@article{hedgespeth_stauffer_robertson_gookin_2020, title={Association of fecal sample collection technique and treatment history with Tritrichomonas foetus polymerase chain reaction test results in 1717 cats}, volume={34}, ISSN={["1939-1676"]}, url={http://dx.doi.org/10.1111/jvim.15727}, DOI={10.1111/jvim.15727}, abstractNote={Fecal polymerase chain reaction (PCR) testing for Tritrichomonas foetus is considered the most sensitive means for diagnosis of infection but results could be influenced by fecal collection technique and prior use of antimicrobial drugs.To establish any association between fecal collection technique or treatment history and results of fecal PCR testing for T. foetus.Fecal samples from 1717 cats submitted by veterinarians between January 2012 and December 2017.This study used a retrospective analysis. T. foetus PCR test results from 1808 fecal samples submitted for diagnostic testing were examined for their association with method of fecal collection and prior antimicrobial treatments. Data were collected from sample submission form.Positive T. foetus PCR test results were obtained for 274 (16%) cats. Fecal samples collected via fecal loop had increased probability of positive PCR test results (odds ratio [OR] 2.04, 95% confidence interval [CI] 1.31-3.17, P = .002) compared to samples collected by colonic flush. There was no association between PCR test results and treatment history, treatment type, or prior treatment with ronidazole. After an initial positive PCR test, 4/19 (21%; 95% CI 2.7%-39.4%) cats treated with ronidazole had a second positive test result.Results of this study support that fecal samples collected by loop might be better for PCR diagnosis of T. foetus infection. Lack of association of ronidazole with PCR test results and a 21% all-potential-causes failure rate of ronidazole in cats with preconfirmed infection are important limitations to use of this drug.}, number={2}, journal={JOURNAL OF VETERINARY INTERNAL MEDICINE}, author={Hedgespeth, Barry A. and Stauffer, Stephen H. and Robertson, James B. and Gookin, Jody L.}, year={2020}, month={Mar}, pages={734–741} }
 @article{tolbert_stauffer_brand_gookin_2014, title={Cysteine Protease Activity of Feline Tritrichomonas foetus Promotes Adhesion-Dependent Cytotoxicity to Intestinal Epithelial Cells}, volume={82}, ISSN={["1098-5522"]}, DOI={10.1128/iai.01671-14}, abstractNote={Trichomonads are obligate protozoan parasites most renowned as venereal pathogens of the reproductive tract of humans and cattle. Recently, a trichomonad highly similar to bovine venereal Tritrichomonas foetus but having a unique tropism for the intestinal tract was recognized as a significant cause of colitis in domestic cats. Despite a high prevalence, worldwide distribution, and lack of consistently effective drugs for treatment of the infection, the cellular mechanisms of T. foetus pathogenicity in the intestinal tract have not been examined. The aims of this study were to determine the pathogenic effect of feline T. foetus on porcine intestinal epithelial cells, the dependence of T. foetus pathogenicity on adhesion of T. foetus to the intestinal epithelium, and the identity of mediators responsible for these effects. Using an in vitro coculture approach to model feline T. foetus infection of the intestinal epithelium, these studies demonstrate that T. foetus promotes a direct contact-dependent activation of intestinal epithelial cell apoptosis signaling and progressive monolayer destruction. Moreover, these pathological effects were demonstrated to be largely dependent on T. foetus cell-associated cysteine protease activity. Finally, T. foetus cysteine proteases were identified as enabling cytopathic effects by promoting adhesion of T. foetus to the intestinal epithelium. The present studies are the first to examine the cellular mechanisms of pathogenicity of T. foetus toward the intestinal epithelium and support further investigation of the cysteine proteases as virulence factors in vivo and as potential therapeutic targets for ameliorating the pathological effects of intestinal trichomonosis.}, number={7}, journal={INFECTION AND IMMUNITY}, author={Tolbert, M. K. and Stauffer, S. H. and Brand, M. D. and Gookin, J. L.}, year={2014}, month={Jul}, pages={2851–2859} }
 @article{tolbert_stauffer_gookin_2013, title={Feline Tritichomonas foetus adhere to the intestinal epithelium by receptor-ligand-dependent mechanisms}, volume={192}, ISSN={["1873-2550"]}, DOI={10.1016/j.vetpar.2012.10.019}, abstractNote={Tritrichomonas foetus (TF) is a protozoan that infects the feline ileum and colon resulting in chronic diarrhea. Up to 30% of young purebred cats are infected with TF and the infection is recognized as pandemic. Only a single drug, characterized by a narrow margin of safety and emerging development of resistance, is effective for treatment. While the venereal pathogenicity of bovine TF is attributed to adherence to uterovaginal epithelium, the pathogenesis of diarrhea in feline TF infection is unknown. The aim of this study was to establish an in vitro model of feline TF adhesion to intestinal epithelium. Confluent monolayers of porcine intestinal epithelial cells (IPEC-J2) were infected with axenic cultures of feline TF that were labeled with [(3)H] thymidine or CFSE and harvested at log-phase. The effect of multiplicity and duration of infection, viability of TF, binding competition, formalin fixation and cytoskeletal inhibitors on adherence of feline TF to IPEC-J2 monolayers was quantified by liquid scintillation counting and immunofluorescence. [(3)H] thymidine and CFSE-labeled TF reproducibly adhered to IPEC-J2 monolayers. Clinical isolates of feline TF adhered to the intestinal epithelium in significantly greater numbers than Pentatrichomonas hominis, the latter of which is a presumably nonpathogenic trichomonad. Adhesion of TF required viable trophozoites but was independent of cytoskeletal activity. Based on saturation and competition binding experiments, adherence of feline TF to the epithelium occurred via specific receptor-ligand interactions. The developed model provides a valuable resource for assessing pathogenic mechanisms of feline TF and developing novel pharmacologic therapies for blocking the adhesion of feline TF to the intestinal epithelium.}, number={1-3}, journal={Veterinary Parasitology}, author={Tolbert, M.K. and Stauffer, S.H. and Gookin, J.L.}, year={2013}, month={Feb}, pages={75–82} }
 @article{ghosh_borst_stauffer_suyemoto_moisan_zurek_gookin_2013, title={Mortality in Kittens Is Associated with a Shift in Ileum Mucosa-Associated Enterococci from Enterococcus hirae to Biofilm-Forming Enterococcus faecalis and Adherent Escherichia coli}, volume={51}, ISSN={["1098-660X"]}, DOI={10.1128/jcm.00481-13}, abstractNote={ABSTRACT Approximately 15% of foster kittens die before 8 weeks of age, with most of these kittens demonstrating clinical signs or postmortem evidence of enteritis. While a specific cause of enteritis is not determined in most cases, these kittens are often empirically administered probiotics that contain enterococci. The enterococci are members of the commensal intestinal microbiota but also can function as opportunistic pathogens. Given the complicated role of enterococci in health and disease, it would be valuable to better understand what constitutes a “healthy” enterococcal community in these kittens and how this microbiota is impacted by severe illness. In this study, we characterized the ileum mucosa-associated enterococcal community of 50 apparently healthy and 50 terminally ill foster kittens. In healthy kittens, Enterococcus hirae was the most common species of ileum mucosa-associated enterococci and was often observed to adhere extensively to the small intestinal epithelium. These E. hirae isolates generally lacked virulence traits. In contrast, non- E. hirae enterococci, notably Enterococcus faecalis , were more commonly isolated from the ileum mucosa of kittens with terminal illness. Isolates of E. faecalis had numerous virulence traits and multiple antimicrobial resistances. Moreover, the attachment of Escherichia coli to the intestinal epithelium was significantly associated with terminal illness and was not observed in any kitten with adherent E. hirae . These findings identify a significant difference in the species of enterococci cultured from the ileum mucosa of kittens with terminal illness compared to the species cultured from healthy kittens. In contrast to prior case studies that associated enteroadherent E. hirae with diarrhea in young animals, these controlled studies identified E. hirae as more often isolated from healthy kittens and adherence of E. hirae as more common and extensive in healthy kittens than in sick kittens.}, number={11}, journal={JOURNAL OF CLINICAL MICROBIOLOGY}, author={Ghosh, Anuradha and Borst, Luke and Stauffer, Stephen H. and Suyemoto, Mitsu and Moisan, Peter and Zurek, Ludek and Gookin, Jody L.}, year={2013}, month={Nov}, pages={3567–3578} }
 @article{gookin_stauffer_dybas_cannon_2010, title={Documentation of In Vivo and In Vitro Aerobic Resistance of Feline Tritrichomonas foetus Isolates to Ronidazole}, volume={24}, ISSN={["1939-1676"]}, DOI={10.1111/j.1939-1676.2010.0534.x}, abstractNote={Background: The mainstays of treatment for clinically important trichomonad infections are the 5-nitroimidazoles. Metronidazole resistance of feline Tritrichomonas foetus is presumed because of common treatment failure, and tinidazole does not consistently eradicate infection. To date, ronidazole is the only drug demonstrated as effective for treatment of cats infected with T. foetus. Objective: To document in vivo treatment failure and identify underlying causes and in vitro conditions of resistance of feline T. foetus to ronidazole. Animals: Two intact male Abyssinians failing ≥5 courses of treatment with increasing doses of 5-nitroimidazole drugs. An intact male Abyssinian documented to clear infection after treatment with a single course of ronidazole. Methods: T. foetus isolates were cultured from feces and tested in vitro for susceptibility to ronidazole under aerobic and anaerobic culture conditions. A urogenital nidus of T. foetus infection was investigated by culture, polymerase chain reaction, or immunohistochemical testing of urogenital specimens. Results: Resistance to ronidazole under aerobic conditions was uniquely identified in T. foetus isolated from cats with well-documented treatment failure. Treatment failure could not be attributed to reinfection, inappropriate treatment protocol, or presence of a urogenital nidus of infection. Conclusions and Clinical Importance: Clinical resistance to metronidazole, low efficacy of tinidazole, and present documentation of in vivo and in vitro resistance to ronidazole in some cats are consistent with a high level of cross resistance of feline T. foetus to 5-nitroimidazole drugs. Current lack of alternative drugs with clinical efficacy against feline T. foetus suggests that active investigation of other treatment approaches is warranted.}, number={4}, journal={JOURNAL OF VETERINARY INTERNAL MEDICINE}, author={Gookin, J. L. and Stauffer, S. H. and Dybas, D. and Cannon, D. H.}, year={2010}, pages={1003–1007} }
 @article{stauffer_birkenheuer_levy_marr_gookin_2008, title={Evaluation of four DNA extraction methods for the detection of Tritrichomonas foetus in feline stool specimens by polymerase chain reaction}, volume={20}, ISSN={["1943-4936"]}, DOI={10.1177/104063870802000518}, abstractNote={Feces are increasingly valued as practical samples for molecular diagnosis of infectious disease. However, extraction of polymerase chain reaction (PCR) quality DNA from fecal samples can be challenging because of coextraction of PCR inhibitors. Because the type and quantity of PCR inhibitors is influenced by diet, endogenous flora, and concurrent disease, it is unlikely that extraction method performance with human feces can be directly extrapolated to that of domestic cats. In the present study, 4 commercially available DNA extraction methods were examined for their influence on the sensitivity of PCR for the detection of Tritrichomonas foetus in feline stool. DNA was extracted from serially diluted feline-origin T. foetus trophozoites in the absence or presence of feline feces. The ZR Fecal DNA kit was identified as affording the greatest analytical sensitivity and reproducibility and was able to detect >or=10 T. foetus organisms per 100 mg feces in 100% of PCR reactions. Further, the identified extraction method could be completed in the shortest time of all kits tested.}, number={5}, journal={JOURNAL OF VETERINARY DIAGNOSTIC INVESTIGATION}, author={Stauffer, Stephen H. and Birkenheuer, Adam J. and Levy, Michael G. and Marr, Henry and Gookin, Jody L.}, year={2008}, month={Sep}, pages={639–641} }
 @article{gookin_stauffer_stone_2008, title={Induction of arginase II by intestinal epithelium promotes the uptake of L-arginine from the lumen of Cryptosporidium parvum-infected porcine ileum}, volume={47}, ISSN={["0277-2116"]}, DOI={10.1097/MPG.0b013e31816f6c02}, abstractNote={ABSTRACT Objectives: To determine the specific transport system activities and expression of transporter genes responsible for uptake of L‐arginine from the lumen of normal and Cryptosporidium parvum –infected neonatal porcine ileum and the influence of L‐arginine catabolic pathways on L‐arginine uptake. Methods: Intact sheets of ileal mucosa from control and C parvum –infected neonatal piglets were mounted in Ussing chambers and the uptake of 14 C‐L‐arginine was determined under initial rate conditions and in the presence of transport system–selective inhibitors. Epithelial expression of L‐arginine transporter genes was quantified by real‐time reverse transcription polymerase chain reaction. L‐Arginine catabolic enzyme expression was examined by immunoblotting epithelial lysates for arginase I and II. The role of intracellular catabolism in promoting the uptake of L‐arginine was determined by pharmacological inhibition of nitric oxide synthase and arginase activities. Results: C parvum –infected ileum transported L‐arginine at rates equivalent to uninfected epithelium despite profound villous atrophy. This was attributed to enhanced uptake of L‐arginine by individual epithelial cells in the infection. There were no differences in L‐arginine transport system activities (y + and B 0,+ ) or level of transporter gene expression (CAT‐1, CAT‐2A, and ATB 0,+ ) between uninfected and C parvum –infected epithelial cells. However, infected epithelia had induced expression of the L‐arginine hydrolytic enzyme arginase II and lower concentrations of L‐arginine. Furthermore, transport of L‐arginine by the infected epithelium was significantly inhibited by pharmacological blockade of arginase. Conclusions: Intracellular catabolism by arginase II, the induction of which has not been described previously for intestinal epithelium, facilitates uptake of L‐arginine by infected epithelium using transport systems that do not differ from those of uninfected cells. Induction of arginase II may limit nitric oxide synthesis by competing with nitric oxide synthase for utilization of L‐arginine or promote use of L‐arginine for the synthesis of reparative polyamines.}, number={4}, journal={JOURNAL OF PEDIATRIC GASTROENTEROLOGY AND NUTRITION}, author={Gookin, Jody L. and Stauffer, Stephen H. and Stone, Maria R.}, year={2008}, month={Oct}, pages={417–427} }
 @article{gookin_foster_coccaro_stauffer_2008, title={Oral Delivery of L-arginine Stimulates Prostaglandin-dependent Secretory Diarrhea in Cryptosporidium parvum–infected Neonatal Piglets}, volume={46}, ISSN={0277-2116}, url={http://dx.doi.org/10.1097/mpg.0b013e31815c0480}, DOI={10.1097/MPG.0b013e31815c0480}, abstractNote={ABSTRACT Objectives: To determine if oral supplementation with L‐arginine could augment nitric oxide (NO) synthesis and promote epithelial defense in neonatal piglets infected with Cryptosporidium parvum . Materials and Methods: Neonatal piglets were fed a liquid milk replacer and on day 3 of age infected or not with 10 8 C parvum oocysts and the milk replacer supplemented with L‐arginine or L‐alanine. Milk consumption, body weight, fecal consistency, and oocyst excretion were recorded daily. On day 3 postinfection, piglets were euthanized and serum concentration of NO metabolites and histological severity of villous atrophy and epithelial infection were quantified. Sheets of ileal mucosa were mounted in Ussing chambers for measurement of barrier function (transepithelial resistance and permeability) and short‐circuit current (an indirect measurement of Cl − secretion in this tissue). Results: C parvum –infected piglets had large numbers of epithelial parasites, villous atrophy, decreased barrier function, severe watery diarrhea, and failure to gain weight. L‐Arginine promoted synthesis of NO by infected piglets, which was unaccompanied by improvement in severity of infection but rather promoted epithelial chloride secretion and diarrhea. Epithelial secretion by infected mucosa from L‐arginine‐supplemented piglets was fully inhibited by the cyclooxygenase inhibitor indomethacin, indicating that prostaglandin synthesis was responsible for this effect. Conclusions: Results of these studies demonstrate that provision of additional NO substrate in the form of L‐arginine incites prostaglandin‐dependent secretory diarrhea and does not promote epithelial defense or barrier function of C parvum –infected neonatal ileum.}, number={2}, journal={Journal of Pediatric Gastroenterology and Nutrition}, publisher={Ovid Technologies (Wolters Kluwer Health)}, author={Gookin, Jody L and Foster, Derek M and Coccaro, Maria R and Stauffer, Stephen H}, year={2008}, month={Feb}, pages={139–146} }
 @article{gookin_stauffer_coccaro_poore_levy_papich_2007, title={Efficacy of tinidazole for treatment of cats experimentally infected with Tritrichomonas foetus}, volume={68}, ISSN={["1943-5681"]}, DOI={10.2460/ajvr.68.10.1085}, abstractNote={Abstract Objective —To determine the efficacy of tinidazole for treatment of cats with experimentally induced Tritrichomonas foetus infection. Animals —8 specific-pathogen-free kittens. Procedures —Tinidazole was tested for activity against a feline isolate of T foetus in vitro. Kittens were infected orogastrically with the same isolate and treated or not with tinidazole (30 mg/kg, PO, q 24 h for 14 days). Amoxicillin was administered 28 weeks after completion of tinidazole administration to induce diarrhea. Feces were repeatedly tested for T foetus by use of PCR assay and microbial culture for 33 weeks. Results —Tinidazole killed T foetus at concentrations ≥ 10 μg/mL in vitro. In experimentally induced infection, tinidazole administered at 30 mg/kg decreased T foetus below the limit of molecular detection in 2 of 4 cats. Recrudescent shedding of T foetus , as elicited by amoxicillin-induced diarrhea, was diminished in cats that received prior treatment with tinidazole. Conclusions and Clinical Relevance —Although tinidazole decreased the detection of T foetus and treated cats were resistant to later efforts to incite the infection, inability of tinidazole to eradicate infection in many cats poses a serious impediment to the drug’s effectiveness in practice.}, number={10}, journal={AMERICAN JOURNAL OF VETERINARY RESEARCH}, author={Gookin, Jody L. and Stauffer, Stephen H. and Coccaro, Maria R. and Poore, Matthew F. and Levy, Michael G. and Papich, Mark G.}, year={2007}, month={Oct}, pages={1085–1088} }
 @article{gookin_stauffer_levy_2007, title={Identification of Pentatrichomonas hominis in feline fecal samples by polymerase chain reaction assay}, volume={145}, ISSN={["1873-2550"]}, DOI={10.1016/j.vetpar.2006.10.020}, abstractNote={Pentatrichomonas hominis is considered to be a commensal protozoan of the vertebrate digestive tract. On the basis of light microscopic examination of feces, some investigators presumptively identified P. hominis as a causative agent of feline diarrhea. However, molecular identification of P. hominis infection in the cat has not been reported. Another trichomonad, Tritrichomonas foetus, is recognized as an intestinal pathogen in cats and often presumptively diagnosed on the basis of the presence of trichomonads in diarrheic feces. It is of importance to determine if cats are natural hosts for P. hominis, as the presence of this organism could result in inaccurate assumption of T. foetus infection. In this study, we used a species-specific PCR assay to identify P. hominis 18S rRNA genes in fecal samples collected from a convenience population of cats in which a high prevalence of T. foetus infection had been previously identified (cat show) or suspected (submitted for T. foetus diagnostic testing). The prevalence of T. foetus infection in these samples was 31% and 28.6%, respectively. P. hominis infection was identified by PCR of DNA extracted from feces of five cats (1.9% and 2.1% of fecal samples, respectively). All cats in which P. hominis was identified were also infected with T. foetus. PCR identification of P. hominis infection in the cat should facilitate future studies to determine the pathogenicity of this species and enable differentiation of P. hominis from other known or as-yet unidentified species of trichomonads that may infect cats.}, number={1-2}, journal={VETERINARY PARASITOLOGY}, author={Gookin, Jody L. and Stauffer, Stephen H. and Levy, Michael G.}, year={2007}, month={Apr}, pages={11–15} }
 @article{gookin_stauffer_coccaro_marcotte_levy_2007, title={Optimization of a species-specific polymerase chain reaction assay for identification of Pentatrichomonas hominis in canine fecal specimens}, volume={68}, ISSN={["1943-5681"]}, DOI={10.2460/ajvr.68.7.783}, abstractNote={To determine the optimum reaction conditions and detection limits of PCR assay for identification of Pentatrichomonas hominis in DNA extracted from canine feces.DNA extracted from feces of 4 dogs with diarrhea from which trichomonads were observed, 81 dogs that had feces submitted to a diagnostic laboratory, and 19 dogs residing in a laboratory animal facility.Optimum reaction conditions and absolute and practical detection limits of 2 P hominis 18S species-specific primer pairs were determined by use of an in vitro cultivated canine isolate of P hominis in the presence and absence of canine feces. The optimized PCR assay was applied to amplification of P hominis 18S rRNA genes from DNA extracted from the feces of dogs.Under optimized conditions, a primer pair was identified as able to detect as few as 1 P hominis organism/180-mg fecal sample. The PCR assay identified P hominis in diarrheic feces of 4 dogs in which trichomonads were seen by light microscopy. The P hominis genes were not amplified from other fecal samples examined.Molecular identification of P hominis in feces of 4 dogs with trichomonosis and diarrhea reported here validates the identity of this species in such infections. Sensitive and specific PCR amplification of P hominis 18S rRNA genes from DNA extracted from feces will directly facilitate studies examining pathogenicity of this trichomonad and enable differentiation of P hominis from other known or novel species of trichomonads that may infect the gastrointestinal tract of dogs.}, number={7}, journal={AMERICAN JOURNAL OF VETERINARY RESEARCH}, author={Gookin, Jody L. and Stauffer, Stephen H. and Coccaro, Maria R. and Marcotte, Miriam J. and Levy, Michael G.}, year={2007}, month={Jul}, pages={783–787} }
 @article{gookin_copple_papich_poore_stauffer_birkenheuer_twedt_levy_2006, title={Efficacy of ronidazole for treatment of feline Tritrichomonas foetus infection}, volume={20}, ISSN={["1939-1676"]}, DOI={10.1892/0891-6640(2006)20[536:EORFTO]2.0.CO;2}, abstractNote={Objectives: To 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.Animals: A cat naturally infected with T foetus infection and diarrhea.Ten specific-pathogen-free (SPF) kittens.Procedure: RDZ, 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.Results: Both RDZ and TDZ killed T foetus at concentrations .0.1 mg/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.Conclusions and Clinical Relevance: Oral 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.}, number={3}, journal={JOURNAL OF VETERINARY INTERNAL MEDICINE}, author={Gookin, Jody L. and Copple, Christina N. and Papich, Mark G. and Poore, Matthew F. and Stauffer, Stephen H. and Birkenheuer, Adam J. and Twedt, David C. and Levy, Michael G.}, year={2006}, pages={536–543} }
 @article{gookin_chiang_allen_armstrong_stauffer_finnegan_murtaugh_2006, title={NF-kappa B-mediated expression of iNOS promotes epithelial defense against infection by Cryptosporidium parvum in neonatal piglets}, volume={290}, ISSN={["1522-1547"]}, DOI={10.1152/ajpgi.00460.2004}, abstractNote={Cryptosporidium sp. parasitizes intestinal epithelium, resulting in enterocyte loss, villous atrophy, and malabsorptive diarrhea. We have shown that mucosal expression of inducible nitric oxide (NO) synthase (iNOS) is increased in infected piglets and that inhibition of iNOS in vitro has no short-term effect on barrier function. NO exerts inhibitory effects on a variety of pathogens; nevertheless, the specific sites of iNOS expression, pathways of iNOS induction, and mechanism of NO action in cryptosporidiosis remain unclear. Using an in vivo model of Cryptosporidium parvum infection, we have examined the location, mechanism of induction, specificity, and consequence of iNOS expression in neonatal piglets. In acute C. parvum infection, iNOS expression predominated in the villous epithelium, was NF-kappaB dependent, and was not restricted to infected enterocytes. Ongoing treatment of infected piglets with a selective iNOS inhibitor resulted in significant increases in villous epithelial parasitism and oocyst excretion but was not detrimental to maintenance of mucosal barrier function. Intensified parasitism could not be attributed to attenuated fluid loss or changes in epithelial proliferation or replacement rate, inasmuch as iNOS inhibition did not alter severity of diarrhea, piglet hydration, Cl- secretion, or kinetics of bromodeoxyuridine-labeled enterocytes. These findings suggest that induction of iNOS represents a nonspecific response of the epithelium that mediates enterocyte defense against C. parvum infection. iNOS did not contribute to the pathogenic sequelae of C. parvum infection.}, number={1}, journal={AMERICAN JOURNAL OF PHYSIOLOGY-GASTROINTESTINAL AND LIVER PHYSIOLOGY}, author={Gookin, JL and Chiang, S and Allen, J and Armstrong, MU and Stauffer, SH and Finnegan, C and Murtaugh, MP}, year={2006}, month={Jan}, pages={G164–G174} }
 @article{zadrozny_stauffer_armstrong_jones_gookin_2006, title={Neutrophils do not mediate the pathophysiological sequelae of Cryptosporidium parvum infection in neonatal piglets}, volume={74}, ISSN={["1098-5522"]}, url={http://europepmc.org/abstract/med/16988224}, DOI={10.1128/IAI.00153-06}, abstractNote={Cryptosporidium parvum is a minimally invasive protozoal pathogen of intestinal epithelium that results in villus atrophy, mucosal lipid peroxidation, diarrhea, and diminished barrier function. Influx of neutrophils is a consistent feature of human and animal cryptosporidiosis, and yet their contribution to the pathological sequelae of infection has not been investigated. Accordingly, we used an established neonatal piglet model of C. parvum infection to examine the role of neutrophils in disease pathogenesis by inhibiting their recruitment and activation in vivo using a monoclonal anti-CD18 antibody. Infected piglets were treated daily with anti-CD18 or isotype control immunoglobulin G and euthanized at peak infection, at which time neutrophil infiltrates, lipid peroxidation, severity of infection, and intestinal barrier function were quantified. C. parvum infection resulted in a significant increase in mucosal neutrophil myeloperoxidase activity that was prevented by treatment of piglets with anti-CD18 antibody. Neutrophil recruitment was dependent on mucosal superoxide formation (prevented by treatment of infected piglets with superoxide dismutase). Neutrophils did not contribute to peroxynitrite formation or peroxidative injury of C. parvum-infected mucosa and had no impact on the severity of epithelial infection, villus atrophy, or diarrhea. The presence of neutrophils in C. parvum-infected mucosa was associated with enhanced barrier function that could not be attributed to mucosal elaboration of prostaglandins or stimulation of their synthesis. These studies are the first to demonstrate that neutrophilic inflammation arising in response to infection by a noninvasive epithelial pathogen results in physiologic rather than pathological effects in vivo.}, number={10}, journal={INFECTION AND IMMUNITY}, author={Zadrozny, Leah M. and Stauffer, Stephen H. and Armstrong, Martha U. and Jones, Samuel L. and Gookin, Jody L.}, year={2006}, month={Oct}, pages={5497–5505} }