Adam J. Birkenheuer

TRITRICHOMONAS FOETUS AND NOT PENTATRICHOMONAS HOMINIS IS THE ETIOLOGIC AGENT OF FELINE TRICHOMONAL DIARRHEA

Recently, several investigators have reported large-bowel diarrhea in cats associated with intestinal trichomonad parasites. These reports have presumptively identified the flagellates as Pentatrichomonas hominis, an organism putatively capable of infecting the intestinal tracts of a number of mammalian hosts, including cats, dogs, and man. The purpose of the present study was to determine the identity of this recently recognized flagellate by means of rRNA gene sequence analysis; restriction enzyme digest mapping; and light, transmission, and scanning electron microscopy (SEM).

Single-Tube Nested PCR for Detection of Tritrichomonas foetus in Feline Feces

ABSTRACT Tritrichomonas foetus, a venereal pathogen of cattle, was recently identified as an inhabitant of the large intestine in young domestic cats with chronic diarrhea. Recognition of the infection in cats has been mired by unfamiliarity with T. foetus in cats as well as misdiagnosis of the organisms as Pentatrichomonashominis or Giardia sp. when visualized by light microscopy. The diagnosis of T. foetus presently depends on the demonstration of live organisms by direct microscopic examination of fresh feces or by fecal culturing. As T. foetus organisms are fastidious and fragile, routine flotation techniques and delayed examination and refrigeration of feces are anticipated to preclude the diagnosis in numerous cases. The objective of this study was to develop a sensitive and specific PCR test for the diagnosis of feline T. foetus infection. A single-tube nested PCR was designed and optimized for the detection of T. foetus in feline feces by using a combination of novel (TFITS-F and TFITS-R) and previously described (TFR3 and TFR4) primers. The PCR is based on the amplification of a conserved portion of the T. foetus internal transcribed spacer (ITS) region (ITS1 and ITS2) and the 5.8S rRNA gene. The absolute detection limit of the single-tube nested PCR was 1 organism, while the practical detection limit was 10 organisms per 200 mg of feces. Specificity was examined by using P. hominis, Giardialamblia, and feline genomic DNA. Our results demonstrate that the single-tube nested PCR is ideally suited for (i) diagnostic testing of feline fecal samples that are found negative by direct microscopy and culturing and (ii) definitive identification of microscopically observable or cultivated organisms.

Canine and feline blood donor screening for infectious disease.

Consensus Statements of the American College of Veterinary Internal Medicine (ACVIM) provide veterinarians with guidelines regarding the pathophysiology, diagnosis, or treatment of animal diseases. The foundation of the Consensus Statement is evidence‐based medicine, but if such evidence is conflicting or lacking, the panel provides interpretive recommendations on the basis of their collective expertise. The Consensus Statement is intended to be a guide for veterinarians, but it is not a statement of standard of care or a substitute for clinical judgment. Topics of statements and panel members to draft the statements are selected by the Board of Regents with input from the general membership. A draft prepared and input from Diplomates is solicited at the Forum and via the ACVIM Web site and incorporated in a final version. This Consensus Statement was approved by the Board of Regents of the ACVIM before publication.

Babesia gibsoni infections in dogs from North Carolina.

The recognition of canine babesiosis in North Carolina caused by Babesia gibsoni documents the expansion of the previously reported endemic area of this disease. Clinical signs ranged from severe hemolytic anemia and thrombocytopenia to subclinical infections. No infected dogs had traveled to endemic areas. Antibabesial treatment failed to eradicate the organism from infected dogs.

Efficacy of Ronidazole for Treatment of Feline Tritrichomonas foetus Infection

OBJECTIVESnTo determine the efficacy of ronidazole (RDZ), tinidazole (TDZ), and metronidazole (MDZ) against Tritrichomonas foetus in vitro and of RDZ for treatment of feline naturally occurring or experimentally induced T. foetus infection.nnnANIMALSnA cat naturally infected with T. foetus infection and diarrhea. Ten specific-pathogen-free (SPF) kittens.nnnPROCEDUREnRDZ, TDZ, and MDZ were tested for activity against 3 different feline isolates of T. foetus in vitro. RDZ then was administered to a naturally infected cat at 10 mg/kg PO q24h for 10 days. SPF kittens were infected orogastrically with feline T. foetus and treated with either placebo or RDZ (10 mg/kg PO q12h for 14 days). Cats with relapsing infection or those receiving placebo were treated subsequently with RDZ (either 30 or 50 mg/kg PO q12h for 14 days). Feces were examined for T. foetus by direct microscopy, culture, and polymerase chain reaction (PCR) testing weekly.nnnRESULTSnBoth RDZ and TDZ killed T. foetus at concentrations >0.1 microg/mL in vitro. In the naturally infected cat, RDZ abolished diarrhea and T. foetus infection for 85 days after treatment, at which time infection and diarrhea relapsed. Retreatment with RDZ eradicated diarrhea and T. foetus infection for over 407 days. In experimentally induced infection, RDZ at 10 mg/kg caused initial improvement, but infection relapsed in all 5 cats 2 to 20 weeks after treatment. At 30 or 50 mg/kg, 10/10 cats were negative for T. foetus infection for follow-up durations of 21 to 30 weeks after treatment.nnnCONCLUSIONS AND CLINICAL RELEVANCEnOral administration of RDZ at 30 to 50 mg/kg q12h for 14 days resolved diarrhea and eradicated infection (on the basis of polymerase chain reaction [PCR] testing) in 1 naturally infected cat and 10 experimentally inoculated cats receiving a different isolate of T. foetus.

Serosurvey of antiBabesia antibodies in stray dogs and American pit bull terriers and American staffordshire terriers from North Carolina.

Stray dogs (n=359) and kennel dogs (n=149) from North Carolina were tested for evidence of antiBabesia antibodies. AntiBabesia antibodies were detected in 21/359 and 22/149 of the stray and kennel dogs, respectively. A total of 57 dogs from both groups were tested for babesiasis by light microscopy and polymerase chain reaction (PCR). Babesia deoxyribonucleic acid (DNA) was detected in 3/28 of the stray dogs and 14/29 of the kennel dogs. When Babesia DNA was detected by PCR, the species-specific PCR results differed from the Babesia species antibody titer results in 6/17 of the PCR-positive dogs. There was no association between antiBabesia antibodies and the presence of ticks. There are currently Babesia gibsoni epizootics affecting American pit bull terrier kennels.

MOLECULAR CHARACTERIZATION OF TRICHOMONADS FROM FECES OF DOGS WITH DIARRHEA

Trichomonads are occasionally observed in the feces of dogs with diarrhea. On the basis of superficial morphological appearance, these infections have been attributed to opportunistic overgrowth of the commensal, Pentatrichomonas hominis. However, molecular characterization of canine trichomonads has never been reported. This study was performed to determine, by means of rRNA gene sequence analysis, the identity of trichomonads observed in feces from dogs with diarrhea. Total DNA was isolated from fecal samples obtained from a 3-mo-old mixed breed dog and litter of German Shepherd puppies having profuse liquid diarrhea containing numerous trichomonads. Total DNA was subject to PCR amplification of partial 18S rRNA gene or 5.8S, ITS1, ITS2, and partial 18S and 28S rRNA genes using species-specific and universal primers, respectively. Products of 642 and 1864 base-pair length were amplified and cloned. On the basis of rRNA gene sequence, the trichomonads observed in the single dog and the litter of puppies shared 100% identity with Tritrichomonas foetus and P. hominis, respectively. The present study is the first to establish the molecular identity of trichomonads infecting dogs with diarrhea. These studies validate the longstanding assumption that canine trichomoniasis may be attributed to P. hominis. Importantly, these studies additionally recognize that canine trichomoniasis may also be caused by infection with T. foetus.

Efficacy of Atovaquone and Azithromycin or Imidocarb Dipropionate in Cats with Acute Cytauxzoonosis

BACKGROUNDnImidocarb or a combination of atovaquone and azithromycin (A&A) has been suggested for treatment of cats with cytauxzoonosis, but neither has been prospectively evaluated for efficacy.nnnHYPOTHESIS/OBJECTIVESnThat survival to hospital discharge is improved by treatment with A&A as compared with imidocarb.nnnANIMALSnEighty acutely ill cats with Cytauxzoon felis infection treated at one of 18 veterinary clinics in 5 states.nnnMETHODSnAn open-label, randomized prospective study compared survival in cats treated with atovaquone (15 mg/kg p.o. q8h) and azithromycin (10 mg/kg p.o. q24h) or imidocarb (3.5 mg/kg i.m.). All received heparin, fluids, and supportive care. Clinical and clinicopathologic data from initial presentation were collated. Parasitemia was quantified (n = 79) and pathogens genotyped (n = 60). Logistic regression was used to determine the impact of treatment group on the primary endpoint, survival to hospital discharge or death. Covariants were analyzed by rank-sum testing.nnnRESULTSnOf 53 cats treated with A&A, 32 (60%) survived to discharge while only 7 of 27 cats (26%) treated with imidocarb survived (P = .0036; odds ratio 7.2, 95% CI 2.2, 24). Cats with a lower parasitemia were more likely to survive, as were cats with higher white blood cell counts and lower total bilirubin. Unique pathogen genotypes were identified from 15 cats, while genotype isolated from 21 cats had been described previously. There were multiple pathogen genotypes identified in 24 cats.nnnCONCLUSIONS AND CLINICAL IMPORTANCEnSurvival to discharge was more likely in cats treated with A&A as compared with imidocarb, although case fatality rate remained high.

Canine Rocky Mountain Spotted fever: a retrospective study of 30 cases.

Rocky Mountain spotted fever (RMSF) was diagnosed in 30 dogs examined at North Carolina State University, Veterinary Teaching Hospital between 1984 and 1997. Historical, physical examination, and laboratory abnormalities were reviewed. Diagnostic criteria included a four-fold rise in antibody titer to Rickettsia rickettsii (R. rickettsii) (n=15) or a single R. rickettsii antibody titer of 1:1,024 or greater (n=15; when this initial titer was determined one week or more after the onset of clinical signs). Fifteen (50%) dogs were greater than seven years of age, and 13 (43%) dogs were between two and seven years of age. There was no sex predilection. Only five (17%) dogs had a history of known tick exposure. Presumably due to delayed diagnosis, dogs with antibody titers of 1:1,024 or greater at the time of presentation had a higher incidence of more severe neurological dysfunction (e.g., ataxia, hyperesthesia, vestibular disease, and seizures) and cutaneous lesions (e.g., hyperemia, edema, petechiae, ecchymoses, and necrosis). Laboratory findings included anemia, leukocytosis accompanied by toxic granulation of neutrophils, hypoalbuminemia, and coagulation abnormalities; signs were generally more severe in the 15 dogs with R. rickettsii antibody titers of 1:1,024 or greater at the time of presentation. Twelve (40%) dogs in this study were severely thrombocytopenic (less than 75 x10(3) platelets/microl; reference range, 200 to 450 x 10(3)/microl), without clinical evidence of fulminant disseminated intravascular coagulation. In this study, the survival rate following R. rickettsii infection was 100%.

Evaluation of brain tissue or cerebrospinal fluid with broadly reactive polymerase chain reaction for Ehrlichia, Anaplasma, spotted fever group Rickettsia, Bartonella, and Borrelia species in canine neurological diseases (109 cases).

BACKGROUNDnVector-transmitted microorganisms in the genera Ehrlichia, Anaplasma, Rickettsia, Bartonella, and Borrelia are commonly suspected in dogs with meningoencephalomyelitis (MEM), but the prevalence of these pathogens in brain tissue and cerebrospinal fluid (CSF) of dogs with MEM is unknown.nnnHYPOTHESIS/OBJECTIVESnTo determine if DNA from these genera is present in brain tissue and CSF of dogs with MEM, including those with meningoencephalitis of unknown etiology (MUE) and histopathologically confirmed cases of granulomatous (GME) and necrotizing meningoencephalomyelitis (NME).nnnANIMALSnHundred and nine dogs examined for neurological signs at 3 university referral hospitals.nnnMETHODSnBrain tissue and CSF were collected prospectively from dogs with neurological disease and evaluated by broadly reactive polymerase chain reaction (PCR) for Ehrlichia, Anaplasma, Spotted Fever Group Rickettsia, Bartonella, and Borrelia species. Medical records were evaluated retrospectively to identify MEM and control cases.nnnRESULTSnSeventy-five cases of MUE, GME, or NME, including brain tissue from 31 and CSF from 44 cases, were evaluated. Brain tissue from 4 cases and inflammatory CSF from 30 cases with infectious, neoplastic, compressive, vascular, or malformative disease were evaluated as controls. Pathogen nucleic acids were detected in 1 of 109 cases evaluated. Specifically, Bartonella vinsonii subsp. berkhoffii DNA was amplified from 1/6 dogs with histopathologically confirmed GME.nnnCONCLUSION AND CLINICAL IMPORTANCEnThe results of this investigation suggest that microorganisms in the genera Ehrlichia, Anaplasma, Rickettsia, and Borrelia are unlikely to be directly associated with canine MEM in the geographic regions evaluated. The role of Bartonella in the pathogenesis of GME warrants further investigation.