Gastric Cryptosporidiosis Research Articles

Evaluating the Effects of Viruses on Eastern Indigo Snakes (Drymarchon couperi) with Gastric Cryptosporidiosis.

A breeding colony of wild-origin eastern indigo snakes (EISs, Drymarchon couperi) that is part of a reintroduction program has been impacted by gastric cryptosporidiosis. Gastric cryptosporidiosis is an insidious disease of squamates caused by an apicomplexan protozoan, Cryptosporidium serpentis. Viral coinfections have been implicated as possible immunosuppressant agents that allow for disease progression and both adenovirus and reovirus have been implicated in allowing for the progression of gastric cryptosporidiosis during coinfection in other snake species. Molecular (PCR) screening for adenoviruses and reoviruses was performed for both C. serpentis-positive and C. serpentis-negative EIS within the breeding colony. No reoviruses were detected in the collection. Adenoviruses were present in 11/68 (16.2%) EISs evaluated, and there was no significant difference between C. serpentis-positive and C. serpentis-negative EISs (p = 0.196). There was no significant difference in adenovirus status between C. serpentis-positive EISs' lifespan (p = 0.191) or survival rates (p = 0.823). These findings suggest that the presence of the adenoviruses found in this study does not contribute to the formation or progression of gastric cryptosporidiosis in EISs.

Comparison of sampling techniques and diagnostic tests for Cryptosporidium serpentis in eastern indigo snakes (Drymarchon couperi).

The purpose of this study was to compare various sampling techniques and commercially available diagnostic tests for Cryptosporidium serpentis. A colony of 80 eastern indigo snakes (Drymarchon couperi) in human care was screened for the presence of C serpentis using endoscopic gastric mucosal biopsies for histologic and molecular analyses. At the time of endoscopic examination and biopsy, a cloacal swab, gastric swab, and gastric lavage sample were also collected. A C serpentis-specific probe hybridization quantitative PCR (qPCR) was performed on each sample. The gastric lavage sample was divided equally for direct microscopy, acid-fast stain, rapid qualitative immunochromatographic assay, direct fluorescent antibody, and 5 different PCR analyses. If a fecal sample was available at the time of endoscopic evaluation, it was also evaluated for Cryptosporidium oocysts by direct microscopy and acid-fast staining. When comparing test results to histologic analyses, the sensitivity of the probe hybridization qPCR of gastric biopsy, gastric lavage, and gastric swab was 100% while the cloacal swab was 72%. When gastric lavage tests were compared, qPCRs outperformed the other tests. Endoscopic biopsy for histologic and qPCR analyses is recommended for disease diagnosis, while gastric lavage or gastric swab samples for qPCR analysis are as sensitive as endoscopic biopsy for screening for the pathogen but cannot diagnose disease. The results from this study allow the veterinary practitioner to select the most appropriate sample and testing methodology when evaluating an ophidian patient for gastric cryptosporidiosis.

Physiological Responses of Eastern Indigo Snakes (Drymarchon couperi) Infected with Cryptosporidium serpentis

Abstract A significant disease of concern in captive populations of snakes is gastric cryptosporidiosis, caused by Cryptosporidium serpentis, a gastrointestinal, protozoal parasite that can cause varying degrees of morbidity and mortality. The purpose of this study was to understand physiological responses of eastern indigo snakes (EIS; Drymarchon couperi) infected with C. serpentis. Body condition index (BCI), heterophil:lymphocyte ratio (HLR), bactericidal ability (BA), hemolysis-hemagglutination assays (SRBC), and plasma corticosterone levels (CORT) were compared between EIS across cryptosporidiosis infection states including cryptosporidia infection positive with clinical signs, infection-positive without clinical signs, infection- recovered, and infection-free snakes. We found snakes that had recovered from C. serpentis had significantly lower SRBC titers than C. serpentis negative snakes (P < 0.05). Recovered snakes had significantly higher BCI than infection positive with clinical signs, infection positive without clinical signs, and infection-free snakes (P = 0.00198). Female EIS had significantly higher CORT levels than males (P = 0.0112), BA had a significant positive relationship with HLR (P = 0.0333), and BA had a significant relationship with SRBC (P = 0.0170). These results give meaningful insight into reptilian physiology of disease and show that snakes recovered from C. serpentis may have remaining negative effects of cryptosporidiosis on their immune system. Results from this study may aid conservation projects in determining suitability for release of EIS that have been infected with C. serpentis.

Open Label, Multi-Arm Randomized Clinical Trial Evaluating Vitamin E-Selenium Injection, Vitamin C Injection, and Hydrogen Peroxide Gavage as a Treatment for Gastric Cryptosporidiosis in Eastern Indigo Snakes (Drymarchon couperi)

Abstract Gastric cryptosporidiosis (GC) is an insidious infection in squamates caused by the protozoan Cryptosporidium serpentis and has impacted the captive breeding colony for the eastern indigo snake (Drymarchon couperi; EIS) reintroduction program. This study investigates a novel treatment of GC in EIS. Seventeen EIS with GC were randomly divided into three groups: A, B, and C. Group A (n = 6) received parenteral administration of 25 mg/kg vitamin C, 0.5 mg/kg vitamin E, and 50 μg/kg selenium and 5 ml/kg 3 % H2O2 gavage; Group B (n = 6) received the same injections but 5 ml/kg water gavage; and Group C received no treatments and served as controls. All EIS from Groups A and B tested negative for C. serpentis for three months following treatment, while only 60% (3/5) in Group C tested negative. EIS testing negative received one 4 mg/kg dexamethasone sodium-phosphate injection. For three months following dexamethasone, 66.7% (4/6) in Group A continued to test negative compared to 83.3% (5/6) in Group B and 20% (1/5) in Group C. EIS testing negative underwent gastric biopsies but only one from Group C was confirmed to be negative for C. serpentis. Although parenteral vitamin C, vitamin E, and selenium with H2O2 gavage decreased shedding of C. serpentis, it did not outperform the vitamins/selenium without H2O2. The parenteral use of 25 mg/kg vitamin C, 0.5 mg/kg vitamin E, and 50 ug/kg selenium once weekly cannot be recommended for treatment of C. serpentis in EIS if complete resolution of the parasite is desired.

Open Label, Multi-Arm Randomized Clinical Evaluation of Clofazimine and Curcumin Treatments for Cryptosporidium serpentis Infection in Eastern Indigo Snakes (Drymarchon couperi)

Squamate gastric cryptosporidiosis (SGC) is a common parasitic disease in captive snakes and is associated with substantial morbidity and mortality. Typical medications used to treat parasitic infections are ineffective against SGC. The purpose of this study was to evaluate new drug regimens for treating SGC in captive eastern indigo snakes (EIS; Drymarchon couperi). Twenty-six EIS naturally infected with Cryptosporidium serpentis were randomly divided into three groups: Group A (n = 9) received 10 mg/kg clofazimine in corn oil twice weekly by oral gavage for 6 weeks, Group B (n = 9) received 5 mg/kg curcumin in corn oil twice weekly by oral gavage for 6 weeks, and Group C (n = 8) received corn oil twice weekly by oral gavage for 6 weeks as a control. Cloacal swabs were collected every 2 months for 6 months to measure C. serpentis shedding by probe hybridization quantitative polymerase chain reaction analysis (qPCR). Snakes that were qPCR negative after 6 months were immunosuppressed with a single dose of 4 mg/kg dexamethasone sodium-phosphate SC. These snakes were then screened by qPCR for an additional 6 months as described above. Prior to immunosuppression, 1/9 (11%) snakes from Group A, 2/9 (22%) snakes from Group B, and 2/8 (25%) snakes from Group C had tested qPCR negative. Following immunosuppression, all snakes were qPCR positive within 6 months. These findings suggest that 10 mg/kg clofazimine and 5 mg/kg curcumin administered orally twice weekly for 6 weeks are ineffective in eliminating C. serpentis in naturally infected D. couperi.

Analytical and Clinical Evaluation of Two Methods for Measuring Erythrocyte Sedimentation Rate in Eastern Indigo Snakes (Drymarchon couperi).

Erythrocyte sedimentation rate (ESR) is a hematological test that can detect inflammatory activity within the body. Although not specific for any particular disease, ESR is often used as a screening "sickness indicator" due to its reliability and low cost. The Westergren method is a manual ESR technique commonly used but requires special graduated pipettes and over 1mL of whole blood, precluding its use in smaller patients where limited sample volumes can be obtained. A modified micro-ESR technique has been described using hematocrit capillary tubes but is used less commonly. ESR has been reported to be a useful inflammatory indicator in gopher tortoises (Gopherus polyphemus) and box turtles (Terrapene spp.) but not in Florida cottonmouth snakes (Agkistrodon conanti). Having an inexpensive screening test for inflammation can help guide medical decisions within conservation efforts of imperiled species. This study evaluated the correlation between these two ESR methodologies in threatened eastern indigo snakes (Drymarchon couperi, EIS) and found a very strong correlation (rs = 0.897), without constant or proportional biases and a reference interval of 0 (90% CI -1-1)-9 mm/h (90% CI 8-11) was defined. Additionally, a significant difference was found between healthy EIS and EIS in mid-ecdysis (p = 0.006) and EIS with gastric cryptosporidiosis (p = 0.006), indicating ESR as a useful inflammatory indicator in EIS.

Gastric Cryptosporidiosis in Snakes, a Review

Cryptosporidium is a genus of unicellular parasites in the phylum Apicomplexa, which includes well-known genera such as Eimeria and Toxoplasma. Cryptosporidium can persist in the environment for prolonged periods as oocysts. Cryptosporidium serpentis can cause severe, life-threatening gastric disease in snakes known as gastric cryptosporidiosis. This review will provide an overview on the existing knowledge of the biology of the parasite, explain the current treatment and management options for gastric cryptosporidiosis, and describe a synopsis of current and next-generation approaches that could spearhead further research in combating C. serpentis in snakes.

An Alternative Technique for Gastric Sampling in Snakes

Abstract Collecting gastric samples is often necessary when evaluating a snake exhibiting clinical signs of gastric cryptosporidiosis. Traditional techniques for obtaining these samples include gas...

Limitations in the screening of potentially anti-cryptosporidial agents using laboratory rodents with gastric cryptosporidiosis.

The emergence of cryptosporidiosis, a zoonotic disease of the gastrointestinal and respiratory tract caused by Cryptosporidium Tyzzer, 1907, triggered numerous screening studies of various compounds for potential anti-cryptosporidial activity, the majority of which proved ineffective. Extracts of Indonesian plants, Piper betle and Diospyros sumatrana, were tested for potential anti-cryptosporidial activity using Mastomys coucha (Smith), experimentally inoculated with Cryptosporidium proliferans Kváč, Havrdová, Hlásková, Daňková, Kanděra, Ježková, Vítovec, Sak, Ortega, Xiao, Modrý, Chelladurai, Prantlová et McEvoy, 2016. None of the plant extracts tested showed significant activity against cryptosporidia; however, the results indicate that the following issues should be addressed in similar experimental studies. The monitoring of oocyst shedding during the entire experimental trial, supplemented with histological examination of affected gastric tissue at the time of treatment termination, revealed that similar studies are generally unreliable if evaluations of drug efficacy are based exclusively on oocyst shedding. Moreover, the reduction of oocyst shedding did not guarantee the eradication of cryptosporidia in treated individuals. For treatment trials performed on experimentally inoculated laboratory rodents, only animals in the advanced phase of cryptosporidiosis should be used for the correct interpretation of pathological alterations observed in affected tissue. All the solvents used (methanol, methanol-tetrahydrofuran and dimethylsulfoxid) were shown to be suitable for these studies, i.e. they did not exhibit negative effects on the subjects. The halofuginone lactate, routinely administered in intestinal cryptosporidiosis in calves, was shown to be ineffective against gastric cryptosporidiosis in mice caused by C. proliferans. In contrast, the control application of extract Arabidopsis thaliana, from which we had expected a neutral effect, turned out to have some positive impact on affected gastric tissue.

An unusual case of miliary PML-IRIS in an HIV+ patient.

A 45-year-old female patient recently diagnosed with HIV infection (CD4+ cell count 64 cells/mm3 and plasma HIV viral load 1.4 × 106 copies/mL at diagnosis) started antiretroviral therapy (ART). At the time, she presented with gastric cryptosporidiosis and an oral candidiasis, which subsided rapidly on treatment. One month after ART initiation, the patient noticed progressive generalized slowness, speech difficulty, and a slight weakness of her right arm and leg. She also reported headache, generalized fatigue, night sweats, and weight loss. She was admitted to the hospital 4 weeks after symptoms onset. The neurologic examination revealed psychomotor slowing, mild memory and executive dysfunction, meningeal signs, dysarthria, mild lower-limb ataxia, and right faciobrachial weakness (Medical Research Council grade 4). Generalized adenopathy and wasting syndrome were also identified. Brain MRI showed a pattern of multiple miliary-enhancing parenchymal nodules (1–4 mm in diameter) more marked in the brain stem and basal ganglia (figure, A–D).

Diagnosis of gastric cryptosporidiosis in birds using a duplex real-time PCR assay

Three species and several genotypes of Cryptosporidium can infect the epithelial surface of the bursa of Fabricius, the respiratory tract, the proventriculus, the intestine, and the urinary tract in birds. There is reason to believe that gastric cryptosporidiosis in birds is caused by Cryptosporidium galli and Cryptosporidium avian genotype III, resulting in a chronic illness of the proventriculus that can lead to a debilitating and fatal clinical condition in birds of the orders Passeriformes and Psittaciformes. The objectives of the present study were to develop a duplex real-time polymerase chain reaction (PCR) that targets the 18S rRNA gene to simultaneously detect C. galli and Cryptosporidium avian genotype III DNA and to compare the duplex real-time PCR results to those of nested PCR targeting a partial fragment of the 18S rRNA gene, followed by sequencing of the amplified products (nPCR/S). A total of 1027 fecal samples were collected from birds of the orders Psittaciformes and Passeriformes originating either from captivity or the wild. Duplex real-time PCR results were positive in 580 (56.47%) and 21 (2.04%) samples, respectively, for C. galli and Cryptosporidium avian genotype III, whereas nPCR/S was positive in 28 (2.73%) and three (0.29%) samples, respectively, for C. galli and Cryptosporidium avian genotype III. Novel host birds were identified for both of the above gastric species, and it was also possible to identify Cryptosporidium baileyi and, for the first time in Brazil, Cryptosporidium avian genotype V. The duplex real-time PCR assay developed in the present study represents a sensitive and specific method for the detection of C. galli and Cryptosporidium avian genotype III in bird fecal samples. Moreover, this method may serve as an alternative to nPCR/S as a gold standard for the diagnosis of gastric cryptosporidiosis in birds.

AIDS-related gastric cryptosporidiosis simulating malignancy

Cryptosporidium localization to the stomach is uncommon. We report a case of a 56-year-old, HIV-positive, male patient with severe immunosuppression after antiretroviral treatment dropout who presented with epigastric discomfort, nausea and vomiting. The abdominal CT with contrast showed an important concentric thickening of the gastric antrum wall. Upper gastrointestinal endoscopic evaluation revealed lack of distensibility of the gastric wall and marked thickening, stiffness, distortion, and erosions of mucosal folds involving the antrum region. The endoscopy was clinically reported as gastric malignancy suggestive of lymphoma. The histopathologic diagnosis was cryptosporidiosis. To our knowledge, gastric cryptosporidiosis simulating malignancy has not been previously described. This type of lesion should be included in the differential diagnosis of AIDS patients with lesions simulating gastric malignancy. La localización del Cryptosporidium en el estómago es infrecuente. Se presenta el caso de un paciente de 56 años de edad, VIH-positivo, con inmunosupresión severa, después de abandonar el tratamiento anti-retroviral, que presentó malestar epigástrico, náuseas y vómitos. La TC abdominal con contraste mostró un engrosamiento concéntrico importante del antro gástrico. La evaluación endoscópica gastrointestinal reveló falta de distensibilidad de la pared gástrica y marcado engrosamiento, junto con rigidez, distorsión y erosión de los pliegues de la mucosa del antro. El estudio endoscópico fue informado como proceso maligno sugestivo de linfoma. El diagnóstico histopatológico fue de criptosporidiosis. De acuerdo con nuestro conocimiento, la criptosporidiosis gástrica simulando un tumor maligno no ha sido descrita. Este tipo de lesión se debe incluir en el diagnóstico diferencial de pacientes VIH+ con lesiones que sugieran malignidad gástrica.

Activated CD8+ T cells contribute to clearance of gastric Cryptosporidium muris infections

The role of CD4+ and CD8+ T lymphocytes in the development of a protective immune response against Cryptosporidium muris infection was studied by the reconstitution of severe combined immunodeficient (SCID) mice with well-defined populations of either naive or immune CD8+ or CD4+ T lymphocytes. Adoptive transfer of both naive and immune CD4+ T lymphocyte subpopulations protects SCID mice against cryptosporidiosis. Moreover, a significant biological impact of activated CD8+ T cells against gastric cryptosporidiosis was observed. The significant difference in the course and intensity of the infection in reconstituted SCID mice was found to be dependent on the protective function of both the CD4+ and CD8+ T-cell populations transferred. While SCID mice reconstituted with either immune or naive CD4+ or immune CD8+ T-cell subpopulations resolved the infection within 29, 37 and 51 days post-infection, respectively, those reconstituted with naive CD8+ T cells suffered from chronic infection similar to control SCID mice. Reconstitution with CD4+ T cells resulted in suppression of oocyst excretion and shortening of patent period in comparison with SCID mice reconstituted with CD8+ T cells. Thus, although CD4+ T cells are considered important in protective immunity, our results are the first to demonstrate the involvement of activated CD8+ T lymphocytes in the protection of mice against gastric cryptosporidiosis.

DIAGNOSTIC EXERCISE

An approximately one-and-a-half-year-old, neutered male, mixed-breed dog was presented for a chronic history of vomiting. Profuse diarrhea was also noted during examination. An exploratory laparotomy was performed, bone chips were removed from the stomach, and a raised, circular area of gastric mucosa was biopsied. Histologically, there was severe gastric cryptosporidiosis as well as numerous spiral bacteria, consistent with Helicobacter spp. Polymerase chain reaction revealed visible bands for the 18S ribosomal RNA gene for Cryptosporidium spp. The polymerase chain reaction product was sequenced and was found to be most similar to Cryptosporidium muris. Both the gastric location and the species of Cryptosporidium are unusual in a dog.

Activation of protective cell-mediated immune response in gastric mucosa during Cryptosporidium muris infection and re-infection in immunocompetent mice

Gastric cryptosporidia only inhabit the glandular part of the stomach of all age categories of their hosts and can cause chronic life-long infections independent of a host's immune status. The immune response in the stomach mucosa during the primary infection and re-infection with Cryptosporidium muris (TS03 and CB03) in immunocompetent BALB/c mice was characterized using flow cytometry analysis and measurement of IFN-gamma and IL10 by enzyme-linked immunosorbent assays (ELISA). Significantly, elevated migration of T lymphocytes (more than 1,000-fold), especially CD8+ T lymphocytes, to the stomach mucosa occurred during primary infection and persisted for more than 2 months after its resolution. The ex vivo cultures of splenocytes revealed very low levels of IFN-gamma production during the course of the primary infection (0.5 ng/ml), whereas in the following re-exposure to the parasites, the concentration of IFN-gamma rapidly increased 22-fold. Although the two parasite strains that were tested were genetically distinct, they yielded similar results in the induction of cellular immune responses, suggesting that these patterns are not unique to a single parasite strain. These results imply that the CD8+ T lymphocytes are involved in the immune response to gastric cryptosporidiosis and could play an important role in the elimination of C. muris infection in mice.

Cryptosporidiosis caused by two distinct species in Russian tortoises and a pancake tortoise

Cryptosporidiosis in squamates is well documented, but there is very limited information available on cryptosporidiosis in testudines. We describe three cases of cryptosporidiosis in tortoises with associated pathology. Two Russian tortoises ( Agrionemys [ Testudo] horsfieldii) and a pancake tortoise ( Malacochersus tornieri), all from separate collections, were found dead. At necropsy, two had histological evidence of intestinal cryptosporidiosis and one had gastric cryptosporidiosis. Consensus Cryptosporidium sp. PCR and sequencing was used to identify the Cryptosporidium sp. present in these three tortoises. In the juvenile Russian tortoise with gastric cryptosporidiosis, the organism had 98% homology with a previously reported sequence from an Indian star tortoise isolate. A second chelonian Cryptosporidium sp. was identified in the pancake tortoise and the second Russian tortoise. This sequence was 100% identical to a shorter gene sequence previously reported in a marginated tortoise. This is the first report coordinating pathology with Cryptosporidium characterization in chelonians. The two Cryptosporidium sp. found in tortoises segregate according to site of infection, and there may be further differences in pathology, host range, and transmission. These Cryptosporidium sp. appear to be able to infect diverse tortoise host species. This may be an under-recognized problem in tortoises.

Isolated Gastric Cryptosporidiosis in an Immunocompetent Patient

Isolated Gastric Cryptosporidiosis in an Immunocompetent Patient

A Case Presentation of Gastric Cryptosporidiosis in an HIV Patient

A 24 yo female, with a history of sexually acquired HIV on no medication, presented complaining of epigastric abdominal pain associated with nausea and non-bloody vomiting for 7 days. She reported watery, non-bloody diarrhea for 3 months with a 10 pound weight loss, but denied any fever, chills, or change in appetite. On exam, she was afebrile with generalized wasting. Laboratory studies revealed 12.9 mg/dl Hgb, with the base line Hgb of 12–13 mg/dl, WBC = 1.7 with 61.6% neutrophils, 18.8% lymphocytes and a low CD4 count (< 20). Stool work up was negative for ova and parasites x 3, including Cryptosporidium parvum, wright stain and stool cultures were negative for WBCs and bacteria. Upper endoscopy and colonoscopy were performed. The antral mucosa was markedly erythematous, edematous and the mucosal surface was markedly friable. Thickened gastric folds were seen. Biopsies were taken of these areas. The colonic mucosa appeared normal. Histopathologic examination of antral and duodenal mucosal biopsies showed chronic active gastritis with extensive cryptosporidiosis and H. pylori organisms. Biopsies from the colonic mucosa were positive for Cryptosporidium parvum as well. In view of these findings, therapy with Alinia 500 mg twice daily was started. After 2 weeks, clinical improvement was observed with reduction of both epigastric pain and diarrhea. Repeat endoscopy 8 weeks later showed normal gastric mucosa. No organisms were identified on repeat antral and duodenal biopsies. Cryptosporidium parvum is a common protozoal infection that has been recognized as a human pathogen since 1976. It causes self-limited diarrhea in immunocompetent hosts, but severe and prolonged diarrhea in immunocompromised individuals, particularly those with HIV infection. Cryptosporidium parvum has been detected with increasing frequency in the gastrointestinal tract but involvement of the stomach is rarely reported. This case report presents the clinical, pathologic and endoscopic features of Cryptosporidium associated erosive gastritis and highlights the importance of upper endoscopy and biopsy in diagnosis of this entity. [figure 1]Figure 1

Gastrointestinal cryptosporidiosis in a puppy

An 8-week-old female Yorkshire terrier with a history of weakness, diarrhea and intestinal isosporiasis was euthanized and a post mortem examination performed. Histologically, there was severe gastrointestinal cryptosporidiosis, severe intestinal isosporiasis and thymic lymphoid depletion. PCR revealed visible bands for the actin and 18S rRNA genes but not for the acetyl CoA synthetase gene for Cryptosporidium spp. The PCR product for the actin gene was sequenced and found to have a 97.6–99.8% similarity to that of Cryptosporidium canis. To our knowledge, this is the first report of gastric cryptosporidiosis in a canine.

Eradication of AIDS-related gastric cryptosporidiosis with azithromycin

Human cryptosporidiosis is a protozoan infection that can produce severe debilitating disease in immunocompromised patients, especially those with AIDS. The efficiency of different treatments is limited and there is no uniform therapy. We report a case of eradication of AIDS-related cryptosporidiosis with azithromycin and discuss new treatment regimens reported in the literature.