution, prevalence, and seasonal infection rates of Culex flavivirus (CxFV) and West Nile virus (WNV) in Culex quinquefasciatus Say in the Southeastern United States, investigate the potential association between CxFV and WNV prevalence in Cx. quinquefasciatus and describe the phylogenetic relationship among CxFV and WNV isolates from the Southeastern United States and around the world. Using ArboNET records, 11 locations were selected across Georgia, Mississippi, and Louisiana that represented a range of WNV human case incidence levels. Cx. quinquefasciatus were trapped weekly throughout the summer of 2009 and pools were screened for flavivirus RNA by reverse transcriptase polymerase chain reaction. Cx. quinquefasciatus from Georgia had significantly higher CxFV infection rates than either Mississippi or Louisiana. CxFV was not detected in Mississippi after July, and no CxFV was detected in Cx. quinquefasciatus in Louisiana. In Georgia, CxFV infection rates were variable between and within counties and over time. WNV infection rates were not significantly different across states or months, and WNV sequences from all three states were identical to each other in the envelope and NS5 gene regions. Phylogenetically, NS5 and E gene sequences from Georgia CxFV isolates clustered with CxFV from Japan, Iowa, and Texas. Multiple CxFV genetic variants were found circulating simultaneously in Georgia. No evidence was found supporting an association between WNV and CxFV infection prevalence in Cx. quinquefasciatus.

Factors associated with mosquito pool positivity and the characterization of the West Nile viruses found within Louisiana during 2007.

BACKGROUND: West Nile virus (WNV) is an arbovirus of public health importance in the genus Flavivirus, a group of positive sense RNA viruses. The NS3 gene has a high level of substitutions and is phylogenetically informative. Likewise, substitutions in the envelope region have been postulated to enable viruses to subvert immune responses. Analysis of these genes among isolates from positive mosquitoes collected in Louisiana illustrates the variation present in the regions and provides improved insight to a phylogenetic model. Employing a GIS eco-regionalization method, we hypothesized that WNV pool positivity was correlated with regional environmental characteristics. Further, we postulated that the phylogenetic delineations would be associated with variations in regional environmental conditions. RESULTS: Type of regional land cover was a significant effect (p < 0.0001) in the positive pool prediction, indicating that there is an ecological component driving WNV activity. Additionally, month of collection was significant (p < 0.0001); and thus there is a temporal component that contributes to the probability of getting a positive mosquito pool. All virus isolates are of the WNV 2002 lineage. There appears to be some diversity within both forested and wetland areas; and the possibility of a distinct clade in the wetland samples. CONCLUSIONS: The phylogenetic analysis shows that there has been no reversion in Louisiana from the 2002 lineage which replaced the originally introduced strain. Our pool positivity model serves as a basis for future testing, and could direct mosquito control and surveillance efforts. Understanding how land cover and regional ecology effects mosquito pool positivity will greatly help focus mosquito abatement efforts. This would especially help in areas where abatement programs are limited due to either funding or man power. Moreover, understanding how regional environments drive phylogenetic variation will lead to a greater understanding of the interactions between ecology and disease prevalence.

Detection of West Nile virus RNA in mosquitoes and identification of mosquito blood meals collected at alligator farms in Louisiana.

Since 2001, alligator farms in the United States have sustained substantial economic losses because of West Nile virus (WNV) outbreaks in American alligators (Alligator mississippiensis). Once an initial infection is introduced into captive alligators, WNV can spread among animals by contaminative transmission. Some outbreaks have been linked to feeding on infected meat or the introduction of infected hatchlings, but the initial source of WNV infection has been uncertain in other outbreaks. We conducted a study to identify species composition and presence of WNV in mosquito populations associated with alligator farms in Louisiana. A second objective of this study was to identify the origin of mosquito blood meals collected at commercial alligator farms. Mosquitoes were collected from 2004 to 2006, using Centers for Disease Control light traps, gravid traps, backpack aspirators, and resting boxes. We collected a total of 58,975 mosquitoes representing 24 species. WNV was detected in 41 pools of females from 11 mosquito species: Anopheles crucians, Anopheles quadrimaculatus, Coquillettidia perturbans, Culex coronator, Culex erraticus, Culex nigripalpus, Culex quinquefasciatus, Mansonia titillans, Aedes sollicitans, Psorophora columbiae, and Uranotaenia lowii. The blood meal origins of 213 field-collected mosquitoes were identified based on cytochrome B sequence identity. Alligator blood was detected in 21 mosquitoes representing six species of mosquitoes, including Cx. quinquefasciatus and Cx. nigripalpus. Our results showed that mosquitoes of species that are known to be competent vectors of WNV fed regularly on captive alligators. Therefore, mosquitoes probably are important in the role of transmission of WNV at alligator farms.

In vitro comparison of Staphylococcus pseudintermedius susceptibility to common cephalosporins used in dogs.

The in vitro activity of 10 cephalosporin antimicrobial agents against 75 isolates of methicillin-susceptible Staphylococcus pseudintermedius derived from dogs was assessed. The lowest minimal inhibitory concentration for 90% of strains (MIC90) values obtained were for cephalothin, cefovecin, and cefazolin (0.12 ug/mL), followed by ceftiofur and cefoxitin (0.25 ug/mL), cefpodoxime (0.5 ug/mL), and cefaclor and cefadroxil (1 ug/mL). The highest MIC90 values were found for cephalexin and cefixime (2 ug/mL). In this in vitro study, sensitivity to cephalothin was indicative of cephalexin susceptibility, although there were marked differences in MICs. Cephalothin susceptibility was not indicative of susceptibility to all tested cephalosporins, nor was there a clear trend in susceptibility based on cephalosporin generation.

Evaluation of surveillance methods for detection of West Nile virus activity in East Baton Rouge Parish, Louisiana, 2004-2006.

A 3-year study was conducted to determine if testing mosquitoes collected in modified sentinel chicken boxes for West Nile virus (WNV) or testing sentinel chickens for WNV antibody would detect WNV activity before onset of human cases in East Baton Rouge Parish, LA. In each year mosquitoes tested positive for WNV before the onset of human cases were detected, but seroconversions of sentinel chickens were detected after the human cases occurred. In 1 year we also compared the effectiveness of Centers for Disease Control and Prevention (CDC) light traps, gravid traps, and sentinel chicken box traps for collecting WNV-positive mosquitoes. Gravid traps collected more WNV-positive mosquitoes than CDC light traps or sentinel chicken box traps. However, WNV was detected earlier in mosquitoes collected from sentinel chicken box traps than in mosquitoes collected with gravid traps or CDC light traps. In total, 1,222 pools containing 19,353 mosquito specimens representing 18 species were tested for WNV. West Nile virus was detected in 59 mosquito pools from 4 species; 87% of the positive pools were detected from Culex quinquefasciatus, which was the most abundant species collected in all 3 years.

West Nile virus detection in mosquitoes in East Baton Rouge Parish, Louisiana, from November 2002 to October 2004.

The prevalence of West Nile virus (WNV) was determined in mosquitoes between November 2002 and October 2004 in East Baton Rouge Parish, LA. A total of 244,374 female mosquitoes were collected and tested by viral isolation. Additionally, 131,896 female mosquitoes were collected in 2003 and tested by VecTest and 167,175 female mosquitoes were collected in 2004 and tested by reverse transcriptase-polymerase chain reaction (RT-PCR). West Nile virus was isolated by cell culture from 17 (47.2%) out of 36 mosquito species collected over the study period. In 2003, WNV was detected in 9 (33.3%) out of 27 species tested by VecTest. In 2004, 14 (50%) out of the 28 mosquito species tested by RT-PCR were positive for WNV. The species with the greatest number of WNV-positive pools detected by all 3 testing methods was Culex quinquefasciatus. A significantly greater proportion of Cx. salinarius pools collected in light traps placed at a 3-m height were positive for WNV by viral isolation than in pools collected in light traps placed at a 1.5-m height.

Association of West Nile virus with lymphohistiocytic proliferative cutaneous lesions in American alligators (Alligator mississippiensis) detected by RT-PCR.

West Nile virus (WNV) is known to affect captive populations of alligators and, in some instances, cause significant mortalities. Alligators have been shown to amplify the virus, serve as a reservoir host, and even represent a source of infection for humans. This study describes a cutaneous manifestation of WNV in captive-reared American alligators (Alligator mississippiensis), previously described as lymphohistiocytic proliferative syndrome of alligators (LPSA), based on the findings of gross examination, histopathologic evaluation, WNV antibody testing, and WNV reverse transcriptase polymerase chain reaction (RT-PCR). Forty alligators with LPSA and 41 controls were examined. There was a significant difference (P = 0.01(-21)) in the WNV serostatus between the treatment group (100%) and the control group (0%, 95% CI: 0-7.3%). In the treatment group, 97.5% (39/40) (95% CI: 92.7-102.3%) of the LPSA skin lesions were positive for WNV via RT-PCR. Of the skin sections within the treatment group that had no LPSA lesions, 7.5% (3/40) (95% CI: 0-15.7%) were positive for WNV. In the control group, all of the skin samples were negative for WNV (41/41) (0%; 95% CI: 0-7.3%). The LPSA skin lesions were significantly more likely to be WNV positive by RT-PCR when compared to control animals (P = 0.07(-20)) and normal skin sections from affected animals (P = 0.08(-16)). There was no significant difference in the WNV RT-PCR results between control animals and normal skin sections from affected animals (P = 0.24). These findings suggest that LPSA is a cutaneous manifestation of WNV in alligators.

Characterization, distribution, antimicrobial resistance and resistance risk factors in staphylococci isolated from cats from 2001 to 2014.

Relatively few studies have been published describing the patterns of staphylococcal isolation and antimicrobial resistance over time in cats. The objective of this retrospective study was to determine the frequency, location, characteristics and antimicrobial resistance profiles of staphylococci isolated by the Louisiana Animal Disease Diagnostic Laboratory between the years 2001 and 2014. All feline staphylococcal isolates were classified phenotypically. Isolates corresponding to known or possibly pathogenic species (Staphylococcus intermedius group (SIG) and Staphylococcus aureus (SA)) as well as Staphylococcus epidermidis (SE) and non-speciated coagulase-negative staphylococci (CNS) were further evaluated to determine antimicrobial resistance patterns. A total of 519 staphylococci were isolated. The largest percentage of isolates was CNS, representing 39.3% of the total, while SIG, SE, SA and non-speciated coagulase positive staphylococci (CPS) represented 18.1%, 10.2%, 8.3% and 7.3%, respectively. Methicillin resistance (MR) was identified in 57.1% of SA and 20.5% of SIG. Resistance to 3 or more antimicrobial classes (multidrug resistance; MDR) was demonstrated in 54.5% of SA and 23.9% of SIG. The prevalence of MDR increased over time in both SIG and SA, while the prevalence of MR increased over time in SIG. An increase in mean antimicrobial resistance score over time was seen in SIG. This study demonstrates a high and increasing prevalence of MDR in SIG and SA, as well as increasing prevalence of MR in SIG isolated from cats.

West Nile virus surveillance in East Baton Rouge Parish, Louisiana.

West Nile virus (WNV) was detected for the first time in Louisiana in the fall of 2001. Surveillance data collected from East Baton Rouge Parish in 2002 were examined to establish baseline data on WNV activity, to support the current design of disease surveillance programs, and to target vector control efforts in the parish. The first indications of WNV activity were from a dead Northern Cardinal collected in February and from a live male cardinal sampled on 14 March. In mosquito pools, WNV was first detected on June 11. The onset of the first human case and the first detection of WNV in sentinel chickens occurred concurrently on June 24. The number of reported human cases and minimum infection rates in mosquitoes peaked in July. WNV prevalence in wild birds increased in late August and was highest in December. WNV-positive wild birds and mosquito pools were detected an average of 31 and 59 days in advance of the onset date of reported human cases, respectively, within 5 km of the residence of a human case. Antibodies to WNV were detected in sera from 7 (Northern Cardinal, House Sparrow, Northern Mockingbird, Blue Jay, Hermit Thrush, Yellow-rumped Warbler, and White-throated Sparrow) of the 42 wild bird species tested. Wild bird serology indicated WNV activity during the winter. Out of 18 mosquito species tested, the only species found positive for WNV was Culex quinquefasciatus, a result suggesting that this species was the primary epizootic/epidemic vector.

Disseminated mycobacteriosis in a bald eagle (Haliaeetus leucocephalus).

A mature bald eagle (Haliaeetus leucocephalus) was diagnosed with mycobacterial infection after being presented for an inability to fly, emaciation, and a swelling of the left tibiotarsal-tarso metatarsal joint. Results of a complete blood cell count revealed a persistent, marked leukocytosis, with heterophilia, monocytosis, and anemia. Radiographs revealed lysis of the left distal tibiotarsus and soft-tissue swelling around the left tibiotarsal-tarsometatarsal joint, multiple pulmonary opacities, and an enlarged liver. Endoscopic evaluation and biopsy of caseated material within the left caudal coelom revealed acid-fast organisms. The eagle was euthanatized, and results of necropsy and histologic evaluation revealed caseated granulomas of the intestine, lungs, air sacs, and subcutaneous regions of the hock. Results of culture, a polymerase chain reaction testing, and direct deoxyribonucleic acid (DNA) sequencing for mycobacterial 16S ribosomal ribonucleic acid DNA determined this organism most likely to be Mycobacterium avium.

A recombinant fusion protein consisting of West Nile virus envelope domain III fused in-frame with equine CD40 ligand induces antiviral immune responses in horses.

West Nile Virus (WNV) is endemic in the US and causes severe neurologic disease in horses since its introduction in 1999. There is no effective pharmaceutical treatment for WNV infection rendering vaccination as the only approach to prevention and control of disease. The purpose of this study was to evaluate a recombinant vaccine containing domain III (DIII) of the WNV envelope glycoprotein with and without a natural adjuvant equine (CD40L) in producing virus neutralizing antibodies in horses. Serum IgG1 concentration in the groups of horses vaccinated with the DIII-CD40L+TiterMax and DIII-CD40L proteins were significantly increased (p<0.05) after the second booster vaccination compared to other groups. Serum IgG4 and IgG7, IgG3 and IgG5 concentrations were not significantly increased among all groups. Western blot results showed that animals immunized with the DIII-CD40L protein (with or without TiterMax) exhibited the highest specific anti-DIII antibody activities after vaccinations. Moreover, animals immunized with the DIII-CD40L protein (with or without TiterMax) exhibited significantly stronger neutralization activity (p<0.05) compared to other groups starting at week eight. The DIII-CD40L protein (with or without TiterMax) stimulated more CD8+T cells, but not CD4+T cells in equine PMBCs. The results demonstrated that vaccination with recombinant WNV E DIII-CD40L protein induced superior humoral and cellular immune response in healthy horses that may be protective against WNV-associated disease in infected animals. CD40L could be utilized as a non-toxic, alternative adjuvant to boost the immunogenicity of subunit vaccines in horses.

Detection of West Nile virus RNA in pools of three species of ceratopogonids (Diptera: Ceratopogonidae) collected in Louisiana.

Light traps were used to collect ceratopogonids in East Baton Rouge parish, Louisiana. In total, 46,496 ceratopogonids were sorted from 4,968 light trap collections from 20 November 2002 through 25 November 2004. Two hundred and nine pools containing specimens of 18 species of Culicoides Latreille, seven pools containing specimens of Atrichopogon Kieffer, and five pools containing specimens of Forcipomyia Meigen were tested for West Nile virus (family Flaviviridae, genus Flavivirus, WNV) RNA using real-time reverse transcriptase polymerase chain reaction. Five out of the 209 pools of Culicoides specimens were positive for WNV RNA.

Evaluation of microbial culture techniques for the isolation of Pythium insidiosum from equine tissues.

The purpose of this study was to evaluate the effects of sample handling, storage, and culture techniques on the isolation of Pythium insidiosum from infected equine tissues. Tissue and kunker samples obtained immediately posteuthanasia from a horse with subcutaneous pythiosis were used to assess the effects of sample type (kunkers vs. tissues), media type (selective vs. nonselective), storage technique, and storage time on P. insidiosum isolation rate. Overall, isolation rates were higher from fresh kunkers (94.6%) and stored kunkers (76.4%) than from fresh tissues (8.3%) or stored tissues (4.6%). Isolation of P. insidiosum also occurred more often on antibiotic-containing media than on nonselective media for both fresh and stored samples. For samples that were stored for 1-3 days prior to culture, P. insidiosum isolation rates were highest for the following techniques: kunkers stored at room temperature and plated on selective media (100%), kunkers stored at 4 C and then plated on either nonselective (91.7%) or selective (95.8%) media, kunkers stored on cold packs and then plated on either nonselective (93.8%) or selective (100%) media, kunkers stored in ampicillin solution and plated on selective media (100%), and kunkers stored in ampicillin/gentocin solution and plated on selective media (87.5%). For samples stored for 4-5 days, P. insidiosum isolation rates were highest for kunkers stored at 4 C and then plated on either nonselective (81.3%) or selective (87.5%) media, kunkers stored in ampicillin solution and then plated on selective media (87.5%), and kunkers stored in ampicillin/gentocin solution and plated on selective media (87.5%). Results of this study suggest that optimal isolation rates of P. insidiosum from infected equine tissues are achieved by culturing fresh kunkers on selective media. For samples that cannot be processed immediately, acceptable handling techniques include storage at room temperature for up to 3 days, refrigeration for up to 5 days, shipping on cold packs, and storage in antibiotic solution, each combined with subsequent inoculation on selective media.

Oral conidiobolomycosis in a dog.

Conidiobolomycosis was diagnosed via culture from an oral lesion in a 1.5-year-old German Shepherd dog. Clinically, the lesion consisted of a large, irregularly shaped, ulcerative focus on the caudal hard palate. Microscopically, the lesion was characterized by an eosinophilic granulomatous stomatitis with hyphal organisms surrounded by eosinophilic sleeves (Splendore-Hoeppli material) suggestive of an entomophthoramycosis. The fungus was cultured and identified with features consistent with Conidiobolus sp. Treatment with itraconazole at 10 mg kg-1 twice daily for 61 days resulted in clinical and radiographic resolution of the lesion.

Experimental Reproduction of Feline Mycobacterium fortuitum Panniculitis.

Resumen- Con el fin de lograr un modelo experimental de la infección por Mycobacterium fortuitum, causante de paniculitis en condiciones naturales en el gato, se inoculóM. fortuitum en el cojinte plantar de ratones o en el tejido adiposo inguinal de conejos y gatos. Los ratones manifestaron una dermatitis crónica y una linfadenitis granulomatosa necrotizante con localizatión intracelular del microorganismo. Los conejos manifestaron inflamaciones granulomatosas supurativas necrotizantes con microrganismos en vacuolas adiposas rodeadas por heterófilos macrófagos epitelioides y/o zonas de necrosis. Los cinco gatos adultos y una de las tres crias mostraron fistulas supurativas, úlceras puntuales o nódulos en el paniculo adiposo de la zona inguinal. La lesión en la región inguinal de estos seis animales consistia en una paniculitis granulomatosa; las otras dos crias presentaban una paniculitis piogranulomatosa necrotizante. Se identificaron bacilos en los cortes histológicos teñidos con Hematoxilina y Eosina en cuatro gatos adultos y en una de las crias. Se aislóMycobacterium fortunitum a partir del tejido adiposo en todos los gatos adultos y en una de las tres crias. La inoculación de 1.4 × 1010 M.fortuitum en el tejido adiposo subcutáneo inguinal en los gatos con grandes masas grasas en esas zonas causó una infección microbacteriana idéntica a la enfermedad felina en condiciones naturales. [Lewis, D. T., Hodgin, E. C, Foil, C. S., Cox, H. U., Roy, A. F., Lewis, D. D. Experimental reproduction of feline Mycobacterium fortuitum panniculitis. (Reproductión experimental de la paniculitis felina por Mycobacterium fortuitum). Abstract- In order to establish an animal model of the naturally occurring feline Mycobacterium fortuitum panniculitis an inoculum of M. fortuitum organisms was injected into the hindlimb footpad of mice or subcutaneous fat of the inguinal area of rabbits and cats. Mice developed chronic dermatitis and necrotizing granulomatous lymphadenitis with intracellular localization of the organism. Rabbits developed necrotizing suppurative granulomatous inflammation with organisms in heterophil-lined fat vacuoles, epithelioid macrophages and/or necrotic areas. All five adult cats and one of three kittens developed draining tracts, punctate ulcers or nodules in the panniculus adiposus of the inguinal area. A pyogranulomatous panniculitis characterized the inguinal region in these six animals; a necrotizing pyogranulomatous panniculitis was present in the remaining two kittens. Rod-shaped bacilli were present on hematoxylin and eosin stained tissue sections in four adult cats and one kitten. Mycobacterium fortuitum was isolated from the inguinal subcutaneous fat in all five adult cats and one of three kittens. Injection of 1.4 × 1010 M. fortuitum organisms into the subcutaneous fat of the inguinal area of cats with extensive inguinal fatpads produced a mycobacterial infection identical to the naturally occurring feline disease.